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Parenterals formulation development is given in this presentation
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Parenterals
Formulation development
Pragati Kumar BAsst ProffNimra College of PharmacyVijayawada
Introduction and importance of the studybull Parenteral products are products that are administered to
the body by injection bull Because this route of administration bypasses the normal
body defense mechanisms it is essential that these products are prepared with a high degree of care and skills than utilized in preparing conventional oral or topical products
bull The finished product must be sterile non-pyrogenic and free from extraneous insoluble materials These products must satisfy a number of requirements for parenteral products
Sterile formulations must meet a number of special criteria such as
bull Sterilitybull Particulate materialbull Pyrogen freebull Stabilitybull pHbull Osmotic pressure
During the formulation of parenteral products the following factors are critical
bull The vehicle in which the drug is dissolved or dispersed
bull Volume (dose) of the injectionbull Adjustment of isotonicitybull Adjustment of pHbull Stabilisersbull Preservativesbull Adjustment of specific gravity (for spinal anaesthesia)bull Concentration units
Difference between parenterals and other products
bull Limits to the level of pyrogens present and of particulate matter
bull The injection route dictates the volume of formulation Hence the solubility of the drug in the selected vehicle is critical in the formulation
Vehicle bull The preferred vehicle is water as it is well tolerated
by the body easy to administer and a large solvent capacity
bull Water for injection must be sterile and free from pyrogens
bull Cellulose glass rubber cores cloth or cotton fibres may constitute the contaminants list
bull Suitable filtration media for removal of particulate material are sintered glass filters or membrane filters with a pore size of 045-12 microns
Ph and Buffersbull As parenteral products are administered directly to tissues
and systemic circulation formulations prepared should not vary significantly from physiological pH which is about 74
bull The acceptable pH range is 3-105 for iv preparations and 4-9 for other routes
bull Buffers are included in injections to maintain the pH of the packaged product
bull The buffers used in the injection must allow the body fluids to change the product pH after injection
bull Acetate citrate and phosphate buffers are commonly used in parenteral products
Osmotic pressurebull The osmotic pressure of the blood is approx
300 milli OsmolesL and ideally any sterile solution would be formulated to have the same osmolarity
bull For eg 09 wv NaCl iv solution has an osmolarity of 308 milli OsmolesL
bull 5 wv Dextrose iv solution has an osmolarity of 280 milli OsmolesL
bull NaCl Mannitol or glucose can be used to adjust osmolarity
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Introduction and importance of the studybull Parenteral products are products that are administered to
the body by injection bull Because this route of administration bypasses the normal
body defense mechanisms it is essential that these products are prepared with a high degree of care and skills than utilized in preparing conventional oral or topical products
bull The finished product must be sterile non-pyrogenic and free from extraneous insoluble materials These products must satisfy a number of requirements for parenteral products
Sterile formulations must meet a number of special criteria such as
bull Sterilitybull Particulate materialbull Pyrogen freebull Stabilitybull pHbull Osmotic pressure
During the formulation of parenteral products the following factors are critical
bull The vehicle in which the drug is dissolved or dispersed
bull Volume (dose) of the injectionbull Adjustment of isotonicitybull Adjustment of pHbull Stabilisersbull Preservativesbull Adjustment of specific gravity (for spinal anaesthesia)bull Concentration units
Difference between parenterals and other products
bull Limits to the level of pyrogens present and of particulate matter
bull The injection route dictates the volume of formulation Hence the solubility of the drug in the selected vehicle is critical in the formulation
Vehicle bull The preferred vehicle is water as it is well tolerated
by the body easy to administer and a large solvent capacity
bull Water for injection must be sterile and free from pyrogens
bull Cellulose glass rubber cores cloth or cotton fibres may constitute the contaminants list
bull Suitable filtration media for removal of particulate material are sintered glass filters or membrane filters with a pore size of 045-12 microns
Ph and Buffersbull As parenteral products are administered directly to tissues
and systemic circulation formulations prepared should not vary significantly from physiological pH which is about 74
bull The acceptable pH range is 3-105 for iv preparations and 4-9 for other routes
bull Buffers are included in injections to maintain the pH of the packaged product
bull The buffers used in the injection must allow the body fluids to change the product pH after injection
bull Acetate citrate and phosphate buffers are commonly used in parenteral products
Osmotic pressurebull The osmotic pressure of the blood is approx
300 milli OsmolesL and ideally any sterile solution would be formulated to have the same osmolarity
bull For eg 09 wv NaCl iv solution has an osmolarity of 308 milli OsmolesL
bull 5 wv Dextrose iv solution has an osmolarity of 280 milli OsmolesL
bull NaCl Mannitol or glucose can be used to adjust osmolarity
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Sterile formulations must meet a number of special criteria such as
bull Sterilitybull Particulate materialbull Pyrogen freebull Stabilitybull pHbull Osmotic pressure
During the formulation of parenteral products the following factors are critical
bull The vehicle in which the drug is dissolved or dispersed
bull Volume (dose) of the injectionbull Adjustment of isotonicitybull Adjustment of pHbull Stabilisersbull Preservativesbull Adjustment of specific gravity (for spinal anaesthesia)bull Concentration units
Difference between parenterals and other products
bull Limits to the level of pyrogens present and of particulate matter
bull The injection route dictates the volume of formulation Hence the solubility of the drug in the selected vehicle is critical in the formulation
Vehicle bull The preferred vehicle is water as it is well tolerated
by the body easy to administer and a large solvent capacity
bull Water for injection must be sterile and free from pyrogens
bull Cellulose glass rubber cores cloth or cotton fibres may constitute the contaminants list
bull Suitable filtration media for removal of particulate material are sintered glass filters or membrane filters with a pore size of 045-12 microns
Ph and Buffersbull As parenteral products are administered directly to tissues
and systemic circulation formulations prepared should not vary significantly from physiological pH which is about 74
bull The acceptable pH range is 3-105 for iv preparations and 4-9 for other routes
bull Buffers are included in injections to maintain the pH of the packaged product
bull The buffers used in the injection must allow the body fluids to change the product pH after injection
bull Acetate citrate and phosphate buffers are commonly used in parenteral products
Osmotic pressurebull The osmotic pressure of the blood is approx
300 milli OsmolesL and ideally any sterile solution would be formulated to have the same osmolarity
bull For eg 09 wv NaCl iv solution has an osmolarity of 308 milli OsmolesL
bull 5 wv Dextrose iv solution has an osmolarity of 280 milli OsmolesL
bull NaCl Mannitol or glucose can be used to adjust osmolarity
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
During the formulation of parenteral products the following factors are critical
bull The vehicle in which the drug is dissolved or dispersed
bull Volume (dose) of the injectionbull Adjustment of isotonicitybull Adjustment of pHbull Stabilisersbull Preservativesbull Adjustment of specific gravity (for spinal anaesthesia)bull Concentration units
Difference between parenterals and other products
bull Limits to the level of pyrogens present and of particulate matter
bull The injection route dictates the volume of formulation Hence the solubility of the drug in the selected vehicle is critical in the formulation
Vehicle bull The preferred vehicle is water as it is well tolerated
by the body easy to administer and a large solvent capacity
bull Water for injection must be sterile and free from pyrogens
bull Cellulose glass rubber cores cloth or cotton fibres may constitute the contaminants list
bull Suitable filtration media for removal of particulate material are sintered glass filters or membrane filters with a pore size of 045-12 microns
Ph and Buffersbull As parenteral products are administered directly to tissues
and systemic circulation formulations prepared should not vary significantly from physiological pH which is about 74
bull The acceptable pH range is 3-105 for iv preparations and 4-9 for other routes
bull Buffers are included in injections to maintain the pH of the packaged product
bull The buffers used in the injection must allow the body fluids to change the product pH after injection
bull Acetate citrate and phosphate buffers are commonly used in parenteral products
Osmotic pressurebull The osmotic pressure of the blood is approx
300 milli OsmolesL and ideally any sterile solution would be formulated to have the same osmolarity
bull For eg 09 wv NaCl iv solution has an osmolarity of 308 milli OsmolesL
bull 5 wv Dextrose iv solution has an osmolarity of 280 milli OsmolesL
bull NaCl Mannitol or glucose can be used to adjust osmolarity
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Difference between parenterals and other products
bull Limits to the level of pyrogens present and of particulate matter
bull The injection route dictates the volume of formulation Hence the solubility of the drug in the selected vehicle is critical in the formulation
Vehicle bull The preferred vehicle is water as it is well tolerated
by the body easy to administer and a large solvent capacity
bull Water for injection must be sterile and free from pyrogens
bull Cellulose glass rubber cores cloth or cotton fibres may constitute the contaminants list
bull Suitable filtration media for removal of particulate material are sintered glass filters or membrane filters with a pore size of 045-12 microns
Ph and Buffersbull As parenteral products are administered directly to tissues
and systemic circulation formulations prepared should not vary significantly from physiological pH which is about 74
bull The acceptable pH range is 3-105 for iv preparations and 4-9 for other routes
bull Buffers are included in injections to maintain the pH of the packaged product
bull The buffers used in the injection must allow the body fluids to change the product pH after injection
bull Acetate citrate and phosphate buffers are commonly used in parenteral products
Osmotic pressurebull The osmotic pressure of the blood is approx
300 milli OsmolesL and ideally any sterile solution would be formulated to have the same osmolarity
bull For eg 09 wv NaCl iv solution has an osmolarity of 308 milli OsmolesL
bull 5 wv Dextrose iv solution has an osmolarity of 280 milli OsmolesL
bull NaCl Mannitol or glucose can be used to adjust osmolarity
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Vehicle bull The preferred vehicle is water as it is well tolerated
by the body easy to administer and a large solvent capacity
bull Water for injection must be sterile and free from pyrogens
bull Cellulose glass rubber cores cloth or cotton fibres may constitute the contaminants list
bull Suitable filtration media for removal of particulate material are sintered glass filters or membrane filters with a pore size of 045-12 microns
Ph and Buffersbull As parenteral products are administered directly to tissues
and systemic circulation formulations prepared should not vary significantly from physiological pH which is about 74
bull The acceptable pH range is 3-105 for iv preparations and 4-9 for other routes
bull Buffers are included in injections to maintain the pH of the packaged product
bull The buffers used in the injection must allow the body fluids to change the product pH after injection
bull Acetate citrate and phosphate buffers are commonly used in parenteral products
Osmotic pressurebull The osmotic pressure of the blood is approx
300 milli OsmolesL and ideally any sterile solution would be formulated to have the same osmolarity
bull For eg 09 wv NaCl iv solution has an osmolarity of 308 milli OsmolesL
bull 5 wv Dextrose iv solution has an osmolarity of 280 milli OsmolesL
bull NaCl Mannitol or glucose can be used to adjust osmolarity
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Ph and Buffersbull As parenteral products are administered directly to tissues
and systemic circulation formulations prepared should not vary significantly from physiological pH which is about 74
bull The acceptable pH range is 3-105 for iv preparations and 4-9 for other routes
bull Buffers are included in injections to maintain the pH of the packaged product
bull The buffers used in the injection must allow the body fluids to change the product pH after injection
bull Acetate citrate and phosphate buffers are commonly used in parenteral products
Osmotic pressurebull The osmotic pressure of the blood is approx
300 milli OsmolesL and ideally any sterile solution would be formulated to have the same osmolarity
bull For eg 09 wv NaCl iv solution has an osmolarity of 308 milli OsmolesL
bull 5 wv Dextrose iv solution has an osmolarity of 280 milli OsmolesL
bull NaCl Mannitol or glucose can be used to adjust osmolarity
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Osmotic pressurebull The osmotic pressure of the blood is approx
300 milli OsmolesL and ideally any sterile solution would be formulated to have the same osmolarity
bull For eg 09 wv NaCl iv solution has an osmolarity of 308 milli OsmolesL
bull 5 wv Dextrose iv solution has an osmolarity of 280 milli OsmolesL
bull NaCl Mannitol or glucose can be used to adjust osmolarity
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Antimicrobial agentsbull Aqueous preparations which are prepared
using aseptic preparations and which cannot be terminally sterilized may contain a suitable antimicrobial preservative in an appropriate concentration
bull Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use
bull Antimicrobial agents must be effective in the parenteral formulation
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Antimicrobial preservativesSNO ANTIMICROBIAL AGENT CONCENTRATION (wv)
1 Benzalkonium chloride 0012 Benzyl alcohol 1- 23 Chlorobutol 025 ndash 054 Phenol 055 Chlorocresol 01 ndash 036 Phenyl mercuric salts 00027 Methyl hydroxy benzoate 01- 02
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Atioxidants
bull Aqueous solutions are more susceptible to oxidation
bull Bisulphites and metabisulphites are commonly used antioxidants in aqueous injections
bull Injection formulations may in addition also contain chelating agents such as EDTA or citric acid to remove trace elements which catalyse oxidative degradation
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Sterilization bull Sterility Absence of life or absolute
freedom from biological contaminationbull Sterilization inactivation or elimination of all
viable organism and their sporesbull Disinfectant substance used on non- living
objects to render them non- infectious kills vegetative bacteria fungi virus but not spores Eg Formaldehyde
bull Bactericide ( Germicide) substance that kills vegetative bacteria and spores
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Advantages and disadvantages
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Moist Heat
bull Spore are killed by moist heatbull Culture media amp other liquids
required to retain their content of water
bull Not applicable to waterproof materials such as oils and greases or dry materials
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Pasteurization
bull Moist heat at temperature below 1000 Cbull Heat labile fluids may be disinfected not
sterilized by heating at 560 C for 30 minbull Sufficient to kill mesophilic bacteria but
not sporesbull For serum or other body fluids containing
proteins temp to rise above 59o Cbull UHT 140o C less than 1 sec
bull Cold Pasteurizationbull High pressure pasteurization
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Washer Disinfectorsbull Washing machines using hot
water steam and detergents may be used
bull Washing action at 710 C for 3 min or 800 C for 1 minute sufficient to kill vegetative organism
bull Recommended for instruments contaminated with HBV and HIV
bull Accordingly it is set at 93o C for 10 min
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Boiling at 100o C
bull Heating in boiling water at 100o C for 5 minutes sufficient to kill all vegetative bacteria HBV and some bacterial spore
bull Used only in case of emergencies to sterilize medical and surgical equipment
bull Heat labile articles and hollow or porous items where water will not penetrate lumen cannot be disinfected this way
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Steaming at 100o C
bull To prevent glass from cracking which happens when it is heated directly
bull Also in case of heat labile culture media
bull Pure steam in equilibrium with boiling water at normal atmospheric pressure
bull Exposure to this temperature for 5 minutes will kill microorganism
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Free Steaming
bull Koch and Arnold steamers are used
bull Useful for selective heat labile media like DCA XLD TCBS and Slenite F broth
bull These media do not support the growth of heat resistant bacteria
bull Tyndallization- 20 min for 3 succesive days
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Steam sterilization
bull Irreversibly coagulates and denatures microbial enzymes and proteins
bull Important parameters for effectiveness
1 Exposure time2 Temperature of process3 Level of moisture4 Pressure
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
1) Advantagesbull 1048708 Non-toxicbull 1048708 Cycle easy to control and monitorbull 1048708 Inexpensivebull 1048708 Rapidly microbiocidalbull 1048708 Least affected by organicinorganic soilsbull 1048708 Rapid cycle timebull 1048708 Penetrates medical packing device lumens2) Disadvantagesbull 1048708 Deleterious for heat labile instrumentsbull 1048708 Potential for burns
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Autoclaves
bull Invented by Charles Chamberland in 1879
bull Precursor was the Steam digester invented by Denis Papin in 1679
bull At correct temp lethal to all bacteria viruses fungi amp protozoa
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
bull Example of usage of autoclaves arebull Hospitals amp OPD- Porous load
autoclavesbull Mortuary- Bench top autoclavesbull Microbiology Lab Media preparators
or fluid cycle steam sterilizersbull Pharmaceutical- Fluid Cycle Sterilizer
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Types of autoclavesbull Simple laboratory autoclavebull Transportable bench top autoclavesbull Large simple autoclavebull Downward displacement autoclavebull Multipurpose laboratory autoclavebull Pre vacuumbull High security autoclavebull Porous load sterilizerbull Low temperature steambull Steam flash pressure pulsing steam sterilization
autoclaves
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
bull Sterilization Hold Timebull Heat Penetration Timebull Condensation of steam 3 effects1 Wets microorganisms2 Liberates latent heat3 Significant contraction of steam
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
bull Time and temperature with pressure required for sterilization by steam under pressure in autoclave
bull 121-124o c at 11 bar for 15 minbull 134-138o c at 22 bar for 3 min1 bar=1 atm pressure=147
pounds per square inch
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
bull Steam which is present must be 1 Saturated 2 Dry3 Pure Steam supply1 Superheated steam ( Dry gas)2 Wet steam
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
bull Removal of air1 Simple transportable (pressure-cooker)
autoclaves2 Downward displacement autoclaves3 Porous load autoclavesbull Items to be put in autoclave1 Unwrapped non porous items2 Porous loads3 Discard loads4 Fluids in sealed containers5 Nutrient media
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Simple laboratory autoclave
1 Considered unsatisfactory2 Not monitor temperature of load
and therefore cannot prevent spoiling nutritive value
3 No safety interlock
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Transportable bench top autoclaves
bull Sophisticated version of air displacement autoclave
bull Possess automatic cycle control with indicator for cycle failure and thermal safety
bull Disadvantages of this are1 Not fitted with vacuum assisted air
removal2 No assisted drying3 Cannot be used for porous loads
packaged items or discard loads4 Cant handle liquid loads
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Large simple autoclave
bull Larger version of simple pressure cooker
bull No means of assisted air removalbull Size too large to permit removal of
airbull Unsuitable for wrapped articles and
make safe loads
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Downward displacement laboratory autoclaves
bull Provision for removal of air from chamber balanced pressure steam trap
bull Other devices assist the drying of the loadbull No vacuum assistance for air removalbull Air removal was found to be inadequatebull Only value is sterilization of unwrapped
non-porous metallic itemsbull Cooling may take many hours to be below
80o C
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Multipurpose laboratory autoclaves
bull Different types of load require different cycles required for these different loads
Special featurebull Presence of efficient means of assisted air
removal and drying assisted cooling and a temperature sensitive probe reading directly from the load
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Multipurpose laboratory autoclaves ( Contd)
bull Aqueous media1 Accelerated cooling required to avoid damage
to nutrient properties of culture medium2 Cooling to below 80o C before autoclave is
opened minimize risk of breakage and explosions
3 Volume of DIN bottle which is used should be not over 80
4 Duration of heating up period should be controlled automatically with a thermocouple placed in one of the largest bottle or simulators which reproduces thermal characteristic of bottle
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Multipurpose laboratory autoclaves ( Contd)
bull 121o c for 15 min Cooling of load should be assisted so that media are cooled to under 80o c in 30 min
bull Methods of coolingbull Cooling time duration set by a
thermocouple placed in simulator bull Air blastingbull Glasswares are satisfactorily
processed as part of a fluid cycle
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
1 High security autoclave
2 Porous load sterilizer
3 Low temperature steam
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Gas sterilization
bull For delicate heat labile equipmentsbull Monitoring their efficacy by biological test
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
1 Low temperature steam formaldehyde
bull Steam at sub atmospheric pressure (temp below 100o C) kills the spores of thermophilic bacteria very slowly
bull Sporicidal at high concentration in presence of moisture
bull Synergism between formaldehyde and steambull Difficulty in combining the two
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
2 Ethylene oxide sterilizerbull Useful also for small
proportion of medical and surgical devices which cannot withstand autoclaving
bull Kills micro organisms by altering their DNA by alkylation
bull only materials with documented ethylene oxide penetration and dissipation properties should be used as wrappers
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Low temperature sterilization by ozone
bull The 125 l ozone sterilizer uses medical grade oxygen water and electricity to generate ozone within the sterilizer to provide efficient sterilants without producing toxic chemicals or using high temperatue
bull End of cycle O2 and H2O are formed
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Liquid Sterilization
bull To sterilize immersible devices like endoscopes etc with 35 liquid Peracetic acid
bull Done using STERIS system 1bull Acid is diluted with sterile filtered
waterbull Commercially available spores can
be used for monitoring sterilizationbull Disadvantage is high cost
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Sterilizing Filter
bull Aqueous liquids sterilized by forced passage through filter of porosity small enough to retain any microorganism
1) Membrane filtersbull Manufactured from variety of
polymeric material such as cellulose diacetate polycarbonate and polyester as disc
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Membrane Filter (Contd)bull Membrane made in 2
ways1 Capillary pore
membranes For viruses2 Labyrinthine pore
membranes Bacteria amp yeast
bull Exact procedure for use varies with form in which filter is supplied
bull Filters may be supplied with plastic holders pre sterilized for single use or mounted in re-usable holders and fitted to filtration vessels
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Sterilizing Filter2)Syringe filters bull Membrane 13-25mm diameterbull Fitted in syringe like holders
of stainless steel or polycarbonate
bull Used for sterilization of small volumes of fluid
3)Vacuum and in- line filtersbull Membranes of 25-45 mm
diameter are used either with in line filter holders of Teflon or stainless steel and aluminum
bull Used for sterilization of large volumes of air and liquid
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
4) Pressure filtrationbull Large membranes 100-
293 mm in diameter housed in pressure filter holders
bull Production of pure water
5) Air Filtersbull Large volume of air
rapidly freed from infection by passage through HEPA (High efficiency particle arrester)
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
Air Filters (Contd)bull Principal use is to render
safe the air withdrawn from an exhaust ventilated safety cabinets used for work with pathogens
bull To decontaminate air input into laminar flow cabinet
bull Fitted with disposable pre filter which reduces load collected by main filter
bull Main HEPA should have 9999 efficiency
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
STERILIZATION BY RADIATIONbull IONIZING radiation - γ
radiation from radioactive elements usually Co60
Eg Sterilization of Disposable Syringes
Bacillus pumilis used for testing
bull ULTRAVIOLET RAYS Mercury vapor lamps emitting radiation in the range of 250-260nm are bactericidal amp to a lesser extent sporicidal
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
References
bull Mackie amp McCartney Practical Medical Microbiology 14 th edition
bull Murray Scot et al Text book of medical microbiology
bull Bailey amp Scot Diagnostic Medical Microbiology Twelfth Edition
bull Ananthnarayan and Paniker Textbook of microbiology 8 th edition
