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Antiseptics and disinfectants
Erzsébet Pászti-Gere
AETIOTROPIC AGENTS
ANTISEPTICS AND DISINFECTANTSCHEMOTERAPEUTICS
• Ignaz SemmelweisPuerperal fever could be reduced
in the form of chlorinated lime.
• Joseph ListerHe introduced phenol to sterilize
hands and surgical instruments and to clean wounds.
TERMINOLOGY
Sterilization – a process that destroys all viable microbes, including viruses and endospores ; microbicidalDisinfection – a process to destroy vegetative pathogens , not endospores; inanimate objectsAntiseptic –applied directly to body surfacesSanitization – any cleansing technique that mechanically removes microbesDegermation – mechanically removing microbes from surface (skin) such as surgical hand scrubbing, or wiping skin with alcohol prior to venapuncture
„CID ” vs „STATIC ”
Sepsis – bacterial contamination
Asepsis – absence of significant contamination, employment of techniques, aim: microbe-free environment!
Bacteriocidal (microbiocidal) - -cidal means kill
Bacteriostatic (microbiostatic) - -static means inhibition of growth and multiplication
EFFICACY OF DISINFECTION AND STERILIZATION
– prior cleaning of the object– organic load– type and level of microbial contamination– concentration of the agents and exposure time– physical nature of the object– presence of biofilms– temperature and pH of the disinfection process– relative humidity of the sterilization process (e.g.,
ethylene oxide)
TYPE AND LEVEL OF MICROBIAL CONTAMINATION
• Number and location of microorganisms• Innate resistance
– Spore coat and cortex: spores– Waxy cell wall: Mycobacteria– Outer membrane: Gram- bacteria
• Antibiotic-resistant bacteria
www.jnanobiotechnology.comwww.nature.com
RESISTANCE OF MICROORGANISMS
SUSCEPTIBILITY OF MICROORGANISMS
Effectiveness of germicides classified as high, intermediate, or low
– High-level kill all pathogens, including endospores
– Intermediate-level kill fungal spores, protozoan cysts, viruses and pathogenic bacteria
– Low-level germicides kill vegetative bacteria, fungi, protozoa, and some viruses
CRITICAL/SEMICRITICAL ITEMS
Mahon: Textbook of Diagnostic Microbiology
BIOFILMS
• thick masses of cells and extracellular materials• up to 1,000 times more resistant• some enzymes and detergents can degrade biofilms
FACTORS AFFECTING THE EFFICACY…
• The more concentrated the disinfectant is, the greater its efficacy is– Exception: iodophors
• QAT and phenol have a concentration exponent effects• temperature, pH• increase in pH improves the antimicrobial activity of:
glutaraldehyde, QAT but decreases the antimicrobial activity of phenols, hypochlorites, and iodine.
• Water hardness• Relative humidity
METHODS OF STERILIZATION
Sridhar Rao www.microrao.com
PHYSICAL METHODS OF STERILIZATION
• Exposure to extremes of heat– Dry heat– Moist heat
• Denaturation of proteins• Interference with integrity of cytoplasmic membrane and cell walls
• Disruption of structure and function of nucleic acids
• Exposure to extremes of cold• Desiccation• Filtration• Radiation
– Non-ionizing rays– Ionizing rays
DRY HEAT
• Types: – read heat – flaming– incineration – hot air oven
• Used for materials that cannot be sterilized with moist heat• Denatures proteins and oxidizes metabolic and structural chemicals• Requires higher temperatures for longer time
MOIST HEAT
• Better conduction of heat• Methods of microbial control using moist heat
– Boiling: Endospores, protozoan cysts, and some viruses can survive boiling!!! time is critical !!!
– Autoclaving– Pasteurization:
• Batch method 30 minutes at 63ºC• Flash pasteurization 72ºC for 15 seconds
– Ultrahigh-Temperature Sterilization: 140ºC for 1 second, then rapid cooling
AUTOCLAVING
Autoclave conditions – 121ºC, 15 psi, 20 minutes•Control:
•BOWIE DICK TAPE•BIOLOGICAL: spores of B. stearothermophilus
RADIATION-NON IONIZING RAYS• UV light causes pyrimidine (thymine) dimers in DNA• UV light does not penetrate well• Suitable for disinfecting air, transparent fluids, and
surfaces of objects
RADIATION- IONIZING RAYS
• Electron beams: sterilization is accomplished in few seconds
• Electromagnetic rays: gamma rays (Co60, Cs137)– More penetrative power– Longer time of exposure– Nucleic acid damage– Impairs the flavour of certain foods
OTHER PHYSICAL METHODS
• Refrigeration and Freezing• Desiccation and Lyophilization• Filtration• Osmotic Pressure
www.pln.gov
purerair.com
AN IDEAL DISINFECTANT
Broad spectrum: should have a wide antimicrobial spectrumFast acting: should produce a rapid kill
Not affected by environmental factors:
should be active in the presence of organic matter and compatible with soaps, detergents
Nontoxic: should not be harmful to the user or patient
Surface compatibility: should not corrode instruments and metallic surfaces
Residual effect on treated surfaces: should leave an antimicrobial film on the treated surface
Odorless: should have a pleasant odor or no odor to facilitate its routine use
Economical: should not be prohibitively high in costSolubility: should be soluble in waterStability: should be stable in concentrate and use-dilutionCleaner: should have good cleaning propertiesEnvironmentally friendly: should not damage the environment on disposal
CHEMICAL AGENTS-HOW DO THEY ACT?
• Reaction with components of the cytoplasmic membrane:– Alcohol– Detergent
• Denaturation of cellular proteins:– Alcohol– Phenol
• Reaction with the thiol groups of enzymes:– H2O2– Halogens
• Damage of RNA and DNA:– ETO– Formaldehyde
Classification of disinfectants
GROUPS OF ANTISEPTICS AND DISINFECTANTS
• Halogen-containing compounds– Iodine-containing agents – Chlorine-containing agents
• Biguanides• Acids and Alkalis• Aldehydes and Reducing agents• Alkylating agents• Alcohols• Peroxides and oxidative agents• Surfactants• Phenols and Related Compounds• Metals• Dyes
ALCOHOLS• Ethyl alcohol, isopropyl alcohol and methyl alcohol (not sporicide) • Isopropyl alcohol can be used to disinfect surfaces and to disinfect
clinical thermometers. • Methyl alcohol kills fungal spores (disinfecting inoculation hoods). • Disadvantages: Skin irritant, volatile (evaporates rapidly), inflammable • Antimicrobial effect is related to their lipid solubility (damages bacterial
membranes) and their ability to precipitate cytoplasmic proteins. However, they do not destroy bacterial spores.
ACIDS AND ALKALIS
ACIDS• Bacteriostatic at pH 3-6, bactericidal at pH<3• Strong mineral acids but corrosive• Skin contaminated by spores: 2.5% HCl• Weak organic acids can be applied as:
– Food preservatives (benzoic acid)
– Antiseptics (acetic acid 0.25-1%) , at 5% bactericidal to many bacteria (otitis externa)
– Fungicides
ALKALIS• pH>9 it inhibits most bacteria and many viruses• Irritant and caustic effects• 2% solution of soda lye: fowl cholera• CaO and Ca (OH)2
HALOGENS
• Oxidizing agents cause damage by oxidation of essential sulfhydryl groups of enzymes.
• Chlorine compounds and iodine compounds– Tincture of iodine (2% iodine in 70% alcohol) and iodine can be combined with neutral
carrier polymers povidone-iodine.
– Household bleach used in a stock dilution of 1:10.
• Disadvantages: – They are rapidly inactivated in the presence of organic matter.
– Iodine is corrosive and staining. Bleach solution is corrosive and will corrode stainless steel surfaces.
IODINE COMPOUNDS
• Germicide with a wide spectrum (G+, G-, spores, fungi, viruses) of activity
• 50 ppm iodine kills bacteria in 1 min. and spores in 15 min.
Mode of action: oxidizing agents
inorganic !broad spectrum (sporicide)
organic (iodophor) � antiseptic+disinfectant
povidone-iodine
1:10-1:100 dilution!
warm
Irritation, pain, allergy!
IODOPHORE
• Applications: in teat dips to control mastitis, as dairy sanitizers, and as a general antiseptic or disinfectant for various dermal and mucosal infections.
• They slowly release iodine as an antimicrobial agent and are widely used as skin disinfectants, particularly before surgery.
CHLORINE COMPOUNDS
• potent germicidal effect• undissociated hypochlorous acid (HOCl) effective against most
organisms at 0.1 ppm• irritant to the skin and mucous membranes• used to disinfect water supplies and inanimate objects (e.g.,
utensils, bottles, pipelines) in dairies, and milk houses.
• Inorganic chlorides • sodium hypochlorite solutions (bleach 3-6%): 10X dil corrosive
and against spores, calcium hypochlorite • Organic chlorides:
– Chloramine T– Chloramine B
HYDROGEN PEROXIDEBroad spectrum: ROS damage proteins and DNA (strong solutions
are sporicidal)
Catalase, organic matters, light
Skin disinfection (peracetic acid synergism!) – Antiseptic?
5-20% � 3%
PERACETIC ACID (40%)Sterilant and antiseptic
Broad antimicrobial spectrum
Lack of harmful decomposition products of hydrogen peroxide
Greater lipid solubility
No inactivation by tissue catalase and peroxidases
Corrosive
CATIONIC SURFACTANTS
• alkyl- or aryl-substituted quaternary ammonium compounds• benzalkonium chloride, benzathonium chloride, cetylpyridinium
chloride, benzoxonium chloride /Bradophen®/) • +halogen, such as bromide, iodide, or chloride. • the major site of action: cell membrane• anionic tenzides, hard water, organic material• Not active against spores and viruses
Resist. S.aureus !• aqueous solutions (1:1,000 to 1:5,000) have good antimicrobial
activity, especially at slightly alkaline pH.
PHENOL AND PHENOLIC COMPOUNDS
They denature proteins– Phenol:
• is expensive• is caustic • has a pungent odor
– Derivatives have:• greater germicidal activity • lower toxicity than phenol
– Bisphenols are combinations of two phenol molecules
• They are commonly used in disinfection and antisepsis
PHENOLICS
• good penetrating power • disinfection of equipment or organic materials• irritant and corrosive• cauterize infected areas, e.g., infected umbilicus of
neonates
HEXACHLOROPHEN
• bacteriostatic, Gram+• absorption of the antiseptic to cause spongiform degeneration
of the white matter in the brain • in medicated soaps 1-3%• fungicidal effect
TRICLOSAN
• antibacterial and antifungal agent • risk of bacterial resistance• in soaps (0.10-1.00%)• no extra benefits: anti-gingivitis effect in toothpaste• bind to both human estrogen and androgen receptors in vitro
NO TRICLOSAN IN TOOTHPASTE!!!!
PHENOLS AND RELATED COMPOUNDS
The bactericidal activity is enhanced by - EDTA- warm temperatures
decreased by - alkaline medium- lipids - soaps
- conc. >0.5% exert a local anaesthetic effect , 5% solution is strongly irritating and corrosive to tissues .
Oral ingestion or extensive application to skin can causesystemic toxicity (CNS and cardiovascular effects).
DYES
• Triphenylmethane dyes (e.g. bromocresol green, malachite/brilliant green, crystal/gentian violet, fuchsine)
- active against Gram-positive bacteria. - lower activity in the presence of blood and tissue debris.
- Usage : 0.5-1 % solution: wounds, burns, 2-5% alcoholic solution: navel.
- Malachite green is fungicide and antiparasitic (significant in fish-medicine)
coal tar residue
gentian violet
The Gram staining method
• They alter the 3-D shape of proteins• Low-level bacteriostatic and fungistatic agents• 1% silver nitrate prevents blindness caused by N. gonorrhoeae• Thimerosal (mercury-containing compound) used to preserve
vaccines…problems?• Copper controls algal growth in reservoirs, fish tanks, swimming pools,
and water storage tanks; interferes with chlorophyll.
HEAVY METALS
academic.pgcc.edu
ALDEHYDESbroad spectrum, acts through alkylation of amino-, carboxyl and hydroxyl group (sporicide)
FORMALDEHYDE
37% cc, 1-6%
15°C !
above 35°C gas
carcinogenic� USA, EU not allowed
GLUTERALDEHYDE
at 4°C it is effective
in 2% sporicidal effect
spray, on surgace, smoke
organic material
VAPOR-PHASE DISINFECTANTS
• alkylating agents such as formaldehyde, ethylene oxide, and propylene oxide
• they are broad-spectrum biocides • active against bacteria, viruses, and fungi, including spores. • highly reactive gaseous fumigants • used for sterilizing animal feed, human food, surgical equipment that
cannot be autoclaved (e.g., endoscopes, gloves, syringes, catheters, tubing, implantable devices)
• noncorrosive
GASEOUS AGENTS
•Ethylene oxide, propylene oxide, and beta-propiolactone used in closed chambers to sterilize items•Denature proteins and DNA by cross-linking functional groups•Used in hospitals and dental /veterinary offices
BIGUANIDES
CHLORHEXIDINE
cell membrane damage
very broad spectrum (not sporicide!)
incomp: anionic detergent!
organic material
concentration: 1-4%
Chlorhexidinefacts.com
CHLORHEXIDINE
• Gram+, Gram – (10-50X conc)• Not sporicidal, but prevents development of spores• Application:
– Dermatitis (Staphylococcus, Streptococcus)– Otitis externa– Dental hygiene, gingivitis– Hygienic hand disinfection– Udder health
PRACTICAL DISINFECTION
• Disinfection of drinking water and swimming pool– Guideline values: Free chlorine: 0.5-1.5 ppm (mg/l) Combined
chlorine: 0-0.5 ppm (mg/l) Total chlorine: 0.5-1.5 ppm (mg/l) Cyanuric acid: 0-20 ppm (mg/l)
• Disinfecting patient’s skin– Alcohols, iodine compounds
• Disinfection of excretion– Phenolic preparations
• Instrument, surface, room disinfection• Hygienic hand disinfection
– Alcohols
• Surgical hand disinfection– Alcohols combined with iodophores, alcohols
standard disinfection (rubbing into hands 3 ml of isopropanol 60% v/v, 30 seconds, twice)
Target Antiseptic/disinfectant Mode of action
Cell envelope (cell wall,
outer membrane)
Glutaraldehyde Protein cross-linking
EDTA other
permeabilizers
Gr- : LPS ↑ Mg2+
removal
Cytoplasmic membrane QAT membrane damage (PL
bilayers)
Chlorhexidine membrane integrity
Phenols leakage, uncoupling
DNA interaction Acridins intercalation between 2
layers of base pairs in
DNA
Cross-linking of
macromolecules
Formaldehyde,
glutaraldehyde
CL of proteins and
nucleic acids
Interaction with thiol
groups
silver compounds membrane-bound
enzymes
Effects on DNA halogens, H2O2, silver
ions
DNA strans breakage
Oxidizing agents halogens, peroxygens ROSClinical Microbiology Reviews McDonnell 1999