Sterilization & DisinfectionPhysical Agents

Hugh B. Fackrell

PhysSter.ppt

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Sterilization & Disinfection

Definitions Bacterial Cell Death Physical methods

– Heat– Radiation– Filtration

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Definitions

Sterilization– Complete Destruction of ALL microbes

Disinfection– Killing of pathogens on inanimate objects– physical or chemical

Antiseptic– A chemical agent for disinfection of living tissue– External skin, mouth vagina

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Definitions

Bacteriocide – kill microbes– also germicide, fungicide, virucide

Bacteriostatic– Prevents or stops microbial growth– also fungistatic, virustatic

Aseptic(Asepsis)– Prevent contamination of person or object by microbes

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Definitions

Sanitize– Removal of pathogens from inanimate objects– Mechanical or chemical cleaning– need not sterilize of disinfect

Contamination– Presence of living microbes on object

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Definitions

Infection– Presence of living multiplying microbes in host

tissues– often pathogenic

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Bacterial Cell Death

Time (min)

Survivors

(log)

0

1

2

3

4

5 1 log decrease = 90% kill

1 log Death rate (D) varies with bacteriocide

D Time(min) to kill 90%

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Physical Methods

Heat Radiation Filtration

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Heat Sterilization

Factors – length of exposure– temperature

“Moist Heat” “Dry Heat”

Method of choice

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10“Moist” heat

Effective quickly against most pathogens – low thermal death point– 60-70% in 10 min

Pasteurization 60-70 oC Sanitizes Hot water 60-100 oC Disinfects Steam 15 psi 121oC Sterilizes

Endospores resist boiling for hours!!

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11Heat: Mechanism of Action

Inactivation of enzymes Structural changes

– coagulation of proteins

Oxidation – membrane disruption

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12Pasteurization Liquids Minimal heat

– 145 oF (62.9 oC) - 30 min

– 162 oF (71.6 oC) - 15-30 sec

Flavour of drink preserved Kills non spore forming pathogens

– TB. Brucella, Salmonella, Streptococcus

Kills spoilage bacteria– Lactobacillus, Streptococcus

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13“Flash” Pasteurization

72 oC - 15 sec 88 oC - 1 sec 90 oC 0.5 sec 96 oC 0.05 sec

Heating coil Cooling coil

Raw milk

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14Boiling Water

100 oC (212 oF) 10- 30 min Disinfectant

– spores & some viruses are resistant

Safe for clean instruments that DO NOT penetrate the tissue

Soiled items -add detergent

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15Steam Under Pressure

Autoclave 15 psi 121 oC 250 oF Time 15 min - dependent on volume Steam displaces air & penetrates

– Packing of autoclave critical!

Airlocks in packing– reduce effective temp & pressure

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16Dry Heat

Longer & hotter Oxidation Carbonization

– cotton plugs

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17Types of Dry Heat

Oven– 160-165 oC 2 hours– 170-175oC 1 hr– Glass, oils, waxes, glycerin

Incineration– destruction of pathological material

Flaming– used as aseptic techniques– scissors, scalpels, loops, needles

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Radiation Sterilization

Ionizing RadiationUltraviolet Radiation

Point mutations involve changes in only one or a few nucleotides

*

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Radiant Energy spectrum

Ionizing

Radiation

UV lightIncreased

energy

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20 Sterilization by

Ionizing Radiation Gamma Rays

– Food– Plastics

Electron Beams– pharmaceuticals

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Ionizing Radiation

Creates free radicalsCauses breaks in two DNA strands

–Results in deletions

–Results in translocations

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Free Radicals

High Energy Radiation–energy absorbed by atoms

–energy added to electrons

–electrons ejected from atoms

–leaves ionized atoms & unpaired electrons

Free radicals can breaks both phosphodiester bonds of a DNA helix

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23Sterilization by Ultraviolet light

Circulating air Surfaces

– benchtops

Small samples of liquid

Surfaces only poor penetration

Photoreactivation

white light reverses effect of ultraviolet

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Ultraviolet Radiation

Pyrimidine dimers

–errors during repairPeroxides, Free radicals

–See ionizing radiationDirect molecular damage

–intense exposure

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Repair of thymine dimer

GTA T GTCCATAACAG

GTA TTGTCCATAACAG

CATAACAG GTA GTC

TT

TTGT GTCCATAACAG

A

GT GTCCATAACAG

TTA G

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26Filtration Sterilization

HEPA filters

– High Efficiency Particulate Air filter

– Remove airborne microbes• Filters particles >0.3 m

– Operating rooms

– “Clean rooms” instrument assembly

– Burn patients

– Immunocompromized

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27Filter sterilization

Membrane filters– cellulose nitrate or synthetic polymers

Pore size– bacteria 0.22m 0.45 m– viruses 0.01 mm

Sterilize heat or energy sensitive materials– bacterial media– drug solutions