CHAPTER 9CHAPTER 9

sterilization & sterilization & disinfectiondisinfection

biocidebiocide

A chemical agent that inactivates A chemical agent that inactivates microorganismsmicroorganisms

Usually broad-spectrumUsually broad-spectrum

bacteriostaticbacteriostatic

A biocide is able to inhibit bacterial A biocide is able to inhibit bacterial multiplicationmultiplication

Multiplication resumes upon removal Multiplication resumes upon removal of the agentof the agent

bactericidalbactericidal

A biocide is able to kill B.A biocide is able to kill B.

The “killed” organism can no longer The “killed” organism can no longer reproduce, even after being removed reproduce, even after being removed from contact with the agentfrom contact with the agent

Sterilization:Sterilization:

Completely destroys or removes all Completely destroys or removes all

microbial life, including spores. microbial life, including spores.

by physical or chemical processby physical or chemical process

disinfectantsdisinfectants

Products or biocides used to kill Products or biocides used to kill microorganisms on inanimate objects microorganisms on inanimate objects or surfaces.or surfaces.

antisepticantiseptic

A biocide or product that destroys or A biocide or product that destroys or inhibits the growth of M. in or on living inhibits the growth of M. in or on living tissuetissue

asepticaseptic

Absence of pathogenic microbesAbsence of pathogenic microbes

DisinfectionDisinfection killing, or removal of killing, or removal of

microorganisms that may cause microorganisms that may cause disease.disease.

Methods for control of Methods for control of microorganismsmicroorganisms

Physical methodsPhysical methods HeatHeat Hot-air sterilizerHot-air sterilizer

Autoclaving Autoclaving

RadiationRadiation FiltrationFiltration UltrasoundUltrasound DrynessDryness Low temperatureLow temperature

Chemical agentsChemical agents

Section 1Section 1

physical physical sterilization and sterilization and disinfectiondisinfection

Physical agentsPhysical agents

heat heat radiationradiation filtration ---- Remove organismsfiltration ---- Remove organisms Ultrasonic and sonic vibrationsUltrasonic and sonic vibrations Freezing Freezing

Impart energyImpart energy

heatheat

denature or coagulate proteins denature or coagulate proteins

damage membranedamage membrane

enzymatic cleavage of DNAenzymatic cleavage of DNA

moist heat sterilizes at a lower temperature moist heat sterilizes at a lower temperature

than dry heatthan dry heat

Two waysTwo ways

1.1. Dry heat Dry heat

2.2. Moist heatMoist heat

IncinerationIncineration

Direct flamesDirect flames

Hot-air sterilizerHot-air sterilizer

160-170160-170C for 2 hours----sporesC for 2 hours----spores

Glass petri dishes and pipettesGlass petri dishes and pipettes

InfraredInfrared

microwavemicrowave

Dry heatDry heat

For glassware and materials For glassware and materials impervious to steamimpervious to steam

Kill of all organisms, including the Kill of all organisms, including the spore formersspore formers

Moist heatMoist heat

Autoclaving:Autoclaving: pressure: 15 1b/inpressure: 15 1b/in2 2

temperature: 121temperature: 121 held: 15-20 minutes held: 15-20 minutes autoclaves or pressure cookersautoclaves or pressure cookers kill both vegetative organisms and endkill both vegetative organisms and end

osporesospores

pasteurizationpasteurization

Two methods:Two methods: 62 for 30 minutes℃62 for 30 minutes℃ 72 for 15 seconds℃72 for 15 seconds℃

to kill particular spoilage organisms or to kill particular spoilage organisms or pathogenspathogens

Primarily for milkPrimarily for milk

fractional sterilizationfractional sterilization间歇蒸气灭菌法 间歇蒸气灭菌法 ::

1) Steam heating to 100 1) Steam heating to 100 °°C for 30 min(C for 30 min( 常压常压 // 流动蒸气流动蒸气消毒法消毒法 ) ) Vegetative cells are destroyed but endospores survVegetative cells are destroyed but endospores survive ive

2) Incubate at 30 2) Incubate at 30 °°C -37 C -37 °°C overnight C overnight Most bacterial endospores germinate Most bacterial endospores germinate

3) Second heat treatment, 100 3) Second heat treatment, 100 °°C, 30 min: GC, 30 min: Germinated endospores are killed. erminated endospores are killed.

4) Second incubation at 304) Second incubation at 30°°C-37 C-37 °°C overnigC overnight: Remaining endospores germinate ht: Remaining endospores germinate

5) Third heat treatment, 100 5) Third heat treatment, 100 °°C, 60 min: LasC, 60 min: Last remaining germinated endospores are killt remaining germinated endospores are killeded

fractional sterilization:fractional sterilization:

radiationradiation

Two types of radiationTwo types of radiation

ultraviolet lightultraviolet light

Ionizing RadiationIonizing Radiation

UV lightUV light

at 250-260 nmat 250-260 nm

absorbed by purine and pyrimidin bases oabsorbed by purine and pyrimidin bases of DNAf DNA

form thymine dimersform thymine dimers

inhibit DNA replicationinhibit DNA replication

very poor penetrating power very poor penetrating power To sterilize the air and any exposed To sterilize the air and any exposed

surfacessurfaces damage the cornea and skin :damage damage the cornea and skin :damage

the eyes, cause burns, and cause the eyes, cause burns, and cause mutation in cells of the skin mutation in cells of the skin

UV lightUV light

filtrationfiltration

for certain solutions for certain solutions that that may be damaged or may be damaged or denatured by high denatured by high temperatures or chemical temperatures or chemical agentsagents

has a pore size of 0.22 μmhas a pore size of 0.22 μm physically trap particles physically trap particles

larger than the pore size larger than the pore size and retain smaller particles and retain smaller particles

Section 2Section 2

chemical chemical sterilization and sterilization and disinfectiondisinfection

Alcohols Alcohols 醇醇 Aldehydes Aldehydes 醛醛 Halogens Halogens 卤素卤素 Heavy metals Heavy metals 重金属重金属 Phenolics Phenolics 酚酚 Detergents Detergents 去污剂 去污剂

Chemical agentsChemical agents

常用的化学消毒剂常用的化学消毒剂类别类别 作用机制作用机制 常用种类常用种类酚类酚类 蛋白变性，细胞膜损伤蛋白变性，细胞膜损伤 洗必泰洗必泰醇类醇类 去除脂类，蛋白变性去除脂类，蛋白变性 乙醇乙醇卤素卤素 蛋白变性蛋白变性 氯气、碘酊 、碘伏氯气、碘酊 、碘伏

重金属盐重金属盐 蛋白变性蛋白变性 红汞、硫柳汞、硝酸银红汞、硫柳汞、硝酸银

醛类醛类 蛋白变性蛋白变性 福尔马林、戊二醛福尔马林、戊二醛

表面活性剂表面活性剂 蛋白变性，细胞膜损伤蛋白变性，细胞膜损伤 新洁而灭新洁而灭酸碱类酸碱类

染料染料

破坏细胞膜、细胞壁，破坏细胞膜、细胞壁，蛋白变性蛋白变性干扰氧化、抑制繁殖干扰氧化、抑制繁殖

十一烯酸十一烯酸

龙胆紫龙胆紫

Nature, concentration and acting Nature, concentration and acting time of the disinfectanttime of the disinfectant

Type and amount of the microbeType and amount of the microbe TemperatureTemperature pHpH Organic matterOrganic matter

Factors affecting the results

Chemical agentsChemical agents

1.1. disruption of the lipid-containing cell disruption of the lipid-containing cell membranemembrane

2.2. modification of proteinsmodification of proteins

3.3. modification of DNA modification of DNA

disruption of the cell membranedisruption of the cell membrane

AlcoholsAlcohols to clean the skinto clean the skin disorganizing the lipid structure in disorganizing the lipid structure in

membranesmembranes denature proteinsdenature proteins far more effective at 70% than at 100% far more effective at 70% than at 100%

DetergentDetergent

SurfactantsSurfactants Disrupt the membraneDisrupt the membrane Widely used for skin antisepsisWidely used for skin antisepsis

phenolsphenols

The first disinfectant used in the The first disinfectant used in the operating roomoperating room

Too causticToo caustic Damage membranes and denature Damage membranes and denature

proteinsproteins

modification of proteinsmodification of proteins

ChlorineChlorine

to purify the water supplyto purify the water supply

to treat swimming poolsto treat swimming pools

an oxidizing agentsan oxidizing agents

iodineiodine

the most effective skin antisepticthe most effective skin antiseptic

tincturetincture: to prepare the skin prior to : to prepare the skin prior to

blood culture blood culture

irritate to the skinirritate to the skin

iodophorsiodophors: to prepare the skin prior to : to prepare the skin prior to

surgerysurgery

less irritatingless irritating

Heavy metalsHeavy metals

mercury and silvermercury and silver: :

have the greatest antibacterial have the greatest antibacterial

activityactivity

bind to sulfhydryl---- block bind to sulfhydryl---- block

emzymatic activityemzymatic activity

Ethylene oxideEthylene oxide

for heat-sensitive materialsfor heat-sensitive materials

alkylate both proteins and nucleic alkylate both proteins and nucleic

acids acids