Experiment three Physical and Chemical agents for the Control of microorganisms Physical and Chemical agents for the Control of microorganisms –Effect

Experiment threeExperiment three

•Physical and Chemical agents for Physical and Chemical agents for

the Control of microorganismsthe Control of microorganisms– Effect of Ultraviolet RadiationEffect of Ultraviolet Radiation

– Effect of chemical agentsEffect of chemical agents

Effect of Ultraviolet RadiationEffect of Ultraviolet Radiation

•Ultraviolet radiationUltraviolet radiation (UV, 240-300nm)(UV, 240-300nm)

– The most effective wavelengths: The most effective wavelengths:

265nm265nm– MechanismMechanism:: destroy bacteria by interfering destroy bacteria by interfering

with DNA replication and transcription with DNA replication and transcription

during protein synthesisduring protein synthesis

– CharacteristicsCharacteristics::

•poor penetrationpoor penetration

•damage of the eyes and the skindamage of the eyes and the skin..

• MATERIALS:MATERIALS: – Media: Media: Nutrient agar plateNutrient agar plate– Culture:18-24 hours cultures of Escherichia Culture:18-24 hours cultures of Escherichia

coli coli – Alcohol burner , inoculating loop, cotton swabAlcohol burner , inoculating loop, cotton swab

. .


• PROCEDURE:PROCEDURE:– Take a nutrient agar plate, streak E.coli intensively on Take a nutrient agar plate, streak E.coli intensively on

the plate. the plate. – Place the plate directly under the ultraviolet lamp. ThPlace the plate directly under the ultraviolet lamp. Th

e plate lid cover half part of the agar surface.e plate lid cover half part of the agar surface.– After exposed to ultraviolet light for 30minutes,close After exposed to ultraviolet light for 30minutes,close

the plate.Incubate the plate in an inverted position at the plate.Incubate the plate in an inverted position at 37℃ for 18 to 24 hours and observe the results.37℃ for 18 to 24 hours and observe the results.

• RESULT:RESULT:– Observe the growth of bacterium in the plate.Observe the growth of bacterium in the plate.

. .


•Mechanisms of actionMechanisms of action-- Destroy cell membranes; -- Destroy cell membranes;

e.g., surfactants/detergents -- e.g., surfactants/detergents -- dissolve lipids dissolve lipids

-- Denature bacterial proteins (e.g., -- Denature bacterial proteins (e.g., enzymes ); enzymes );

Interfere bacterial metabolismInterfere bacterial metabolism

-- Damage genetic materials (DNA, -- Damage genetic materials (DNA, RNA);RNA);

e.g., formaldehydee.g., formaldehyde. .

Effect of chemical agentsEffect of chemical agents

• Commonly used chemical agentsCommonly used chemical agents ---Phenol---Phenol---Soaps and detergents ---Soaps and detergents ---Alcohols: 70-75%(medical use) ---Alcohols: 70-75%(medical use) ---Heavy metals: Silver nitrate (1%) ---Heavy metals: Silver nitrate (1%) ---Chlorine ---Chlorine ---Iodine ---Iodine ---Aldehydes: 37%, Formalin---Aldehydes: 37%, Formalin---Dyes: crystal violet---Dyes: crystal violet


• Reagents:Reagents: 1. 1% crystal violet1. 1% crystal violet 2. 2% mercurochrome2. 2% mercurochrome 3. 5% iodine3. 5% iodine 4. 0.1% bromogeramine4. 0.1% bromogeramine 5. normal sodium5. normal sodium

• SpeciesSpecies ：：– S.aureus S.aureus 1 agar plate1 agar plate– E.coli E.coli 1 agar plate1 agar plate


• PROCEDURE:PROCEDURE: – Using Sterile techniques, incubate the agar plate with tUsing Sterile techniques, incubate the agar plate with t

he culture of S.aureus (or E.coli) by streaking the agar plhe culture of S.aureus (or E.coli) by streaking the agar plate in horizontal and vertical directions with an inoculaate in horizontal and vertical directions with an inoculating loopting loop

– Using a flamed and cooled forceps, take each saturated Using a flamed and cooled forceps, take each saturated disc and place it in the appropriately labeled area respedisc and place it in the appropriately labeled area respectively to ensure that each one ie at the middle point of ctively to ensure that each one ie at the middle point of radius and of them are equidistant from each other. Geradius and of them are equidistant from each other. Gently press each disk down so that they adhere to the surntly press each disk down so that they adhere to the surface of the agar.face of the agar.

– Label and incubate all plate culture in an inverted positiLabel and incubate all plate culture in an inverted position at 37℃ for 18-24 hours. on at 37℃ for 18-24 hours.


crystal violet

mercurochrome

iodine

bromogeramine

normal sodium


RESULT:RESULT:Measure inhibition zone

– diameter or 2× radius

CulturesCulturesDiameter of Inhibition Zone (mm)Diameter of Inhibition Zone (mm)

crystacrystal l violetviolet

mercurochromercurochromeme

iodineiodine bromogeraminbromogeraminee

normal normal sodiumsodium

S.AureusS.AureusE.coliE.coliSusceptibiliSusceptibilityty

degreedegree

Record:

Inhibition Inhibition zone zone diameter diameter (mm)(mm)

>15mm>15mm 10~15m10~15mmm

<10mm<10mm 0mm0mm

susceptibility susceptibility degreedegree

ExtremelExtremely y sensitivesensitive

MediumMedium sensitivesensitive

Slight Slight sensitivsensitivee

restistanrestistantt

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Experiment three Physical and Chemical agents for the Control of microorganisms Physical and Chemical agents for the Control of microorganisms –Effect