DEFINITIONS Disinfectant Chemicals used to kill harmful microorganisms or inhibit their growth on non-living surfaces and objects.
Ineffective against bacterial spores.
Antiseptic
Chemicals that can kill or inhibit growth of
microorganisms on living tissues like skin, oral cavities, and open wounds.
Not effective against spores.
TO BE LIVING OR NOT TO BE !!!
DIFFERENCE
DEFINITIONS Biocide Chemical agents that kill all living microorganisms including spores.
Biostat Chemical agents that inhibit microbial growth but do not kill microorganisms.
Sporicide
Chemical that kills all living microorganisms including spores.
Synonym to biocide
DEFINITIONS Germicide
Chemical agents that kills pathogenic organisms.
Not sporicidal.
Used on both living tissues and non-living objects
Antiseptics Disinfectants
Disinfection Process that reduces or eliminates microorganisms with the exception of spores.
PROCESSES THAT PROVIDE A 5 LOG (105) REDUCTION IN MICROBIAL CONTENT.
DEFINITIONS Sanitization
Process by which the bioburden of area is taken to a safe level.
Applied to substances used on inanimate substances.
PROCESSES THAT PROVIDE A 3 LOG (103) REDUCTION IN MICROBIAL CONTENT.
Disinfection: 5 log reduction
DIFFERENCE
DEFINITIONS Decontamination
Process by which the bioburden of area is removed.
Can be done by sterilization, disinfection, sanitization or mechanical processes.
DISINFECTANTS IN PHARMACEUTICAL INDUSTRY
Excellent cleaning and sanitization program is necessary
To prevent microbial contamination of sterile and non-sterile products
Pharmaceutical ingredients (2) Process water (1) Packaging components Manufacturing environment Processing equipment (3) Operators (4)
SOURCE OF CONTAMINATION To prevent contamination
Disinfectants
Antiseptics
FREQUENTLY USED DISINFECTANTS IN PHARMACEUTICAL INDUSTRY
Alcohols Phenols
Quaternary Ammonium Compunds Sodium Hypochloride
Peracetic acid Hydrogen peroxide Gluteraldehyde Formaldehyde Chlorine dioxide
Peracetic acid/Hydrogen peroxide
I. ALCOHOLS 70 % ethanol and 70 % isopropyl alcohol.
Act on cytoplasmic membrane and solubilize lipids, denature membrane proteins
DEHYDRATE CELLS
Absolute alcohol is not antimicrobial !!
NEED WATER FOR
ANTIMICROBIAL ACTICITY
Fast acting disinfectant ACTIVITY REDUCED BY ORGANIC MATERIAL
II. PHENOLS Phenol, irgasan, hexaclorophen, chlorocresol, chloroxylenol
Target sites cytoplasmic membrane, cell wall and intracytoplasmic coagulation
Microbicidal and microbiostatic
Activity reduced in alkaline pH and organic matter
Not compatible with QAC, iodophors and detergents
III. QUATERNARY AMMONIUM COMPOUNDS
Benzalkonium chloride
Cetrimide
Act on cell membrane, increase permeability and intracytoplasmic coagulation.
Cationic, surface active substance
Inhibited by anions, acidic pH and organic matter.
Incompatible with phenols, detergents and anionic compunds.
IV. SODIUM HYPOCHLORIDE
Act on cell wall, enzymes and amino groups
Weak sporicidal
Not compatible with hydrogen peroxide, detergents or organic matter.
Corrosive
Bleach
V. PERACETIC ACID
Sporicidal
Toxic and corrosive.
Not compatible with organic matter.
VI. HYDROGEN PEROXIDE
Bactericidal and fungicidal
Antiseptic at 3% concentration
Sporicidal at 30%.
Acts on enzymes and ribosomes
VII. GLUTARALDEHYDE 2% concentration Excellent disinfectant in
few minutes
Sporicidal in at least 10 hours.
Target sites Cell wall, enzymes, cogulation, amino grops
VIII. FORMALDEHYDE
Sporicidal
Liquid and gas
Highly toxic
Not compatible with temperature below 22 °C and
humidity below 60-80%.
Acts on cell wall, cell membrane, enzyme, ATP, ribosome, nucleic
acid, coagulation
IX. CHLORINE DIOXIDE Gas
Sporicidal at low concentration and room temperature
Non-carcinogenic, non-flammable
Incompatible with copper and uncoated alluminium.
X. PERACETIC ACID/HYDROGEN PEROXIDE
Disinfectant
Sporicidal 3 h
15 min Nontoxic, noncorrosive liquid
SPORICIDAL DISINFECTANTS
Peracetic acid
Gluteraldehyde
Formaldehyde
Chlorine dioxide
Hydrogen peroxide at 30%
Peracetic acid/hydrogen peroxide
POINTS TO CONSIDER DURING THE SELECTION OF DISINFECTANTS
Bioburden
The type and the number of microorganisms.
The higher the number; the more difficult for the disinfectant to be effective.
Resistance varies according to microorganism type (vegetative bacteria, spore, virus, Mycobacteria...)
POINTS TO CONSIDER DURING THE SELECTION OF DISINFECTANTS
Surface
The type of surface may react with disinfectants.
Compatibility
Hypochloride Peracetic acid
Corrosive
Organic matter Organic material like blood, soil, clinical material of the patient negatively affect performance of some disinfectants.
Alcohol, phenol, QAC, bleach, peracetic acid
Clean the surface before
Chlorine dioxide Copper and alluminium
Contact time
The time required for the disinfectant to be effective.
Must make the surface exposed to disinfectant for at least contact time period.
POINTS TO CONSIDER DURING THE SELECTION OF DISINFECTANTS
Disinfectants should be choosed considering their contact time and the time that the surface/device should be ready.
Goal of the process (Endpoint results)
POINTS TO CONSIDER DURING THE SELECTION OF DISINFECTANTS
I. TO MAINTAIN THE MICROBIAL POPULATION AT THE CURRENT LEVEL OR TO ENSURE THAT THE CONCENTRATION OF MICROORGANISMS DOESNOT INCREASE SIGNIFICANTLY
USE A DISINFECTANT THAT HAS STATIC EFFECT
II. TO SIGNIFICANTLY REDUCE MICROBIAL POPULATION
USE A DISINFECTANT THAT HAS CIDAL EFFECT
POINTS TO CONSIDER DURING THE SELECTION OF DISINFECTANTS
Preparation of disinfectant
Dilute the disinfection accurately to use-dilution to get desired result.
Generally diluted with water.
Dilute using aseptic thecniques and sterile purified water.
If not, sterilize with filtration
Choose disinfectants that are easy to prepare.
POINTS TO CONSIDER DURING THE SELECTION OF DISINFECTANTS
Shelf life Concentrate of a disinfectant Shelf life
determined by manufacturer
Shelf life of concentrate ≠ Shelf life of diluted disin.
Dilution affects the properties and stability of compund
It is user’s responsibility to determine the shelf life of diluted disinfection
POINTS TO CONSIDER DURING THE SELECTION OF DISINFECTANTS
Safety and Residues
Is the disinfectant safe ?
Inflammable ?
Toxic residues ?
Explosive ?
Remove
Use combined disinfectants
BE CAREFUL ABOUT
COMPATIBILITY
Phenol QAC
Phenol iodophore
QAC detergent
TESTS FOR THE EFFICACY OF DISINFECTANTS
1. Suspension tests
2. Use-Dilution test
3. Surface Challenge Tests
4. In situ tests
Companies who register a public health disinfectant must ensure the safety and effectiveness of their product
It is the user’s responsibility to provide evidence that the disinfectant is effective at reducing the microbiological load of environmental isolates on the surfaces found in their facility.
Inappropriate use, ineffective cleaning and ineffective disinfectants
Affect the quality of pharmaceuticals manufactured
in the environment
DISINFECTANT EFFICACY TESTS
TESTS FOR THE EFFICACY OF DISINFECTANTS
1. Suspension test A sample of the bacterial culture is directly suspended into the disinfectant solution.
After contact time, a sample from suspended disnfectant is transferred to agar containing neutralizant
Qualitative or quantitative
TESTS FOR THE EFFICACY OF DISINFECTANTS
1. Suspension test Quantitative Suspension method
Number of surviving microorganism is compared to initial inoculum.
loginoculum - logrecovery
Log reduction must be equal to or greater than log5.
Acceptance criteria
Suspension test is used as a simple screening to determine the type of disinfectant most effective against a specific set of organisms before performing a complete (full) disinfectant qualification.
Advantage Bacteria are uniformly exposed to the disinfectant
TESTS FOR THE EFFICACY OF DISINFECTANTS
1. Suspension test
Disadvantage Not related with ‘’real life’’ disinfection process (no surface)
Use of Suspension test ?
USP doesnot imply suspension tests.
Use-dilution tests Surface challenge tests.
For disinfection efficacy testing, USP recommends
DISINFECTANT EFFICACY TESTS
TESTS FOR THE EFFICACY OF DISINFECTANTS
2. Use-Dilution test
To determine the effectiveness of the disinfectant at specific concentrations and exposure times by inoculating the specific concentrations of disinfectant with the determined organisms.
TESTS FOR THE EFFICACY OF DISINFECTANTS
2. Use-Dilution test
Carriers Stainless steel cylinders
Transferred into suspensions of test culture
Dried in incubator
Transferred into tubes containing disinfectants
After contact time transferred into broths containing neutralizant
Incubate 37 C, 48 h
1. The carrier is contaminated by submersion in a liquid culture of the test organism.
TESTS FOR THE EFFICACY OF DISINFECTANTS
2. Use-Dilution test
2. The carrier is then dried and is brought in contact with the disinfectant for a given contact time.
3. Transfer the carrier into broth containing NEUTRALIZING AGENTS.
4. No growth (turbidity) indicates activity of the disinfectant; whereas turbidity indicates failing.
Salmonella S aureus P. aeruginosa
TESTS FOR THE EFFICACY OF DISINFECTANTS
2. Use-Dilution test
Test strains
Acceptance Criteria 10 out of 10 carriers negative
59 out of 60 carriers negative
No turbidity in the broth
In addition to test strains frequently isolated ENVIRONMENTAL
STRAINS should also be tested.
LIMITATIONS OF USE-DILUTION TEST
1. The number of bacteria dried on a carrier is hard to standardize
2. The survival of the bacteria on the carrier
during drying is not constant.
3. It does not mimic the real life disinfection method.
In real life disinfection, the surface is not submerged into disinfectant. Disinfectant is sprayed or mopped or wiped onto surface.
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests Surface challenge testing involves the creation of a dried organism film onto representative surface types.
Surface chalenge tests both mimics the contaminated environment and the disinfection procedure performed in facility
Imitates ‘’real life’’ disinfection procedure
Coupons Representatives (miniatures) of the actual facility surfaces that are disinfected
5 cm x 5 cm
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests
Plastic, metal, glass, rubber, latex, vinyl, aluminium, steel
Choose the coupon that represents the surface to be disinfected
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests
Challenge (test) organisms
Staphylococcus aureus Escherichia coli Pseudomonas aeruginosa Candida albicans Aspergillus brasiliensis Bacillus subtilis
+ ENVİRONMENTAL STRAINS
Gr (+) coccus
Gr (-) bacilli
Spore forming
Fungi
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests – Steps 1 1. Prepare challenge organisms and environmental
organisms
2. Inoculate coupons with each of the challenge and environmental organisms.
3. After the coupons have dried, treat them with disinfection.
Treat the coupons with disinfectant using the same application method that you will use in the ‘’real life’’.
To make the procedure closest to real procedure
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests
during application of disinfectant to coupons, use the same method
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests – Steps 2 4. After treating the coupon with disinfectant, wait
for contact time
In order to determine the best contact time;
Use the recommended contact time One or two times below the recommended time One or two times above the recommended time
t= 2, 4, 10, 12, 15m t= 5, 10, 15m 1 time above and below 2 times above and
below
Recommended
The time during which the disinfectant does its job.
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests – Steps 3 5. Take samples from coupon that is treated with
disinfectant and test strain and from coupon that is untreated with disinfectant but treated with test strain
Sampling Methods
Swabbing Contact plates
Surface rinse
Flat surfaces
Irregular surfaces
Samples should be processed in the laboratory immediately.
If more than 60 min; refrigarate the sample.
Swabs must be moistened before taking the sample and remain moistened through the holding time.
Use transport media
Prevent both the drying and proliferation of microorganisms.
Phosphate buffer Ringer solution
6. Determine the number of cfu/ml of disinfectant treated and untreated coupon by plate count methods.
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests – Steps 3
The media that the samples (swaps ) are streaked onto or contact plates must contain neutralizer.
Determine log reduction by subtracting logbeforefrom logafter
logbefore - logafter
TESTS FOR THE EFFICACY OF DISINFECTANTS
3. Surface Challenge Tests
Acceptance criteria
2 log reduction bacterial spores 3 log reduction vegetative
bacteria, yeast, mold spores
TESTS FOR THE EFFICACY OF DISINFECTANTS
4. IN SITU TESTS After evaluating the efficacy of disinfectants under
laboratory conditions;
Perform the tests in real areas and on equipments to be routinely disinfected.
In sıtu
tests
Perform bioburden determination before disinfection, do disinfection and determine bioburden again.
Show lower count after disinfection
TESTS FOR THE EFFICACY OF DISINFECTANTS
4. IN SITU TESTS Before and after in situ tests are not performed
routinely after the disinfection method has been validated.
If during environmental monitoring new
microorganisms are isolated or increase in the number of
microorganisms is detected, it must be carried out before and
after disinfection.
IMPORTANCE OF ENVİRONMENTAL
MONITORING
As part of an environmental monitoring program, comparison of the frequency and number of organisms recovered would be performed within the actual facility.
If a new organism is routinely being recovered
If not, test disinfectant efficacy against this microor.
determine whether or not this type of organisms was covered in the disinfectant efficacy.
If a new disinfectant is introduced into the cleaning regimen, efficacy tests should be repeated.