Disinfectant Qualification and Cleaning Procedures · PDF fileDisinfectant Qualification and Cleaning Procedures Workshop IVT's Cleaning Validation and Critical Cleaning Processes

Disinfectant Qualification

and Cleaning Procedures

Workshop

IVT's Cleaning Validation and Critical Cleaning Processes

in Philadelphia, PA - August 18-20, 2015

Presented by

Ziva Abraham

President

Microrite, Inc.

5019 New Trier Avenue

San Jose, CA 95136

Phone: 408-445-0507 Fax: 408-445-0507

Email: [email protected] www.microrite.com

mailto:[email protected]

Ziva Abraham is the President and Founder of Microrite, Inc., a California based

consulting firm providing consulting and training services to pharmaceuticals,

biotechnology, medical devices and in vitro diagnostics in the areas of quality

assurance, quality control, microbiology, and validation. Ziva has over 25 years

of academic, research, clinical and industrial experience in microbiology, and

quality assurance. Ziva has received her Master’s Degree in microbiology and has

conducted research on developing microbial insecticides using entomogenous

bacteria and fungi towards a Ph.D degree. Her career also includes founding and

managing clinical laboratories for Maccabi Medical in Israel. She has trained

personnel from various industries in microbiology techniques and methods. She

uses her extensive experience to teach why assessing risk of microbial

contamination should be in the forefront of any company that has products for

human/veterinary use. Her experience in clinical laboratories has provided her

with the framework to understand the effects of microbial contamination in

products from a patient safety perspective.

About the Speaker

Who is Microrite?

Goal Microrite’s goal is to provide practical solutions

through consulting and training

Consulting Services Microrite is a San Jose, CA based Consulting Company

helping Pharmaceuticals, Biotechnology, Medical

Devices, In-Vitro Diagnostics and Combination products

in the areas of Quality Assurance, Microbiology, Process

Development, Process Validation, Facility and Utility

Validation

Visit www.microrite.com to learn more about us

About Disinfectants, Sanitizers and Sterilants

• Definitions

• Mode of Action

• Advantages/Disadvantages

• Governing Bodies/Regulations

• Testing for Manufacturers

• Planning Disinfectant Qualification

• Methods generally used in Pharmaceuticals

Topics

• Tube Dilution Method

• Different Coupon/Hard Surface Methods

• Expiry Dating Studies

• Neutralization Study

• Method Details

• Calculations

• Disinfectant Qualification Report

Topics

About Cleaning Procedures

• Challenges in Cleaning

• Procedure for Receiving, Storage, Preparation of

Disinfectants

• Disinfecting/ Sanitizing Procedure

• Training

Topics

Plan of Attack on Planktonic Microbes

Know your enemy before you attack!

http://www.clipartguide.com/_pages/0511-1102-2413-2657.html

http://www.clipartguide.com/_pages/0511-1101-2813-3216.html

Sporicide: Chemical agent that kills all microorganisms including

bacterial and fungal spores

Disinfectant: Chemical agent that kills microorganisms but not

necessarily spores

Germicide: Chemical agent that kills microorganisms

Bactericide: Chemical agent that kills bacteria but not

necessarily spores

Fungicide: Chemical agent that kills fungi

Virucide: Chemical agent that kills viruses

Cleaning agent: Agent with surfactant

Surfactant: Agent that breaks surface tension

Sanitizer: Reduces the number of microbes

Definitions

Do not kill spores, fungal or bacterial, bactericidal and virucidal claims, leaves residue Examples: Vesphene, LpHse

Do not kill spores, fungal or bacterial, bactericidal and virucidal claims, leaves residue Example: Intercept

Kills spores, very unstable, available chlorine may vary depending upon time on shelf

3% kills vegetative bacteria, greater than 6% sporicidal, safe enough to use for food industry

H202

Bleach

QUCS

Phenolics

Disinfectant Facts

Mixture of peracetic acid and H2O2 forms a potent sporicidal agent

Formaldehyde and Gluteraldehyde potent sporicides, highly carcinogenic

Sanitizer, 30% water required to seep through cell wall barrier and disrupt cell contents, some bacteria may use it as carbon source (pseudomonads)

PAA

Aldehydes

Alcohol

Disinfectant Facts

Bioguinide compounds are used as antiseptics in hand

wash solutions and for surgical gloves

Proven to have upto 1.5 log reduction

Iodides, and other combinations are used to clean

before surgical procedures

Ability to lower the surface tension of water by

addition of surfactants makes some disinfectants good

cleaning agents

Other Antimicrobials and Surfactants

Phenolic residue

Courtesy of Veltek Associates, all rights reserved

Regulatory Bodies

• Disinfectants are classified by FDA, EPA,AAMI and CDC mainly for medical device industry

• OSHA oversees the health and safety aspects

• Disinfectants used for hard surfaces are registered with EPA

Required testing before marketing disinfectants:

• Simulated Use Testing

• Actual Use Testing

• Biocompatibity data-evaluation of residue

• Toxicity data

• Material Compatibility Data

• Chemical Indicator

Regulatory Bodies

Regulations

• Required by EPA/FDA to confirm label claims

• Disinfectants must pass the tests described in the

AOAC (Association of Official Analytical Chemists)

in order to be approved by EPA or FDA

• No harmonized consensus on methodology

• AOAC: Association of Official Analytical Chemists

• AFNOR: Associacion Francais de Normalization

• BSI: British British Standards Institution

• DGHM: Deutshe Gesellschaft fur Hygeine und Mikrobiologie

• CEN-TC 216: European Committee for Standardization

Regulatory Bodies

AOAC Methods

Disinfectants must pass the tests described in the

AOAC Manual in order to be approved by the

Environmental Protection Agency.

AOAC Tests Include:

• Spray Products

• Phenol Coefficient

• Use Dilution

• Hard Surface Carrier

• Tuberculocidal Activity

• Sporicidal Activity

• Fungicidal Activity

Testing for Manufacturers

T. mentagrophytes for fungicidal claim

EU methods require a mold A. niger and a yeast C.

albicans

AOAC for bacterial is S. cholerasuis, S. aureus and P.

aeruginosa also common for many different regions

and agencies

AOAC for sporicidal is an aerobic B. subtilis and an

anaerobic C. sporogenes

What Organisms are tested for label claim?


AOAC for tuberculocidal is M. bovis; M. smegmatis can

be used as a presumptive test but its resistance is

much weaker to germicides than M. bovis or M. terrae

AOAC for virucidal is not set, they recommend

following the EPA method which generally uses HBV,

HCV, Herpes simplex, HIV and influenza

FDA uses polio virus II and an enveloped virus one

selects while EU calls for polio virus I and adenovirus

V


Label claim testing does not address all

microorganisms found in cleanrooms

Cleanroom flora much diverse

Clinically relevant organisms such as S. aureus,

Salmonella or Trichophyton not commonly

encountered in cleanrooms

Bacterial Contamination

Bacteria

Gram Positive

Aerobic

Gram Negative

Aerobic

Cocci Rods Cocci Rods

Mainly human

borne

Staph aureus (bad

bug) (<61>, <62>)

Staph epidermidis

Staphylococcus

warneri

Enterococcus

Streptococcus

Bacillus:

Live in soil

Bad gowning and

cleanroom practices

Disinfection program

nor adequate

Facility design flawed

Concern: product

contamination (

preservative system

may not be adequate

and biofilms

B. subtilis

B. licheniformis

B. cereus (serious

problem)

Neisseria

meningitidis, N.

gonorrhoeae,

Moraxella

catarrhalis

Cocco-bacillus:

Haemophilus

influenzae,

Acinetobacter

Live in soil

Flourish in water

systems

Cause endotoxin

Detrimental to patient

Bad gowning and

cleanroom practices-

soil

Concern:

Endotoxin , Biofilm

E.Coli

Enterobacter

Pseudomonas

http://www.buddycom.com/bacteria/gnc.html

http://www.buddycom.com/bacteria/gnr.html

Fungi Found in Cleanrooms

http://comenius.susqu.edu/bi/202/Fungi/ASCOMYCOTA/Ascomycota/ascocarps-bold.jpg

Zygomycota

Ascomycota



Fungal Mycelia

Fungal Mycelia

The Bacterial Spore Challenge

Bacillus cereus is a serious problem

Paenibacillus has been a continuous issue once sets in

Biofilms formed by Bacillus is not uncommon in

fermentation equipment

Biofilm organisms are your planktonic ones-just

smarter

United we stand, divided we fall

Biofilms

Use Dilution Method

• Cylinders introduced into test culture for 15 minutes

• Dried for 40 minutes

• Dropped in diluted disinfectant

• Contact time achieved

• Dropped in neutralizing broth and incubated

• No growth in 10 out of 10 - Pass

• Growth in any - Fail

• Acceptance Criteria-Maximum dilution of disinfectant which

kills test organisms in a 10 minute interval

AOAC Tests

Carrier Test Method

• Fire polished borosilicate glass carriers used

• Culture suspension is sonicated and 1 mL of culture

introduced per carrier

• Carriers inverted on filter paper after 15 minutes and dried

for 40 minutes

• One carrier dropped into one tube of disinfectant every 30

seconds

• At 10 minute contact time, each carrier is removed at

interval of 30 seconds

• Excess disinfectant is removed and carriers dropped in tubes

with letheen broth

• Broth with carriers are incubated at 37° C for 48 to 54 hours

• 2 positive out of 60 is the acceptance criteria

AOAC Tests

Suitability of Methods

• AOAC Methods not suitable for Pharmaceutical

Industry

• No one method used or recommended

• Minimum guidance in Pharmacopeial Forum Vol. 28

(1) Jan-Feb 2002 <1072> Disinfectants and

Antiseptics

Methods commonly used in Industry:

• Tube Dilution Method

• Coupon Methods

Planning Disinfectant Qualification

Common factors no matter which method you choose

• Choice of organisms

• Purity of organisms

• Maintenance of organisms

• Media growth promotion and sterility for buffers performed

• Enumeration for target inoculum

• Maintenance of target inoculum

• Daily inoculum verification

• Method validation

• Recovery study and recovery loss calculation-in case of hard

surfaces

• Calculating log reduction


When is it applicable?

Initial screening of disinfectants

Testing disinfectants for hard to kill microorganisms

before using for routine cleaning

Tube Dilution Method

Disinfectant dilution(s) challenged with inoculum

After contact time is attained, the disinfectant with

inoculum my be filtered

Results may be qualitative or quantitative

Consider the target inoculum to avoid multiple

dilutions and filtration

Positive controls considered as target inoculum

Consider running negative controls be run per lot of

media and buffer


Two tubes per organism for each microorganism per

disinfectant dilution tested

Two negative controls of dilution of disinfectant to be

tested

Two positive controls of organisms to be tested-

positive controls may be prepared in water or saline


Planning

• Number of organisms

• Saline tubes for enumeration

• Number of plates for enumeration

• Number of disinfectants

• Number of dilutions

• Number of time points per dilution

• Number of tubes

• Number of filters

• 400mL buffer per filter

• Number of plates for filters


Planning - Example

Organism - S. aureus

Disinfectant tested – LpHse

1 dilution - MRC

1 time point - 10 minutes

Always test one microorganism at a time


Planning for Enumeration - Agar plates and saline

tubes

• Microbial Identification

• Two slants for working culture

• Two plates for purification of culture

• Two plates for purification of culture

• 8 plates for enumeration

• 4 plates for controls

• 1 saline tube for harvest

• 8 saline tubes for enumeration dilutions

• 2 saline tubes for target inoculum

• Streakers


Planning for Filtration Method – Efficacy – S. aureus

with LpHse for one dilution and one Contact Time

• 2 filters

• 800mL buffer for method validation of

• 800 mL for positive control

• 800mL buffer for negative control

• 2 TSA plates for challenge

• 2 filters for negative control

• 2 plates for negative control

• 2 filters for positive control

• 2 plates for positive control


Efficacy Data Analysis

With target inoculum of 1 X 103 results can be quantitative with

1 or two log reduction

1 log reduction- 100 CFUs can be enumerated on filter

2 log reduction 10 CFUs can be enumerated

TNTC data (>200 CFUs cannot be enumerated)

2 X 103 results may be quantitative with 1 or two log reduction




Efficacy Data Analysis

3 X 103 results may be quantitative with 1 or two log

reduction

1 log reduction 300 CFUs may not be enumerated


The inoculum strength falls after enumeration

The enumeration reading is not the exact value of

target inoculum

Positive control may be treated as Time “0” count


Efficacy Study-Quantitative

Dilutions of Disinfectants made

Each dilution to be tested in duplicate or triplicate for each contact time for each organism

Each dilution inoculated with test organism-106-8

After contact time is attained serial dilutions are filtered

The higher the target inoculum the more dilutions needed

106 - 5 dilutions to be made - last 4 filtered




Benefits

Less time consuming than testing on coupons

Drawbacks

Does not prove that the disinfectant is effective on

hard surfaces

Materials used for higher inoculum as expensive as

coupon study


This Coupon Method can be used in a manner similar to the tube

dilution study

The only difference would be to inoculate coupons instead of

directly inoculating the disinfectant dilution

Inoculum is applied to the coupon, dried and then sprayed or

flooded with the disinfectant

After the contact time is achieved the coupon may be placed in

a tube containing buffer

The tube with the coupon and the buffer is vortexed and then

filtered

Multiple dilutions may be made depending in the target inoculum

Coupon Methods 1

The difference between this method and the Coupon

Method 1 is that instead of using filtration as a way of

neutralizing the disinfectant, the coupons are placed

in a tube with neutralizing broth

Pour plate method or steak plate method may be

used

If the neutralizing broth used is in large quantities,

the recovery may not be adequate

Coupon Methods 2

Points to consider

Calculate a target inoculum whereby many dilutions

are not required yet a 3 log reduction or target log

reduction can be achieved

Ensure that the coupon sizes are small so as to avoid

using large quantities of neutralizing broth-getting

adequate dilutions to yield useable results may be

difficult

Perform method validation

Coupon Methods 1 and 2

Advantages

Coupon Method 1 is a quantitative study

Coupon Method 2 is less time consuming considering

large quantities of broth is not used

Disadvantages

Only sterilizable coupons such as stainless steel and

glass can be used for both methods

Coupons like wall material will introduce

contamination which may make reading surviving

CFUs a challenge in both methods

Coupon Method 1 and 2

Microorganism inoculated on the coupon and dried

Contact plates used to recover surviving

microorganisms from coupon

Recovery considerably lower than swab recovery

method

Coupon Method 3 (contact plates)

This study is both qualitative and quantitative depending upon

the effectiveness of the disinfectant and inoculum size

For achieving a 1-3 log kill an approximately 2 X 103 target

inoculum can be used the results can be quantified

~200 CFUs can be counted on a contact plate

Less than a log of kill will result in TNTC where the results will

be qualitative

If care is taken to adjust the inoculum strength with the

knowledge of disinfectant capability this study can save a lot of

time and cost

Due to variability in microbiological methods attaining accurate

inoculum strength is difficult


Challenges

Recovery may be inconsistent due to:

• Nature of surfaces

• Technique

If loss of recovery is close to a log it may become

difficult to prove 1-3 log reduction

Contact plates may not contain adequate neutralizers

DE Neutralizing agar which neutralizes all

disinfectants, is not used for routine EM


Other challenges

Positive control for efficacy may be difficult to read

with a 103 or 104 challenge

Plating dilutions of the target inoculum may help to

assess time “0” count


Benefits

Less time consuming among the representative Hard

Surface Studies

Drawbacks

Very inconsistent and may be able to prove a 3 log

reduction due to limited number of colonies that can

be counted on 25 cm. sq. area

Hard to enumerate fungi and spreaders

Not always quantitative (with TNTC growth, the CFUs

cannot be counted)


Coupons inoculated with target inoculum

Dried and treated with disinfectant

Recovered using swab, preferably two swabs into

tube containing buffer

Buffer with swabs vortexed and appropriate number

of dilutions made

Each dilution filtered, filters plated and incubated

Positive controls represent the recovery loss due to

drying of technique

Coupon Method 4 (swab recovery method)

Rinsing of coupons may be used instead of swabbing

Both recoveries and efficacies can me much lower than swab method

Coupon Method 5 (Rinse Method)

Using plastic bags and manual dislodging is not

practical for wall surfaces

Wall coupons cannot be steam sterilized

Mold growth may take over

Coupon Method 6 (Manual Dislodging)

Benefits

All representative hard surfaces can be tested

Complete quantitative study

Drawbacks

Extensive and time consuming

Coupon Method 4 (swab recovery method)

Benefits

All representative hard surfaces can be tested


Drawbacks


Inconsistent recoveries

Coupon Method 5 (rinse method)

Benefits

All representative hard surfaces cannot be tested

Coupons that cannot be sterilized may add

contaminants


Drawbacks


Low and inconsistent recoveries in dislodging method

Loss of CFUs as they can adhere to coupons and bags

Coupon Method 6 (manual dislodging method)


• Choice of test microorganisms

• Choice of surfaces

• Enumeration

• Calculating target inoculum for method validation and

efficacy

• Method Validation

• Efficacy

• Ensuring viability of inoculum-Time “0” count

• Calculation log reduction

• Expiry Study

• Neutralization Study

• Translation of results to cleaning and EM Procedures

Performed for:

• Undiluted and stored disinfectants

• Disinfectants that are diluted in-house and stored in

containers with an expiry date

Tests performed include:

• Efficacy: To verify no drop in efficacy on around

expiry

• Chemical Stability: Where applicable, to verify

no drop in chemical stability has occurred on or

around expiry

Expiry Dating Study

Neutralization Study

• To verify that the residue on surface does not interfere with

recovery of organisms during environmental monitoring

• The residue from disinfectants does not exert bacteriostatic /

fungistatic effect on surviving microorganisms

• To verify that the neutralizers in the contact plates are

adequate to neutralize the active ingredients in the

disinfectant residue

• The time lapse between cleaning and sampling is appropriate

for recovery of organisms


• Coupons - Stainless steel

• Disinfectants - highest concentration used for

disinfectant qualification

• Time points - 0, 2hrs, 4 hrs, 6 hrs, 12 hrs, 24 hrs

(aligned with cleaning practices)

• Organisms - Used for disinfectant qualification


• Duplicate coupons are treated with disinfectant

• At each time point coupons re sampled using

appropriate contact plates

• Contact plates are inoculated with ~ 100 CFU of

test organism

• Positive control contact plates are inoculated

with ~ CFU or test organism

• Contact plates are appropriately incubated

Acceptance criteria- The time point that yields 70% or factor

of 2 CFU count as compared to positive control is appropriate

for sampling of surfaces after disinfection / sanitization


• ATCC Strains: USP challenge organisms

• In-house Isolates: Predominant and objectionable

• Gram Positive Cocci - ATCC Strain and/or In-house

organism

• Gram Negative Baccilli - ATCC Strain and/or In-

house organism

• Gram Positive Bacilli In-house organism

(predominant)

• Yeast - ATCC Strain and/or In-house organism

• Mold - ATCC Strain and/or In-house organism

• Include objectionable and process microorganisms

Choice of Microorganisms

Choice of Microorganisms

Hard Surfaces Evaluation

• Plastic

• Glass

• Stainless Steel (316L, 316S etc.)

• Painted Surfaces

• Tile

• Aluminum

• Rubber

• Wood

• Flooring

• Latex

• Epoxy

• Vinyl

Choosing Hard Surface Coupons

• TSA settle and contact plates- tested for growth

promotion and sterility

• SDA settle and contact plates - tested for growth

promotion and sterility

• Rinse Fluids: Fluid A, Peptone Water or Phosphate Buffer

- tested for sterility

• Suspension Fluid for Inoculum: Sterile Saline - tested for

pH and sterility

• Letheen Broth or DE Neutralizing Broth - tested for

sterility

• Manifolds

Media and Supplies

• Dacron swabs

• Filters-correct size and material

• Sterile tubes

• Sterile saline

• McFarland standards

• Coupons-correct size and number

• Totes

• Disinfectants with short and extended expiry

• Water for dilution of disinfectants

• Hood

Media and Supplies


• Small coupons of surfaces that may be autoclaved

may be used for immersion into broth medium ~1”

X 3”

• For efficacy tests performed using contact plates,

coupons should be appropriately ~3” X 3” in size,

up to 4” X 4” not larger - multiple coupons to be

tested under a hood-hood size is important

• Coupons for efficacy tests using swab recovery ~2 ‘

X 2” up to 4” X 4” not larger - multiple coupons to

be tested under a hood


Cleaning or Sterilization of Hard Surfaces

• Hard surfaces coupons may be re-used.The coupons

need to be clean and free of micro-organisms

Choice of Sterilization Method

• Autoclave - For hard surfaces e.g. plastic, metal,

steel, etc.

• Chemical - Using a sporicidal compound for hard

surfaces that cannot be autoclaved, e.g. wall, floor

etc. followed by a wipe down with 70% IPA or sterile

water

Cleaning and Sterilization of Coupons

• Contact time: Elapsed time between application of

the disinfectant and its removal

• Test at manufacturer recommended contact time

• Understand the time disinfectants are active after

drying-study to show activity

Contact Time

Covering the entire area where inoculum is applied

may be attained by:

• Spraying

• Flooding

• Spreading without removal of inoculum

Special attention should be paid to surface tension

which prevents the disinfectant from completely

covering a hard surface

Application of Disinfectant

Swabbing

• Two swab technique

• Consistency in swabbing technique is critical

• Swabbing is accomplished by ten strokes in one

direction and additional ten strokes at 90 degrees

to the first ten. Each stroke is a full forward and

backward motion with the swab

Contact plates

• Contact plates with neutralizers are used

Sampling Method For Coupons


Rinse

Develop a method for rinsing and be consistent

Manual Dislodging

Be as consistent as possible


Disinfectant Neutralizer(s)

Gluteraldehyde Sodium Sulphite, Bisulphite

Chlorine Sodium Thiosulphate

Phenolics Tween (polysorbate)

QAC Lecithin, Polysorbate

Hydrogen peroxide Catalase

TSA and SDA contact plates contain lecithin and polysorbate 80

Letheen Broth contains lecithin and polysorbate 80

DE neutralizing agar and broth contain all above mentioned neutralizers - DE is not used in industry

Neutralization of Disinfectant

Incubation Time

• Bacteria generally up to 48 hours

• Yeast generally up to 72 hours

• Mold until profuse sporulation occurs-use

microscopy to ensure sporulation

Harvesting Organisms

• Avoid scraping agar in case of bacteria and fungi

• Lightly roll wet swab over sporulation of mold to

avoid mycelia

Enumeration

Storage of Harvest

• Bacteria and yeast stored at 2-4°C for 7 days

• Mold and spores stored at 2-4°C for 30 days

Dilutions

• Use Mc. Farland Standards equivalent suspensions

as starting point for dilutions

• 3.0 X 107-8 bacteria in 1 Mc. Farland equivalent

• Use 2 – 4 Mc. Farland Standard equivalent for mold-

determine during dry run

Enumeration

Filtration Method

• This applies to methods where swab recovery, rinse with

buffer or manual dislodging is buffer is used

• To verify that the filtration method and the amount of rinses

remove all disinfectant

• Filter the disinfectant dilutions to be tested, followed by the

number of rinses used in the method (3 X 100mL)

• Filter a suspension of ~100 CFU of test organism

• Incubate and compare to positive control

• Acceptance criteria - Within factor of 2 or 70% of positive

control

Method Validation

Neutralizing Broth

• This method can be applied only when using coupons that are

immersed in a neutralizing broth

• Neutralizing broth cannot be filtered

• Spray or apply disinfectant to coupon

• After contact time place the coupon inside the broth tube

• Inoculate with 102 microorganisms

• Positive Controls of broth without disinfectant

(enumeration may be a challenge)

• Negative control of broth only

• Incubate and compare test with positive control

Method Validation

Contact Plates

• Spray coupons with disinfectant

• Take samples using contact plate

• Inoculate contact plate with ~ 102 CFU of test

organism

• Positive controls without disinfectant

• Negative controls of media plates

Method Validation

• Stored inoculum may drop in viability

• Time ‘0’ Study verifies the inoculum count on the

day of testing

• Time ‘0’ is performed at the end of efficacy when

the same target inoculum is used over days

• Efficacy must be repeated if Time ‘0’ drops too low

(where 3 log reduction cannot be calculated)

Time ‘0’ Study or Inoculum Verification

• Each organism is tested by method of choice in

duplicate or triplicate

• Different surfaces dry or soak disinfectant at

different rates

• 15 minute time point not attainable in practice

• 10 minute time point recommended but difficult to

attain

Efficacy Study

Calculation of Target Inoculum

Calculate target inoculum in the enumeration study

by counting CFU on the plate where the count is less

than 300 CFU and multiplying with the dilution factor

Calculation of Time “0” Inoculum

Calculate Time “0” inoculum by counting CFU on the

plate where the count is less than 300 CFU and

multiplying with the dilution factor

Calculations

Calculation of Log Reduction

The log of the average CFU recovered from the

efficacy study is subtracted from the log of those

recovered on the positive control

Calculations

Test Matrix

• Disinfectants

• Dilutions

• Time points

• Surfaces

• Organisms

Disinfectant Qualification Report

• Enumeration methods summary

• Method Validation summary and results

• Microbiological efficacy results

• Expiry dating results

• Chemical stability results for efficacy and expiry

dating

• Conclusions and discussions

• Deviations


Tabulating results for easy review Organism

• Disinfectant

o RC

o Time Point

o Hard Surface

o Target inoculum

o Time ‘0’ count

o Efficacy count

o Log of efficacy count

o Positive control

o Log of positive control

o Log reduction


• Target inoculum un attainable for certain fungi

• Drop in inoculum

• Disinfectant does not cover organisms of hard

surface due to surface tension

• Variability within replicates

• Data does not represent disinfectant’s capability

Common errors

Challenges in Cleaning and Disinfecting

• Nature of Surface

• Smooth - Surface tension, needs multiple strokes to

cover entire surface

• Porous - Absorbs disinfectant fast, dries out fast,

needs multiple strokes to cover surface

• Soft - Absorbs disinfectant fast, dries fast, needs

multiple strokes to keep wet for contact time

• Hard - Surface tension, needs multiple strokes to

cover entire surface

Cleaning and Disinfection

Aluminum Surface

Nature of Surfaces


Stainless Steel Surface

Nature of Surfaces


Epoxy Coated Surface

Nature of Surfaces


Cleanroom Curtain

Nature of Surfaces

Above picture is courtesy of Veltek Associates

Cleanroom Curtain

Nature of Surfaces


Vinyl Surface

Nature of Surfaces


Points to consider

• Cleanroom surfaces are irregular in nature

• Such surfaces trap residues and contaminants and

makes surfaces more difficult to disinfect

Nature of Surfaces

Procedure for Receiving, Storage, and Preparation

of Disinfectants

Points to consider

• Safety Considerations

• Definitions

• List of all agents used in Manufacturing Faculty

• Procurement, Receipt, and QC Release

Procedure

• Preparation and use

Cleaning Procedures

Procedure for Disinfecting/Sanitizing of

Manufacturing Facility

Points to consider

• Responsibility

• Definitions

• Safety Considerations

• Supplies and storage

• Sequence of Cleaning

• Schedule or Frequency of Cleaning

Disinfection / Sanitization Procedure

Procedure for Disinfecting/Sanitizing of

Manufacturing Facility

Points to consider

• Disinfectants/Sanitizers

• Rotation of Agents

• Cleaning Procedure

• Removal of Residue

• Documentation

• Review

• Training

Disinfection / Sanitization Procedure

Chemical Kill vs Physical Removal

Chemical Kill

• Choice of disinfectants

• Rotation of disinfectants

• Wipe down procedures

• Disinfectant qualification

Physical Removal

• Choice of mops - ergonomic and reaching hard to clean areas

• Choice of mop heads - capacity to hold and dispense

adequate disinfectant to attain contact time

• Choice of adequate wipes to attain contact time and

physically remove particulates and residue

• Disinfection needs both chemical kill and physical

removal

• Surfaces are irregular and differ in time taken to

absorb or dry

• Adequate application of disinfection without

soaking is the key

• Ability of supplies to cover all surfaces evenly

• Ability of supplies facilitate physical removal

• Is contact time a myth?

• Good coverage and maximum contact time=

reduction of contamination

Significance of contact time

Mops and Mop Handles

• Hold and deliver disinfectant

• Reach hard to clean areas

• Physically remove debris and residue

• Ergonomic

• Can be used on all cleanroom surfaces-ceilings,

walls floors, behind equipment

Wipes

Non linting

Remove debris and residue

Cannot promote microbial proliferation

Points to consider

Is contamination removed or added?

Can you access the hard to clean areas?

Can you access all the critical surfaces?

If you can access these surfaces can you apply

adequate amount of disinfectant uniformly?

Can you remove residue buildup?

Do you generate particulates by way of cleaning?

If so, do you know what is the amount of

contamination generated?

Are wipes made of organic or inert materials?

Disinfectant Qualification using Biofilms






What is the measure of cleaning/disinfecting efficacy?

Disinfectant qualification proves the chemical efficacy

and contact time for the desired kill

Environmental monitoring data validates the chemical

kill and physical removal

Also understand the lapse time between and

monitoring

Your disinfectant qualification for (b)(4) and (b)(4) bi-spore

disinfectants documented that the log reduction criteria

(Bacteria ≥ 4, Fungi ≥ 3) was not met when challenged with

multiple organisms in a variety of surfaces. After disinfection,

you recovered Micrococcus luteus on vinyl, (b)(4), stainless

steel, glass, and wall laminate and Enterobacter cloacae,

Rhodococcus sp, Burkholderia cepacia, Pseudomonas aeruginosa,

Methylobacterium mesophilicum and, Acinetobacter lwoffi on

glass. However, your procedures for routine cleaning of the

aseptic manufacturing area continue to require the use of

unqualified disinfectants during days (b)(4) through (b)(4) of

your disinfectant program.

FDA 483 Observations

Consistent with SOP XXX, Requirements for Cleaning/Disinfecting

Procedures in Controlled Areas in that surfaces were not wiped

with a sterile wipe after disinfectant dwell time. There is no

assurance that the disinfectants applied without wiping do not

inhibit growth on RODAC surface samples. Qualification studies

used to demonstrate the ability of [redacted] to support that the

growth of microorganisms is not inhibited by potential exposure

to residual antimicrobial agents found on surfaces following

cleaning did not specify how surfaces were cleaned for the study

(disinfectants allowed to dry on or wiped with sterile wipe after

dwell).


“The disinfectant ___ was replaced by _____ as

cleaning agent in September 1998, however was used

to clean Laminar Airflow unit ____ in April 1999”


“Contact time for disinfectants not documented in

cleaning log”

“Daily cleaning failed to remove objectionable

material”

“End of process failed to remove objectionable

material”

“Operators failed to assure satisfactory room

cleanliness during filling operations”


Also, your environmental monitoring program does not provide

meaningful information on the quality of the aseptic processing

environment and ancillary areas used in the manufacture of [redacted] .

For example, your sampling and testing procedure does not incorporate

the use of positive or negative controls; the frequency of routine

environmental monitoring was changed from [redacted] per DCR 05-678

in February of 2006 without adequate justification or rationale or the

change; and an evaluation of environmental monitoring sampling

locations had not been conducted to assure the locations provide a true

and accurate assessment of microbial flora and in the production areas

they represent locations of microbiological risk to the product.

Furthermore, no evaluation had been performed to assure cleaning

solution residues do not negatively impact environmental monitoring

sampling or testing.


Disinfection, Sterilization, and Preservation; Seymor S. Block

Ed., Lippincott, Williams and Wilkins

PDA Publications

Survey Volume 51, Number 6, Nov-Dec 1997

Current Practices in the Use of Disinfectants within the

Pharmaceutical Industry; Denny and Marsik

Survey Volume 53, Number 3, May-June 1999

Elements of successful disinfection program in the

pharmaceutical environment. Denny, Kopis and Marsik

Some Suggested Reading

Questions??

Documents

Disinfectant Qualification and Cleaning Procedures · PDF fileDisinfectant Qualification and Cleaning Procedures Workshop IVT's Cleaning Validation and Critical Cleaning Processes