Basic’s of Contamination Sources of Contamination Environment Specification Elements of Cleanroom Design and Qualification Definitions Control of Contaminations People, Cleaning, Environment & Material Operation, Monitoring and Control Documents and Records

1. Contamination Control
In Cleanrooms
Dr. A. Amsavel, M.Sc., B.Ed., Ph.D.,
In Cleanrooms
GMP

2. An Overview
 Basic’s of Contamination
 Sources of Contamination
 Environment Specification
 Elements of Cleanroom Design and Qualification Elements of Cleanroom Design and Qualification
 Definitions
 Control of Contaminations
 People, Cleaning, Environment & Material
 Operation, Monitoring and Control
 Documents and Records

3. Contamination Control
Final stage of API as well as drug product is manufactured in the
cleanroom, hence prevention or minimization of Contamination in
cleanroom is necessary
 To ensue the Patient safety
 To ensure quality, purity, safety and efficacy of the products To ensure quality, purity, safety and efficacy of the products
 To ensure GMP and Regulatory Requirement
 Minimize the risk of product defective
 Quality of product should be built into design and systems during manufacturing, NOT
BY TESTING
 Contamination may pose risk to the patients
 Set the systems, practices and procedures to eliminate or minimise the introduction of
contaminant into a product / process.

4. 1. How does the manufacturing environment affect quality,
contamination and cross-contamination?
2. How do we arrive at an optimal environment ?
The following will ensures the Product Quality AndThe following will ensures the Product Quality And
prévention of contamination and cross-contamination.
• Best Clean Room design and maintenance- HVAC
• Prevention / restriction of source of contaminants
• Monitoring and Control procedures
• Effective Cleaning
• Training and practices.

5. What are the possible Contaminants
 Contaminants : Any unwanted substance present in or on a
material or any surface within a Clean Area
Physical Chemical Biological Control/ Removal
Dust Organic Bacteria • Air FiltrationDust Organic
Compounds
Bacteria • Air Filtration
• Thermal
• Disinfectant
• Electro-magnetic
• Electrostatic
discharge
• Electrical radiation
•Cleaning &
operational control
Dirt Inorganic Salts Fungus
Grit Vapour Spore
Fibre Mist Virus
Fly ash,
shoots
Fume / Smoke Human skin
cells

6. Sources of Contamination
Major Sources of Contamination and Controls
# Source Reduce / Removal
1 Personnel Training, Practice, PPEs,
Gowning, Hygiene Equip
ment
10%
Air
flow
5%
Materi
al
5%
2 Environ-
ment
Facility , HVAC design,
Clean room, Air filtration,
Cleaning, Operation and
Maintenance
3 Equipment Cleaning, Maintenance
4 Material Restriction, de-dust/ de-
contamination
People
80%
10%
Contamination

7. Particle Size of Matters in the Air

8. Selection of Filters for HVAC

9. Cleanroom
 Cleanroom Requirement
 To translate the qualitative concept of “clean” to a
quantifiable parameters for environment control
,environment specification / classification is required.,environment specification / classification is required.
 For classification we measure the suspended
contaminant density or numbers of suspended particles
per unit volume. The lower the contaminant density, the
cleaner the environment.

10. Definition
 Cleanroom:[ISO 14644-1:1999, 2.1.1].
Room in which the concentration of airborne particles is controlled,
and which is constructed and used in a manner to minimize the
introduction, generation, and retention of particles inside the room,
and in which other relevant parameters, e.g. temperature, humidity,
and pressure, are controlled as necessaryand pressure, are controlled as necessary
 Changing Room:
Room where people using a cleanroom may change into, or out of,
cleanroom apparels.
 Airlock:
Intermediate room or area that is usually ventilated, and used to
minimise the transfer of airborne contamination from one area to
another

11. Definition
 Contaminant:
Any particulate, molecular, non-particulate and biological entity that
can adversely affect the product or process
 Disinfection:
Removal, destruction or de-activation of micro-organisms on objectsRemoval, destruction or de-activation of micro-organisms on objects
or surfaces
 HEPA: (High Efficiency Particulate Air)
Filter element rated between 85% and 99.995% removal efficiency.
 Cleanliness:
Condition of a product, surface, device, gas, fluid, etc. with a
defined level of contamination
 NOTE: Contamination can be particulate, non-particulate, biological, molecular or of other
consistency

12. Definition
 Filter leakage test:
test performed to confirm that the final filters are properly installed
by verifying that there is absence of bypass
leakage in the installation, and that the filters and the grid system
are free of defects and leaks
 Airflow:
The average airflow velocity, volume and uniformity in a cleanroom
or an installation, as well as to determine air supply volume flow
rate.
 Air Change: A measure of the amount of air moving in or out of a
space because of leakage or mechanical ventilation. One air
change is a volumetric flow of air equal to the cubic content of the
space.

13. Cleanroom Design
 Following parameters should be taken into account while designing
of Cleanroom:
 Room classification (ISO Norms)
 Temperature & Humidity
 Air changes & Make-up Air Air changes & Make-up Air
 Filter grades to meet particles (Viable & non-viable)
 Differential pressure cascade
 Airflow velocity patterns
 Flow patterns (turbulent and uni-directional)
 GMP criteria & critical process requirements
 Noise & Vibration in Duct and Electrostatic discharge
Ref guideline ISO-14644-1,2,& 4 Clean room standard design, and construction

14. Cleanroom Design
Ensure the following are considered while commissioning of
cleanroom;
Identification of the clean zones and size of control zones
Air locks & Change rooms, interlocks, air curtains
Temperature & RH control, dehumidifier,Temperature & RH control, dehumidifier,
Number of AHUs, sélection of filters , Air flow pattern
HEPA, Insulation of Structure & ducts
Effectively balance air quantities in area needing exhaust
Floors, walls, ceilings should be smooth finish and easy to clean.
Drains, coving, area for washing and storage of equipments and
accessories
Note: Recommended pressure differential between two adjacent zones is 15Pa, but pressure
differentials of between 5Pa and 20Pa may be acceptable.

15. Cleanroom Class Limits
ISO
Classifi-
cation
Number
Maximum concentration limits( Particles/m3 of air)
for particles equal to and larger than the considered
sizes shown below
>=
0.1µm
>= 0.2m >= 0.3µm >= 0.5µm >= 1µm >= 5.0µm
ISO Class 1 10 2
Ref : ISO Standard 14644-1
ISO Class 1 10 2
ISO Class 2 100 24 10 4
ISO Class 3 1000 237 102 35 8
ISO Class 4 10000 2370 1020 352 83
ISO Class 5 100000 23700 10200 3520 832 29
ISO Class 6 1000000 237000 102000 35200 8320 293
ISO Class 7 352000 83200 2930
ISO Class 8 3520000 832000 29300
ISO Class 9 35200000 8320000 293000

16. Cleanroom Requirement
ISO Class Air flow (fpm)@
Air Changes
M3/m2 per hour
HEPA Coverage
as % of Ceiling**
1 70 - 100 >750 ** 100
2 70 - 100 >750 ** 100
3 70 - 100 >750 ** 100
4 70 – 100 500 – 600 ** 100
5 70 - 100 225-275 @ 100
6 NA 70-160 # 33 - 40
7 NA 30-70 # 10 - 15
8 NA 10 – 20 # 05 -10
@ ASHRE; # ISO14644-4; ** Industrial standardFor ISO Class-8 :HEPA filters or 95% ASHRE HEPA’s recommended

17. Cleanroom Monitoring
Test Parameter Class
Maximum Time
Interval
Test
Procedure
<= ISO 5 6 Months
> ISO 5 12 Months
Schedule of Tests to Demonstrate Continuing Compliance
Particle Count
Test
ISO 14644-1
Annex A
ISO 14644-2 Specifications for testing and monitoring to prove
continued compliance to ISO-14644-1
> ISO 5 12 Months
Air Pressure
Dif f erence All Classes 12 Months
ISO 14644-1
Annex B5
Airf low All Classes 12 Months
ISO 14644-1
Annex B4
Test Annex A
Test Parameter Class
Maximum Time
Interval
Test
Procedure
Installed Filter
Leakage All Classes 24 Months
ISO 14644-3
Annex B6
Containment
Leakage All Classes 24 Months
ISO 14644-3
Annex B4
Recovery All Classes 24 Months
ISO 14644-3
Annex B13
Airf low
Visualization All Classes 24 Months
ISO 14644-3
Annex B7
Schedule of Additional Optional Tests

18. Qualification of HVAC
Ensure that the following parametres are
considered, tested, qualified and documented during qualification of
cleanroom and HVAC system;
 Differential pressure between rooms
Temperature and %RH level and uniformity test Temperature and %RH level and uniformity test
 Determination of differential pressure on filters
 Determination of air flow velocity
 Measurement of air volume and uniformity – air exchange rate
 Airflow parallelism test
 Determination of airflow patterns
 Determination of room classification (airborne particle count mapping)

19. Qualification of HVAC
Filter installation leak test (challenge test):
 Qualification test shall confirm that HEPA and ULPA filters are
properly installed. There is no by-pass leakage in the installation
(frame, gasket seal, and filter bank framework) and the filters are(frame, gasket seal, and filter bank framework) and the filters are
free of defects and small leaks in the filter medium and frame seal.
 Tests are performed by introducing an aerosol challenge upstream
of the filters and scanning immediatly downstream of the filters and
support frame or sampling in a downstream duct.

20. Qualification of HVAC
 Determination of the recovery time:
This test is not recommended for unidirectional airflows. Perform thet test
that clean room or clean zone is capable of returning to its specified
cleanliness class within a finite time, minimum hold “hold“ time to addrres
the account of power failure, start (recovery), mode change, use of
changing rooms, etc.changing rooms, etc.
The key factors for a successful HVAC qualification are
 The understanding of interfaces beween product purity / characteristic,
process, clean zones, HVAC functions and clean rooms requirements,
 The knowledge concerning general and HVAC specific tests,
 The structured identification of critical functions and operations,
appropriate measures (design, qualification, calibration, and validation
activities) in a documented way.

21. Qualification Documents
 Document the qualification with supported by appropriate
documents.
 Protocols & reports of DQ, IQ, OQ & PQ consists of
 Conceptual documents, URS, Conceptual documents, URS,
 PO, approved diagrams, SAT, FAT
 OQ & PQ, Test parameters with acceptance criteria, test results,
test methods, raw data, supporting data (printouts, electronic
data, visual etc), MOC, calibration certificates, manuals, etc..
 Drawings: Layout, as-built drawings with zone classification, men
and material flow, etc.

22. Operational Controls
Consider the following for effective operational controls and monitoring to
prevent the contaminations.
 HVAC system and environment
 Production Process
 Cleaning & Maintenance Cleaning & Maintenance
 Training & Personnel Practices: Gowning, Clothing & Hygiene etc
 Seasonal Effects – influence of variation in temperature & RH
 Disinfection- type, concentration, rotations
 Non-Product contact Equipment
 Raw materials & Components
 Tools & Utensils

23. Personnel
 Personnel should be trained in a manner that minimizes the possibility of
contamination being generated or transferred or deposited on or into the
product.
 A policy concerning jewelry, cosmetics or similar material to control the
contamination
 Training on the following areas , but not limited to ;
 GMP, Procedure, Processes and Operation, cleaning of area & equipment
 Clean room practices , gowning, aseptic technique, health, Safety risk, and relevant
intervention procedure,
 Also train on the cause and consequences , if it is not followed
 Assess the effectiveness of training
 Validation /verification and Monitoring as required. eg bio-burden In gown, finger etc

24. Personnel
 Establish procedure for Practice good sanitation, personal
hygiene, health habits and operations in the cleanroom.
 Avoid direct contact with intermediates or APIs or drug products
 Instruction to follow cleanliness, use of clean clothing, proper
washing, bathing, nail cut, and use of disinfectant, gowning, change overwashing, bathing, nail cut, and use of disinfectant, gowning, change over
and all clean room practices.
 Use of PPEs, covers for head, face, hands and arms, proper gowning to
reduce the exposure and contamination. Eg dead skin cells , hair
fall, sweat, etc
 Personal and other items shall be restricted to the cleanroom
 Cleaning of equipment as per procedure and ensure the line clearance.
 Ensure the above is followed always; even no one is watching…

25. Definition: Type of Disinfectants
 Antiseptic: An agent that inhibits or destroys
microorganisms on living tissue including skin, oral cavities,
and open wounds.
 Disinfectant: A chemical or physical agent used on
inanimate surfaces and objects to destroy or removeinanimate surfaces and objects to destroy or remove
infectious fungi, viruses, and bacteria, but not necessarily
their spores.
 Cleaning Agent: An agent for the removal from facility and
equipment surfaces of product residues that may inactivate
sanitizing agents or harbor microorganisms.

26. Definition: Type of Disinfectants
 Sanitizing Agent: An agent for reducing, on inanimate
surfaces, the number of all forms of microbial life including
fungi, viruses, and bacteria.
 Sporicidal Agent: An agent that destroys bacterial andSporicidal Agent: An agent that destroys bacterial and
fungal spores when used in sufficient concentration for a
specified contact time. It is expected to kill all vegetative
microorganisms.
 Sterilant: An agent that destroys all forms of microbial life
including fungi, viruses, and all forms of bacteria and their
spores. Sterilants are liquid or vapor-phase agents

27. Methods of Cleaning
Vacuum cleaning:
 Use HEPA/ULPA installed vacuum cleaners to remove larger particles and
other debris.
 Use in unidirectional strokes to minimize air turbulence.
 Also can be used to removing excess water and suspended particles and
faster drying after wet mopping.
Wet cleaning
 Mopping is an effective method in gross or intermediate cleaning for removing
particulate & residues.
 Establish procedure and clear instruction for mopping;
 Disinfectant preparation, type of mops, no. of buckets, instruction for stroking,
overlapping, direction of mopping, rinsing & squeezing with water, frequency of
changing solutions

28. Methods of Cleaning
Wet cleaning
 Scrubbing: Use machine or manual cleaning to remove stains or
heavily soiled areas / equipment surface by scrubbing and then
clean by mopping or wiping.
 Damp cleaning
Wiping techniques are used in most phases of cleaning. Wiper
should be dampened with the appropriate cleaning solvent/solution.
Wiping should always be done in unidirectional, overlapping
strokes, proceeding from most critical to least critical areas.

29. Cleaning of Cleanrooms
 Determine the procedure for cleaning for its Suitablility, efficacy,
limitations of disinfectants and procedures should be
assessed,
 Establish the cleaning procedure, for preparation or disinfectant,
work sequence, contact time . Alos define the cleaning surfaces,work sequence, contact time . Alos define the cleaning surfaces,
method of cleaning and frequency of cleaning
 Cleaning of Surfaces:
Identify the surfaces according to criticality to the product or process and
establish the cleaning techniques to the required level of cleanliness.
Validate the cleaning & verify the efficacy of disinfectant.
 Floors, Walls, doors, grills, windows and vertical surfaces:
Clean the upstream surfaces during at-rest state. Remove products from
the area or cover the items.

30. Cleaning of Cleanrooms
 Ceilings, diffusers and lamp fixtures & tables :
Wipe thoroughly as per damp the wiper in cleaning solution as per frequency. Clean
after repair or replacement. Eg. Bulbs
 Chairs, furniture and ladders:
Wipe these surfaces from top to bottom. Include cushions, supports, and wheels if
appropriate.appropriate.
 Cross-over benches, garment and supply cabinets, lockers:
periodically empty and clean interiors and surfaces by wiping
 Rubbish bins and containers
Rubbish bins and containers can be lined with plastic bags to remove the refuse and
protect container surfaces. All bins should be removed to general, non-critical areas
and remove the rubbish.
 Cleanroom mats and sticky flooring:
Cleanroom mats and sticky flooring should be cleaned by wet mop or maintained on
a regular basis. Use vacuum cleaner as required.

31. Cleaning Frequency
Schedule / Frequency for cleaning:
Most routine cleaning operations should be performed as per established frequency
at regular basis. Other cleaning may require on certain frequency or when needed.
 Daily cleaning (once in 24hours):
Clean room floors, walls, doors etc. Air locks, changing & operational areas should
be cleaned at least daily. Vacuuming and or mopping floors, and wiping the surfacesbe cleaned at least daily. Vacuuming and or mopping floors, and wiping the surfaces
 Periodic cleaning
Surfaces not cleaned on a daily basis should be cleaned periodically like
weekly, bimonthly or monthly etc eg storage areas, service areas, pipes and fittings.
 Intensive cleaning efforts should be taken after holidays or planned shutdowns

32. Non-routine Cleaning
 Cleaning after modification, construction or maintenance
Effective cleaning after construction is essential to control and
eliminate contamination sources.
 Cleaning during emergency situations
Procedures should be instituted for cleaning in the case of a gross
contamination event.
 Special cleaning during shut down
 environmental incident like major equipment, utility failures or
spills contamination due to failure or ineffective routine cleaning

33. Cleaning of Area & Equipment
Surface Cleaning agent Frequency
Floors, Spill areas
Around drains
Path men movement
Access ports, passbox
Use Dettol, lysol or savlon
as Disinfectant ate
recommended strength.
Surfactant if required
Daily, change over
Wipe or mob
,
Doors& Handles
Sinks, Benches, tools
Trash containers
Surfactant if required
( solvent / water for spill
area)
Walls, ceilings, grills
Trolley, pallets
Disinfectant
Surfactant if required
Weekly
Wipe or mob
Equipment accessories
Pipelines, sampling tools
Utensils,
Water /Solvent
Surfactant if required
Batch to batch/
change over.
As per procedure

34. Microbial Contaminants and Control
Microbial
Contaminants
Sources / enhances Control
Human Skin Flora
Staphylococcus
Propionibacterium acnes
Operator contamination :
 Gowning Material
 Traced back to one operator
 Skin infection
 Non-sterile drug product
 Gowning control
 Proper disinfectant
 Rotation & Rinsing
 Cleaning and
sanitization Non-sterile drug product sanitization
 Cleaning Validation of
Floors, Walls &
Equipment
Fungal Spores
Penicillium
Aspergillus
Cladosporium etc
Items brought to Cleanroom
 Bags, Boxes, Intervention Equipment,
Pallets, Pallet Jacks, Shoes, Shoe
Covers
 Raw Materials, high RH &
temperature
Bacterial Spores
Bacillus cereus Bacillus
circulansgroup
Paenibacillus glucanolyticus
 Cleanroom Shoe Cover
 Process Vessels
 Raw Material

35. Contaminants from Equipment
Possible Contaminants from Equipment
 Product residues
 Previous API’s
 Intermediates
 Side products & degradants
 Raw materials / Solvent
 Cleaning agent residues and breakdown
 Airborne matter
 Lubricants, ancillary material
 Possibly Bacteria, mould and pyrogens on long holding.

36. Cleaning of Equipment
 Equipment and utensils shall be cleaned, maintained and sanitized at
appropriate intervals to prevent contamination that would alter the
safety, identity, strength, quality or purity of the drug product. (21 CFR
§211.67)
 Establish Cleaning / sanitization procedures for cleaning of equipment. Establish Cleaning / sanitization procedures for cleaning of equipment.
 Validate the cleaning procedures and Analytical methods with recovery
used for testing residue,
 Sampling procedures and Sampling locations (clearly defined)
 Acceptance criteria and rationale
 Frequency - for Between batches, product changeover
 Periodic re-evaluation and revalidation
 Routine monitoring equipment/ prior to start up

37. Monitoring & Testing of Cleaning
 Monitoring cleaning effectiveness and testing
Monitoring and testing of cleanliness have to be determined and
established to ensure the effectiveness and prevention of contamination.
 Routine visual inspection for surface cleanliness ie absence of strain,
residue, soiling etcresidue, soiling etc
 Method of checks: Use high-intensity white light for visual check, use wet
wipes by swap the surfaces and check for colouration and other methods
like tape lift method and surface particle detector method.
 Bio-burden: Use Contact plates or Surface swabbing

38. Monitoring of Cleanrooms
 Monitor the following periodically & trend with alert and Action Levels.
Deviation reporting in case of out of limit
 Recommended pressure differential between two adjacent zones is
15Pa, but pressure differentials of between 5Pa and 20Pa may be
acceptable.
 Temperature 25°C for controlled room temperature or as required and
%RH based on the requirement
 Viable & Non-viable particles- ISO-8 :

Viable particles cfu/m3 Non-viable / Bio-burden
0.5µ < 3,520,000
1µ < 832,000
5µ < 29,300
Air sample: <200 CFU/m3 (EU)
<100 CFU/m3 (USP-1116)
Settle plate: 100 CFU 90 mm/4hrs (EU).
Not specified in USP

39. Material
Source of contamination:
 Product/ process materials, Packaging materials, Bags, drums,
Intervention Equipment, Pallets, Pallet Jacks are handled at clean room
Action to prevent contamination:
 Take all precaution and procedure that material does not compromise the
cleanliness of the product or process
 Restrict the transfer of unwanted items
 Minimise the quantities of materials stored in the cleanroom,
 Protective storage or isolation., where necessary
 Cover the items before transfer and De-dust/ de-contamination as required.
 Collect all used and waste materials and remove frequently

40. Documents and Records
Establish procedure operation and maintenance . SOP for routine and
non-routine monitoring activities in cleanrooms.
 Pressure differential between rooms between filters
 Temperature & RH, recorder or time & frequency for manual recording
 Cleaning and Sanitization: Records for cleaning of floors other surface
areas including disinfectant preparation & rotation,
 Log & record for equipment use, repair, replacements, maintenance
followed by cleaning
 Calibration of monitoring devices
 Personal hygiene- Verification /self –declaration

41. Docuements and Records
 SOP for environment monitoring;
 Particulate count
 Bio-burden air filtration method & plate exposure method
 Sampling details, how many. where and how?
 Sample Site Maps ,
 Test method, Incubation Temperature, Test Results, date of results
 Microbial Identifications if required
 Environmental Trending- control chart with alert and action Levels,
 Deviation Reporting, investigation and CAPA procedures & records
 Quality Review

42. Reference
 ISO/DIS 14644 Cleanrooms and associated controlled
environments:
 Part 1: Classification of air cleanliness
 Part 2: Specifications for testing and monitoring to prove
continued compliance with ISO 14644-1continued compliance with ISO 14644-1
 Part 4: Design, construction and start up
 Part 5: Operations
 WHO TRS- 961 Annex-5: Heating, ventilation and air-conditioning
systems for non-sterile pharmaceutical
dosage forms

43. Thank youThank you
Dr. A. Amsavel
Contact: