This session from the Institute of Validation Technology's Contamination and Control Week discusses regulatory expectations and industry drivers for aseptic cleaning and environmental monitoring, regulatory expectations for cleanrooms, and current FDA and EU expectations during inspection of sterile and aseptic operations.

1. Session 5 • Understand the Evolving
Regulations for Aseptic Cleaning and
Environmental Monitoring
IVT’s Contamination Control Week
June 25-27, 2012
Boston, Massachusetts

2. For more information on aseptic cleaning and environmental monitoring,
please visit www.ivtnetwork.com.
Use the promo code SLIDE1 for a 10% discount on a membership!

3. CONTACT INFORMATION
for Course Leader:
David Vincent, CEO
Validation Technologies, Inc.
San Diego, CA
Office: 800-930-9222
Fax: 858-638-5532
Email: davidv@validation.org

4. Topics Addressed
§ Regulatory requirements & expectations
§ Potential sources and types of microbial
contamination
§ Implementation of microbial control
measures
§ Disinfection/Sanitization
§ Environmental monitoring

5. Part 1:
Microbiological control is a
regulatory requirement

6. CFR 21- 211.113 (a)
Control of Microbiological Contamination Microbial
“Appropriate written procedures, designed to
prevent objectionable organisms in drug products
not required to be sterile, shall be established.”
§ Control of bioburden
§ Absence of objectionable organisms

7. WHAT IS
CONTAMINATION?
§ There are two types of contamination.
Non-viable (Particulate)
– Air Filtration
– Materials shedding
Viable (Microbiological or Bioburden)
– Molds
– Bacteria
– Viruses
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8. Viable Contaminants
§ Viruses
§ TSEs (transmissible Spongiform encephalopathies
§ Fungi
§ Bacterial
§ Pyrogens (Endotoxin)
§ Mycoplasma
§ Biological Based Components (DNA, foreign proteins,
etc.)

9. NONVIABLE
CONTAMINATION
§ This is any type of contamination that is not
viable. It is usually filterable. For example:
§ Dirt, sand, powders, etc.
§ Particulate
§ Debris
§ Molding flash
§ Chemical contamination such as crystals, etc.
§ Oils and Mold Release agents
§ Insect parts
§ Controlled room classifications are defined by
particulate contamination sizes and numbers.
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10. What is Contamination
Control?
§ Contamination control is not simply a task or
function. It is a science and applied
technology that interacts continuously with all
products, processes, materials, equipment,
and personnel entering the manufacturing
areas.

11. Microbiological
Control
§ Identify possible sources of
contamination
§ Identify possible types of contamination
likely to occur
§ Implement and validate/qualify
preventative and control measures

12. Possible Sources of
Contamination
§ Raw Materials/Excipients
§ Equipment and Process
§ Environment/Facility
§ Personnel

13. Raw Materials/Excipients
§ Bioburden in raw materials and excipients
v natural vs synthetic products
v water activity level
§ Bioburden in water systems
v potable water, purified water, WFI
v biofilm

14. Equipment and Process
§ Process and Equipment § Carbon filters/DI
Design resins/membrane
§ Material/Surface filters
§ Unprotected storage tanks § Sampling/transfer
hoses
§ Back flow
§ Valves
§ Perforated heat exchangers
§ Dead legs

15. Facility
§ Facility design
§ Air handling system
§ Construction materials
§ Finish materials

16. Personnel
§ Major potential source of microbial
contamination
v Exposed skin (flakes, oils)
v Hair
v Clothing fibers
v Dirt
v Cosmetics
v Tobacco smoke
v Behavior

17. Part 2:
Regulatory Expectations
Related to Cleanroom Operations

18. Facilities: Cleanroom
Classification
FS209 ISO 14644-1 Viable Ave Airflow
Cleanroom Cleanroom ≥0.5um Microbes Velocity Air
classification classification particles/m3 (cfu/m3) (fpm) changes/hr
100,000 8 3,520,000 100 5-10 5-48
10,000 7 352,000 10 10-15 60-90
1000 6 35,200 7 25-40 150-240
100 5 3,520 1 40-80 240-480
§ a- All classifications based on data measured in the vicinity of exposed materials/articles during periods of
activity.
§ b- ISO 14644-1 designations provide uniform particle concentration values for cleanrooms in multiple
industries. An ISO 5 particle concentration is equal to Class 100 and approximately equals EU Grade A.
§ c- Values represent recommended levels of environmental quality. You may find it appropriate to establish
alternate microbiological action levels due to the nature of the operation or method of analysis.
§ d- The additional use of settling plates is optional.
§ e- Samples from Class 100 (ISO 5) environments should normally yield no microbiological contaminants.

19. Equipment
and Process
§ CFR 211.63, “Equipment used in the manufacture,
processing, packing, or holding of a drug product shall
be of appropriate design, adequate size and suitably
located to facilitate operations for its intended use and
for its cleaning and maintenance.”
§ Q7A, 5.15, “Closed or contained equipment should be
used whenever appropriate. Where open equipment is
used, or equipment is opened, appropriate precautions
should be taken to minimize the risk of contamination.”

20. Equipment and Process
§ Equipment
v easily disassembled and cleaned
v use sanitary fittings and valves/avoid dead legs
v product contact surfaces resistant to corrosion
v store cleaned equipment in a sanitary condition

21. Equipment
and Process
§ Equipment flow and location
v no cross flow of clean and dirty and contaminated
equipment/materials
§ Process
v evaluate each step in a process for the potential of
microbial contamination
v use closed systems whenever possible
v open operations should be contained within HEPA
stations

22. Environment & Facility
§ Flooring and wall materials must be easily
cleaned
§ Surfaces must be resistant to chemicals,
abrasion, and flaking
§ Ensure concave coving at the junction of walls
and floors
§ Floors must be sloped toward the drain
§ Clean/sanitize floors daily
§ Cover floor drains when not in use
§ Avoid standing water

23. Environment & Facility
§ Evaluate traffic patterns and room capacity
§ Production and packaging areas need to be
under positive pressure
§ Establish and monitor operating parameters and
environmental controls
§ Use HEPA filtration systems
§ Clean up spills immediately
§ Cover air ducts, pipes, and light fixtures
§ Avoid build-up of dust and moisture
accumulation

24. Part 3:
Cleaning and Disinfection

25. Disinfection/Sanitization
Program
• Cleaning/Sanitization is a regulatory requirement
• Cleaning should not be confused with disinfection
• A cleaning program is not complete without
contamination control

26. Disinfection/Sanitization
Program
§ CFR211.56 (b)
“ There shall be written procedures assigning
responsibilities for sanitation and describing in
sufficient detail the cleaning schedules, methods,
equipment, and materials to be used in cleaning
the buildings and facilities; such written
procedures shall be followed.”

27. Preparation and Use of
Disinfectants
Preparation and Storage
• EC Guide, Annex 1,
“Disinfectants and detergents should be monitored for
microbial contamination: dilutions should be kept in
previously cleaned containers and should only be
stored for defined periods unless sterilised.
Disinfectants and detergents used in grade A and B
areas should be sterile prior to use.”

28. Current regulatory expectations
for use of disinfectants
Rotation
• EC Guide, Annex 1, 37
“Where disinfectants are used, more than one type
should be employed. Monitoring should be undertaken
regularly in order to detect the development of resistant
strains.”
• ISO Recommendation
• Industry Practice

29. Product Classification
§ Sanitizers
v proper use results in 99.9% (3 log) reduction of bacteria
v cannot handle soil; use on pre-cleaned surfaces
§ Disinfectants
v proper use results in 100% (> 4 log) reduction of bacteria and
yeast. Limited reduction for mold.
v most have surfactants/cleaning ability
§ Sterilants
v proper use results in 100% (> 6-7 log) reduction of all
microorganisms, including bacterial spores.
v require application on pre-cleaned surfaces

30. Disinfection efficacy
§ Suitability, efficacy & limitations of
disinfectant agents and procedures should
be assessed.
§ The disinfection program should include the
use of a sporicidal agent used according to a
written schedule and when environmental
data suggests presence of spore forming
agents (Baccilus spp.).

31. Disinfectant Efficiency
Most Resistant
Endospores
Mycobacteria
Fungal Spores
Small Non-enveloped viruses (polio, rotavirus, rabies)
Vegetative Fungal Cells
Enveloped Viruses (Herpes, Hepatitis B, Hepatitis C, HIV)
Vegetative Bacteria
Least Resistant

32. Equipment and bacteria
Even seemingly smooth
surfaces can harbor
bacteria !
Scanning electron micrograph of Listeria monocytogenes forming a biofilm
in soy on a stainless steel chip. Courtesy of Professor Amy Wong.

33. Cleaning/Sanitization
Program
An effective cleaning and sanitization
process is attained by:
§ Treatment with strong acids and bases
§ Use of high velocity hot water or steam
§ Use of detergents and/or sanitizers
§ Rinsing with high quality water (Purified or WFI)
§ Use of solvent rinses
§ Drying at elevated temperatures

34. Disinfectant Selection
§ Population and types of organisms
§ Spectrum of activity of disinfectant
§ EPA registrations
§ Method of application
§ Contact time
§ Nature and surface to be disinfected
§ Compatibility of surface with disinfectant

35. Disinfectant Selection
§ Corrosiveness of disinfectant
§ Organic compounds present on surface
§ Operator safety
§ Compatibility with cleaning agents
§ Planned rotation of disinfectants
§ Needed steps to avoid contamination of
pharmaceutical products by disinfectant
§ Need for residual bactericidal activity

36. Disinfectant Selection -
Other Factors to Consider
§ Quality, sterility, and stability of product
§ Ease of application
§ Cleaning ability
§ Supporting vendor documentation
§ Cost and availability
§ Factors that may affect performance (temp.,
organic matter, contact time, pH, etc…)
§ Regulatory expectations/regulations

37. Current regulatory expectations
for use of disinfectants
§ Disinfectants Qualification
v Studies to evaluate the effectiveness of the
disinfectants as they are used and prepared
v Studies to evaluate the storage conditions of
disinfectants for possible loss of efficacy
v Use of environmental isolates in the qualification
studies
v Neutralization studies

38. Disinfectant Qualification
- Studies
In-Situ
§ Use actually cleaning procedures
§ Monitoring pre and post cleaning at worst-case conditions/
document activities
§ Increased number of sample sites
§ Compare EM data before & after
In-Vitro
§ Surface Tests (use of coupons or pieces of equipment/
material)
§ Carrier Tests (AOAC method)
§ Use-Dilution Tests (modified AET )

39. Disinfectant Qualification -
Studies
Typical Test Organisms
§ Escherichia coli - ATCC 8739
§ Pseudomonas aeruginosa - ATCC 9027
§ Staphylococcus aureus - ATCC 6538
§ Bacillus subtilis - ATCC 6633
§ Candida albicans - ATCC 10231
§ Aspergillus niger - ATCC 16404
§ Environmental isolates

40. Disinfectant Qualification
Surface Test
§ Preferred method by inspectors
§ No guidance document available
§ Must select types of surfaces (coupons)
§ Variable microbial challenge level
§ Quantitative method
§ Must select contact times (1-5-10 minutes)
§ Variable test conditions (wet vs dry)
§ Sampling/recovery technique (swab, rinse, contact
plate)
§ Need neutralization studies

41. Disinfectant Qualification -
User’s Approach/Decisions
§ Type of agent and materials/surfaces and sites
to be evaluated.
§ Testing protocol (In-vitro vs In-situ)
§ Types of challenge organisms
§ Contact times
§ Sampling methods (swab, rinse, or contact
plate)

42. Disinfectant Qualification -
Acceptance Criteria
§ Surface Tests
v ≥ 3-log reduction vegetative bacteria and fungi
v ≥ 2-log reduction spore-forming bacteria
§ Use-Dilution Tests
v ≥ 4-log reduction (disinfectant properties)
v ≥ 6-log reduction (sporicidal properties)
§ Carrier Test
v AOAC criteria
§ In-situ Test
v counts not to exceed alert levels

43. Part 4:
Environmental Monitoring
Program

44. Monitoring and
Trending
§ Establish an EM program
§ Employee training and monitoring
§ Enforce personal hygiene
§ Enforce cGMPs
§ Schedule regular PMs
§ Product bioburden testing/trending
§ Re-qualification vs on-going verification of
cleaning (bioburden and endotoxin)
§ Change control

45. EM Program
§ Regulatory requirement
§ Must be tailored to a facility
§ Must be manageable and efficient
§ Must be well documented
§ Must be defendable
§ Must not add possible sources of contamination
into the environment
§ Do not rely solely on EM data for assurance of
product quality

46. EM Program
EM is a tool to assist a company
manage the microbiological
control program

47. EM Program
Choice of Equipment
§ Active vs passive
§ Air and surface monitoring devices
§ Detection of potential problems must be
done in a timely manner
§ Sampling activities should not contribute to
contamination of the process

48. EM Program
Test Methods
§ EM procedures must be qualified
§ Use same methods used during qualification studies
§ Viable and non-viable monitoring
§ Use of specific media for fungi: is it really needed?
§ Monitoring for anaerobes: is it really needed?
§ Levels are established based on industry guidelines
and/or historical data

49. EM Program
Frequency and Sites
§ Depends on area being monitored
§ More intensive monitoring for “cleaner”
areas
§ Usually frequency is reduced once
qualification is complete
§ Trend towards continuous monitoring
§ Site selection: depends on room design,
activities, equipment and personnel flow

50. EM Program
Data Management
§ Results are retrospective
§ Useful information can be obtained by trending
data
§ Evaluate data based on Alert and Action Levels
§ Identify isolates
§ Frequent isolates should be maintained and used
in challenge studies

51. EM Program - Hot Topics
§ EM data linked to batch release
EC Annex 1, 5 “Where aseptic operations are
performed, monitoring should be frequent…
Results from monitoring should be considered
when reviewing batch documentation for finish
product release…”
§ EM requirements for non-sterile operations
§ Use of PAT and Rapid Methods

52. EM Program – Focus of
Regulatory Inspections
§ Create presentation describing program
v area classifications
v methods
v frequency
v rationale for choice of sites
v rationale for setting alert and action levels
v describe data management and trending
§ Create trend reports for each area
v tabular and graphical format
v trend by values and organisms
§ Create executive summary reports
v address deviations, adverse trends, CAPA

53. Thank You!!
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