STANDARD OPERATING PROCEDURE

Department: Quality Assurance SOP No.:

Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

1.0 OBJECTIVE:
To lay down a Procedure to define the Standard Operating Procedure for “Qualification of Air
filtration system”.

2.0 SCOPE:
2.1 This SOP is applicable for Qualification of Air filtration system at ……………….
2.2 This SOP is applicable for Validation of HVAC unit.
2.3 This SOP is also applicable to Validation of HEPA unit installed in the Equipment such as (Laminar
air flow units, sterile garments cabinets, Dynamic pass box, but not limited to).

3.0 RESPONSIBILITY :
3.1 Quality Assurance shall be responsible for:
3.1.1 Executive QA shall be responsible for preparation of validation protocol.
3.1.2 Head QA shall be responsible for review of validation protocol & Report
3.1.3 Executive QA shall be responsible for review of calibration certificates of measuring
Instruments received from external agency.
3.1.4 Executive QA shall provide the guidance during execution of validation activity.
3.1.5 Preparation of validation report after execution of validation activity.
3.2 Head-EG/designee shall be responsible for:
3.2.1 Arrange the External Agency to performed validation activity
3.2.2 Review of validation protocol and report of Air Filtration systems.
3.2.3 Co-ordination with external agency and user departments to execute the validation activity.
3.2.4 Ensuring the Online data recording required for the report compilation during validation.
3.3 Head/Designee MF, WH, QC shall be responsible for:
3.3.1 Concerned department head shall be responsible for review of validation protocol and report
of Air Filtration system.
3.3.2 To provide support in execution of validation activity.
3.4 Head Quality/Designee shall be responsible for approval of validation protocol and report.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

4.0 ACCOUNTABILITY:
Head-Quality shall be accountable for ensuring over all compliance of this Standard Operating
Procedure.

5.0 DEFINITIONS:
5.1 As-built: Condition where the installation is complete with all services connected and functioning but
with no production equipment, materials, or personnel present.
5.2 At-rest: Condition where the installation is complete with equipment installed and operating in a
manner agreed upon by the customer and supplier, but with no personnel present.
5.3 Operational: Condition where the installation is functioning in the specified manner, with the
specified number of personnel present and working (Simulated for working) in the manner agreed
upon.
5.4 Clean zone: Dedicated space 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 zone, and in which other relevant parameters, e.g. temperature, humidity, and pressure, are
controlled as necessary.
5.5 Clean room: 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 necessary.
5.6 Particle : Solid or liquid object which, for the purposes of classification of air cleanliness, falls within
a cumulative distribution that is based upon a threshold (lower limit) size in the range from 0.1 mm to
5 mm.
5.7 Particle size: Diameter of a sphere that produces a response, by a given particle-sizing instrument that
is equivalent to the response produced by the particle being measured.
NOTE: For discrete-particle-counting, light-scattering instruments, the equivalent optical diameter is
used.
5.7.1 Aerosol generator: instrument capable of generating particulate matter having appropriate
size range (e.g. 0.05 μm to 2 μm) at a constant concentration, which may be produced by
thermal, hydraulic, pneumatic, acoustic or electrostatic.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

5.7.2 Test aerosol: Gaseous suspension of solid and/or liquid particles with known and controlled
size distribution and concentration.
5.7.3 Ultrafine particle: Particle with an equivalent diameter less than 0.1 mm.
5.7.4 Macro particle: Particle with an equivalent diameter greater than 5 mm.

6.0 PROCEDURE:
6.1 Validation of the Air filtration system includes following test parameters:
 Air velocity (ACPH) measurement
 HEPA Filter Integrity Test
 Air flow pattern test (Smoke test)
 Non-viable particulate counts measurement
 Recovery test
 Pressure Differential Monitoring
 Temperature and Relative Humidity monitoring
 Viable Particle count (Active & Passive air sampling)
6.2 All the test shall be done as per given Annexure-I and accordingly protocol & report shall be prepared.
6.3 Air velocity (ACPH) measurement:
6.3.1 The objective of this test is to demonstrate that:
6.3.1.1 The air system is balanced and capable of delivering sufficient air velocities to maintain a
minimum cross sectional velocities as per design.
6.3.1.2 To demonstrate that the number of air changes are equal to or more than the designed number
of air changes and should not be less than designed number of air changes
6.3.1.3 The air velocity and air changes test shall be performed by qualified and trained person only.
6.3.1.4 Measuring instrument being used shall be calibrated with reference to NPL traceable
reference standard.
6.3.1.5 Measure the air velocity by taking five reading at approx 6" distance, (i.e. at four corners and
one at centre), with calibrated anemometer below the grill / diffuser by selecting anemometer
in feet per minute.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

V1 V2 Grill/Diffuser

* Location indicated for

V5air velocity.

V3 V4

6.3.1.6 Calculate the average velocity of each grill present in room as per below mention formula:

Average Velocity = V1+V2+V3+V4+V5

5
6.3.1.7 Calculate the CFM of each grill by as per below mention formula:
CFM = Average velocity X Face area of grill (Length X width)
6.3.1.8 Calculate the room volume in Cubic Feet as per below mention formula.
Volume= Length (in ft) x width (in ft) x Height (in ft)
6.3.1.9 Calculate the Air change per hour of particular cubical by using below formula.

Air changes per hour = Total CFM of the room x 60

Volume of the room in ft3

6.3.1.10 Acceptance criteria:

6.3.1.10.1 Air Changes per hours for a clean rooms as per given below
ISO class ISO-6 ISO-7 ISO-8
Acceptance criteria
NLT 60 NLT 40 NLT 20
for ACPH

6.3.1.10.2 Average velocity should be 90±20% feet/min for equipment like Pass box, sterile
cabinet, LAF, dispensing & Sampling booth.

6.4 HEPA Filter Integrity Test:

6.4.1 The objective of HEPA filter integrity test is to ensure that the HEPA filters installed are not
damaged during, transit, installation or operation and fixed properly inappropriate place such
that there is no leakage in the filter and periphery.
6.4.2 The filter integrity test shall be performed by qualified and trained person only.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

6.4.3 Measuring and testing instrument being used shall be calibrated with reference to NPL
traceable reference standard.
6.4.4 Filter testing shall be performed after measuring the air velocities of that supply.
6.4.5 Poly Alpha Olefin (PAO) shall be used for the generation of particles/fumes. MSDS of the
same shall be provided by the external agency.
6.4.6 Position the aerosol generator such that aerosol is produced into the upstream of the
subjected HEPA filter.
6.4.7 Provide Compressed air to the fume generator at pressure of 1.5-2 kg/cm2.
6.4.8 The fume concentration shall be in between 20 mg/m3 to 80 mg/m3. (Calibrate upstream
conc. up to 100% scale).
6.4.9 Actual fume concentration shown in aerosol photometer shall be recorded.
6.4.10 Select the downstream scanning mode for the control panel. Scan the subjected HEPA filter
by holding the photometer probe approximately 1 inch from the filter face and passing the
probe in the slightly overlapping strokes, at rate of NMT 15 cm2/s, so that entire filter face is
sampled.
6.4.11 Similarly scan the entire periphery of the filter along the bond between the filter medium and
frame and along all other joints in the installation through which leakage might by pass.
6.4.12 Acceptance Criteria: The leakage through HEPA filter should not be more than 0.01%.
6.5 Non-Viable Particle count measurement:
6.5.1 The objective of this test is to demonstrate that the critical work stations within the clean
rooms / equipments comply with their designed conditions or the cleanliness class as per the
ISO standard 14644-1 & EU–cGMP.
6.5.2 The particle count test shall be performed by qualified and trained person only.
6.5.3 Measuring instrument being used shall be calibrated with reference to NPL traceable
reference standard.
6.5.4 Particle count shall be performed at rest condition & in operation condition.
6.5.5 Operation condition shall be simulated during the particle count sampling if no such activity
is performing in area.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

6.5.6 Ensure that the sampling locations are evenly distributed throughout the area of the clean
room.
6.5.7 Selection of sampling location for non viable particle count with justification shall be
prepared as Annexure- III and also Schematic diagram of the sampling location shall be
prepared as per annexure-IV.
6.5.8 Minimum sampling time for each location shall be 1 minute. The sampling probe shall be
positioned at working level and air flow probe shall be directed vertically upward (Take
sampling volume as per ISO 14644-1).
6.5.9 Switch “ON” the particle counter and set the particle size channel at 0.5 micron and 5 micron
and set sampling time as per sample volume calculation (Take sampling volume as per ISO
14644-1).
6.5.10 Hold the airborne particle counter and then take print of test.
6.5.11 While taking the room particle count, the isokinetic probe shall be positioned at the height of
working activity (30” to 40”) above the floor.
6.5.12 Minimum 3 samples shall be taken in case of low volume equipments i.e. pass box, sterile
cabinets etc.
6.5.13 Acceptance Criteria: All the clean rooms should comply as per the designed Class in rest
and in operation condition.

Limit for non viable particle count monitoring for clean room.
S.No. EU -cGMP Acceptance
Acceptance criteria
Particle size criteria
(At Rest)
(In operation)
≥ 0.5 µ Particle 3520 3520
1. Grade- A
≥ 5.0 µ Particle 20 20
≥ 0.5 µ Particle 3520 352000
2. Grade- B
≥ 5.0 µ Particle 29 2,900
≥ 0.5 µ Particle 352000 3520000
3. Grade- C
≥ 5.0 µ Particle 2,900 29,000
≥ 0.5 µ Particle 3520000 ND
4. Grade- D
≥ 5.0 µ Particle 29,000 ND

6.6 Recovery Test

6.6.1 For Non-Viable particle count:
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

6.6.1.1 The objective of the recovery test is to determine the amount of time that is necessary for
clean rooms or clean spaces or equipment and their systems to reach a specified steady state
cleanliness level after a brief particle generation event within the clean space.
6.6.1.2 The recovery test shall be performed by qualified and trained person only.
6.6.1.3 Measuring instrument being used shall be calibrated with reference to NPL traceable
reference standard.
6.6.1.4 Locate the particle counter in the highest particle count location at rest condition identified in
particle count test and take the reading before switch off the equipments / HVC of the room.
6.6.1.5 Immediate Switch off Equipment/ HVC of the room under test and adjacent rooms.
6.6.1.6 Generate the particles 100 times multiply with the normal limit with the help of Aerosol
generator.
6.6.1.7 Take particle count reading in HVC/Equipment OFF condition and when limit exceed the
100 times then turn on the HVC/Equipment for ensuring the recovery of Particle count
(beyond the limit as per the designed ISO class).
6.6.1.8 Record the time taken for particles to drop from initial to the base line level obtained in
unmanned conditions.
6.6.1.9 Acceptance Criteria: Recovery time shall be established in order to ascertain the time
required to switch on the air handling device prior to start of work or not more than 15
minutes.

7.0 Air flow pattern test (Smoke test):

7.1.1 The objective of this test is to ensure that the air flow pattern in the classified areas is uniform,
unidirectional and is evenly distributed; there should not be any turbulence in the air.
7.1.2 Air flow pattern test shall be done as per below mentioned procedure.
7.1.3 The Air flow pattern test shall be performed by trained and qualified person only.
7.1.4 This test shall be carried out by using fog generator used to generate aerosols (mist), utilizing
phase transition between gases to liquid by cooling steam boiled DI water.
7.1.5 Place the probe near to supply air terminal and return riser and record the air flow pattern with
the help of video camera.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

7.1.6 To record the flow of air between two rooms by placing the probe at the door entrance and see
the direction of the fog into or out of the room.
7.1.7 Track the fog direction and record the observation with help of video camera and save in the
compact disc as a backup.
7.1.8 Acceptance Criteria:
7.1.8.1 Fog should flow through these critical areas in unidirectional pattern. If the fog return or back
flow due to turbulence, system cannot be accepted and must be rebalanced or readjusted.
7.1.8.2 There should not be any short-circuiting of airflow; dead pocket and flow of air should be
unidirectional from supply to return.
7.1.8.3 Fog should flow from high-pressure zone to low-pressure zone.
7.2 Pressure Differential Monitoring:
7.2.1 The objective of this test is to demonstrate that the HVC system/ equipment are capable of
maintaining the desired designed differential pressure to avoid flow reversal to avoid
contamination and cross contamination.
7.2.2 This test shall be applicable for area where monitoring of differential pressure is required
7.2.3 The differential pressure monitoring shall be performed by trained person only.
7.2.4 Measuring instruments being used shall be calibrated.
7.2.5 Operate the HVAC system about 24 hrs. prior to performing these tests in order to stabilize
the system.
7.2.6 To avoid unexpected changes in pressure and to establish a baseline, all doors in the aseptic
facility must be closed and no traffic is to be allowed through the facility during the test.
7.2.7 Record the pressure differentials of all rooms after every 2 hours for 3 Days.
7.2.8 Acceptance Criteria: The Pressure differential of all the areas / equipments shall be within
limit as applicable.
7.3 Temperature and Relative Humidity Monitoring:
7.3.1 The objective of this test is to demonstrate that the HVAC system is capable of maintaining
room conditions i.e. temperature and relative humidity as per the designed room conditions.
7.3.2 This test shall applicable for that room where temperature and RH monitoring is required.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

7.3.3 This test shall not applicable for Air filtration system like pass box, sterile cabinets, air shower,
LAF, dispensing and sampling booth.
7.3.4 HVAC system should be in running condition at least from last 24 hrs. All lights in the
subjected room should be kept on during the testing as well as during the pre-conditioning
period.
7.3.5 All doors in the facility must be closed and the test should be carried at rest condition.
7.3.6 Measure and record the temperature and RH for three days with frequency of every Two
hour.
7.3.7 Acceptance Criteria: Temperature shall be NMT 25oC and Relative Humidity shall be NMT
55% (or as per requirement)

7.4 Viable particle Count (Active & Passive air sampling):

7.4.1 Viable particle count by Settling Plate Method:
7.4.1.1 The objective of this test is to demonstrate that the clean room and the critical work stations
within the clean rooms comply with their designed conditions by settling plate method.
7.4.1.2 The microbial count testing shall be performed by trained person only.
7.4.1.3 Measuring instruments being used shall be calibrated.
7.4.1.4 Check the microbial count at rest & in operation condition continues 03 days.
7.4.1.5 Inform the manufacturing pharmacist regarding plate exposure activity before commencing
the activity. Enter in the clean area and clean the containers with 70 % IPA then remove the
plates from the container, mark number to the sterile pre-incubated Soya bean casein digest
agar (SCDA) plates and expose the plates at all locations for 4 hours on the plate exposure
stand. Incubate the exposed plates along with positive and negative controls at 20-25 oC
for 72 hours and followed by 30-35oC for 48 hours. After completions of incubation
period record the colony forming units (CFU) per plate in the formats. After completion
of incubation period, count the number of colonies on each agar plate and record.
7.4.2 Viable particle count by Air Sampling Method:
7.4.2.1 The objective of this test is to demonstrate that the clean room and the critical work stations
within the clean rooms comply with their designed conditions by Air sampling method.
7.4.2.2 The microbial count testing shall be performed by trained person only.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

7.4.2.3 Measuring instruments being used shall be calibrated.

7.4.2.4 Check the microbial count at rest & in operation condition continues 03 days.
7.4.2.5 Inform the manufacturing pharmacist regarding plate exposure activity before commencing
the activity. Enter in the clean area and clean the containers and air sampler with 70 % IPA
then remove the plates from the container, mark number to the sterile pre-incubated
Soyabean casein digest agar (SCDA) plates and at the location of air sampling open the top
lid of pre incubated SCDA plate immediately remove the aluminum foil of perforated sieve
and set it with head of air sampler over the SCDA plate. Vertically put the air sampler at the
location and carry out the air sampling of 1000 liter.
7.4.2.6 Incubate the exposed plates along with positive and negative controls at 20-25oC for 72 hours
and followed by 30-35oC for 48 hours. After completions of incubation period record the
colony forming units (CFU) per plate in the formats. After completion of incubation period,
count the number of colonies on each agar plate and record
7.4.2.7 Acceptance Criteria:
Recommended Limits for microbial contamination
Grade Active air Sampling Settle Plate (Diameter 90 mm)
(CFU/m3) CFU/4 Hours
Grade A <1 <1
Grade B NMT 10 NMT 5
Grade C NMT 100 NMT 50
Grade D NMT 200 NMT 100

7.5 Revalidation Criteria: Re-Qualification of Air filtration system shall be performed under the following
conditions:
7.5.1 The Air Filtration system is not performing well within the pre-defined acceptance criteria.
7.5.2 If any of the test results are not within the limits defined in the acceptance criteria.
7.5.3 The revalidation Schedule of the test parameters shall be defined as annexure-IV.
7.5.4 If any major modification/change in facility.
7.5.5 If any of the critical components of air filtration system is replaced due to technical problem.
7.5.6 Any revalidation trigger through change control.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

8.0 ABBREVIATION:
SOP Standard Operating Procedure
QA Quality Assurance
HVAC Heating Ventilation air conditioning
HEPA High Efficiency Particulate Air
% Percent
Mg Milligram
CFM Cubic Feet per minute
ft3 Cubic feet
CFU Colony Forming Unit
RH Relative Humidity
QC Quality Control
MF Manufacturing
EG Engineering

9.0 ANNEXURES:
ANNEXURE No. TITLE OF ANNEXURE FORMAT No.
Annexure-I Revalidation Frequency
Annexure-II Justification for selection of sampling location for non-
viable particle count
Annexure-III Schematic Diagram: Sampling Location for Non viable
count.

10.0 DISTRIBUTION DETAILS:

Master Copy Quality Assurance Department
Controlled Copy No. 01 Quality Assurance Department.
Controlled Copy No. 02 Quality Control Department.
Controlled Copy No. 03 Production Department.
Controlled Copy No. 04 Engineering Department.
Controlled Copy No. 05 Warehouse Department (Store).
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

11.0 REFERENCE:
 PIC’s “Manufacture of sterile medicinal Products” Annexure-1,
 ISO 14644 PART I, II & III.
 EU Guideline Annexure-1,
 In-house

12.0 REVISION HISTORY

Change Details of Reason of Effective

Revision No. Done By
Control No. Changes Changes Date
Not Not
00 New SOP
Applicable Applicable
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

ANNEXURE- I
REVALIDATION FREQUENCY

S.No. Test Frequency

HEPA Filter Integrity Test
1. 12 months ± 30 days
(By using poly alpha olefin)
2. Air velocity (ACPH) measurement 6 months ± 30 days
3. Air flow pattern test (Smoke test) 24 months ± 30 days
12 months ± 30 days (FOR ISO CLASS 8)
4. Nonviable particulate counts measurement
6 months ± 30 days (FOR ISO CLASS 5, 6,7)
5. Recovery test 12 months ± 30 days
During Area Qualification / as per routine
6. Pressure Difference*
monitoring.
During Area Qualification / as per routine
7. Temperature and % Related Humidity*
monitoring.
During Area Qualification / as per routine
8. Viable Particle count*
monitoring.

Note ‘*’ are tests to be performed on routine/regular basis as per frequency specified in the respective SOP
and at the time of Qualification and Re-Qualification activities.
 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

ANNEXURE-II
JUSTIFICATION FOR SELECTION OF SAMPLING LOCATION FOR NON-VIABLE PARTICLE
COUNT

HVAC Room ISO Area in No. of Identified Considered Location Location Justification
No. Name Class Square Location for Total No of details
Meter (A) non-viable Location locations
Count
(√A )

Prepared By Reviewed By Approved By

Quality Assurance Quality Assurance Quality Assurance

 STANDARD OPERATING PROCEDURE
Department: Quality Assurance SOP No.:
Title: Validation of Air Filtration System Effective Date:
Supersedes: Nil Review Date:
Issue Date: Page No.:

ANNEXURE-III
SCHEMATIC DIAGRAM: SAMPLING LOCATION FOR NON VIABLE COUNT

Name of the Room/Equipment.: ______________

Room/Equipment ID.: ______________

Prepared by: Reviewed by Approved by: