Journal of Pharma Analytica & Validation Guide

( JPAVG )
Guide Guide

HVAC Validation
 INTRODUCTION
 HVAC stands for Heating, Ventilating & Air Conditioning
 It includes a variety of active mechanical/electrical systems employed
to provide thermal control in buildings.
 H- Heating system adds thermal energy to a space or building in order
to maintain some selected air temperature that would otherwise not be
achieved due to heat loss to the exterior environment.
 V- Ventilating system is intended to introduce air to or remove air from
a space to move air without changing its temperature. Ventilating
systems may be used to improve indoor air quality or to improve
thermal comfort.
 AC- Air Conditioning system removes thermal energy from a space or
building to maintain some selected air temperature that would
otherwise not be achieved due to heat gain from interior heat sources
and the exterior environment.
 As per ASHRAE (American Society of Heating, Refrigerating & Air
conditioning Engineers), HVAC system should fulfil four objectives:
o Control air temperature
o Control air humidity
o Control air circulation
o Control air quality

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 Journal of Pharma Analytica & Validation Guide
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Guide Guide

 HVAC Validation includes following Tests:

 Air Velocity Test
 Filter Leak Test
 Air Flow Pattern Test
 Non viable Particle Count
 Viable Particle Count
 Recovery Test
 Differential Pressure Monitoring
 Temperature & Relative Humidity Monitoring
 Noise & Light Level Measurement

 AIR VELOCITY TEST

 It expresses that air is delivered at required velocity by AHU through HEPA
filters and it helps in calculation of ACPH (Air Changes Per Hour).
 The airflow velocity should be measured at approximately 150 mm to 300 mm
from the filter face. The number of measuring points should be sufficient to
determine the supply airflow rate in cleanrooms and clean zones and should be
the square root of 10 times of area in square metres but not less than 4.
 Calculate the average velocity of each filter.
 General practice for number of measuring points:
Four Corners & One Centre point of Filter as shown below:

1 2

3 4

 Apparatus used for Test:

Ultrasonic anemometers, thermal anemometers, vane-type anemometers,
Capturehood or their equivalent can be used.

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 Calculation of ACPH:
ACPH = (Airflow discharge in room  60)  Room volume
Air flow discharge (m3/minute) = Air velocity of HEPA filter (m/s)  Filter area
(m2) x 60
 Acceptance Criteria:
o For Unidirectional airflow system (Grade A):
 As per EU & WHO (Technical Report Series No. 961) : 0.36 – 0.54 m/s
 As per Schedule M : 0.3 m/s ± 20% (for vertical flows) and
0.45 m/s ± 20% (for horizontal flows)
o For Non-unidirectional airflow system (Grade B, C & D), the ACPH should
be appropriate for the size of the room and the equipment and personnel
present in it.
 As per Schedule M, the minimum ACPH for Grade B and Grade C areas
shall not be less than 20 air changes per hour in a room with good air
flow pattern and appropriate HEPA filters.

 FILTER LEAK TEST

 This test is performed to confirm that the filter system is properly installed and
that leaks have not developed during use.
 The test is performed by introducing the specific challenge aerosol upstream of
the filter(s) and searching for leaks by scanning the downstream side of the filter.
 The concentration of the aerosol challenge upstream of the filter should be
between 20 mg/m3 and 80 mg/m3.
 Measure the downstream concentration by holding the probe approximately
3 cm from the face of filter.
 Scanning rate: 15 / X (cm/s)
Where X = probe dimension perpendicular to scan direction (expressed in cm)
For example: If probe size is 3 x 3 cm, the scan rate should be 5 cm/s.
 Scanning should be performed over the entire downstream face of each filter, the
perimeter of each filter, the seal between the filter frame and the grid structure,
including its joints.
 Apparatus used for Test:
o Aerosol Photometer: to measure the concentration of aerosol
o Aerosol Generator: capable of generating particle of 0.05 µm to 2.0 µm
size.
o Aerosol Source Substances:
 PAO (Poly Alfa Olefin)

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 DOP (Di-Octyl Phthalarte) : Not in use now as considered carcinogenic

 DOS (Di-Octyl Sebacate) or DEHS (Di-2-Ethyl Hexyl Sebacate)
 PSL (Poly Styrene Latex)
 Acceptance Criteria:
o As per ISO 14644-3, Leakage should not be greater than 0.01% of the
challenge aerosol concentration.
o If any leakage is more than 0.01% of the upstream aerosol concentration,
repair it.
o Repair patches on filter should not exceed maximum of 5% of the total
filter face area & maximum width/ length of each patch should not be
more than 1.5 inches.
o Total no of patches should not exceed 5 number/ filter.

 AIR FLOW PATTERN TEST

 The purpose of airflow direction test and visualization is to confirm that the
airflow direction and its uniformity conform to the design and performance
specifications.
 Apparatus used for Test:
o Smoke/Fog Generator
o Reagents used to generate smoke:
 Glycerine + Water
 Ice Water
 Titanium tetrachloride (Carcinogenic, used in case of emergency only)
 Place the smoke generator and generate the smoke below the supply HEPA filter
and observe the smoke Pattern. Same way observe the smoke pattern near
return riser.
 Airflow study is also performed to see the airflow direction between adjacent
rooms. Smoke is generated at the door (Slightly Open) between two rooms and
airflow direction is observed.
 Carry out the Videography of entire Airflow study.
 Acceptance Criteria:
o Air Flow should be Unidirectional
o Air should flow towards return riser
o Air should flow from room having higher pressure to adjacent room
having less pressure.

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 NON VIABLE PARTICLE COUNT

 It measures concentration of non viable air borne particle and confirms that
respective area/rooms meet predefined condition of air borne particles.
 Apparatus used for Test:
o Non viable particle counter

Portable Particle Counter Online Particle Counter

 Measurement shall be done at working height and in two states i. e. “at rest” &
“in operation”.
 No. of sampling location= X i. e. square root of Room area in square meter
 Rationale for sampling location shall be dependent on man movement, material
movement, area of product exposure, manual intervention and location likely to
have more air borne particles like return riser.
 Sample Volume and Sampling time per location:

Sample Time in minutes

Sample Volume
Grade with 100 LPM counter
At rest In operation At rest In operation
A 1000 L 1000 L 10 min 10 min
B 700 L 7 L* 7 min 1 min
C 7 L* 2 L* 1 min 1 min
D 2 L* ND 1 min 1 min

* Note: As per ISO 14644-1, the volume sampled at each location shall be at least
2 L with minimum sampling time at each location of 1 minute.

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 Acceptance Criteria:
o As per EU Guideline & WHO (TRS No. 961)

At rest In operation
Grade
0.5µ 5.0 µ 0.5µ 5.0 µ
A 3520 20 3520 20
B 3520 29 352000 2900
C 352000 2900 3520000 29000
D 3520000 29000 Not defined

o As per Schedule M:

At rest In operation
Grade
0.5µ 5.0 µ 0.5µ 5.0 µ
A 3520 29 3520 29
B 35200 293 352000 2930
C 352000 2930 3520000 29300
D 3520000 29300 Not defined

 When the number of locations to be sampled is more than one and less than ten,
compute 95% upper confidence limit (UCL) from the average particle
concentrations for all locations as per formula given below.

UCL = X + (t0.95 * SD / n )
Where,
X is the overall mean of the location averages.
t0.95 represents the 95th percentile of the t distribution, with (n-1) degrees
of the freedom.
SD is the standard deviation of the location averages.
n is square root of number of individual location averages.

t0.95 can be calculated from below table:

Number of individual 2 3 4 5 6 7-9
location averages (n)
t0.95 6.3 2.9 2.4 2.1 2.0 1.9

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 VIABLE PARTICLE COUNT

 It measures concentration of viable air borne particle and confirms that
respective area/rooms meet predefined condition of microbial contamination.
 Microbes in air are generally associated with solid or liquid particles. These
particles may consist of a single unattached cell or more commonly as clumps of
organisms.
 Sampling locations are selected based on activity in critical area and related
movement of man and material. These locations should be monitored on regular
basis.
 Preincubated SCDA Plates are generally used for microbial monitoring
 Sampling Techniques:
o Settle Plates (Viable Passive air monitoring)
o Air Sampling (Viable Active air monitoring)
o Contact Plates (Surface monitoring)
 After sampling, all the plates are incubated and no. of Colony Forming Units
(CFU) is observed.
 Acceptance Criteria:
o As per EU Guideline & WHO (TRS No. 961):

Settle Plates Contact Plates

Air Sample (diameter 90 mm) (diameter 55 mm)
Grade
(CFU/m3)
(CFU/4 hours) (CFU/plate)
A <1 <1 <1
B 10 5 5
C 100 50 25
D 200 100 50

o As per Schedule M:

Settle Plates Contact Plates

Air Sample (diameter 90 mm) (diameter 55 mm)
Grade
(CFU/m3)
(CFU/4 hours) (CFU/plate)
A <1 <1 <1
B 10 5 5
C 100 50 25
D 500 100 50

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Guide Guide

 RECOVERY STUDY
 This test is performed to determine the ability of the installation to eliminate
airborne particles.
 This test is only important and recommended for non-unidirectional airflow
systems because the recovery performance is a function of air re-circulation
ratio, inlet-outlet airflow geometry, thermal conditions and the air distribution
characteristics within the controlled zone.
 This test should be carried out in at rest state.
 As per ISO 14644-3, this test is not recommended for class 8 (Grade D) and 9.

 Procedure:
 Measure the clean room or clean zone air borne non-viable particle counts at rest
condition.
 The cleanroom area to be examined should be then contaminated with an
aerosol while the air-handling units are in operation.
 Commence measurements at 1 min intervals. Note the time when the particle
concentration reaches the 100 x target concentration threshold (t100n).
 Note the time when the particle concentration reaches the target cleanliness
level (tn).
 Recovery Time: tn - t100n
 Acceptance Criteria:
As per WHO (TRS No. 961), the recovery period should not be more than
15-20 min.

 DIFFERENTIAL PRESSURE TEST

 This test is carried out to determine the capability of air system to provide
pressure gradient among the different rooms.
 Apparatus used for Test:
Electronic micro manometer, Inclined manometer or mechanical differential
pressure gauge can be used to measure the air pressure difference between two
areas.
 Acceptance Criteria:
o The differential pressure should be as per predefined room design.
o As per EU Guidelines & WHO (TRS No. 961), adjacent rooms of different
grades should have a pressure differential of approximately 10–15 Pascal.
o As per Schedule M, differential pressure between areas of different
environmental standards shall be at least 15 Pascal.

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 TEMPERATURE & RELATIVE HUMIDITY TEST

 Temperature and relative humidity are monitored to demonstrate that
respective rooms contain predefined temperature and RH.

 Apparatus used for Test:

Thermometer, Thermistor, Psychrometer, Dew point sensor, Humidity monitor
capacitive and digital thermo hygrometer can be used to monitor temperature
and RH.

 Acceptance Criteria:
o The temperature and relative humidity depend on the product and nature
of the operations carried out.
o As per Schedule M, temperature and humidity in the aseptic areas shall
not exceed 27 °C and relative humidity 55%, respectively unless there are
product specific requirements.

 NOISE & LIGHT LEVEL

 Noise level and light level should be monitored to demonstrate that respective
area is well lighted and meeting predefined noise level criteria.

 Apparatus used for Test:

Sound meter and Lux meter

 Acceptance Criteria:
o Light level: NLT 300 Lux at working height
o Noise level: NMT 85 db at working height

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Guide Guide

 FREQUENCY OF REVALIDATION

Frequency
Test
As per ISO As per Schedule M
Air Velocity 12 months 06 months

Filter Leak Test 24 months 12 months

Air flow pattern 24 months -

Grade A: 06 months
Non Viable Particle count 06 months
Grade B,C & D: 12 months

Recovery 24 months -

 Environment monitoring (Viable and Non viable particle count) shall be

performed daily in operation on specified sampling locations.

 Differential pressure, Temperature & Humidity shall be monitored on daily basis

at specified time intervals.

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Guide Guide

 REFERENCES
 ISO 14644-3, Cleanrooms and associated controlled environments- Test methods
 EU Guidelines to Good Manufacturing Practice Medicinal Products for Human and
Veterinary Use, Volume 4, Annexure 1
 WHO Technical Report Series, No. 961 (2011), Annexure 6
 Schedule M

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