Standard operating procedure to ensure the sterility of the Pharmaceutical products.
1.0
OBJECTIVE: The objective of this SOP is to ensure that the batch of product is sterile.
2.0 SCOPE:
This procedure is applicable to all parenteral pharmaceutical dosage forms and
any specific material for which this test is applicable.
3.0
RESPONSIBILITY:
Microbiologist /Designee: To operation and temperature monitoring of
Refrigerator as per SOP.
Head/ Designee - QC: To review and ensure compliance of SOP.
Head/ Designee - QA: To approve, implement and ensure compliance of SOP.
4.0
PROCEDURE:
4.1 Prepare
Fluid Thioglycollate medium and Soybean casein digest medium as per SOP forMedia Preparation.
4.3 Sterility Testing Procedure of Products other than Insulin:
4.3.1
Sterilize all required accessories for sterility test in autoclave at 121°C for
30 minutes as per SOP and glassware and forcep/cutter by DHS at 180°C for 2
hrs. as per SOP for Cleaning and Sterilization of Glassware.
4.3.2 Enter
in sterility testing room and follow gowning procedure as per SOP.
4.3.3
Switch “OFF” the UV light of sterility LAF and Switch “ON” the tube light.
4.3.4
Operate the LAF as per SOP and clean the LAF bench with 70% filtered IPA.
4.3.5
Disinfect the outer surface of sample container with 70% filtered IPA prior to
testing and allow it to dry.
4.3.7 Sample Quantity Required for Sterility for Sterility Test:
For finish
product: 20 containers of every batch.
For Bulk
sample: 2 X 50 ml sample in sterile container.
4.3.8
Arrange the filtration assembly in LAF and connect it with vacuum pump.
4.3.9 Place
sterile 0.45µ membrane on filter holder with the help of sterile forcep and
clamped it properly.
4.3.10 Collect
the sample solution from ampoules / vials / bottles in a sterile flask with
help of sterile syringe. For dry powder or lyophilized container, add the
sterile water / 0.1% peptone dissolve and then collect it in sterile flask.
4.3.11
Filter the collected solution aseptically through 0.45µ membrane filter with
help of vacuum.
4.3.12 If
the test sample have any anti-microbial properties / preservatives, filter with
3 X 100 ml of freshly prepared sterile 0.1% peptone as rinsing solution.
4.3.13 Simultaneously
perform the sterility test of 0.1% peptone water or any other diluent used for
rinsing the filter.
4.3.14
After completion of filtration, open the assembly and cut the membrane filter
in two equal parts with sterile cutter.
4.3.15
Inoculate aseptically one half membrane filter in fluid Thioglycollate medium and
other half membrane filter in Soybean casein digest medium with help of sterile
forcep. Keep one tube of each medium without inoculated as a negative control.
4.3.16
Incubation Fluid Thioglycollate medium at 30 – 35°C for not less than 14 days
and incubate Soyabean casein digest medium at 20 - 25°C for not less than 14
days.
4.3.17 The
products terminally sterilized by a validated moist heat process, incubate the
test specimen for not less than 7 days.
4.3.18
Daily observe the incubated media for presence or absence of microbial growth in
form of turbidity and note down the observation as per annexure.
4.4 Sterility Testing Procedure for Insulin:
4.4.1 All
steps to be followed as per 4.3.1 to 4.3.15.
4.4.2 In
case of suspension, dissolve the insulin crystals in sterile 20% ascorbic acid
(Approx 50ml).
4.4.3
Incubate Fluid Thioglycollate medium at 30 – 35°C for NLT 14 Days and Soybean
casein digest medium at 20 – 25°C for NLT 14 Days.
4.4.4 Daily
observe the incubated media for presence and absence of microbial growth in terms
of turbidity and note down the observation as per annexure.
4.4.5 After
completion of incubation period, distribute the each medium in three tubes.
Inoculate the test organisms (approx 100 cfu/ml) in respective medium as
follow.
1) Fluid
Thioglycollate medium: (For aerobic & anaerobic bacteria)
P.aeruginosa
- ATCC – 9027
S.aureus -
ATCC – 6538
Cl.sporogenes
- ATCC – 1437
Incubated
at 30 – 35°C for 2 – 3 Days.
2) Soyabean
casein digest medium: (For bacteria & fungus)
S.aureus -
ATCC – 6538
C.albicans
- ATCC – 10231
A.niger -
ATCC – 16404
Incubated
at 20 – 25°C for 5 Days
4.4.6
Acceptance criteria: Medium should promote the growth within its incubation
period.
4.4.7
Observed the incubated media tubes for growth promotion and note down the
observation.
4.5 If
evidence of Microbial growth is found, reserve the container showing this, and
unless and it is demonstrated by any other means that their presence is due to
causes unrelated to the preparation being examined, then the test for sterility
is invalid and perform a retest on the same number of the sample.
4.6 If no
evidence of microbial growth is found on retesting or the preparation being
examined passes the test for sterility.
4.7 If the
evidence of microbial growth is found on retesting, the preparation being
examined fails the test for sterility.
4.8 As per
IP If no evidence of microbial growth is found, the preparation being examined
complies with the test for sterility.
4.9 If
evidence of microbial growth is found the preparation being examined does not
complies with tests for sterility. Do not repeat the test unless it can be
clearly shown the test was invalid for causes unrelated to the preparation
being examined.
4.10 The
test may consider invalid only when one or more of the following conditions are
fulfilled.
4.10.1 If
the microbial growth found in negative control.
4.10.2 Data
on microbial monitoring of the sterility testing facility show a fault.
4.10.3 A
review of the testing procedure used for the test in question reveals a fault.
4.10.4
After identifying the micro organisms isolated from the containers showing
microbial growth, the growth may be ascribed without any doubt to faults with
respect to the material and / or the techniques used in conducting the test
procedure.
4.10.5 If
the test is declared to be invalid repeat with the same numbers of unit as in
original test. It no evidence of microbial growth is found in the repeat test
the preparation being examined complies with the test for sterility. It
microbial growth is found in the repeat test and confirmed microscopically, the
preparation being examined does not comply with the tests for sterility.
5.0
ABBREVIATIONS
ATCC =
American Type culture collection
LAF =
Laminar Air Flow
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