ZAFA Pharmaceutical Laboratories (Private) Ltd
TITTLE: SOP No. ZP/MIC/GEN/009
Issue No. 05
SOP FOR LIMIT TEST FOR
SPECIFIED MICRORGANISM Supersedes 04
Issue date 22-05-2024
Effective date 24-05-2024
Next Review date 22-05-2027
Pages Page 1 of 6
1.0 PURPOSE
To describe the procedure of testing procedure of specified microorganism.
2.0 SCOPE
This sop is applicable for media fill operation of sterile area of all sites of ZAFA pharmaceuticals
Laboratories Pvt. Ltd.
3.0 ABBRIVATION / GLOSSARY
Not applicable
4.0 RESPONSIBILITY
4.1 GM Plants
Responsible for authorization of this SOP
4.2 Sr. Manager Quality Operations
Responsible for the approval and implementation of this SOP
4.3 QC Manager
Responsible for the implementation of this SOP.
4.4 QA Manager
Responsible to check this SOP.
4.4 Microbiologist
Responsible to follow this SOP
5.0 PROCEDURE
5.1 Escherichia coli
5.1.1 ZAFA Pharmaceutical Laboratories (Private) Ltd
5.1.1
5.1.1
TITTLE: SOP No. ZP/MIC/GEN/009
5.1.1
5.1.1 FOR LIMIT TEST FOR Issue No. 05
SOP
5.1.1
SPECIFIED MICRORGANISM Supersedes 04
5.1.1
5.1.1 Issue date 22-05-2024
5.1.1
5.1.1 Effective date 24-05-2024
5.1.1
Next Review date 22-05-2027
5.1.1
5.1.1 Pages Page 2 of 6
5.1.1
Prepare a sample using a 1 in 10 dilution of not less than 1 g of the product to be examined and use 10 mL
or the quantity corresponding to 1 g or 1 mL, to inoculate a suitable amount (determined as described
under Suitability of the Test Method) of Soybean–Casein Digest Broth, mix, and incubate at 30° to 35°
for 18 to 24 hours.
5.1.2 Shake the container, transfer 1 mL of Soybean–Casein Digest Broth to 100 mL of MacConkey Broth, and
incubate at 42° to 44° for 24 to 48 hours. Subculture on a plate of MacConkey Agar at 30° to 35° for
18 to 72 hours.
5.1.3 Growth of colonies indicates the possible presence of E. coli. This is confirmed identification by
tests. The product complies with the test if no colonies are present or if the identification tests
are negative.
5.2 Salmonella
5.2.1 Prepare a sample using a 1 in 10 dilution of not less than 1 g of the product to be
examined and use 10 mL or
5.2.2 the quantity corresponding to 1 g or 1 mL, to inoculate a suitable amount (determined as described
under
5.2.3 Suitability of the Test Method) of Soybean–Casein Digest Broth, mix, and incubate at 30° to 35° for 18
to 24
5.2.4 hours.
5.2.5 Transfer 0.1 mL of Soybean–Casein Digest Broth to 10 mL of Rappaport Vassiliadis Salmonella
Enrichment Broth, and incubate at 30° to 35° for 18 to 24 hours. Subculture on plates of Xylose
Lysine Deoxycholate Agar. Incubate at 30° to 35° for 18 to 48 hours.
5.2.6 The possible presence of Salmonella is indicated by the growth of well-developed, red colonies, with
or without black centers.
5.2.7 This is confirmed by identification tests.
5.2.8 The product complies with the test if colonies of the types described are not present or if the
confirmatory identification tests are negative.
5.3 Pseudomonas aeruginosa
5.3.1 Prepare a sample using a 1 in 10 dilution of not less than 1 g of the product to be examined and use
10 mL or the quantity corresponding to 1 g or 1 mL, to inoculate a suitable amount (determined as
described under Suitability of Soybean–Casein Digest Broth, mix, and incubate at 30° to 35° for 18
to 24 hours.
5.3.2 Subculture on a plate of Cetrimide Agar, and incubate at 30° to 35° for 18 to 72 hours.
5.3.3 Growth of colonies indicates the possible presence of P. aeruginosa. This is confirmed by identification
tests.
5.3.4 The product complies with the test if colonies are not present or if the confirmatory identification tests
are negative.
ZAFA Pharmaceutical Laboratories (Private) Ltd
TITTLE: SOP No. ZP/MIC/GEN/009
Issue No. 05
SOP FOR LIMIT TEST FOR
SPECIFIED MICRORGANISM Supersedes 04
Issue date 22-05-2024
Effective date 24-05-2024
Next Review date 22-05-2027
Pages Page 3 of 6
5.4 Staphylococcus aureus
5.4.1
5.4.2 Subculture on a plate of Cetrimide Agar, and incubate at 30° to 35° for 18 to 72 hours.
5.4.3 Subculture on a plate of Mannitol Salt Agar, and incubate at 30° to 35° for 18 to 72 hours.
5.4.4 The possible presence of S. aureus is indicated by the growth of yellow or white colonies
5.4.5 surrounded by a yellow zone. This is confirmed by identification tests.
5.4.6 The product complies with the test if colonies of the types described are not present or if the
5.4.7 confirmatory identification tests are negative.
5.5 Clostridia
5.5.1 Prepare a sample using a 1 in 10 dilution (with a minimum total volume of 20 mL) of not less than 2
g or 2 mL of
5.5.2 the product to be examined divide the sample into two portions of at least 10 ml. Heat one portion at
80° for
5.5.3 minutes, and cool rapidly. Do not heat the other portion.
5.5.4 Use 10 mL or the quantity corresponding to 1 g or 1 mL of the product to be examined of both
portions to
5.5.5 inoculate suitable amounts (determined as described under Suitability of the Test Method) of
Reinforced
Medium for Clostridia. Incubate under anaerobic conditions at 30° to 35° for 48 hours. After
incubation, make
subcultures from each container on Columbia Agar, and incubate under anaerobic conditions at 30°
to 35° for 48 to 72 hours.
5.5.6 The occurrence of anaerobic growth of rods (with or without endospores) giving a negative catalase
reaction indicates the presence of Clostridia.
5.6 Candida albicans
5.6.1 Prepare the product to be examined as described in Microbiological Examination of Nonsterile Products:
Microbial Enumeration Tests 61, and use 10 mL or the quantity corresponding to not less than 1 g
or 1 mL, to inoculate 100 mL of Sabouraud Dextrose Broth, and mix. Incubate at 30° to 35° for 3 to
5
ZAFA Pharmaceutical Laboratories (Private) Ltd
TITTLE: SOP No. ZP/MIC/GEN/009
Issue No. 05
SOP FOR LIMIT TEST FOR
SPECIFIED MICRORGANISM Supersedes 04
Issue date 22-05-2024
Effective date 24-05-2024
Next Review date 22-05-2027
Pages Page 4 of 6
days.
Subculture on a plate of Sabouraud Dextrose Agar, and incubate at 30° to 35° for 24 to 48 hours.
5.6.2 Growth of white colonies may indicate the presence of C. albicans.
6.0 PROCESS FLOW
Not applicable
7.0 TRAINING
Training on this SOP shall be provided to QC managers of ZAFA sites and recorded as per SOP of
training and development.
8.0 DISTRIBUTION
S. No Dist Distribution List Copy No.
1. Compliance Cell Master
2. QC Manager – B-22 Copy 01
3. QC Manager – A-46 Copy 02
4. QC Manager – B-21 Copy 03
5. QC Manager – B-10 beta Copy 04
6. QC Manager – B-10 ceph Copy 05
7. QC Manager – A-79 Copy 06
ZAFA Pharmaceutical Laboratories (Private) Ltd
TITTLE: SOP No. ZP/MIC/GEN/009
Issue No. 05
SOP FOR LIMIT TEST FOR
SPECIFIED MICRORGANISM Supersedes 04
Issue date 22-05-2024
Effective date 24-05-2024
Next Review date 22-05-2027
Pages Page 5 of 6
9.0 ANNEXURES/ ATTACHED DOCUMENTS
Not applicable
10.0 DOCUMENT HISTORY
Version No. Effective Date Reason for Revision
02 20-04-2018 Sop revised as per review period
01 24-05-2024 i. Revise as per sop of preparation and management
ii. Revise as per revision date
02 iii.
11.0 REFERENE:
ZAFA Pharmaceutical Laboratories (Private) Ltd
TITTLE: SOP No. ZP/MIC/GEN/009
Issue No. 05
SOP FOR LIMIT TEST FOR
SPECIFIED MICRORGANISM Supersedes 04
Issue date 22-05-2024
Effective date 24-05-2024
Next Review date 22-05-2027
Pages Page 6 of 6
<62> Microbiological examination of non-sterile products, test for specified microorganism
End of Document
