SOP for Microbial Testing in Ointment Development – V 2.0

Procedure for Microbial Testing in Ointment Development

Department	Microbiology/Quality Control (QC)/Research and Development (R&D)
SOP No.	FD-018
Supersedes	V 1.0
Page No.	Page X of Y
Issue Date	[Insert Issue Date]
Effective Date	[Insert Effective Date]
Review Date	[Insert Review Date]

1. Purpose

The purpose of this Standard Operating Procedure (SOP) is to establish a standardized procedure for microbial testing of ointments during formulation development. This ensures the product meets microbial safety standards and complies with pharmacopeial requirements.

2. Scope

This SOP applies to the Quality Control (QC) and Microbiology departments responsible for microbial limit testing (MLT) of ointments. It includes procedures for sample preparation, microbial enumeration, pathogen detection, and data recording.

3. Responsibilities

Microbiologist: Conducts microbial testing and records observations.
Quality Control (QC) Analyst: Performs confirmatory tests and ensures compliance.
Quality Assurance (QA) Personnel: Reviews and approves microbial testing reports.
R&D Head: Approves final microbial test results before product development proceeds.

4. Accountability

The QC and Microbiology Managers are accountable for ensuring microbial testing is conducted per regulatory guidelines and that results are accurately documented.

5. Procedure

5.1 Sample Preparation

Collect representative samples from different points in the batch (top, middle, bottom).
Weigh 10 g of the ointment sample in a sterile container.
Prepare a 1:10 dilution by suspending the sample in sterile buffered peptone water (BPW) or another validated diluent.
Vortex the sample for uniform dispersion.

5.2 Total Aerobic Microbial Count (TAMC)

Inoculate 1 mL of the sample dilution onto Soybean Casein Digest Agar (SCDA) plates.
Spread the sample evenly using a sterile spreader.
Incubate at 30-35°C for 48-72 hours.
Count and record the colony-forming units (CFU) per gram of ointment.

5.3 Total Yeast and Mold Count (TYMC)

Inoculate 1 mL of the sample dilution onto Sabouraud Dextrose Agar (SDA) plates.
Spread the sample evenly using a sterile spreader.
Incubate at 20-25°C for 5-7 days.
Count and record the CFU per gram of ointment.

5.4 Pathogen Detection

Test for specific pathogens using selective media:
- Escherichia coli: MacConkey Agar (Incubate at 35-37°C for 24-48 hours).
- Salmonella spp.: Xylose Lysine Deoxycholate (XLD) Agar (Incubate at 35-37°C for 24-48 hours).
- Pseudomonas aeruginosa: Cetrimide Agar (Incubate at 35-37°C for 24-48 hours).
- Staphylococcus aureus: Mannitol Salt Agar (Incubate at 35-37°C for 24-48 hours).
Confirm pathogen identity using biochemical or molecular techniques if necessary.

5.5 Acceptance Criteria

Total Aerobic Microbial Count (TAMC): Not more than 10³ CFU/g.
Total Yeast and Mold Count (TYMC): Not more than 10² CFU/g.
Pathogens: Absent in 1 g of the sample.

5.6 Documentation and Reporting

Record all test results in the Microbial Testing Log.
Summarize findings in the Microbial Testing Report.
Obtain approval from the QA department before proceeding with further formulation development.

6. Abbreviations

TAMC – Total Aerobic Microbial Count
TYMC – Total Yeast and Mold Count
QC – Quality Control
QA – Quality Assurance
CFU – Colony-Forming Unit
BPW – Buffered Peptone Water
MLT – Microbial Limit Test

7. Documents

Microbial Testing Log (Annexure-1)
Microbial Testing Report (Annexure-2)

8. References

USP <61> Microbial Enumeration Tests
USP <62> Tests for Specified Microorganisms
ICH Q6A Specifications for Microbial Quality
WHO GMP Guidelines for Pharmaceutical Manufacturing
21 CFR Part 211 – Current Good Manufacturing Practices

9. SOP Version

Version 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name
Designation
Department

11. Annexures

Annexure-1: Microbial Testing Log

Date	Batch Number	Test Parameter	Result	Specification	Verified By
01/02/2025	OINT-001	TAMC	500 CFU/g	≤1000 CFU/g	QA Officer
02/02/2025	OINT-002	TYMC	80 CFU/g	≤100 CFU/g	QA Officer

Annexure-2: Microbial Testing Report

Date	Batch Number	Test Parameter	Specification	Final Result	Final Status
01/02/2025	OINT-001	Pathogens	Absent	Absent	Pass
02/02/2025	OINT-002	TAMC	≤1000 CFU/g	700 CFU/g	Pass

12. Revision History:

Revision Date	Revision No.	Details of Revision	Reason for Revision	Approved By
01/01/2024	1.0	Initial Version	New SOP	QA Head
01/02/2025	2.0	Updated SOP Format	Standardization of Document	QA Head