Antimicrobial preservative effectiveness test & its SOP

Antimicrobial preservative effectiveness test; Purpose

Antimicrobial preservative effectiveness test; To confirm that the concentration of Antimicrobial Preservatives used in different drug preparations are able to kill or destroy or inhibit or prevent or terminate the growth of microorganisms and thus make the product stable throughout its declared shelf life.

Antimicrobial preservative effectiveness test; Scope

This SOP is applicable for Oral liquids tests at Microbiology Laboratory in XX Pharmaceuticals Ltd.

Definition/Abbreviation:  

Preservative:

Antimicrobial preservatives are the substances added to different dosage forms or products to protect or defend them from microbial growth or from different microorganisms that are introduced unintentionally during or succeeding in the manufacturing process.

[] ATCC: American Type Culture Collection

[] SDA: Sabouraud Dextrose Agar

[] TSA: Tryptone Soya Agar

Responsibilities:

The roles and responsibilities are as follows:

Lab Attendant

Sample collection

Executive/ Sr. Executive, Microbiology

Sample collection, analysis of preservative test & respective test report preparation.

Manager, QC/Microbiology

• Ensure sampling, analysis, documentation & application of sound technical information.
• Review of this SOP and confirm that the whole procedure is technically sound.

Head of Quality Assurance

• Take initiative to Approval of this SOP.
• To ensure the overall implementation of this SOP

Procedure:

Instructions

• Use Bio-safety Cabinet to Perform tests of Antimicrobial preservative effectiveness of products.
• Before preparation of media and conduct antimicrobial preservative effectiveness test, Wear a mask, Hand gloves and headgear Bring the media to the boil to dissolve completely before autoclave.
• Mix well before pouring the substances.
• Cool the broth media at room temperature (25⁰C) and the agar media at 50⁰C & for use as per the requirement of the test.

Media & Biochemical Preparation

As per Media preparation for Media, preparation SOP prepare the same.

Inoculum Preparation:

• Remove the vial of pellets from refrigerated storage & allow equilibrating to the room condition.
• Warm hydrating & diluting fluids to [34 to 38]⁰C, before use.
• To achieve a concentration of about 10⁸ cfu per ml, transfer pellets to hydrating fluid.
• Immediately place the microbial suspension into a [34 to 38]⁰C incubator for 30 minutes
• To assure complete hydration, immediately following incubation, vortex the hydrated material to achieve a homogenous suspension.

Inoculation of Microbial Suspension to Product

• To give inoculums of 10⁵ to 10⁶ microorganisms per ml of product and mix well, inoculate the product to be examined, each with a suspension of the test organisms.
• The volume of the suspension of inoculums does not exceed 1 percent of the volume of the product.
• Maintain the inoculated product at the temperature range from [20 to 25]⁰C and protected it from light.
• According to the type of the product, Remove 1 ml sample from each container at suitable intervals & determine the number of viable microorganisms using the Pour plate method.

Suitability of counting method for non-sterile products

As per suitability of counting method for suitability of Microbial count, method SOP perform the same.

Test Sample Preparation

• Aseptically accurately measured 1 ml sample to be transferred from each inoculated product container to a market sterile capped dilution tube containing 9 ml of sample diluent & mix well. The ratio of this dilution is 1:10.
• Aseptically accurately measured 1 ml sample to be transferred from 1:10 dilution to a second dilution tube containing 9 ml of sterile diluent and mix well. This is the ratio of this dilution is 1:100.
• Continue this dilution up to 10-5 or as essential levels.

Plating and incubation:

• Take two sterile petri plates then Take 1 ml quantity sample from each dilution.
• Maintain the temperature not more than 45⁰C, then For Bacterial count pour 15 to 20 ml of sterile TSA medium into each plate being at and mix well.
• Maintain the temperature not more than 45⁰C, For Fungal count, pour 15 to 20 ml of sterile SDA medium into each plate being and mix well.
• Pour two plates with TSA & SDA medium each with 1 ml diluents as the negative control.
• After solidification incubate them at [20 to 25]⁰C for 5 to 7 days for the fungal count and [30 to 35]⁰C for 3 to 5 days for the bacterial count.

Acceptance Criteria

• For Oral Liquid & Other than Antacids
• For Bacteria (S. aureus, E.coli & P. aeruginosa)
• Not less than 1.0 log reduction from the initial count at 14 days.
• No increase from the 14 days counts at 28 days.

For Fungi (C. albicans, A. brasiliensis)

• No escalation from the initial calculated count at 14 and 28 days.

For Oral Liquid Antacids Preparation

• For Bacteria (E.coli, P. aeruginosa, S. aureus)
• No upsurge from the initial calculated count at 14 and 28 days.

For Fungi (C. albicans, A. brasiliensis)

• No increase from the initial calculated count at 14 and 28 days.

[Reference of acceptance criteria: USP 41]

Download All Annexure Here:

Annexure I: Preservative Efficacy Test Report Oral Liquid Preparation
Annexure II: Preservative Efficacy Test Report Oral Liquid Antacids Preparation