Antimicrobial Effectiveness Testing (AET) or Preservative Effectiveness Testing (PET) is performed to help evaluate a products ability to withstand microbial contamination during use. It is used for all injections packaged in multiple-dose containers, as well as for products containing antimicrobial preservatives.
Products such as aqueous-based, multiple-dose topical and oral dosage forms, and pharmaceutical dosage forms including otic, ophthalmic, irrigation, nasal and dialysis fluids.
This type of test is performed according to USP <51> Antimicrobial Effectiveness Testing, EP 5.1.3 Efficacy of Antimicrobial Preservation and CTFA Preservative Challenge Testing. Challenge microorganisms are introduced into non-sterile products during testing to determine levels of antimicrobial effectiveness related to the addition of preservatives. Microorganisms are selected based on possible contaminants to the drug product before or during the manufacturing process.
When sterile articles packaged in multiple-dose containers are tested, preservatives are added to a product to restrain growth of microorganisms that may be introduced from withdrawing individual doses multiple times.
The following microorganisms are required per USP <51>: Escherichia coli, Staphylococcus aureus, Candida albicans, Aspergillus brasiliensis and Pseudomonas aeruginosa.
The test consists of two parts:
Suitability Counting Method
The test neutralizes residual preservatives present in the recovery agar to obtain accurate plate counts of survivor microorganisms. It is determined after adding challenge microorganisms to product dilutions and saline controls to examine if the diluted product exhibits antimicrobial properties. If biocidal activity is detected, chemical neutralizers may be incorporated into the testing scheme or further product dilution.
Testing of Products
Challenge microorganisms will be added to a product and the microorganisms that survive the preservative system are enumerated at different time intervals lasting over 28 days.
The following techniques are used during the testing of bioburden
Membrane Filtration Method
An effective quantitative technique that isolates colonies of bacteria and fungi from a fluid sample using a membrane filter and vacuum.
Standard Plate Count
Pour Plate Method
This method is used to isolate colonies of bacteria and fungi by plating a sample preparation directly to an empty Petri dish and adding nutrient agar that allows growth of any microorganisms that might be present.
Spread Plate Method
Isolate colonies of bacteria and fungi by plating a sample preparation directly to a nutrient agar plate.
Antimicrobial activity of the preparation in its final container is evaluated over the period of validity to ensure that such activity has not been impaired by storage conditions. While the antimicrobial effectiveness test is not a batch release test, it is expected that these products comply with the specifications set forth in the compendial method. Because of this, it is critical to have some level of post-market stability program in place.
Whitehouse Laboratories contains GMP-compliant stability chambers for storing client products, which are monitored for temperature and humidity.
Antimicrobial Effectiveness Testing – Method Suitability
Antimicrobial Effectiveness Testing – Routine Test