Optimizing neutralization strategies for microbial testing of non-sterile pharmaceutical finished products with...

Method suitability for microbial limit tests during microbiological quality control (QC) can be a complicated process that requires multiple steps of optimization and plays a central role in ensuring reliable QC results. When antimicrobial activity of a given product cannot be neutralized, it is assumed that the inhibited microorganism is not present in the product, as specified by the U.S. pharmacopeia. This can potentially lead to contaminants that can multiply during storage or use, resulting in potential health risks or even death.

Method suitability testing was performed on a total of 133 pharmaceutical finished products as part of microbiological QC testing performed prior to product marketing. For total microbial count, depending on the pharmaceutical dosage form, testing included sequential trials of different dilution factors up to 1:200, the addition of 1-5% tween (polysorbate) 80, 0.7% lecithin, and/or filtration using different membrane filter types until the optimal method was selected for each product. Method suitability for microbial limit tests as well as for absence of specific pathogens was verified using a range of bacteria and fungi including Burkholderia cepacia.

There was an acceptable microbial recovery of at least 84% for all standard strains with all neutralization methods, demonstrating minimal to no toxicity. Forty of 133 finished products required multiple steps of optimization. Of these, 18 were neutralized through 1:10 dilution with diluent warming. Another 8 had no inherent antimicrobial activity to their API and were neutralized through dilution and the addition of tween 80. Neutralization of the remaining 13 products (mostly antimicrobial drugs) was achieved through variations of different dilution factors and filtration with different membrane filter types with multiple rinsing steps.

Our results provide detailed protocols that can help guide method suitability testing of finished pharmaceutical products for which neutralization proves challenging. We also include testing for B. cepacia complex in aqueous dosage form, often overlooked in the QC process. Since the current QC practice mostly assumes that these products do not require further testing, similar studies are especially needed to ensure the safety of finished pharmaceutical products during marketing, storage, and use.