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USP 51: Antimicrobial Effectiveness Testing

By February 26, 2022Indoor Air Quality

Antimicrobial effectiveness testing is a protocol which tests the effectiveness of the antimicrobial compounds in non-sterile, aqueous pharmaceutical products. This type of testing is done to ensure that the antimicrobial compounds present in the product are capable of inhibiting the growth of microorganisms that may be introduced during or after the manufacturing process. Microorganisms may be introduced to these types of products in any number of ways – one of the most common is reusing a jar of lotion which requires the consumer to dip their fingers into the jar. Even just-washed hands aren’t sterile, so it is inevitable that microbes will be introduced. What the testing here aims to do is provide evidence that the antimicrobial compounds can inhibit those microbes over a period of time.

So, where do we start? If you haven’t read our blog post about Method Suitability Testing, head there first to understand the true beginning of the process. If you’re already versed in method suitability, keep reading…

Once Daane Labs has performed method suitability and understands the strength of your preservative system, we are ready to begin challenging that system by introducing known levels of bacteria and fungus. The challenge organisms used in USP 51 Antimicrobial Effectiveness Testing are the same as those used for method suitability testing: Staphylococcus aureus (ATCC 6538), Escherichia coli (ATCC 8739), Pseudomonas aeruginosa (ATCC 9027), Aspergillus brasiliensis (ATCC 16404), and Candida albicans (ATCC 10231). 

Ideally this testing is performed in five original containers (i.e., the final manufactured package) to mimic real-world conditions as closely as possible. The challenge organisms are inoculated directly into the five containers and we perform a “Day 0” plate count which gives us our baseline. The inoculated containers are incubated at 20-25C for up to 28 days and are plated periodically during that time. 

USP has categorized non-sterile aqueous products into four categories which are tested at different intervals and have to meet slightly different criteria for antimicrobial effectiveness testing. The four categories are summarized in the table below:

Category Product Description
1 Injections; other parenterals including emulsions, otic products, sterile nasal products, and ophthalmologic products made with aqueous bases or vehicles 
2 Topically used products made with aqueous base or vehicles; nonsterile nasal products and emulsions, including those applied to mucous membranes
3 Oral products other than antacids, made with aqueous bases or vehicles
4 Antacids made with an aqueous base

What we expect to see with antimicrobial effectiveness testing is, at minimum, no increase in microbial growth and, at best, a microbial population decline over time. The four categories indicated above have unique requirements for the antimicrobials to be considered effective. The table below describes the criteria:

Category 1
Bacteria No less than (NLT) 1.0 log reduction from the initial calculated count at 7 days, NLT 3.0 log reduction from the initial count at 14 days, and no increase from the 14 days’ count to 28 days 
Yeast and Mold No increase from the initial calculated count at 7, 14, and 28 days
Category 2
Bacteria NLT 2.0 log reduction from the initial count at 14 days, and no increase from the 14 days’ count at 28 days
Yeast and Mold No increase from the initial calculated count at 14 and 28 days
Category 3
Bacteria NLT 1.0 log reduction from the initial count at 14 days, and no increase from the 14 days’ count at 28 days
Yeast and Mold No increase from the initial calculated count at 14 and 28 days
Category 4
Bacteria, Yeast, and Mold No increase from the initial calculated count at 14 and 28 days

As you can see, there is a lot that goes into antimicrobial effectiveness testing from method suitability to final calculations. Luckily, we’ve been doing this for a long time and we are experts at compendial USP methods, USP 51 included. If this is a type of testing you need, or think you need, reach out! Daane Labs staff are always happy to help new and existing clients with antimicrobial effectiveness testing, method suitability, and routine testing.

References

https://latam-edu.usp.org/wp-content/uploads/2021/01/USP-NF-51.pdf (USP 51)