The Antimicrobial Preservative Efficacy test is designed to verify the efficacy of antimicrobial preservatives added to non-sterile dosage forms or sterile multi-dose containers in order to inhibit the growth of microorganisms that may be introduced inadvertently, during the manufacturing process or product use. Antimicrobial effectiveness must be demonstrated on all injectables packaged in multiple-dose containers and other products such as ophthalmics containing antimicrobial preservatives.
The USP Chapter <51> Antimicrobial Effectiveness Test (AET) outlines and specifies the use of stated organisms as challenge organisms for evaluating the efficacy of the antimicrobial preservative.
Bacteria are grown on Soybean Casein Digest Agar plates (SCDA). Yeast and Mold are grown on Sabouraud Dextrose Agar (SDA). All organisms are adjusted to approximately 1.0 x 10˄8 colony-forming units (CFU)/mL by diluting in sterile saline.
Individual test samples and a positive control are prepared for each organism. At Time 0, the test samples and positive controls are individually inoculated with each organism to reach a concentration of 1.0 x 10˄5 – 1.0 x 10˄6 CFU/mL. The positive controls and samples are assayed immediately after inoculation to determine the challenge level and to provide confirmation that the samples were inoculated.
The inoculated samples are stored in a 20 – 25°C incubator at 7, 14 and 28 days. After incubation the samples are removed from the incubator and assayed for surviving organisms. The organisms recovered from each time point are compared to those seen in the Time 0 positive control. Changes in the concentrations of organisms are converted to Log reduction values in order to assess the antimicrobial effectiveness of the samples. Acceptance criteria are established based on the type of product (ex: injectable, oral dose) and are specified in USP Chapter <51>.
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