The rabbit pyrogen test, per USP <151>, is a method used to detect the presence of pyrogens. Pyrogens are fever-inducing substances that may be in pharmaceutical products intended for parenteral (injectable) administration. Pyrogens can cause serious adverse reactions in patients, so it is critical to ensure injectable products are free of these contaminants.

How the USP <151> Pyrogen Test is Performed

If running the test in compliance with the USP, the test involves measuring the rise in temperature of rabbits following the intravenous injection of a test solution and is designed for products that can be tolerated by the test rabbit in a dose not to exceed 10 mL per kg injected intravenously within a period of not more than 10 minutes.

The test is conducted using a minimum of 3 rabbits. Once dosed, temperatures are recorded for 3 hours post-injection. The test fails if any rabbit shows a rise in temperature > 0.5°C.

For products that require preliminary preparation or are subject to special conditions of administration, there may be additional directions given in the individual monograph or, for antibiotics or biologics, the additional directions given in federal regulations.

Variations of the USP <151> Pyrogen Test

While Nucro-Technics conducts the pyrogen test most commonly against USP requirements for injectable drug products, there are many permutations of the test. The British Pharmacopoeia (BP), European Pharmacopoeia (EP), and Japanese Pharmacopoeia (JP) have their own variations that are very similar – typically differing on the number of test animals required, the extent of testing time, and the rise in temperature threshold.

Medical devices are also commonly tested for pyrogenicity in alignment with ISO 10993-11. Because there is no injectable drug substance or drug product, extracts of the medical device are used, but the testing is otherwise similar in conduct to the USP <151> variation.

There are in vitro variations of this test as well. Most testing that would have previously been conducted in vivo is now conducted in an in vitro setting. The bacterial endotoxin (BET) or Limulus Amebocyte Lysate (LAL) test is the most common and accepted approach. In more recent years, the Monocyte Activation Test has also gained popularity as a promising alternative and has been accepted by the European Pharmacopoeia as a valid alternative for the testing of pyrogenicity in drugs. Nucro-Technics also routinely performs these alternative tests.

Which Approach to Take?

The choice of pyrogen test method depends on factors like the sensitivity required, the nature of the product, and regulatory requirements. Pharmaceutical companies must validate and use the appropriate pyrogen testing method as part of their quality control procedures.

If this pyrogen test is used as a release test, it would be conducted in alignment with Good Manufacturing Practice (GMP) Regulations. This test can also be performed non-GLP or in compliance with Good Laboratory Practice Regulations if being used to support a regulatory submission.