USP <61> and <62> are the two chapters of the United States Pharmacopeia governing the microbiological examination of nonsterile products: the first defines the enumeration tests (TAMC and TYMC), the second the tests for specified microorganisms. In the lab they are often cited together, yet they answer two different questions: "how many microorganisms are there?" and "are there microorganisms that are unacceptable for this route of administration?". Confusing them — or applying only one when the monograph requires both — is one of the most frequent causes of incomplete specifications and audit findings. In this guide we compare the two chapters, with the operational requirements of each and the mistakes to avoid.
USP <61> vs <62>: two chapters, one system
The two chapters are designed to be used as a pair and are harmonized through the Pharmacopeial Discussion Group (PDG) with the European Pharmacopoeia (Ph. Eur. 2.6.12 and 2.6.13) and the Japanese Pharmacopoeia: a method compliant with USP <61>/<62> therefore maps onto the equivalent European chapters — a concrete advantage for sites releasing to multiple markets. The third element of the system is USP <1111>, the informational chapter that sets acceptance criteria by dosage form: <61> and <62> tell you how to run the test, <1111> tells you which result is acceptable.
| Aspect | USP <61> | USP <62> |
|---|---|---|
| Question it answers | How many viable microorganisms the product contains (quantitative count) | Whether specified "objectionable" microorganisms are present (qualitative outcome: presence/absence) |
| Test output | TAMC and TYMC in CFU/g or CFU/mL | Absence (or presence) of the target microorganism in the defined sample quantity |
| Methods available | Membrane filtration, pour plate, spread plate, MPN | Broth enrichment followed by isolation on selective media |
| Main media | Soybean–Casein Digest Agar (TAMC), Sabouraud Dextrose Agar (TYMC) | Dedicated selective media: MacConkey, XLD, Cetrimide, Mannitol Salt, VRBG, Columbia |
| Incubation | TAMC: 30–35 °C for 3–5 days; TYMC: 20–25 °C for 5–7 days | Target-specific (e.g. E. coli enrichment in MacConkey Broth at 42–44 °C) |
| Typical use | Release and stability specifications, raw material monitoring | Compliance with route-of-administration requirements per <1111> |
USP <61>: microbial enumeration (TAMC and TYMC)
Chapter <61> measures total microbial load through two separate counts: the Total Aerobic Microbial Count (TAMC) on Soybean–Casein Digest Agar, incubated at 30–35 °C for 3–5 days, and the Total Combined Yeasts and Molds Count (TYMC) on Sabouraud Dextrose Agar, incubated at 20–25 °C for 5–7 days. The laboratory chooses among membrane filtration, pour plate, spread plate or Most Probable Number (MPN) depending on the nature of the sample and the limit to be verified: membrane filtration is preferable when the limit is low or the product has antimicrobial activity removable by rinsing, while MPN is reserved for cases where the other methods are not applicable.
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USP <62>: tests for specified microorganisms
Chapter <62> does not count: it searches. The logic is enrichment in a non-selective or selective broth, followed by subculture on selective media and identity confirmation. The targets covered by the chapter are:
- Bile-tolerant Gram-negative bacteria — enrichment in Enterobacteria Enrichment Broth Mossel, isolation on Violet Red Bile Glucose Agar;
- Escherichia coli — enrichment in MacConkey Broth at 42–44 °C, isolation on MacConkey Agar;
- Salmonella — on at least 10 g of product, with Rappaport Vassiliadis Salmonella Enrichment Broth and Xylose Lysine Deoxycholate Agar;
- Pseudomonas aeruginosa — isolation on Cetrimide Agar;
- Staphylococcus aureus — isolation on Mannitol Salt Agar;
- Clostridia — Reinforced Medium for Clostridia and Columbia Agar under anaerobic conditions;
- Candida albicans — Sabouraud Dextrose Broth and Sabouraud Dextrose Agar.
Which of these targets to test is not the laboratory's call: it is dictated by the product monograph and the route-of-administration requirements. An aqueous oral product typically requires absence of E. coli; a topical product absence of S. aureus and P. aeruginosa; a rectal preparation, as a rule, no specified microorganisms.
Method suitability: where labs fail
Both chapters require demonstration of method suitability in the presence of the product, and this is where findings concentrate. There are three key requirements. First: the inoculum of the reference strains must not exceed 100 CFU, with strains inoculated individually, never as a mixture. The strains listed in the harmonized chapter include S. aureus ATCC 6538, P. aeruginosa ATCC 9027, B. subtilis ATCC 6633, C. albicans ATCC 10231 and A. brasiliensis ATCC 16404 (or equivalents from other recognized collections such as NCIMB, CIP, NBRC). Second: for <61>, recovery in the presence of the product must not differ by a factor greater than 2 from the control without product; if the product is inhibitory, neutralizers, dilutions or filtration with rinses must be applied — and the choice documented. Third: a negative control must accompany every execution of the test to demonstrate the absence of procedural contamination. A suitability study performed once and never re-evaluated after formulation changes is a classic finding: the rationale for its continued validity must be written down, not assumed.
Acceptance criteria: the role of USP <1111>
The results of <61> and <62> are interpreted against the criteria of USP <1111>, harmonized with Ph. Eur. 5.1.4. For aqueous oral dosage forms the typical reference is TAMC 102 CFU/g or mL and TYMC 101, with absence of E. coli in 1 g or mL; for non-aqueous oral forms TAMC 103 and TYMC 102; for cutaneous, nasal and auricular routes TAMC 102 and TYMC 101 with absence of S. aureus and P. aeruginosa; for the inhalation route, absence of bile-tolerant Gram-negatives is added. Watch out for the interpretation of the limits, often wrong in SOPs: a criterion of 102 CFU corresponds to a maximum acceptable count of 200 CFU, not 100, because the chapter accounts for the inherent variability of the counting method. And meeting the <1111> limits does not close the topic: the evaluation of objectionable microorganisms for the specific product, process and patient remains the manufacturer's responsibility, as FDA systematically reminds industry in non-sterile inspections.
GuideGxP recommendation
In specifications and SOPs, treat <61> and <62> as a single system: a product specification citing TAMC, TYMC and the specified microorganisms with the sample quantity; documented suitability for each matrix, with a re-evaluation rationale at every change of formulation or supplier; the interpretation of limits written in black and white (102 = maximum 200 CFU) to avoid OOS results declared out of excessive zeal; and a documented rationale on objectionable organisms beyond the <62> targets. In an audit, the question will not be "do you run the test?" but "why is your method suitable for this product?".
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