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Adding a cleaning step to the field-testing protocol, and combining it with the data generated to register sanitizing and disinfectant agents under FIFRA and the CEN TC 216 work program, produces a sanitation-and-disinfection validation methodology that is cost-effective, simple, and time-saving.
Although we commonly talk about "disinfectant validation," the US Food and Drug Administration validates only processes (1). Disinfectants themselves are qualified—that is, found to be effective in the context of a given process, just as we qualify the clean steam supply for an autoclave and then validate the steam sterilization process. The approach to disinfection should be similar, so that a working definition for disinfection process validation would be "establishing documented evidence that a disinfection process will consistently remove or inactivate known or possible pathogens from inanimate objects."
The working definition becomes critical as process operators attempt to comply with the current good manufacturing practice (CGMP) requirements of 21 CFR 211.56 (Sanitation) and 21 CFR 211.67 (Equipment cleaning and maintenance). The Sanitation clauses require that "any building used in the manufacture, processing, packing, or holding of a drug product shall be maintained in a clean and sanitary condition." And, the Cleaning and Maintenance provisions stipulate that "Equipment and utensils shall be cleaned, maintained, and sanitized at appropriate intervals to prevent malfunctions or contamination that would alter the safety, identity, strength, quality, or purity of the drug product beyond the official or other established requirements."
Even though the regulators have never formally defined "disinfectant validation," FDA Form 483 observations and Warning Letters frequently cite failures to ensure proper disinfection.
USP draft General Chapter ‹1072›, Disinfectants and Antiseptics
In 2002 the United States Pharmacopoeia (USP) published the draft General Chapter ‹1072›, "Disinfectants and Antiseptics" (2). This chapter addressed several key factors: selecting chemical disinfectants and antiseptics; demonstrating the effectiveness of disinfectants and antiseptics as bactericidal, fungicidal, and sporicidal agents; and applying disinfectants in manufacturing areas—along with the relevant regulations and safety considerations. The draft did not, however, clearly address disinfectant validation, per se, focusing instead on disinfectant effectiveness, which is a necessary prerequisite of disinfectant validation, but not sufficient in itself to ensure a valid disinfection process.
Draft chapter ‹1072› does say that demonstrating a disinfectant's effectiveness within a pharmaceutical manufacturing environment may require a battery of tests. Two of the tests listed are the "use-dilution test" and the "surface challenge test." As is so often the case, these tests may seem trivial at first glance, but they are not. In practice, they can be complicated and variable, which can make industry process validation personnel apprehensive every time they attempt a disinfection process validation.
Disinfectant effectiveness tests
Title 7 of the United States Code, the Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA), mandates that disinfectants be registered with the US Environmental Protection Agency (EPA) for sale and distribution in the United States. Chapter 6, subchapter II, Section 3(c)(5) of FIFRA requires that the composition of a pesticide product (the category that includes disinfectants) is such to guarantee the claims made on the label. Registrants must submit data demonstrating effectiveness to the EPA, which reviews effectiveness data before registration of public health antimicrobial pesticides. Subdivision G of the Pesticide Assessment Guidelines requires that effectiveness test data for submission must be obtained by methods accepted by the Association of Official Analytical Chemists (AOAC). The AOAC analyses include carrier tests, and use-dilution tests for bactericidal, mycobactericidal, and sporicidal activity—better known collectively as Disinfectant Effectiveness Tests or DETs (see Table I).
Table I: Association of Official Analytical Chemists (AOAC ) test methods.
The AOAC's disinfectant effectiveness tests are not the only means of gathering antimicrobial performance data. The United Kingdom accepts the Kelsey-Sykes Capacity test. In most parts of Europe the recognized DET standards come from the systematic approach of the European Committee for Normalization (CEN) and Technical Committee (TC) 216 work program, "Chemical Disinfectants and Antiseptics."
The CEN TC 216 methodology has three phases to its Disinfectant Effectiveness Test—European Tiered Test Approach (see Table II).
Table II. European Committee for Normalization TC 216 Committee; Disinfectant Effectiveness Tests-European Tiered Test Approach.
As a result of the CEN TC 216 work program, and with the concurrence of the EU Biocides Directive, several BS EN (European Standards as adopted by the British Standards Institute, test methods have been issued. These pass–fail test methods detail how disinfectants should be tested against selected cultures under controlled laboratory conditions. They also require an evaluation of the laboratory test results against field test results (see Table III).
Disinfectant effectiveness tests are not yet routine, so most drug-industry microbiologists are unfamiliar with them. The protocols take a long time to learn and a long time to perform, which makes them expensive. In addition, they are difficult to do and are done infrequently, which makes it hard to maintain laboratory personnel proficiency and hard to produce reproducible results.
Reproducibility is critical to proper interpretation of results. Variability in AOAC`s tests is mostly caused by errors in preparing test solutions and media. Some studies have shown, however, that the AOAC testing technique (3, 4) and inoculum preparation (5) may also be at fault. Variations in conditions during cold storage (i.e., refrigeration, freezing, freeze-drying) can affect the predominant genotype in the stock culture or source culture (6). Thus, when pharmaceutical companies try to validate disinfection processes using AOAC protocols, they may be unable to reproduce the disinfectant manufacturers' bactericidal label claims. CEN TC 216 tests are also liable for reproducibility problems caused by variation in the inoculum size and preparation (7, 8).
Table III: British Standards (BS) EN test methods.
In the United States, the EPA shares regulatory authority with FDA, which has jurisdiction under 21 CFR 880.6890, General purpose disinfectants. This Subpart (concerned with medical devices), defines a general purpose disinfectant as a "germicide intended to process noncritical medical devices and equipment surfaces." (Noncritical medical devices make only topical contact with intact skin.) Liquid chemical sterilizers intended for use on critical or semicritical medical devices are defined and regulated by FDA under 21 CFR 880.6885, Liquid chemical sterilants/high-level disinfectants.
Sanitation process validation
A review of regulatory definitions (see sidebar, "Definitions and Distinctions") quickly leads to the realization that "disinfectant validation" is actually equivalent to sanitation process validation. This analysis leads to the following proposal for improving the validity of the validation process while making it easier and less time-consuming to perform.
Definitions and distinctions
This approach is based on the CEN TC 216 work program methodology for registering disinfectants in the European Union and on the quantitative carrier tests (QCT) developed by the University of Ottawa for environmental surfaces and medical devices (9). Like the CEN TC 216 method, the Ottawa QCTs first assess microbicidal performance under ideal conditions, and then move on to assess field performance with more stringent tests.
Here, we define sanitation process validation as "establishing documented evidence that a sanitation process will consistently reduce microorganism populations on inanimate surfaces to preestablished levels that are considered safe." The preestablished microbial bioburden levels must be based on public health codes, regulations, industry guidelines, or a scientifically sound rationale.
In the pharmaceutical and food industries, the sanitation process typically follows a cleaning stage. Cleaning with a detergent usually removes or kills more than 90% of vegetative bacteria present on surfaces. The surviving microorganisms are mostly surface-attached, and the sanitation process will inactivate them in-situ.
Disinfectant effectiveness tests do not mirror in-service conditions. The test microorganism inoculum used in DETs does not mimic the behavior of environmental growth (e.g., biofilms and surface-adhered microorganisms). In addition, DETs fail to consider the effects of the preceding cleaning stage. Therefore, the testing conditions used by agents manufacturers for disinfection and sanitation have little relationship to the sanitation processes actually used in the pharmaceutical industry. Manufacturers rely on use-dilution and carrier tests to provide data for registration in the United States and for recommended in-use concentrations.
Validation of a sanitation process should instead be based on empirical measurements of sanitizer effectiveness in working conditions, suggesting the logical equation:
successful field tests under in-use conditions = validation.
For chemical disinfectant and sanitizing agents already registered under FIFRA in the United States or the CEN TC 216 work program in the EU, there would be no need to perform in-house effectiveness tests. The DETs already performed by the manufacturer for registration should be sufficient. Abundant experience and USP draft chapter ‹1072› show that sanitizers and disinfectants are more effective in actual use than the DETs indicate. Furthermore, the high inoculum counts used in the laboratory studies represent a "worst-case" scenario. We should, therefore, be able to rely on a second logical equation:
registration = qualification.
This approach would make in-house DETs unnecessary, and would greatly simplify the sanitation validation process by removing the most difficult step; performing DETs. Only sanitizing or disinfecting agents that have not been registered for the intended purpose would then require additional qualification with DETs.
Surface tests would still be required to develop procedures for sanitation processes. These tests would assess the effectiveness of the selected sanitizer against surface-adhered microorganisms. In these assays, microorganisms are dried onto surfaces, sanitized, and then removed for counting by conventional techniques or rapid microbiological methods (RMMs). Established surface tests are straightforward and inexpensive, and can thus be carried out by most microbiology laboratories. Surface tests can reflect in-use conditions like contact times, temperatures, use-dilutions, and surface properties. These tests can be modified to follow a representative cleaning stage, and so will better mimic real in-use conditions. The proposed sanitation process would be developed from the surface tests. Finally, the proposed sanitation process would be validated via challenge in field tests.
The sanitation process validation would then be performed following a simple methodology:
The validation of agents for sanitizing and disinfecting seems like a major undertaking, but does not need to be. These agents are not validated; they are qualified for the intended purpose, and then the sanitation process itself is validated. The most difficult part in sanitation process validation would be the execution of disinfectant effectiveness tests by the user. In-house DETs are superfluous, however, when the selected sanitizer or disinfectant has been registered in accordance with FIFRA or the CEN TC 216 work program. The sanitation process validation methodology proposed here includes cleaning as part of the sanitation process, and sanitation process validation becomes the successful execution of field tests under actual in-use conditions. The resulting methodology would be cost-effective, simple, and time-saving.
My gratitude to Daniel Y. C. Fung, professor of food science at Kansas State University, for his kindness in reviewing a draft of this article. And special thanks to Douglas McCormick, Advanstar Communications, for his assistance in the final editing of the manuscript.
José E. Martinez is a consultant for JEM Consulting Services Inc., PMB 652 Box 4956; Caguas, PR 00726, tel. 787.349.3857, email@example.com
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10. Center for Food Safety and Applied Nutrition, "Comprehensive List of Terms," (FDA, Rockville, MD, 2001), http://www.cfsan.fda.gov/~dms/a2z-s.html, accessed Feb. 15, 2006.
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