From a microbiological perspective, pharmaceutical products fall into two categories, nonsterile and sterile. For either category, manufacturers must eliminate or minimize potential health risks to patients related to microorganisms and the toxins they produce, while also maintaining product quality. Many contributing factors may affect the quality of a medicine or its ingredients, but microbial bioburden control and proper sterilization methods are critical considerations for the manufacturer throughout the product’s lifecycle.

The U.S. Pharmacopeial Convention (USP) is a global health organization that develops standards for the identity, strength, quality and purity of medicines, foods and dietary supplements and their ingredients, which are published in the United States Pharmacopeia and National Formulary (USP–NF). USP has no role in enforcement of these, or other provisions that recognize USP–NF standards, which is the responsibility of the U.S. FDA and other government authorities in the United States and elsewhere. Standards for drugs or drug ingredients are expressed in USP–NF monographs, general chapters and General Notices. Monographs are developed for single articles (e.g., drug substance, drug product, excipient, etc.) and general chapters can be applied across multiple articles. General chapters numbered above <1000> in USP–NF typically are informational and contain no mandatory requirements, unless specifically referenced in a monograph, General Notices or a general chapter numbered below <1000>. General chapters designated as below <1000> contain tests and procedures that are intended to apply to items recognized in USP or NF when called out in a monograph, General Notices or other applicable general chapters. The FDA may also require manufacturers to conform to USP standards that may not otherwise apply by the terms of USP–NF, if determined by the Agency to be within the scope of cGMPs. USP–NF is available for use globally. Besides this role, USP’s standards are also recognized and used around the world.

Through its Microbiology Expert Committee (Microbiology EC), USP develops and revises general chapters for the advancement of pharmaceutical microbiology. The Microbiology EC has an established work plan for its five-year operating cycle. The work plan is intended to help meet USP’s standards-setting goals. Major initiatives in the current USP 2010–2015 cycle regarding microbiological contamination include sterilization processes and sterility assurance. These key activities are discussed below.

Sterility Assurance and Sterilization

While all products purported to be sterile have to meet the requirements of General Chapter <71> Sterility Tests, sterility assurance can only be achieved by the use of robust sterilization processes. USP’s current General Chapter <1211> Sterilization and Sterility Assurance of Compendial Articles addresses principles of sterility assurance and provides information on various sterilization processes. In response to stakeholder feedback that greater detail is needed to address specific sterilization methods, USP initiated a two-stage revision approach in 2009. Phase 1 focused on updating content with current information. The general chapter became official in USP 33–NF 28 (2010). Phase 2 involves a significant rewrite of the general chapter. The initial focus of this second phase is on sterilization, to be followed by sterility assurance. In determining how to update the sterilization material in the general chapter (which originated in the late 1980s), the Microbiology EC decided to split the content of the general chapter into two major parts—sterility assurance and sterilization processes. Information related to sterilization processes was removed from the existing General Chapter <1211> and developed separately as the <1229.x> series. In a future revision, General Chapter <1211> will be renamed Sterility Assurance of Compendial Articles. The focus of the revised General Chapter <1211> will be limited to sterility assurance and will include aseptic processing, environmental monitoring, as well as a brief review of sterility testing and parametric release.

The new series of general chapters (the “<1229.x> series”) is dedicated to individual sterilization processes, with an overarching general chapter (<1229>), covering the overall concept of sterilization. In the development and organization of these general chapters, the Expert Committee has decided to develop a parallel series of chapters on depyrogenation separate from sterilization, in alignment with current industry practices. Depyrogenation processes will be addressed in a manner similar to those for sterilization processes. Moist heat sterilization will be divided into hard goods (direct contact approach) and aqueous liquids. Gas and vapor sterilization will be addressed in separate general chapters.

To date, the Microbiology EC has planned twelve general chapters, which will provide valuable information and guidance on distinct methods of sterilization (listed below). Currently, nine of these (shown in bold in the list below) have already been approved for inclusion in USP–NF.

<1229> Sterilization of Compendial Articles

<1229.1> Steam Sterilization by Direct Contact

<1229.2> Moist Heat Sterilization of Aqueous Liquids

<1229.3> Monitoring of Bioburden

<1229.4> Sterilizing Filtration of Liquids

<1229.5> Biological Indicators for Sterilization

<1229.6> Liquid Phase Sterilization

<1229.7> Gaseous Sterilization

<1229.8> Dry Heat Sterilization

<1229.9> Physicochemical Integrators and Indicators for Sterilization

<1229.10> Radiation Sterilization

<1229.11> Vapor Phase Sterilization

The other general chapters will be proposed for public comment in 2014 in Pharmacopeial Forum (tinyurl.com/mskdna8)—USP’s free access, online tool for public comment on USP’s quality standards and monographs.

Some of the new official and proposed general chapters are detailed below.

Steam Sterilization

Although many steam sterilization approaches related to hard goods are relevant to aqueous liquids, <1229.1> Steam Sterilization by Direct Contact and <1229.2> Moist Heat Sterilization of Aqueous Liquids have been written separately for greater clarity and for distinction between the two. For example, when using the direct contact approach, overkill is the method of choice because overprocessing of parts or hard goods usually is not a concern. On the other hand, when developing sterilization processes for liquid-filled containers, consideration of the overkill method will depend on the impact of the cycle on the fluids and container attributes. If the impact is too high, then alternatives to overkill cycles may be developed.

Sterilizing Filtration of Liquids

General Chapter <1229.4> Sterilizing Filtration of Liquids describes how filtration is a retentive, nondestructive process relative to the microbial cell. Due to the methodology’s nondestructive nature, critical factors that can impact retention of microorganisms include the conditions of filtration, characteristics of the filter, and bioburden. A good understanding of these parameters will enhance the potential for successful retention.

Radiation Sterilization

General Chapter <1229.10> Radiation Sterilization is not meant to repeat what is already described in the ANSI/AAMI/ ISO 11137 standards. Due to the importance of determining radiation dose, however, the general chapter will briefly discuss dose setting per ANSI/AAMI/ISO standards. This is the only commonly accepted sterilization method that does not require biological indicators for validation due to the accuracy of dose measurement and its extensive correlation to microbial destruction. It is important to understand the bioload or bioburden of material being irradiated.

Vapor Phase Sterilization

General Chapter <1229.11> Vapor Phase Sterilization focuses on the use of vaporized chemical agents for sterilization and decontamination, which is common practice, especially in isolators used in manufacturing. Currently, the technology of vapor sterilization mandates that two phases can potentially occur simultaneously—a gas phase and a liquid phase. Kill rates for each phase appear to be quite different. Vapor phase processes require careful process control to achieve reproducible efficacy.

Monitoring of Bioburden

General Chapter <1229.3> Bioburden Monitoring discusses bioburden control and monitoring as they relate to sterilization in particular. Since an understanding of bioburden is crucial to appropriate sterilization design, this chapter will enhance the understanding of key elements in detection and identification of microorganisms with regard to survival, population and related patient health risks.

Depyrogenation

Parenteral products not only need to be sterile, but also free from harmful levels of pyrogens, or fever causing agents. For the purposes of this chapter series, “depyrogenation” refers to the destruction or removal of bacterial endotoxin, the most prevalent pyrogen. Depyrogenation may be accomplished by a variety of methods and processes which may be combined to assure endotoxin reduction to a safe level. The intent and processes for depyrogenation are very different from those of sterilization. Thus, for greater clarity of understanding the methods intended for depyrogenation have been separated into the <1228.x> series of general chapters. Commonly used depyrogenation processes and associated control measures will be the subject of this chapter series:

<1228> Depyrogenation

<1228.1> Depyrogenation by Dry Heat

<1228.2> Depyrogenation by Chemical Inactivation

<1228.3> Depyrogenation by Filtration

<1228.4> Depyrogenation by Physical Means

<1228.5> Endotoxin Indicators for Use in Depyrogenation

<1228.6> Endotoxin Control and Monitoring

<1228.7> Other Endotoxin Reduction Methods

Monitoring of Aseptic Environments

Another major consideration for manufacturers with regard to microbial presence is contamination control. Keeping pace with changes in regulations and technologies, General Chapter <1116> Microbiological Control and Monitoring of Aseptic Processing Environments has recently undergone a major revision and became official in 2012. By changing the focus from evaluation of cleanrooms to key guidelines that support aseptic pharmaceutical processing environments, revised General Chapter <1116> stresses prevention of contamination rather than merely attempting to measure contamination using methods unlikely to correlate with process outcome.

Recommendations in the general chapter as well as monitoring parameters given for microbiological evaluation should be applied only to cleanrooms, restricted-access barrier systems (RABS) and isolators used for aseptic processing. Changes to <1116> include clarification of limitations of counting methods used in microbiological evaluation, including sampling, recovery, data tracking and trend analysis. The use of microbial recovery frequency provides a more practical and useful concept for indicating that an environment is in a state of “control.”

The general chapter provides a key description of microbiological incubation conditions relative to intended recovery.

Bioburden Control of Nonsterile Drugs

There is very little information available regarding bioburden control for nonsterile pharmaceutical products, in the pharmacopoeias or regulatory guidance documents. Clearly, the quality of raw materials, the microbiological cleanliness of processing equipment, and proper facility design are just some of the factors that can contribute to the bioburden of a product. General Chapter <1115> Bioburden Control of Nonsterile Drug Substances and Products outlines a risk-based approach to control potential contamination in nonsterile product manufacturing. It is important to understand that the manufacture and management of microbiological content of nonsterile products are distinctly different from what is required for sterile products. Therefore, the contamination recovery rates defined in <1116> Microbiological Control and Monitoring of Aseptic Processing Environments are not intended for nonsterile environments. By looking at specific factors that may affect product quality and patient safety and considering the best approaches to dealing with them, manufacturers can identify the risks associated with a product and apply appropriate methods for bioburden control.

Replacing Conventional Microbial Tests

Conventional microbiology tests currently found in pharmacopoeias, such as sterility tests, rely on the demonstration of microbial growth. Limitations of these tests include their low sensitivity as well as their time- and labor-intensive nature. Certain cytotherapy or regenerative medicine products and radiopharmaceuticals are administered to patients prior to results from sterility testing. A more rapid result test would be very beneficial in those cases. USP is seeking to identify new referee tests or procedures (used by FDA or third parties to assess regulatory compliance) based on modern methods that can detect and enumerate microorganisms in a more rapid and sensitive manner. Objectives for such test method development include broad application, multisourced and nonpatented instrumentation and supplies and the method applicability in any laboratory setting.

Protocols are being developed to conduct proof-of-concept studies to demonstrate the reliability and suitability of the proposed methods. Based on these studies, General Chapter <71.1> Rapid Sterility Tests will be developed and published in Pharmacopeial Forum for public comments.

USP also is currently in the process of revising General Chapter <1223> Validation of Alternative Microbiological Methods, which provides guidelines for validating alternative microbiological methods, including those based on modern technologies.

On Sept. 8–9, USP will host a workshop titled, Alternative Microbiological Methods—A Workshop on Current Status and Future Directions of Compendial Standards, which will take place at USP’s headquarters in Rockville, Md. (tinyurl.com/mtuoedo).

Conclusion

USP is committed to continuous revision and improvement of its standards and values the input of users in the field. Indeed, this input is critical to the success, not only of USP, but of the industry as a whole. Stakeholder feedback helps to ensure that USP’s standards are sufficiently comprehensive in application or scope and reflect current and emerging practices in industry.

About the Author

Radhakrishna Tirumalai, PhD, is currently a Principal Scientific Liaison in the Global Science and Standards Division.