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Technologies in use and on the horizon could change aseptic processing in ways that seemed inconceivable years ago but approaches to aseptic process validation still need to move beyond their 1970s roots.
Since 1988, when the Parenteral Drug Association first reviewed industry practices (1) and expectations, and 2004, when the authors last took stock of changes in technology and practices (2), the aseptic processing landscape has changed dramatically, with advances in equipment design, altered perspectives, and regulatory updates. (3-9) This article reviews some of the extensive changes that have taken place in aseptic processing within the past 15 years, many of which surpass “revolutionary” and might be considered “extraordinary.”
Despite this progress, one problem that the authors noted in 2004 persists: the idea that contemporary microbiological methods can recover all microorganisms with a limit of detection that approaches zero. As a result, contemporary approaches to aseptic process validation still reflect their 1970’s genesis far more than they should. This is particularly evident with excessive lethality requirements for terminal sterilization, as well as environmental monitoring, media-fill sampling, and continued over-reliance on visualization methods such as smoke studies.
At a time when aseptic processing technology is automating processes and reducing or eliminating the potential for contamination, it is time to revise the industry’s approach to validation and its expectations for aseptic processing. This article highlights challenges that are emerging for 2020 and beyond, in light of improvements that have been made.
Many in the industry continue to believe that most drugs and biologics cannot withstand a sterilization process in their final containers, so most of the parenterals and other products labeled sterile are manufactured using aseptic processing. New thinking about sterilization processes might change that, at least for some products. For the last 40 or so years, sterilization has focused primarily on concerns for microbial destruction, and sterilization validation had a singular focus on inactivation of a highly resistant, high population biological indicator. This resulted in the unintended consequence that terminal sterilization processes became increasingly, and even unreasonably, arduous.
Overly aggressive, scientifically unnecessary approaches to sterilization resulted in excessively lethal processes that exposed products to extreme conditions. Inevitably, these conditions affected products’ essential quality attributes, and increased the industry’s reliance on aseptic processing as opposed to terminal sterilization processes (9-12).
For more than 20 years, the industry and regulatory communities have come into increasing conflict with respect to aseptic processing. The dilemma arises directly from the seemingly logical, but scientifically flawed notion that it is possible to prove that each and every lot of aseptically manufactured product contains only “sterile” material. Sterile is, if taken in its most absolute meaning, a word that offers no room for uncertainty in measurement or outcome. Sterile means free of any viable organisms. The ability to measure microbes in the environment is not pertinent to risk assessment when the process operates consistently below practical assay capabilities. Processes operating below the limit of microbial detection are desirable. Yet, when they reach that level, existing monitoring requirements actually increase risk by requiring operator interventions that can introduce contamination.
The introduction of advanced aseptic processing systems that exclude gowned personnel from the critical environment has fostered a new understanding, which defines a new reality grasped by some, but actively opposed by others. Sterility is attained by the proper implementation of practices that, when combined, provide the protection from contamination that sterile products require. Sterility by design (13-15) describes a state where confidence in the output is realized by specific intent across the many contributing systems. As a result, improvements in aseptic process performance have created a conundrum. Microbial test methods are no longer useful because contamination levels have fallen below the levels of detection. This should be recognized as a problem modern microbiological methods can’t readily resolve. Advanced aseptic processing systems have become so robust that in many closed, state-of-the-art systems, contamination is no longer a concern, while, in others, contamination is so well controlled (when assessed by methods that were intended for use in traditional cleanrooms) that it is irrelevant. Nevertheless, present day regulatory beliefs and expectations continue, not only to fixate on sampling and testing using methods that have little utility, but to demand that their use increase.
Perhaps the industry erred by labeling aseptically manufactured products “sterile.” However, little if any harm has come from this approach. No current standard explicitly calls for zero contamination in media-fill process simulation tests or in environmental monitoring, yet regulatory documents expect zero or near zero contamination levels (16,17). Zero may be valid as an aseptic processing target, but it is important to remember that no aseptic environment or aseptically produced product can be proven to be sterile.
In no other segment of the pharmaceutical industry is the control of manufacturing processes as critical as in the production of aseptically produced products. That recognition has led to the development of advanced production and quality assurance systems. Firms have continued to develop and implement more rigorous methods for the validation of aseptic processes, summarized in the following sections.
Sterilization of components is a negligible risk factor in aseptic processing. Equipment suppliers, clean room suppliers and the pharmaceutical industry itself have responded by systematically reducing the risk of contamination. Growing implementation of automation and advanced aseptic processing technologies will incrementally reduce risks associated with human contamination (3,8,9) in aseptic processing, as shown in Table I. The primary source of contamination disappears in advanced aseptic technologies in which operators are no longer present!
The methods used for component sterilization in the manufacture of glass and elastomeric-stopper container systems have not changed. However. increasingly, suppliers of these materials are supplying them in either a ready-to-sterilize or ready-to-use condition. This trend is partially due to increased use of outsourcing, and the emergence of smaller firms that lack the infrastructure to prepare these materials on a limited basis.
Moist-heat sterilization continues to be the most widely used sterilization method (18). For glassware, dry heat continues to be the method of choice for sterilization and depyrogenation. Regulatory expectations for depyrogenation are a minimum three-log reduction of reference standard bacterial endotoxin. Changes in the US Pharmacopeia’s (USP’s) standards suggest that use of naturally occurring endotoxin is a more appropriate challenge given that there is no standardized resistance for dry heat endotoxin indicators (19, 20).
Industry scientists are aware that glass containers are manufactured at high temperatures that reduce the pyrogen burden on glass below the limit of detection. Also, present day water systems are so well controlled that the potential for pyrogen contamination of glass during washing steps is exceedingly low.
Process filter validation
The basic principles involved in product filtration have remained unchanged for more than 30 years (21). Firms are placing greater emphasis on assessment of pre-filtration bioburden and use these data to confirm the appropriateness of their filtration processes on a lot-by-lot basis. The highly specialized nature of filter validation generally requires considerable participation by filter vendors, but it remains the responsibility of the user to ensure each filtration process is appropriate for its intended purpose.
Process simulation—media fills
Regulatory interest in media fills has increased over the last decade with expectations that a firm’s media-fill program adequately supports its everyday aseptic processing practices. Intervention frequency and their alignment with operating procedures are key areas of concern (22–25). In operations where fill speeds of 200 or more units/minute are attainable, the duration of a media fill in which the target population was only 5000 would result in a media fill that might last roughly 30 minutes, not including set-up. However, requiring media-fill tests that are a high percentage of the total number of units ordinarily filled in a batch is not necessary to provide an assessment of process capability. Media-fill populations of more than 10,000 units are rarely if ever required even for high throughput operations. With advanced technologies operating so effectively, this historically important test (26–28) has far less utility than it formerly did (Table II).
One reason given for longer duration fills is to evaluate the effect of operator fatigue. Abundant means exist to qualify personnel for aseptic operations without the requirement for their participation in a media-fill test. Employees can be individually evaluated in terms of gowning proficiency and laboratory simulations can be used to evaluate their aseptic technique.
Additionally, operators should be comprehensively trained on equipment operations, relevant operating procedures and work instructions. The adoption of detailed intervention practices, closely followed by all operators, is preferable. A firm should develop a rationale for its container/closure system selection based upon a careful analysis of risk, and establish a media-fill program that supports the operational variations of container, closure, and line speed. It is important to remember that a media fill test is a snapshot in time and is not predictive of future outcome or informative regarding previously manufactured product (29, 30). In the future, the industry must develop validation and process-control methods built around engineering principles relating to the statistical demonstration of process capability against process parameters rather than overly relying on microbiological evaluations and invasive sampling practices.
Environmental control and facility management programs
The basic elements of an aseptic processing environmental control program (i.e., viable air, surface, and personnel monitoring as well as particle monitoring) have not changed meaningfully since their introduction.
The classification of each room or module within an aseptic processing area must be appropriate for its intended use (16,17, 31,32). The highest level of control will be directed to critical zones in which aseptic manipulation of sterile materials and surfaces occurs. These areas are designed to comply with Class 5 of the International Standard Organization’s (ISO’s) 14644 (ISO Class 5 is functionally equivalent to traditional US Federal Standard [FS] 209 E Class 100, and to European Union Grade A in terms of total particulate air quality). ISO-5 environments are customarily equipped with total coverage high-efficiency particulate air (HEPA) filtration, and unidirectional airflow is maintained to the extent that it is technically possible. Isolators that can maintain ISO-5 conditions without unidirectional air do not require full HEPA coverage (33). European and US aseptic processing area zoning differs in that the area immediately adjacent to the critical zone in the USA is typically Class 7 [FS 209 Class 10,000] while in Europe this area is Grade B, for which there is no precise analog in either ISO 14644 or the withdrawn FS 209E classification schemes. These classification schemes are essentially equivalent. Firms may use different approaches, provided they have a rationale that is based upon good scientific and engineering practices (34).
Other parameters typically considered in the design of an aseptic processing area are direction of airflow, air balance, air changes/hour and air velocity. Numerous engineering guidelines exist that offer sound design recommendations for aseptic processing areas. Today’s regulatory environment tends to overemphasize the importance of both air velocity and uniformity of airflow. The 90-feet/minute or 0.45m/s air velocity expectation is a reasonable target value. However, it is unreasonable to conclude that these values are sacrosanct (35).
High air-exchange rates that are necessary in manned cleanrooms are of much less value in isolators, where the sources of contamination are absent. Isolators operate quite well at air velocities of 0.2m/s or less. When air velocity was last included as a cleanroom design specification in FS 209C, the measurement was taken approximately one foot from the face of the HEPA filter. There is no basis to require that any specific air velocity be attained at the work surface. Because the work surface is located perpendicular to the direction of airflow and cannot be made aerodynamic, maintenance of strict unidirectional airflow at the work surface is not possible and cannot be reasonably expected (36).
Visualization of airflow may provide for optimization of air movements (37). Visualization of air flow (commonly called “smoke studies”) is entirely subjective, however, because it is based on human observations of the outcome rather than objective criteria. Regulators have overemphasized its importance, which should not take precedence over objective data.
This is one area in which aseptic processing may actually have regressed. Given vastly improved equipment design, the utility of environmental monitoring has diminished. Comprehensive environmental monitoring programs continue to be required for classified environments, however, and they are becoming increasing expansive, even though there is no evidence that today’s programs are any better than those of the past.
Sampling methods and approaches have remained largely unchanged since 1970; what has changed is the amount of monitoring performed and the actions expected by inspectors in the event of excursions. Environmental excursions (i.e., results that exceed action levels) occur at a much lower frequency than ever before.
In response to regulatory pressure, the industry is misusing environmental monitoring and increasingly ignoring the uncertainty and inaccuracy inherent in the practice. In too many instances, sampling results are being considered like a product release microbiological quality assay. In the most extreme cases, samples taken of “critical surfaces,” environmental monitoring has evolved into a de facto product release sterility test. This is scientifically inappropriate. Environmental monitoring excursions should not trigger the application of out-of-specification (OOS) test interpretation and resolution (38,39). The FDA’s OOS guidance was never intended for application to microbial testing, and this position has often been reaffirmed by FDA personnel.
Environmental monitoring must consider the realities of microbiological growth and recovery. The measurement accuracy of active air samples, erroneously called “quantitative” air samplers, is limited, and variability among commonly used active air samplers can exceed five-fold (37). Clearly then, setting an action level of 3 colony forming units (CFU) and an alert level of 1 CFU is illogical. Additionally, microbiological enumeration, particularly at low organism titers, is statistically unreliable. It is more appropriate, scientifically, to monitor trends by evaluating contamination incidence rates rather than by emphasizing colony counts (40).
Environmental sampling plans should include representative sites in the ISO Class 5 critical zone as well as the surrounding environment in conventional manned cleanrooms.The more critical sites should be sampled daily. Non-critical surfaces such as walls and floors should be sampled weekly to ensure that the firm’s disinfection program is performing adequately. Routine environmental sampling of personnel should be limited to glove testing at the conclusion of their stay in the aseptic environment. Sampling of personnel during operations is ill advised because it increases operator activity and risks increasing contamination on gowns and gloves. Post-production sampling of product contact surfaces (e.g., parts, hoppers, and filling needles) can be done, but results should not be considered indicative of sterility or asepsis, and certainly not the lack thereof. Sampling is a manual intervention and subject to adventitious contamination, independent of the process activity. Studies at PDA’s Training and Research Institute (41) show that surface contamination is a poor predictor of media-fill outcome. The industry must focus on product safety, rather than an absolute, abstract expectation that a perfect sterile environment must exist (42,43).
Advanced aseptic processing includes technologies that reduce the risk of human-borne contamination through automation and/or environmental designs, such as isolators, closed restricted access barrier systems (RABS), blow-fill-seal (BFS) or form-fill-seal (FFS) technologies, and various types of machine automation. In 1988, the first isolator-based aseptic filling systems were just being implemented; today isolator systems represent the state of the art for commercial scale aseptic filling. BFS and FFS have been used for nearly 50 years and undergo continuing improvement.
RABS, which were all the rage in 2004, vary significantly in design and operation and have lost some of their appeal. The industry continues to be cautious in adopting isolators, despite overwhelmingly positive experience with the technology. This can be traced to archaic design and monitoring expectations derived from manned cleanrooms (42). Improvements in process equipment have reduced the dependency on operators in many aseptic filling systems. Automatic check weighing, integrated clean and steam-in-place, in-line automated inspection equipment, remote fill adjustment and other advances have dramatically improved filling equipment. Robots are being used for transfer of components, loading and unloading of lyophilizers and even, paradoxically, environmental monitoring. The emergence of closed aseptic processing systems provides a variety of opportunities for enhanced product and worker protection. Existing aseptic technologies, and certainly those just coming into use, provide such superior results when judged by the long-standing monitoring means of sterility testing (i.e., environmental sampling and process simulation) that the means for their evaluation must be reconsidered. In short, today’s advanced aseptic technologies have exceeded past expectations, but unless validation practices catch up, the industry risks making the perfect the enemy of the excellent.
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James Agalloco, firstname.lastname@example.org, is principal of Agalloco & Associates Inc.; James Akers, PhD, is principal of Akers Kennedy & Associates Inc.; Russell Madsen is director of The Williamsburg Group, LLC.
All tables courtesy of James Agalloco.
Vol. 33, No. 8
When referring to this article, please cite it as J. Agalloco, J.Akers, R. Madsen, “Aseptic Processing Practices: Reviewing Three Decades of Change ,” BioPharm International33 (8) 2020.