The time is rapidly approaching when a process will be filed for a high-pressure-processed, shelf-stable low-acid food in a hermetically sealed container.
High-pressure-sterilized (i.e., shelf-stable) low-acid foods are not yet available, but high-pressure-pasteurized, high-value-added specialty products such as guacamole and oysters are currently being marketed in the United States, and jams, jellies, fish, meat products, sliced ham, salad dressing, rice cakes, juices, and yogurt are being marketed in Japan and Europe. As the technology matures (and cost comes down), we will see introduction of high-volume commodity products such as milk and orange juice.
Although the Food and Drug Administration’s regulation for “Thermally Processed Low-Acid Foods Packaged in Hermetically Sealed Containers” found in Title 21, Part 113 of the Code of Federal Regulations was not intended for pressure-processed foods, it is likely that it is applicable, since high-pressure processing (HPP) of low-acid foods has a strong thermal component. Thus, for pressure-sterilized low-acid foods, all of the requirements in 21 CFR 113 must be fulfilled before a product can enter commercial production.
One of the key requirements of the regulation is the establishment of a scheduled controlled process by an individual or organization with expert knowledge of the process. A process authority must establish the process using the appropriate scientific methods to demonstrate the safety.
A low-acid, high-pressure sterilization process must be able to reproducibly produce a safe product under carefully defined conditions. The sterilization process design can then be translated to production on a routine basis to assure that the process provides the required assurance of sterility (Pflug, 1995). Successful HPP validation will require a multidisciplinary approach integrating basic microbiological, physical, chemical, and engineering principles to demonstrate the uniformity and sterility of the process. Traditional thermal processes are described by the application of steam heat for a designated length of time. Both time and temperature are factors in HPP sterilization, but the unique contribution of pressure must also be considered. Product characteristics such as viscosity, density, composition, pH, and water activity may affect the physical sterilization process and the microbial inactivation kinetics.
Food Composition Considerations
The efficacy of HPP treatments for inactivation of vegetative bacteria in foods has been demonstrated (Cheftel, 1995; Smelt, 1998; Farkas and Hoover, 2000). However, there are limited studies on the efficacy of HPP on spore inactivation (Gola et al., 1996; Rovere et al., 1996; Reddy et al., 1999; Meyer et al., 2000; Okazaki et al., 2000).
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The kinetics of HPP have been accurately refined to mathematically predict microbiological lethality for some spores under defined conditions. However, there is insufficient experience to predict the behavior of these spores across the range of microorganisms, foods, and conditions that would be encountered under commercial conditions. Significant questions remain unanswered as to the mechanism of action of the pressure, spore injury, and uniformity of the treatment delivered. Often, food composition can have a protective effect, and it is important to evaluate HPP microbial resistance in the actual foods rather than in traditional buffer solutions. Thus, each type of food must be investigated separately on a case-by-case basis until sufficient experience is gained to build a generalized model.
The picture for HPP is further complicated, since much of the initial pasteurization work did not consider the thermal component of pressure inactivation. As a food is pressurized, compression of the food results in heating of the product. As soon as the pressure is released, the temperature returns to its starting value. This temperature transient for water is in the range of 2–3°C for every 100 MPa and depends on the composition of the food. For oils and fats, the compression heating value can be as high as 9°C/100 MPa (Rasanayagam et al., 2001). A process conducted at 600 MPa with water-based inocula will thus be increased 12–18°C. This compression heating can often account for an initial decrease of 2–3 logs of the bacteria being studied. This effect is transient as a result of the rapid equilibration under the compression conditions. Thermal conductivity is also greatly accelerated, since the molecules are in close proximity, facilitating vibrational energy transfer. The resulting uniformity should contribute to the consistency and reproducibility of microbial inactivation studies.
Proposed Validation Criteria
Commercial sterility is defined in 21 CFR 113 as “a process, which renders a product free of pathogens and spoilage organisms under normal conditions of storage and distribution.” In the absence of prescriptive regulatory standards, two strategies have historically been used to demonstrate the safety component of commercial sterility: (1) a process that delivers a 12-decimal (12-D) reduction of the heat-resistant spores of Clostridium botulinum (common industrial practice); and (2) a calculated process that results in probability of a nonsterile unit having a pathogen of 10–9.
With the historical background of thermal processing, the question remains as to how to validate a pressure-accelerated thermal process to satisfy the safety requirements of 21 CFR 113. The following approaches could be used for validation of low-acid HPP processes:
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• Consider HPP a Conventional Thermal Process. This approach utilizes compression heating to take the product from the minimum initial temperature to the final process temperature. The lethality of the process would be calculated according to conventional thermal processing standards, such as by the Ball method (Pflug, 1995). This method does not take into account the enhanced le thality due to the contribution of pressure. The process would be established as follows:
1. Determine the compression heating value for the food using a compression-heating apparatus.
2. Determine the appropriate thermal process required (example F0 = 5). Compression heat the food to the process temperature, and hold for the calculated cook time. Establish a thermal process for the food product.
3. Confirm the process with a biological validation.
An example would be a potato packaged in water in a flexible container. The potato would be preheated to a temperature of 90°C in a water bath, then transferred to a high-pressure vessel, pressurized to 700 MPa, and held for 30 sec. The final temperature of the potato after compression would be 132°C (assuming a compression heating value of 6). The lethality delivered by the process would be F0 = 5.0 (assuming a Z value of 10°C). This method is considered the simplest approach to validation and would be easy to implement.
Caveat: There is very little scientific evidence or theory to support increased resistance, i.e., that pressure provides any protective effect on bacterial spores.
• Demonstrate a 12-D Process with a Biological Validation Using C. botulinum Spores. As in thermal processing, C. botulinum spores also appear to be the most resistant to pressure. In theory, a 12-D process can be validated by harvesting 1012 spores and then subjecting them to the process. In practice, this approach can be difficult because of the inherent risk associated with culturing a large number of C. botulinum spores and the practical limitations in working with essentially a pure suspension of spores.
To date, only a limited number of C. botulinum strains (types E, A, and B) have been tested. Nonproteolytic type B spores have been identified as the most-pressure-resistant spore-forming pathogens found to date (Reddy et al., 2001). No inactivation has been observed using conditions of less than 600 MPa and 90°C. Studies of this organism have not shown any pressure sensitivity. Further studies are necessary to verify that this is actually the most-resistant strain of C. botulinum. A combination of temperature (in excess of 100° C) and pressure (up to 700 MPa) may be required to exceed the threshold conditions for inactivation of this organism.
Caveats: The process should be demonstrated with the most-resistant strain of C. botulinum associated with that food. At present, there is inadequate knowledge of C. botulinum resistance to select the spore with the greatest resistance. Microbiological validation using multiple strains of C. botulinum spores will be necessary to address this issue.
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• Demonstrate a 12-D Process Using the Inactivation Kinetics for C. botulinum. If a suitable kinetic model can be developed which will demonstrate that inactivation of C. botulinum is linear over a range of values, then it may be possible to determine a process by establishing a D value and calculating a 12-D process. Modeling of conventional thermal processes has been ongoing since the Ball method for doing process calculations was advanced. Process calculations are relatively easy for retort processes, since the exact temperature of the coldest point in the container can be accurately determined.
The most straightforward approach for getting a high-pressure process accepted would be to demonstrate that HPP behaves according to the equations established for conventional thermal processing, with a third term, pressure, added to the equation. Several investigators have taken that approach, defining the lethality in terms of a pressure-dependent log-linear constant called Zp and using transition-state volume changes to model process lethality. The fundamental basis for this approach is the time-dependent exponential decrease in the number of surviving bacteria under defined conditions.
Ignoring studies that utilize pulsing, a convincing argument can be made that HPP processes can be modeled using classical microbiological or chemical kinetics. Under defined conditions of temperature and pressure, an exponential decrease in number of bacterial spores can be predicted. Thus, to validate a process, a demonstrated 6-log reduction of the most-pressure-resistant type of C. botulinum under defined conditions for X length of time can be extrapolated to a 12-log reduction in 2X time.
Caveats: This approach will account for the lethality contributed by the pressure component. The initial validation approaches may be based on biological challenge studies, and mathematical modeling of the process will follow. In all likelihood, scientific consensus on linearity of HPP spore inactivation kinetics will not be accomplished prior to commercialization of the technology. The applicability of first-order kinetics for HPP spore inactivation is not yet conclusively verified. Accordingly, extrapolation of a 6-log reduction to a 12-D process requires systematic scientific demonstration.
• Demonstrate a 12-D Process Using a Surrogate Organism. An alternative approach for demonstrating a 12-D “bot” process would be to identify a surrogate organism with pressure resistance significantly greater than that of C. botulinum. Surrogate organisms are nonpathogenic microorganisms selected from the population of well-known organisms that have well-defined characteristics and a long history of being nonpathogenic. At present, a suitable surrogate organism having a pressure resistance greater than that of C. botulinum has not been identified.
The HPP resistance of the surrogate and the C. botulinum would have to be accurately known so that comparisons could be established. It would then be possible to show that an X-log reduction of the surrogate is equivalent to a 12-log reduction of C. botulinum. Since the relative resistance would also depend on the recovery characteristics of the food product, both the C. botulinum and the surrogate resistance studies would need to be done in the intended food.
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• Consider the Probability of a Non-Sterile Unit Pathogen <10–9. This method requires a precise mathematical model of bacterial lethality to demonstrate that the initial load of pathogens in a product can be sufficiently reduced to yield a product with a 10–9 probability of a pathogen surviving. A historical record of the incidence and level of that pathogen in the raw material is required to have accurate calculations.
This method requires that an accurate model exists for calculating pathogen inactivation. In the absence of detailed scientific knowledge of microbial resistance using a specific product, screening of the raw product for the most-resistant spore is a common practice in traditional thermal processing industrial practice. Pflug (1995) suggested heating the raw product at 100°C for up to 15 min. This will eliminate vegetative bacteria and allow spores to survive. From this surviving-spore list, the most resistant test organism can be identified for process validation. Over time, a similar biological validation approach could be used for validation of the first generation of HPP low-acid foods.
Caveats: There are no historical data on the microbiological load of pressure-resistant spores in any food product. Therefore, extensive sampling would need to be conducted to determine the anticipated maximum spore load in the product prior to calculating the needed process. Existing spore data on thermal process resistance would not be applicable to a pressure-accelerated thermal process. Methodology has not been developed for enumerating the number of pressure-resistant spores in incoming raw materials.
Proposed Validation Approach
Each of the approaches proposed above has advantages and disadvantages. Whether a processor chooses a single-pulse or multiple-pulse (Meyer et al., 2000) pressure treatment, the above questions remain to be discussed. With the existing and not-unlimited resources of the food industry for the National Center for Food Safety and Technology’s high-pressure, low-acid sterilization research program (see sidebar, facing page), it is imperative to choose one or more pathways to focus the efforts. We propose the following options:
• Establish the Process as a Thermal Process. Filing of the process as a purely thermal process will be the first priority. The process for mashed potato will be investigated and the necessary data gathered.
The thermal process will be characterized using the thermo-couples in the high-pressure vessel. Temperature distribution, compression-heating values, medium temperature, loading factors, preheat minimum initial temperature, varietal characteristics, basket heating, barrel temperature, and process controls will be determined. An F0 = 5 process will be filed. This process is consistent with what is required for commercial sterility (spoilage and public health).
At this time, we do not have adequate data to verify whether HPP spore inactivation is similar to thermal spore inactivation. To elucidate the behavior of spores under HPP+thermal vs thermal alone, the lethality under HPP (at the same F0 value calculated on the basis of temperature/time alone) will be compared against positive thermal-only control.
A microbiological validation will be done using the most-resistant C. botulinum spores at a level of 105 per container, distributed homogeneously throughout the product. The product will be brought to the process minimum value for times representing F0 values of 0.5, 1.0, 1.5, 2.0, and 2.5 min.
The survivor curve will be developed using a most-probable-number calculation based on the fraction negative. The packages will be incubated for 90 days past the last blown package, and each package will be subcultured to confirm that it was negative. The expected results of the incubated pack will be calculated prior to conducting the study, and a protocol will be agreed on. A thermal D value at 121°C will be used as a reference process. This test will establish a Dp-value at the process minimum under pressure as well. The results will then be used for the next round of process filing.
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• Establish the Process Using a Kinetic Approach. This involves the following steps:
1. Identify the product (mashed potatoes).
2. Inoculate the product with relevant bacterial spore strains of concern (C. botulinum) in a specific product. A cocktail approach to determine the resistant strain may be used. Ascertain the resistance and verify the harvesting procedure. Verify that the most-resistant strain has been chosen for the product by testing a broad spectrum of spores from various laboratories.
3. Inoculate the product with 106 spores per package. Process at various times at the predetermined minimum process. Determine the D value and calculate the 12-D process. Establish the resistance of pertinent spoilage organisms and calculate a process to also address the spoilage. Surrogate resistance can proceed in parallel with this study.
4. Choose the desired F0 value at the process pressure (based on food safety or spoilage criteria).
5. Conduct biological validation runs over a range of pressures and temperatures to validate the designed process.
The kinetic approach will require extensive systematic scientific studies. In the case of thermal processing, expert scientific knowledge has been accumulated over a period of a hundred years. However, with several laboratories working on the project, it should be possible to make rapid progress toward a conclusion. It will be possible to distribute the task over various worldwide labs and significantly reduce time required to generate the information. Still, it may require at least two years of research. In the interim, food processors may choose to validate the process for specific product(s). The processed product may be distributed as an extended-shelf-life (ESL) product until sufficient confidence about the process is attained.
High-pressure processing of low-acid foods offers unique opportunities and challenges to the food industry. Like thermal processing, HPP may be used commercially before it is completely understood scientifically. Research is still being conducted on thermal processing. The first generation of commercial high-pressure-processed products may be shelf stable, but optionally could be distributed under refrigeration as ESL products. The technology would enable processors to make novel, minimally processed, ESL, convenient food items with fresh-like attributes and natural-looking colors. Identification of commercially viable products would be a major challenge. Development of validation criteria for safe processing of shelf-stable foods will evolve as experience is gained by the first generation of products.
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Consortium Studies High-Pressure Processing
A consortium of researchers from industry, government, and academia is studying the feasibility of producing high-quality, shelf-stable, low-acid foods using high-pressure processing, with the goal of submitting a process filing to the Food and Drug Administration.
In 2000, the National Center for Food Safety and Technology—the nation’s only research consortium which addresses the food safety implications of emerging technologies in food processing, packaging, biotechnology, and HACCP concepts—and Flow International Corp., manufacturer of high-pressure equipment, formed, with other partners, a Dual-Use Science and Technology (DUST) consortium to pursue regulatory acceptance, market development, and equipment demonstration for commercialization of high-pressure processing of low-acid foods.
During its first year, the consortium conducted tests to determine the effect of pressure, temperature, and time on selected food components and packaging materials. Flow International installed a pilot 35-L high-pressure sterilization system at NCFST for DUST member use. The 8-metric-ton unit, shown in the photos below, can process up to 17 L of samples. DUST will focus its second- and third-year efforts validation of a high-pressure process for low-acid food, using the sterilization system.
In the process, a food prepackaged in flexible containers is placed into a stainless-steel cylinder and preheated in a water bath. The cylinder is then raised, inserted into the pressure vessel, and processed. After the desired pressure and time treatment, the cylinder is placed in a cooling bath, then the contents are removed for analysis.
NCFST is located at Illinois Institute of Technology’s Moffett Campus in Summit-Argo, Ill. Flow International is located in Kent, Wash. The current consortium partners are Basic American Foods, Con Agra Grocery Products Co., Hormel Food Engineering Div., Kraft Foods, the U.S. Army Soldier and Biological Chemical Command at Natick, Mass., Unaka Business Development, and Washington Farms. FDA scientists located at NCFST serve as advisers. Edmund Ting of Flow International heads the consortium efforts, and V.M. (Bala) Balasubramaniam (shown in left photo) is the principal investigator for NCFST efforts.
This work was supported in part by a grant from a consortium of companies participating in a U.S. Army Dual Use Science and Technology Program on highpressure processing of low-acid foods.
by Charles E. Sizer, V.M. (Bala) Balasubramaniam, and Edmund Ting
Author Sizer is Professor and Director, and author Balasubramaniam is Associate Professor of Food Process Engineering, National Center for Food Safety and Technology, Illinois Institute of Technology, Moffett Campus, 6502 S. Archer Road, Summit-Argo, IL 60501. Author Ting is Vice President, Flow International Corp., 23500 64th Ave. S., Kent, WA 98032. Authors Sizer and Balasubramaniam are Professional Members of IFT. Send reprint requests to author Balasubramaniam.
Edited by Neil H. Mermelstein, Editor
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