Heat and Light Stability of Pumpkin-Based Carotenoids in a Photosensitive Food: A Carotenoid-Coloured Beverage.

This study aimed to evaluate carotenoid degradation kinetics in a beverage coloured with pumpkin juice concentrate during storage at dark and illuminated conditions at four temperatures (10, 20, 35 and 45 °C). Carotenoids were quantified by HPLC-DAD, and kinetic parameters for carotenoid degradation were estimated by one-step nonlinear regression analysis. During dark storage, degradation kinetics was modelled by fractional conversion (all-trans-ß-carotene) and zero-order equations (all-trans-antheraxanthin, all-trans-lutein, all-trans-violaxanthin and all-trans-neoxanthin). Storage of samples in a climatic chamber with intense light intensity (1875-3000 lux) accelerated the carotenoid losses. At illuminated conditions, degradation followed a first-order (all-trans-lutein, all-trans-violaxanthin and all-trans-neoxanthin) and fractional conversion model (all-trans-ß-carotene and all-trans-antheraxanthin). Carotenoid degradation followed an Arrhenius temperature-dependency, with Ea values lower than 50 kJ/mol. Degradation was shown to be mainly by oxidative reactions. Packaging under minimal oxygen conditions, use of antioxidants (e.g., ascorbic acid), and proper choice of light sources at retail shelves may be considered to optimize the pigment retention in a carotenoid-coloured beverage during storage.

Impact of processing on the production of a carotenoid-rich Cucurbita maxima cv. Hokkaido pumpkin juice.

Pumpkin juice with high carotenoid content can be attractive natural alternative for artificial food colourants. We evaluated the impact of processing treatments on aqueous carotenoid extraction from pumpkins, aiming to enhance carotenoid transfer into the juice fraction. Crushed whole pumpkins were processed by high pressure homogenization (HPH) for mechanical cell disruption, by enzymatic treatment for cell wall polysaccharide degradation or by a pulsed electric field (PEF) treatment for cell membrane electroporation. Processed purees were separated into juice and pomace and carotenoids were quantified by HPLC-DAD. Whereas only 54-60% of the carotenoids in non-processed puree was transferred into the juice, HPH- and enzyme-assisted processing of purees significantly increased juice yields and total soluble solids, and consequently, carotenoid concentrations in these juices up to 90-98% and 72-90%, respectively. No significant improvement was observed for PEF-treated samples. Results obtained can be industrially useful in producing natural colouring plant concentrates as clean-label ingredients.

Time-temperature-resolved functional and structural changes of phycocyanin extracted from Arthrospira platensis/Spirulina.

Arthrospira platensis, commonly known as Spirulina, gains increasing importance as alternative protein source for food production and biotechnological systems. A promising area is functional high-value algae extracts, rich in phycocyanin, a protein-pigment complex derived from A. platensis. This complex has proven functionality as the only natural blue colorant, fluorescent marker and therapeutic agent. The structure-function relationship is heat sensitive, making thermal processing in its production and its subsequent application a crucial aspect. In continuous high-temperature short-time treatments, it was shown how a purified phycocyanin (mixture of allophycocyanin and c-phycocyanin) disassembled and denatured between 50 and 70 °C. Three characteristic transition temperatures were allocated to specific quaternary aggregates. In contrast to sequential chemical denaturation, phycocyanin's chromophore and protein structure were simultaneously affected by thermal processing. Through a functionality assessment, the findings help optimize the efficiency of raw material usage by defining a processing window, enabling targeted process control resulting in desired product properties.

Endospore Inactivation by Emerging Technologies: A Review of Target Structures and Inactivation Mechanisms.

Recent developments in preservation technologies allow for the delivery of food with nutritional value and superior taste. Of special interest are low-acid, shelf-stable foods in which the complete control or inactivation of bacterial endospores is the crucial step to ensure consumer safety. Relevant preservation methods can be classified into physicochemical or physical hurdles, and the latter can be subclassified into thermal and nonthermal processes. The underlying inactivation mechanisms for each of these physicochemical or physical processes impact different morphological or molecular structures essential for spore germination and integrity in the dormant state. This review provides an overview of distinct endospore defense mechanisms that affect emerging physical hurdles as well as which technologies address these mechanisms. The physical spore-inactivation technologies considered include thermal, dynamic, and isostatic high pressure and electromagnetic technologies, such as pulsed electric fields, UV light, cold atmospheric pressure plasma, and high- or low-energy electron beam.

Mechanisms of enhanced bacterial endospore inactivation during sterilization by ohmic heating.

During ohmic heating, the electric field may additionally inactivate bacterial endospores. However, the exact mechanism of action is unclear. Thus, a mechanistic study was carried out, investigating the possible target of electric fields inside the spore. Bacillus subtilis spores were heated by conventional and ohmic heating in a capillary system under almost identical thermal conditions. Wild-type (PS533) spores were used, as well as isogenic mutants lacking certain components known for their contribution to spores' heat resistance: small-acid soluble proteins (SASP) protecting DNA (PS578); the coat covering the spore (PS3328); and the spore germination enzyme SleB (FB122(+)). Treatment-dependent release of the spore core's depot of dipicolinic acid (DPA) was further evaluated. Up to 2.4 log10 additional inactivation of PS533 could be achieved by ohmic heating, compared to conventional heating. The difference varied for the mutants, with a decreasing difference indicating a decreased effect of the electric field and vice versa. In particular, mutant spores lacking SASPs showed a behavior more similar to thermal inactivation alone. The combination of heat and electric field was shown to be necessary for enhanced spore inactivation. Thus, it is hypothesized that either the heat treatment makes the spore susceptible to the electric field, or vice versa.

Effects of High Pressure on Bacillus licheniformis Spore Germination and Inactivation.

Bacillus and Clostridium species form spores, which pose a challenge to the food industry due to their ubiquitous nature and extreme resistance. Pressurization at <300 MPa triggers spore germination by activating germination receptors (GRs), while pressurization at >300 MPa likely triggers germination by opening dipicolinic acid (DPA) channels present in the inner membrane of the spores. In this work, we expose spores of Bacillus licheniformis, a species associated with food spoilage and occasionally with food poisoning, to high pressure (HP) for holding times of up to 2 h. By using mutant spores lacking one or several GRs, we dissect the roles of the GerA, Ynd, and GerK GRs in moderately HP (mHP; 150 MPa)-induced spore germination. We show that Ynd alone is sufficient for efficient mHP-induced spore germination. GerK also triggers germination with mHP, although at a reduced germination rate compared to that of Ynd. GerA stimulates mHP-induced germination but only in the presence of either the intact GerK or Ynd GR. These results suggests that the effectiveness of the individual GRs in mHP-induced germination differs from their effectiveness in nutrient-induced germination, where GerA plays an essential role. In contrast to Bacillus subtilis spores, treatment with very HP (vHP) of 550 MPa at 37°C did not promote effective germination of B. licheniformis spores. However, treatment with vHP in combination with elevated temperatures (60°C) gave a synergistic effect on spore germination and inactivation. Together, these results provide novel insights into how HP affects B. licheniformis spore germination and inactivation and the role of individual GRs in this process.IMPORTANCE Bacterial spores are inherently resistant to food-processing regimes, such as high-temperature short-time pasteurization, and may therefore compromise food durability and safety. The induction of spore germination facilitates subsequent inactivation by gentler processing conditions that maintain the sensory and nutritional qualities of the food. High-pressure (HP) processing is a nonthermal food-processing technology used to eliminate microbes from food. The application of this technology for spore eradication in the food industry requires a better understanding of how HP affects the spores of different bacterial species. The present study provides novel insights into how HP affects Bacillus licheniformis spores, a species associated with food spoilage and occasionally food poisoning. We describe the roles of different germination receptors in HP-induced germination and the effects of two different pressure levels on the germination and inactivation of spores. This study will potentially contribute to the effort to implement HP technology for spore inactivation in the food industry.

Impact of surface structure and feed gas composition on Bacillus subtilis endospore inactivation during direct plasma treatment.

This study investigated the inactivation efficiency of cold atmospheric pressure plasma treatment on Bacillus subtilis endospores dependent on the used feed gas composition and on the surface, the endospores were attached on. Glass petri-dishes, glass beads, and peppercorns were inoculated with the same endospore density and treated with a radio frequency plasma jet. Generated reactive species were detected using optical emission spectroscopy. A quantitative polymerase chain reaction (qPCR) based ratio detection system was established to monitor the DNA damage during the plasma treatment. Argon + 0.135% vol. oxygen + 0.2% vol. nitrogen as feed gas emitted the highest amounts of UV-C photons and considerable amount of reactive oxygen and nitrogen species. Plasma generated with argon + 0.135% vol. oxygen was characterized by the highest emission of reactive oxygen species (ROS), whereas the UV-C emission was negligible. The use of pure argon showed a negligible emission of UV photons and atomic oxygen, however, the emission of vacuum (V)UV photons was assumed. Similar maximum inactivation results were achieved for the three feed gas compositions. The surface structure had a significant impact on the inactivation efficiency of the plasma treatment. The maximum inactivation achieved was between 2.4 and 2.8 log10 on glass petri-dishes and 3.9 to 4.6 log10 on glass beads. The treatment of peppercorns resulted in an inactivation lower than 1.0 log10. qPCR results showed a significant DNA damage for all gas compositions. Pure argon showed the highest results for the DNA damage ratio values, followed by argon + 0.135% vol. oxygen + 0.2% vol. nitrogen. In case of argon + 0.135% vol. oxygen the inactivation seems to be dominated by the action of ROS. These findings indicate the significant role of VUV and UV photons in the inactivation process of B. subtilis endospores.

Impact of different water activities (a w) adjusted by solutes on high pressure high temperature inactivation of Bacillus amyloliquefaciens spores.

Much research has been conducted to comprehend the mechanisms of high pressure (HP) inactivation of spores in aqueous systems but for food model systems these information are scarce. In these systems spores can interact with ingredients which then could possibly lead to retarded or reduced inactivation, which can cause a problem for the sterilization process. The protective mechanism of a reduced a w-value is still unclear. HP processing might prove valuable to overcome protective effects of solutes and achieve shorter process times for sterilization under HP. To gain insight into the underlying mechanisms five a w-values (0.9, 0.92, 0.94, 0.96, 1) were adjusted with two different solutes (NaCl, sucrose). Solutions were inoculated with spores of Bacillus amyloliquefaciens and treated at 105, 110, and 115°C at 600 MPa. Further a thermal inactivation was conducted at the same temperatures for a comparison with the HP data. Afterward, the influence of HP high temperature treatment on the inactivation, the dipicolinic acid (DPA)-release and membrane constitution was assessed by plate count, HPLC and flow cytometry (FCM). The results show that during HP treatments sucrose and salt both have a protective effect, in which the influence of sucrose on the retarded inactivation is higher. The threshold water activities (a w), which is 0.94, here salt and sucrose have a significant influence on the inactivation. The comparison of thermal (105-115°C) and HP and high temperature (600 MPa, 105-115°C) treated samples showed that the time needed to achieve a 4-5 log10 inactivation is reduced from 45 (a w = 1) to 75 (a w = 0.9) min at 105°C to 3 (a w = 1) to 15 (a w = 0.9) minutes at 600 MPa and 105°C. The release of DPA is the rate limiting step of the inactivation and therefore monitoring the release is of great interest. The DPA-release is slowed down in high concentrated solutions (e.g., sucrose, salt) in comparison to a w 1. Since there is a difference in the way the solutes protect the spore it could be seen as an inner spore membrane effect. Maybe as shown for vegetative microorganism the solutes can interact with membranes, e.g., the inner spore membrane. Flow cytometry (FCM) measurement data show a similar trend.

High pressure thermal inactivation of Clostridium botulinum type E endospores - kinetic modeling and mechanistic insights.

Cold-tolerant, neurotoxigenic, endospore forming Clostridium (C.) botulinum type E belongs to the non-proteolytic physiological C. botulinum group II, is primarily associated with aquatic environments, and presents a safety risk for seafood. High pressure thermal (HPT) processing exploiting the synergistic effect of pressure and temperature can be used to inactivate bacterial endospores. We investigated the inactivation of C. botulinum type E spores by (near) isothermal HPT treatments at 300-1200 MPa at 30-75°C for 1 s to 10 min. The occurrence of heat and lysozyme susceptible spore fractions after such treatments was determined. The experimental data were modeled to obtain kinetic parameters and represented graphically by isoeffect lines. In contrast to findings for spores of other species and within the range of treatment parameters applied, zones of spore stabilization (lower inactivation than heat treatments alone), large heat susceptible (HPT-induced germinated) or lysozyme-dependently germinable (damaged coat layer) spore fractions were not detected. Inactivation followed first order kinetics. Dipicolinic acid release kinetics allowed for insights into possible inactivation mechanisms suggesting a (poorly effective) physiologic-like (similar to nutrient-induced) germination at ≤450 MPa/≤45°C and non-physiological germination at >500 MPa/>60-70°C. Results of this study support the existence of some commonalities in the HPT inactivation mechanism of C. botulinum type E spores and Bacillus spores although both organisms have significantly different HPT resistance properties. The information presented here contributes to closing the gap in knowledge regarding the HPT inactivation of spore formers relevant to food safety and may help industrial implementation of HPT processing. The markedly lower HPT resistance of C. botulinum type E spores compared with the resistance of spores from other C. botulinum types could allow for the implementation of milder processes without endangering food safety.

Involvement of Coat Proteins in Bacillus subtilis Spore Germination in High-Salinity Environments.

The germination of spore-forming bacteria in high-salinity environments is of applied interest for food microbiology and soil ecology. It has previously been shown that high salt concentrations detrimentally affect Bacillus subtilis spore germination, rendering this process slower and less efficient. The mechanistic details of these salt effects, however, remained obscure. Since initiation of nutrient germination first requires germinant passage through the spores' protective integuments, the aim of this study was to elucidate the role of the proteinaceous spore coat in germination in high-salinity environments. Spores lacking major layers of the coat due to chemical decoating or mutation germinated much worse in the presence of NaCl than untreated wild-type spores at comparable salinities. However, the absence of the crust, the absence of some individual nonmorphogenetic proteins, and the absence of either CwlJ or SleB had no or little effect on germination in high-salinity environments. Although the germination of spores lacking GerP (which is assumed to facilitate germinant flow through the coat) was generally less efficient than the germination of wild-type spores, the presence of up to 2.4 M NaCl enhanced the germination of these mutant spores. Interestingly, nutrient-independent germination by high pressure was also inhibited by NaCl. Taken together, these results suggest that (i) the coat has a protective function during germination in high-salinity environments; (ii) germination inhibition by NaCl is probably not exerted at the level of cortex hydrolysis, germinant accessibility, or germinant-receptor binding; and (iii) the most likely germination processes to be inhibited by NaCl are ion, Ca(2+)-dipicolinic acid, and water fluxes.

Sterilization of liquid foods by pulsed electric fields-an innovative ultra-high temperature process.

The intention of this study was to investigate the inactivation of endospores by a combined thermal and pulsed electric field (PEF) treatment. Therefore, self-cultivated spores of Bacillus subtilis and commercial Geobacillus stearothermophilus spores with certified heat resistance were utilized. Spores of both strains were suspended in saline water (5.3 mS cm(-1)), skim milk (0.3% fat; 5.3 mS cm(-1)) and fresh prepared carrot juice (7.73 mS cm(-1)). The combination of moderate preheating (70-90°C) and an insulated PEF-chamber, combined with a holding tube (65 cm) and a heat exchanger for cooling, enabled a rapid heat up to 105-140°C (measured above the PEF chamber) within 92.2-368.9 µs. To compare the PEF process with a pure thermal inactivation, each spore suspension was heat treated in thin glass capillaries and D-values from 90 to 130°C and its corresponding z-values were calculated. For a comparison of the inactivation data, F-values for the temperature fields of both processes were calculated by using computational fluid dynamics (CFD). A preheating of saline water to 70°C with a flow rate of 5 l h(-1), a frequency of 150 Hz and an energy input of 226.5 kJ kg(-1), resulted in a measured outlet temperature of 117°C and a 4.67 log10 inactivation of B. subtilis. The thermal process with identical F-value caused only a 3.71 log10 inactivation. This synergism of moderate preheating and PEF was even more pronounced for G. stearothermophilus spores in saline water. A preheating to 95°C and an energy input of 144 kJ kg(-1) resulted in an outlet temperature of 126°C and a 3.28 log10 inactivation, whereas nearly no inactivation (0.2 log10) was achieved during the thermal treatment. Hence, the PEF technology was evaluated as an alternative ultra-high temperature process. However, for an industrial scale application of this process for sterilization, optimization of the treatment chamber design is needed to reduce the occurring inhomogeneous temperature fields.

Comparative study on the high pressure inactivation behavior of the Shiga toxin-producing Escherichia coli O104:H4 and O157:H7 outbreak strains and a non-pathogenic surrogate.

Enterohemorrhagic Escherichia coli strains cause each year thousands of illnesses, which are sometimes accompanied by the hemolytic uremic syndrome, like in the 2011 outbreak in Germany. For preservation thermal pasteurization is commonly used, which can cause unwanted quality changes. To prevent this high pressure treatment is a potential alternative. Within this study, the 2011 outbreak strain O104:H4, an O157:H7 and a non-pathogenic strain (DSM1116) were tested. The cells were treated in buffer (pH 7 and pH 5) and carrot juice (pH 5.1) in a pressure temperature range of 0.1-500 MPa and 20-70 °C. Flow cytometry was used to investigate the pressure impact on cell structures of the strain DSM1116. Both pathogenic strains had a much higher resistance in buffer and carrot juice than the non-pathogenic surrogate. Further, strains cultivated and treated at a lower pH-value showed higher pressure stability, presumably due to variations in the membrane composition. This was confirmed for the strain DSM1116 by flow cytometry. Cells cultivated and treated at pH 5 had a stronger ability to retain their membrane potential but showed higher rates of membrane permeabilization at pressures <200 MPa compared to cells cultivated and treated at pH 7. These cells had the lowest membrane permeabilization rate at around 125 MPa, possibly denoting that variations in the fatty acid composition and membrane fluidity contribute to this stabilization phenomenon.

In situ investigation of Geobacillus stearothermophilus spore germination and inactivation mechanisms under moderate high pressure.

Bacterial spores are a major concern for food safety due to their high resistance to conventional preservation hurdles. Innovative hurdles can trigger bacterial spore germination or inactivate them. In this work, Geobacillus stearothermophilus spore high pressure (HP) germination and inactivation mechanisms were investigated by in situ infrared spectroscopy (FT-IR) and fluorometry. G. stearothermophilus spores' inner membrane (IM) was stained with Laurdan fluorescent dye. Time-dependent FT-IR and fluorescence spectra were recorded in situ under pressure at different temperatures. The Laurdan spectrum is affected by the lipid packing and level of hydration, and provided information on the IM state through the Laurdan generalized polarization. Changes in the -CH2 and -CH3 asymmetric stretching bands, characteristic of lipids, and in the amide I' band region, characteristic of proteins' secondary structure elements, enabled evaluation of the impact of HP on endospores lipid and protein structures. These studies were complemented by ex situ analyses (plate counts and microscopy). The methods applied showed high potential to identify germination mechanisms, particularly associated to the IM. Germination up to 3 log10 was achieved at 200 MPa and 55 °C. A molecular-level understanding of these mechanisms is important for the development and validation of multi-hurdle approaches to achieve commercial sterility.

High-pressure thermal sterilization: food safety and food quality of baby food puree.

The benefits that high-pressure thermal sterilization offers as an emerging technology could be used to produce a better overall food quality. Due to shorter dwell times and lower thermal load applied to the product in comparison to the thermal retorting, lower numbers and quantities of unwanted food processing contaminants (FPCs), for example, furan, acrylamide, HMF, and MCPD-esters could be formed. Two spore strains were used to test the technique; Geobacillus stearothermophilus and Bacillus amyloliquefaciens, over the temperature range 90 to 121 °C at 600 MPa. The treatments were carried out in baby food puree and ACES-buffer. The treatments at 90 and 105 °C showed that G. stearothermophilus is more pressure-sensitive than B. amyloliquefaciens. The formation of FPCs was monitored during the sterilization process and compared to the amounts found in retorted samples of the same food. The amounts of furan could be reduced between 81% to 96% in comparison to retorting for the tested temperature pressure combination even at sterilization conditions of F0-value in 7 min.

Mechanisms of endospore inactivation under high pressure.

It is well known that spore germination and inactivation can be achieved within a broad temperature and pressure range. The existing literature, however, reports contradictory results concerning the effectiveness of different pressure-temperature combinations and the underlying inactivation mechanism(s). Much of the published kinetic data are prone to error as a result of unstable process conditions or an incomplete investigation of the entire inactivation pathway. Here, we review this field of research, and also discuss an inactivation mechanism of at least two steps and propose an inactivation model based on current data. Further, spore resistance properties and matrix interactions are linked to spore inactivation effectiveness.

The release of dipicolinic acid--the rate-limiting step of Bacillus endospore inactivation during the high pressure thermal sterilization process.

High pressure combined with elevated temperatures can produce low acid, commercially sterile and shelf-stable foods. Depending on the temperature and pressure levels applied, bacterial endospores pass through different pathways, which can lead to a pressure-induced germination or inactivation. Regardless of the pathway, Bacillus endospores first release pyridine-2,6-dicarboxylic acid (DPA), which contributes to the low amount of free water in the spore core and is consequently responsible for the spore's high resistance against wet and dry heat. This is therefore the rate-limiting step in the high pressure sterilization process. To evaluate the impact of a broad pressure, temperature and time domain on the DPA release, Bacillus subtilis spores were pressure treated between 0.1 and 900 MPa at between 30 and 80 °C under isothermal isobaric conditions during dwell time. DPA quantification was assessed using HPLC, and samples were taken both immediately and 2 h after the pressure treatment. To obtain a release kinetic for some pressure-temperature conditions, samples were collected between 1s and 60 min after decompression. A multiresponse kinetic model was then used to derive a model covering all kinetic data. The isorate lines modeled for the DPA release in the chosen pressure-temperature landscape enabled the determination of three distinct zones. (I) For pressures <600 MPa and temperatures >50 °C, a 90% DPA release was achievable in less than 5 min and no difference in the amount of DPA was found immediately 2 h after pressurization. This may indicate irreversible damage to the inner spore membrane or membrane proteins. (II) Above 600 MPa the synergism between pressure and temperature diminished, and the treatment temperature alone dominated DPA release. (III) Pressures <600 MPa and temperatures <50 °C resulted in a retarded release of DPA, with strong increased differences in the amount of DPA released after 2 h, which implies a pressure-induced physiological like germination with cortex degradation, which continues after pressure release. Furthermore, at 600 MPa and 40 °C, a linear relationship was found for the DPA release rate constants ln(k(DPA)) between 1 and 30 min.

The impact of high pressure and temperature on bacterial spores: inactivation mechanisms of Bacillus subtilis above 500 MPa.

UNLABELLED: High-pressure thermal sterilization (HPTS) is an emerging technology to produce shelf stable low acid foods. Pressures below 300 MPa can induce spore germination by triggering germination receptors. Pressures above 500 MPa could directly induce a Ca+2-dipicolinic acid (DPA) release, which triggers the cortex-lytic enzymes (CLEs). It has been argued that the activated CLEs could be inactivated under HPTS conditions. To test this claim, a wild-type strain and 2 strains of Bacillus subtilis spores lacking germinant receptors and one of 2 CLEs were treated simultaneously from 550 to 700 MPa and 37 to 80 C (slow compression) and at 60 to 80 C up to 1 GPa (fast compression). Besides, an additional heat treatment to determine the amount of germinated cells, we added TbCl3 to detect the amount of DPA released from the spore core via fluorescent measurement. After pressure treatment for 120 min at 550 MPa and 37 °C, no inactivation was observed for the wild-type strain. The amount of released DPA correlated to the amount of germinated spores, but always higher compared to the belonging cell count after pressure treatment. The release of DPA and the increase of heat-sensitive spores confirm that the inactivation mechanism during HPTS passes through the physiological states: (1) dormancy, (2) activation, and (3) inactivation. As the intensity of treatment increased, inactivation of all spore strains also strongly increased (up to -5.7 log10), and we found only a slight increase in the inactivation of one of the CLE (sleB). Furthermore, above a certain threshold pressure, temperature became the dominant influence on germination rate. PRACTICAL APPLICATION: The continuous increase of high-pressure (HP) research over the last several decades has already generated an impressive number of commercially available HP pasteurized products. Furthermore, research helped to provoke the certification of a pressure-assisted thermal sterilization process by the U.S. FDA in February 2009. However, this promising sterilization technology has not yet been applied in industrial settings. An improved understanding of spore inactivation mechanisms and the ability to calculate desired inactivation levels will help to make this technology available for pilot studies and commercialization at an industrial scale. Moreover, if the synergy between pressure and elevated temperature on the inactivation rate could be identified, clarification of the underlying inactivation mechanism during HP thermal sterilization could help to further optimize the process of this emerging technology.