Microbial inactivation by high pressure homogenization: Effect of the disruption valve geometry

Microbial inactivation by high pressure homogenization (HPH) was studied in two very different disruption chambers, one based on a piston micrometric valve and the other on an orifice valve, in order to relate the geometrical and fluid dynamics aspects with the rate of death of Escherichia coli, Lactobacillus delbrueckii and Saccharomyces cerevisiae.     

Ascospore inactivation and germination by high pressure processing is affected by ascospore age

The effects of age on high pressure resistance of the ascospores of heat resistant moulds Byssochlamys fulva, B. nivea, Neosartorya fischeri and N. spinosa were determined. Ascospores were harvested from cultures grown for 3–15 weeks at 30 °C on malt extract agar. Following filtration and determination of concentration, the ascospores were subjected to high pressure processing (HPP) at 600 MPa for 10 min in 0.1 M citrate phosphate buffer (pH 4 and 6) and mango puree (pH 5). The results supported our hypothesis that age (maturity) affects high pressure resistance of ascospores of heat resistant moulds. A reduction of log₁₀ 2.5 cfu mL^− 1 was achieved for three week old ascospores ofB. nivea whereas for nine week old ascospores only a half log reduction was achieved. Similar results were observed for B. nivea and N. fischeri. The HPP treatment caused activation of ascospores of N. spinosa, with older ascospores showing increased activation.

Industrial relevance

The observation of activation of some ascospores by HPP, indicates that HPP alone is insufficient for elimination of these problematic spoilage microorganisms. HPP would need to be combined with other hurdles in order to produce high quality pressure-treated shelf-stable fruit products.     

超高压诱导食品中微生物失活的研究进展

超高压处理(high pressure processing,HPP)作为一种新型的非热杀菌技术,具有最大限度保留食品中营养物质的优势,已在食品工业中广泛应用。但由于加工环境以及食品基质的复杂性,不同微生物在HPP下敏感度不同,且会形成部分亚致死微生物,引发食品安全的潜在隐患。基于HPP设备原理及其应用于食品加工中的杀菌效果,该文对影响超高压杀菌效果的内外因子以及导致细菌失活(致死或亚致死效应)的作用机制进行了综述,并分析了超高压与其他技术联用可能是抑制微生物亚致死效应的有效途径。     

Aqueous matrix compositions and pH influence feline calicivirus inactivation by high pressure processing

The individual effects of pH (pH 3 to 8), NaCl (0 to 21%), sucrose (0 to 70%), and whey protein (0 to 2%) on pressure resistance of feline calicivirus (FCV) in Dulbecco's modified Eagle medium with 10% fetal bovine serum were determined. At pH 3 through 8, the virus was more resistant to pressure at a pH of < or = 5.2. For FCV samples with sucrose (up to 40%) or NaCl (up to 12%), the amount of FCV inactivated by pressure was inversely proportional to the sucrose or NaCl concentration. For example, a treatment of 250 MPa at 20 degrees C for 5 min reduced the FCV titer by 5.1 log PFU/ml without added sucrose and by 0.9 log PFU/ml with 40% sucrose. Reduced pressure sensitivity with increasing NaCl and sucrose concentrations was not a simple function of water activity. Different PFU reductions were observed for NaCl and sucrose samples with equivalent water activity. When protein at concentrations up to 2% did not provide a protective effect. The combined effect of NaCl and sucrose at 4 and 20 degrees C on pressure resistance of FCV also was examined. When both NaCl and sucrose were added to the FCV stock, they had an additive effect on increasing the pressure resistance of FCV. The individual (6% NaCl or 20% sucrose) and combined (6% NaCl plus 20% sucrose) resistance effects did not abrogate enhanced inactivation for pressure treatments at 4 degrees C compared with those at 20 degrees C. Aqueous matrix compositions, in particular different concentrations of NaCl and sucrose or different pH values, can substantially alter the efficiency of virus inactivation by high pressure processing.     

Model for Listeria monocytogenes inactivation on dry-cured ham by high hydrostatic pressure processing

The aim of the work was to develop and validate a model of the inactivation of Listeria monocytogenes on dry-cured ham by high hydrostatic pressure (HHP) processing, as a function of the technological parameters: intensity, length and fluid temperature. Dry-cured ham inoculated with L. monocytogenes was treated at different HHP conditions (at 347-852 MPa; for 2.3 to 15.75 min; at 7.6 to 24.4 °C) following a central composite design. Bacterial inactivation was assessed in terms of logarithmic reductions of L. monocytogenes counts on selective media. According to the best fitting and most significant polynomial equation, pressure and time were the most important factors determining the inactivation extent. The significance of the quadratic term of pressure and time indicated that little effect was observed below 450 MPa, whereas holding time longer than 10 min did not result in a meaningful reduction of L. monocytogenes counts. Temperature did not show significant influence at the range assayed. The model was validated with results obtained from further experiments and bibliographical data within the range of the experimental domain. The accuracy factor and bias factor were within the proposed acceptable values indicating the suitability of the model for predictive purposes, such as prediction of the process criteria to meet the Food Safety Objectives. The results of this work may help food processors to select optimum processing conditions of HHP.     

Environmental factors influencing the inactivation of Cronobacter sakazakii by high hydrostatic pressure

The effect of High Hydrostatic Pressure (HHP) on the survival of Cronobacter sakazakii was investigated. Deviations from linearity were found on the survival curves and the Mafart equation accurately described the kinetics of inactivation. Comparisons between strains and treatments were made based on the time needed for a 5-log₁₀ reduction in viable count. The ability of C. sakazakii to tolerate high pressure was strain-dependent with a 26-fold difference in resistance among four strains tested. Pressure resistance was greatest in the stationary growth phase and at the highest growth temperatures tested (30 and 37 °C). Cells treated in neutral pH buffer were 5-fold more resistant than those treated at pH 4.0, and 8-fold more sensitive than those treated in buffer with sucrose added (a_w = 0.98). Pressure resistance data obtained in buffer at the appropriate pH adequately estimated the resistance of C. sakazakii in chicken and vegetables soups. In contrast, a significant protective effect against high pressure was conferred by rehydrated powdered milk. As expected, treatment efficacy improved as pressure increased. z values of 112, 136 and 156 MPa were obtained for pH 4.0, pH 7.0 and a_w = 0.98 buffers, respectively. Cells with sublethal injury to their outer and cytoplasmic membranes were detected after HHP under all the conditions tested. The lower resistance of C. sakazakii cells when treated in media of pH 4.0 seemed to be due to a decreased barostability of the bacterial envelopes. Conversely, the higher resistance displayed in media of reduced water activity may relate to a higher stability of bacterial envelopes.     

Fermented minced pepper by high pressure processing,high pressure processing with mild temperature and thermal pasteurization

High pressure processing (HPP) and thermal pasteurization (TP) of fermented minced pepper (FMP) were comparatively evaluated by examining their impacts on microbial load, titratable acid (TA), pH, a_w, firmness, color, capsanthin, ascorbic acid (AA), and biogenic amines (BAs) after processing and during 12 weeks of storage at 25 and 37 °C. The total plate count (TPC) in FMP samples was reduced by 1.48, 0.12 and 1.58 log₁₀ CFU/g after TP (83 °C/15 min), HPP1 (500 MPa/20 °C/5 min) and HPP2 (500 MPa/50 °C/5 min), respectively. The population of spores was reduced by 1.21 log₁₀ CFU/g only after HPP2. During storage at 25 or 37 °C, the TPC in TP, HPP1, and HPP2 samples increased by 0.88/1.21, 0.41/0.62 and 0.60/0.86 log₁₀ CFU/g, respectively, while the spores decreased below the detection limit. The retention of firmness after TP, HPP1 and HPP2 was 36.91, 91.15 and 66.48% respectively, and HPP-treated samples exhibited more retention during the storage. Color of FMP samples was not changed by TP, but slightly changed by HPP1 and HPP2. The content of capsanthin retained 78.99, 93.71 and 88.19% after TP, HPP1 and HPP2, it showed a small decrease during storage. Levels of biogenic amines (BAs) in HPP2 samples were lower than that of TP and HPP1 ones. There were better sensory quality and lower microbial level in HPP-treated samples during storage, indicating that HPP is a better choice for the preservation of FMP.Industrial relevanceConsumption of fermented minced pepper (FMP), as a traditional Chinese food, is becoming increasingly popular. Considering that heat treatment may destroy some heat-sensitive quality of the products, this study evaluated the effects of high pressure processing (HPP) on quality of FMP. Findings of this study could help processors commercialize HPP to replace current thermal processing in industrial production.     

Compression heating influence of pressure transmitting fluids on bacteria inactivation during high pressure processing

Compression heating characteristics of different pressure transmitting fluids [three different concentrations (75:25, 50:50, 25:75) of water–glycol mix and sodium benzoate (2%) solutions] and their influence on inactivation of spores of Bacillus subtilis in phosphate buffer (0.067 M, pH 7.0) during high pressure processing (HPP) were studied. Experiments were conducted using a pilot scale food processor. Pressure transmitting fluids containing highest percentage of glycol (25:75 water–glycol mix) showed highest temperature increase while 2% sodium benzoate solution showed least temperature increase during high pressure processing. The target pressure, holding time, compressibility, initial temperature, and the rate of heat loss to the surroundings primarily influenced the apparent temperature increase of pressure transmitting fluid in a vessel during HPP. The temperature change was further influenced by the fluid properties such as viscosity, specific heat and thermal conductivity. Use of sodium benzoate solution as pressure-transmitting fluid resulted in highest inactivation of B. subtilis spores. Change in pressure transmitting fluid temperature as a result of compression heating and subsequent heat transfer should be considered in inactivation of bacterial spores by HPP.     

Bacterial inactivation by high-pressure homogenisation and high hydrostatic pressure

The resistance of five gram-positive bacteria, Enterococcus faecalis, Staphylococcus aureus, Lactobacillus plantarum, Listeria innocua and Leuconostoc dextranicum, and six gram-negative bacteria, Salmonella enterica serovar typhimurium, Shigella flexneri, Yersinia enterocolitica, Pseudomonas fluorescens and two strains of Escherichia coli, to high-pressure homogenisation (100-300 MPa) and to high hydrostatic pressure (200-400 MPa) was compared in this study. Within the group of gram-positive bacteria and within the group of gram-negative bacteria, large differences were observed in resistance to high hydrostatic pressure, but not to high-pressure homogenisation. All gram-positive bacteria were more resistant than any of the gram-negative bacteria to high-pressure homogenisation, while in relative to high hydrostatic pressure resistance both groups overlapped. Within the group of gram-negative bacteria, there also existed another order in resistance to high-pressure homogenisation than to high hydrostatic pressure. Further it appears that the mutant E. coli LMM1010, which is resistant to high hydrostatic pressure is not more resistant to high-pressure homogenisation than its parental strain MG1655. The preceding observations indicate a different response of the test bacteria to high-pressure homogenisation compared to high hydrostatic pressure treatment, which suggests that the underlying inactivation mechanisms for both techniques are different. Further, no sublethal injury could be observed upon high-pressure homogenisation of Y. enterocolitica and S. aureus cell population by using low pH (5.5 7), NaCl (0 6%) or SDS (0-100 mg/l) as selective components in the plating medium. Finally, it was observed that successive rounds of high-pressure homogenisation have an additive effect on viability reduction of Y. enterocolitica and S. aureus.     

A novel approach to predicting microbial inactivation kinetics during high pressure processing

The inactivation kinetics of Escherichia coli (ATCC 25922) during high pressure processing (HPP) was examined from 200 to 400 MPa in 50 MPa increments at 15 degrees C. Although the time course of HPP-induced E. coli inactivation in 0.1% peptone water successfully fitted the Weibull function, this procedure involved curve fitting, and not prediction. The objective of this study was to develop a novel HPP-induced microbial inactivation model to simulate the inactivation kinetics under various pressure conditions. The maximum inactivation rate during HPP was calculated from the inactivation curves at different pressure conditions on a semi-log plot. The relationship between the square root of the absolute value of the inactivation rate (k(max)) and treatment pressure was linear (R(2)=0.99). The linear relationship between k(max) and treatment pressure also successfully described independent data from other studies in the literature. Overall, the newly developed differential equation model, into which was substituted the square root function of the inactivation rate, was capable of simulating the inactivation kinetics during HPP at constant pressure. Additionally, the model could successfully describe the inactivation kinetics during HPP using other researchers' data. The accuracy of prediction of the new model was comparable to that derived from Weibull or modified Gompertz fitting to the observed data. Furthermore, the new model could successfully simulate the inactivation kinetics during dynamic pressure conditions, which included come-up time, changes in holding pressure during treatment, and pressure-release time. Moreover, the effect of pulsed pressure treatment was also simulated successfully using this model. Therefore, the modeling procedure presented in this study will contribute to the advancement of predictive modeling for HPP-induced microbial inactivation.     

Food processing by high hydrostatic pressure

The use of high hydrostatic pressures (HHP) for food processing is finding increased application within the food industry. One of the advantages of this technology is that because it does not use heat, sensory, and nutritional attributes of the product remain virtually unaffected, thus yielding products with better quality than those processed traditional methods. HHP have the ability to inactivate microorganisms as well as enzymes responsible for shortening the life of a product. In addition to lengthening the shelf-life of food products, HHP can modify functional properties of components such as proteins, which in turn can lead to the development of new products. Equipment for large-scale production of HHP processed products are commercially available nowadays. Guacamole, sliced ham, oysters, and fruit juices are some of the products currently available on the market. HHP technology is one of the most promising nonthermal processes.     

超高压杀菌效果及机制研究进展

超高压作为一种新型的食品非热加工技术, 处理过程温度低、对食品营养成分破坏小,能在有效杀菌的同时显著提升加工食品品质,是未来食品加工技术发展的热点方向。近年来,超高压技术被广泛应用于食品加工,并在国内外实现了商业化应用。杀菌作为超高压加工食品过程中最重要的环节,是保证食品安全、延长产品货架期的关键点,因此一直是本领域研究的重点。本文介绍了超高压技术的设备和作用原理, 总结了超高压或超高压联合其他手段对微生物营养体、细菌芽孢、真菌孢子及病毒的杀灭效果和杀菌机制, 归纳了超高压杀菌中存在的杀菌机制不清、缺乏杀菌指示菌以及深休眠芽孢等问题, 以期为进一步完善超高压杀菌理论、推动超高压技术在食品加工中的产业化应用指明方向。     

Lytic and nonlytic mechanism of inactivation of gram-positive bacteria by lysozyme under atmospheric and high hydrostatic pressure

A different behavior was observed in three gram-positive bacteria exposed to hen egg white lysozyme by plate counts and phase-contrast microscopy. The inactivation of Lactobacillus johnsonii was accompanied by spheroplast formation, which is an indication of peptidoglycan hydrolysis. Staphylococcus aureus was resistant to lysozyme and showed no signs of peptidoglycan hydrolysis, and Listeria innocua was inactivated and showed indications of cell leakage but not of peptidoglycan hydrolysis. Under high hydrostatic pressure, S. aureus also became sensitive to lysozyme but did not form spheroplasts and was not lysed. These results suggested the existence of a nonlytic mechanism of bactericidal action of lysozyme on the latter two bacteria, and this mechanism was further studied in L. innocua. Elimination of the enzymic activity of lysozyme by heat denaturation or reduction with beta-mercaptoethanol eliminated this bactericidal mechanism. By means of a LIVE/DEAD viability stain based on a membrane-impermeant fluorescent dye, the nonlytic mechanism was shown to involve membrane perturbation. In the absence of lysozyme, high-pressure treatment was shown to induce autolytic activity in S. aureus and L. innocua.     

Activation and inactivation of Talaromyces macrosporus ascospores by high hydrostatic pressure

The effect of high hydrostatic pressure treatment (with pressures of up to 700 MPa) on Talaromyces macrosporus ascospores was investigated. At 20 degrees C, pressures of > or = 200 MPa induced the activation and germination of dormant ascospores, as indicated by increased colony counts for ascospore suspensions after pressure treatment and the appearance of germination vesicles and tubes. Pressures of > 400 MPa additionally sensitized the ascospores to subsequent heat treatment. At pressures of > 500 MPa, activation occurred in a few minutes but was followed by inactivation with longer exposure. However, even with the most extreme pressure treatment, a fraction of the ascospore population appeared to resist both activation and inactivation, and the maximal achievable reduction of ascospores was on the order of 3.0 log10 units. Pressure-induced ascospore activation at 400 MPa was temperature dependent, with minimum activation at 30 to 50 degrees C and > or = 10-fold higher activation levels at 10 to 20 degrees C and at 60 degrees C, but it was not particularly pH dependent over a pH range of 3.0 to 6.0. Pressure inactivation at 600 MPa, in contrast, was pH dependent, with the inactivation level being 10-fold higher at pH 6.0 than at pH 3.0. Observation of pressure-treated and subsequently dried spores with the use of light and scanning electron microscopy revealed a collapse of the spore structure, indicating a loss of the spore wall barrier properties. Finally, pressure treatment sensitized T. macrosporus ascospores to cell wall lytic enzymes.     

Effect of water activity on inactivation of Listeria monocytogenes and lactate dehydrogenase during high pressure processing

The aim of this study was to investigate the effect of water activity (a_w) on the inactivation of Listeria monocytogenes and lactate dehydrogenase (LDH) during high pressure processing (HPP). For microbial inactivation lyophilized cells of L. monocytogenes 19,115 were left dry or were suspended in 10 ml of 0.1% peptone water, 10 ml of glycerol, or mixtures of glycerol and peptone water. All samples of various a_ws were high pressure (HP) processed at ambient temperature at 600 MPa for 300 s. Following HPP, samples were serially diluted in 0.1% peptone and spread-plated on Tryptic Soy agar supplemented with Yeast Extract. For enzyme inactivation, 4.2 mg of lyophilized LDH was suspended in 2 ml of 100 mM phosphate buffer (pH 7.4), 2 ml of peptone water or glycerol, or in 2 ml mixtures of glycerol and peptone water. A lyophilized sample with no added liquid was also included. All enzyme samples were subjected to HPP as described above. After HPP, LDH was diluted to 0.28 μg/ml in 100 mM phosphate buffer (pH 7.4). LDH activity was assessed by measuring the change in concentration of β-NADH as a function of time. Dynamic light scattering analysis (DLS) was performed to examine the size distribution, polydispersity, and hydrodynamic radius of LDH before and after HPP. No significant difference in CFU/g was observed between lyophilized cells not subjected to HPP and lyophilized cells subjected to 600 MPa for 300 s (P < 0.05). However, lyophilized cells that were suspended in 100% to 60% peptone water showed a ~ 7.5-log₁₀ reduction when subjected to HPP. Survival of L. monocytogenes following HPP significantly increased (P < 0.05) when the peptone water concentration was decreased below 60% (a_w ~ 0.8). DLS results revealed that LDH suspended in buffer underwent aggregation following HPP (600 MPa, 300 s). Inactivation rate constants obtained using a first-order kinetic model indicated that untreated and HP processed lyophilized LDH had similar activities. When LDH was subject to HPP in solutions containing glycerol, enzyme activity decreased as the water content increased (r² = 0.95). Lyophilization completely protected L. monocytogenes and LDH from inactivation by high pressure. Furthermore, enzyme activity and cell survival increased as water activity was decreased. We postulate low a_w results in protein stabilization, which prevents protein denaturation and cell death during HPP.     

Bacillus spore inactivation differences after combined mild temperature and high pressure processing using two pressurizing fluids

Spores of six species (28 strains) of dairy Bacillus isolates were added to sterile reconstituted skim milk and pressure processed (600 MPa for 60 s at 75 degrees C) using either a water-based pressurizing fluid or silicon oil. Processing temperatures peaked at 88 and 90 degrees C, respectively, for both fluids. For all strains, the log inactivation was consistently higher in the silicon oil than in the water-based fluid. This has potential implications for food safety assessment of combined pressure-temperature processes. High pressure processing causes mild heating during pressurization of both the target sample (i.e., spores) and the pressurizing fluid used for pressure delivery. Primarily, the adiabatic heat of compression of the fluids as well as other heat-transfer properties of the fluids and equipment determines the magnitude of this heating. Pressure cycles run with silicon oil were 7 to 15 degrees C higher in temperature during pressurization than pressure cycles run with the water-based pressurizing fluid, due to the greater adiabatic heat of compression of silicon oil. At and around the target pressure, however, the temperatures of both pressurizing fluids were similar, and they both dropped at the same rate during the holding time at the target pressure. We propose that the increased spore inactivation in the silicon oil system can be attributed to additional heating of the spore preparation when pressurized in oil. This could be explained by the temperature difference between the silicon oil and the aqueous spore preparation established during the pressurization phase of the pressure cycle. These spore-inactivation differences have practical implications because it is common practice to develop inactivation kinetic data on small, jacketed laboratory systems pressurized in oil, with extensive heat loss. However, commercial deployment is invariably on large industrial systems pressurized in water, with limited heat loss. Such effects should be considered in food safety assessments of combined pressure-temperature processes.     

Critical factors determining inactivation kinetics by pulsed electric field food processing

The potential to commercialize the non-thermal pulsed electric field (PEF) technology as a new method to preserve food products has caught the attention of the food industry that wishes to fulfil consumers demands for fresh products. In recent years, numerous research groups have demonstrated the possibility to inactivate a range of microorganisms both in buffer systems and in food products using different PEF systems. In this review, we survey the critical process factors and main characteristics of food products that determine the microbial inactivation kinetics.     

Influence of kinetic parameters of high pressure processing on bacterial inactivation in a buffer system

High pressure processing is recently applied in the food industry to inactivate spoilage and pathogenic microorganisms. Bacterial cells exhibit various barosensibility, and the role of pressurization, depressurization and constant pressure stage remain unknown. We investigated the effect of high pressure processing on Salmonella typhimurium and Listeria monocytogenes cells at 400 and 500 MPa respectively in buffer pH 7 at 20 degrees C. We applied various pressurization/depressurization kinetic rates (1, 5 and 10 MPa/s for pressurization and 250, 20 and 5 MPa/s for depressurization), and various pulse series or pressure holding times. Results show that high pressure pulses reduced linearly the number of bacterial cells according to the product of pressure and time: we defined this product as a Barometric Power (BP). Reduction of both microorganisms increased when holding time increased from 5 to 20 min, and better results were obtained when the rate of pressurization and depressurization were increased.     

Inactivation of Staphylococcus aureus by high pressure processing: An overview

Food safety is a major concern of consumers, food industry and governments, with 25 million foodborne diseases occurring annually worldwide. Staphylococcus aureus, is an extremely versatile opportunistic pathogen being responsible for staphylococcal food poisoning due to enterotoxic strains. With increasing demands for safer food, new food preservation technologies are increasingly gaining interest. In the last two decades, high pressure processing (HPP) appeared as an alternative non-thermal food preservation method promoting inactivation of some spoilage and pathogenic microorganisms, while maintaining food characteristics. Factors that modulate its efficiency will be revised, firstly based on the state-of-the art described for bacteria in general and afterwards, when studies exist, for S. aureus specifically. S. aureus inactivation by HPP, like in other microorganisms, is conditioned by cell structures and biomolecules, matrix, HPP processing conditions, the use of antimicrobials and is also dependent of the strain and growth phase. Cell membrane is the most pressure sensitive structure of S. aureus, being the lipids and proteins the most important target molecules. However, monomeric proteins such as staphylococcal enterotoxins (SE) are not affected by HPP, and strains with SE appear to be more efficiently inactivated than those without. Other phenotypic and genotypic characteristics of S. aureus strains, such as pigmentation and the presence of σ^B factor are extremely important factors determining the efficacy of HPP treatments. Inactivation of S. aureus by HPP to ensure food safety still remains a current challenge regarding the understanding of its particular barotolerance and its inactivation kinetics profile that often deviates from the simpler first order decay. Thus, this review provides state-of-the-art information for researchers interested in studying HPP inactivation of S. aureus.Industrial relevanceThis review gives an insight on the importance of Staphylococcus aureus as a foodborne versatile opportunistic pathogen and its importance from the food safety point, its barotolerance and the main reasons for this resistant behavior to high pressure processing and the mechanisms of S. aureus inactivation by HPP.     

天然淀粉的超高压糊化压力研究

在悬浮液浓度为5%(w/v),温度为(20±2)℃时,对8种不同淀粉进行高压处理5 min使淀粉发生糊化,采用X射线衍射测试技术得到了各种淀粉完全糊化的压力:小麦淀粉和木薯淀粉约为500 MPa,玉米淀粉、荸荠淀粉、糯小麦淀粉和糯米淀粉均为550 MPa,糯玉米淀粉约为650 MPa,马铃薯淀粉为750 MPa.