EFFICACY OF HIGH HYDROSTATIC PRESSURE AND MILD HEAT TO REDUCE GEOBACILLUS STEAROTHERMOPHILUS AS 1.1923 SPORES IN MODEL FOOD SYSTEMS

The objective of this investigation was to evaluate the efficacy of high hydrostatic pressure and mild heat against spores of Geobacillus stearothermophilus AS 1.1923 in model food systems. The pressure-processing conditions were fixed at 625.0 MPa and 86C for 14 min, which have been determined as the optimum processing conditions considering six-log-cycle reductions of G. stearothermophilus spores. Based on the results, response surface methodology was performed in the present investigation, the effects from food ingredients, such as soybean protein, soybean oil and sucrose, as well as pH of the food matrix on the inactivation of G. stearothermophilus spores by high pressure and mild heat was explored, and a quadratic predictive model for the effects of food ingredients and pH on the reduction levels of G. stearothermophilus spores by high-pressure processing was built. The predictive model is significant because the level of significance was P  < 0.0001 and the calculated F value is much greater than the tabulated F value. Moreover, the adequacy of the model equation for predicting the reduction of G. stearothermophilus spores was verified effectively.

PRACTICAL APPLICATIONS

This paper clearly demonstrated the contributions to pressure resistance from food ingredients such as soybean protein, soybean oil, sucrose and pH of the food matrix to the inactivation of Geobacillus stearothermophilus spores by high hydrostatic pressure and mild heat. The predictive model for predicting the reduction of G. stearothermophilus spores in model food systems was built which can be beneficial to targeted process development toward high-pressure sterilization of foods.     

超高(静)压处理对果蔬理化性质的影响

超高（静）压（以下简称超高压）技术用于乳制品、果蔬汁等的报道已有很多，而超高压用于固态食品是近几年研究的新课题。综述了超高压对果蔬质构的影响。超高压可以引起果蔬变软、褐变。但是超高压比热烫对果蔬的质构影响小。     

HIGH HYDROSTATIC PRESSURE EFFECT ON SACCHAROMYCES CEREVISIAE,ESCHERICHIA COLI AND LISTERIA INNOCUA IN PEAR NECTAR

ABSTRACT

An isostatic pressure system was used for processing fresh‐prepared pear nectar, inoculated with Saccharomyces cerevisiae, Escherichia coli and Listeria innocua, at selected pressures (0 to 241 MPa), and times (2 s, and 0 to 15 min). The come up time (time to reach the working pressure) had an important microbial inactivation effect in view of the fact that the initial counts of 6.0 × 10⁵, 1.02 × 10⁷ and 2.4 × 10⁷ cfu/mL were reduced to 2.4 × 10⁵, 6.3 × 10⁵ and 2.2 × 10⁷ cfu/mL, for S. cerevisiae, E. coli and L. innocua, respectively, in nectar. Decimal reduction time values, in the range of 2.0–35.3, 0.6–20.6 and 9.2–588.2 min, were calculated from the first order kinetics modeling of S. cerevisiae, E. coli, and L. innocua survivors, respectively. z_p values of 120.5, 92.6 and 75.2 MPa were obtained for the three types of microorganisms, respectively.

PRACTICAL APPLICATIONS

The application of high hydrostatic pressure may deliver a “cold pasteurized” pear nectar. This “cold pasteurization” may inactivate microorganisms and enzymes as thermal pasteurization does; however, high hydrostatic pressure does not generate unwanted chemical compounds that may change the nectar fresh sensory characteristics. Thus, a fresh‐like fruit product, free from chemicals and with high quality and sensory characteristics, could be offered to consumers.

     

INACTIVATION OF BACTERIAL SPORES BY COMBINED ACTION OF HYDROSTATIC PRESSURE AND BACTERIOCINS IN ROAST BEEF

Foodborne bacterial spores are normally resistant to high hydrostatic pressure; however, at moderate pressure, they can be induced to germinate and outgrow. At this stage, they can be killed by bacteriocin-based biopreservatives (BP-containing pediocin and nisin at 3:7 ratio; BP_X, BP + 100 μg/mL lysozyme; BP_Y, BP_X+ 500 μg/mL Na-EDTA). Based on this principle, spores of the meat spoilage organism, Clostridium laramie (1–2 × 10² spores/bag) alone or a mixture of four clostridial spores (5 × 10³ spores/bag), Clostridium sporogenes, Clostridium perfringens, Clostridium tertium, and Clostridium laramie, were inoculated in roast beef in the presence of 5000 AU/g of bacteriocin-based biopreservatives. The roast beef samples were subjected to hydrostatic pressure (HP) at 345 MPa for 5 min at 60C and stored at 4 or 12C for 84 days or at 25C for 7 days. The HP treatment of roast beef samples inoculated with a mixture of clostridial spores could be stored for 42 days at 4C. The HP in combination with either BP_X or BP_Y extended the shelf-life of roast beef up to 7 days at 25C. The combined treatment of HP and BP controlled the growth of C. laramie spores and extended the shelf-life of roast beef for 84 days when stored at 4C.     

EFFECT OF CONCURRENT HIGH HYDROSTATIC PRESSURE, ACIDITY AND HEAT ON THE INJURY AND DESTRUCTION OF LISTERIA MONOCYTOGENES

The effect of concurrent use of high hydrostatic pressure, heat and acidity on Listeria monocytogenes Scott A and CA was investigated. In general, lethality was enhanced when cells were pressurized at higher temperatures or lower pH. Strain CA demonstrated an additional 3-log₁₀ reduction when pressurized at pH 4.0 as compared with pH 6.0 at 353 MPa, 45C for 10 min. Scott A was reduced an additional 1 log₁₀ by increasing the temperature from 25C to 45C with pressurization at 252 MPa, pH 6.0 for 30 min. Exposure to 404 MPa at 45C for 30 min demonstrated complete injury or death of CA cells with an initial concentration of >10⁸ CFU/mL. At least an 8-log₁₀ reduction was observed for both L. monocytogenes strains Scott A and CA when exposed to the combined treatments of 252 MPa, 45C, pH 4.0 for 30 min .     

EFFECT OF HIGH PRESSURE ON LACTOCOCCAL BACTERIOPHAGES

Four different host-specific lactococcal bacteriophages were subjected to high hydrostatic pressure and heat treatments. Pressure treatments were done at room temperature at 300 and 350 MPa for 5–40 min. Complete inactivation of bacteriophages was observed starting at 350 MPa for 20-min treatment at room temperature. The effect of heat on the bacteriophages was analyzed by heat treatment at 71.7C for predetermined lengths of time (1–5 min). Decrease in bacteriophage number was observed after 3 min of heat treatment at 71.7C. Pressure treatment at 350 MPa/5 min and heat treatment at 71.7C/3 min were both found to be effective for the inactivation of lactococcal bacteriophages. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis indicated that protein profiles of pressure-treated (350 MPa, 25 min) bacteriophages were altered.

PRACTICAL APPLICATIONS

Bacteriophages are still a problem for the production of fermented dairy products, as there has not been a process to eliminate them completely from the fermentation environment. Processes such as pasteurization are not adequate to eliminate bacteriophages. However, new food preservation methods have been developed, one of which is high hydrostatic pressure (HHP) processing. HHP has potential application for the inactivation of viruses. Here, we demonstrate the application of HHP to inactivate the bacteriophages of dairy starter culture Lactococcus in comparison with heat treatment.     

EFFECT OF HIGH HYDROSTATIC PRESSURE PROCESSING ON RHEOLOGICAL AND TEXTURAL PROPERTIES OF PROBIOTIC LOW-FAT YOGURT FERMENTED BY DIFFERENT STARTER CULTURES

The effect of milk processing on rheological and textural properties of probiotic low‐fat yogurt (fermented by two different starter cultures) was studied. Skim milk fortified with skim milk powder was subjected to three treatments: (1) thermal treatment at 85C for 30 min; (2) high hydrostatic pressure (HHP) at 676 MPa for 5 min; and (3) combined treatments of HHP (676 MPa for 5 min) and heat (85C for 30 min). The processed milk was fermented using two different starter cultures containing Streptococcus thermophilus, Lactobacillus delbrueckii ssp. bulgaricus, Lactobacillus acidophilus and Bifidobacterium longum at inoculation rates of 0.1 and 0.2%. Rheological parameters were determined and a texture profile analysis was carried out. Yogurts presented different rheological behaviors according to the treatment used, which could be attributed to structural phenomena. The combined HHP and heat treatment of milks resulted in yogurt gels with higher consistency index values than gels obtained from thermally treated milk. The type of starter culture and inoculation rate, providing different fermentation pathways, also affected the consistency index and textural properties significantly. The combined HHP and heat treatment of milks before fermentation, and an inoculation rate of 0.1% (for both cultures), led to desirable rheological and textural properties in yogurt, which presented a creamy and thick consistency that does not require the addition of stabilizers.     

EFFECT OF HIGH HYDROSTATIC PRESSURE, GAMMA-IRRADIATION AND COMBINATION TREATMENTS ON THE MICROBIOLOGICAL QUALITY OF LAMB MEAT DURING CHILLED STORAGE

The effect of gamma irradiation (1.0 kGy) and high hydrostatic pressure (200 MPa for 30 min), either alone or in combination on the shelf-life of lamb mince meat at 0–3C was studied. Untreated control samples initially had total microbial counts of 10⁵ CFU/g, 10² CFU/g of coliforms and 10⁴ CFU/g of Staphylococcus spp. Coliforms were eliminated by all the treatments . Staphylococcus spp. however, were reduced only by 1 log cycle when treated with irradiation alone and high pressure alone. These species were a mixture of mannitol-fermenting and mannitol-nonfermenting strains. In samples subjected to the combination treatment , Staphylococcus spp. appeared only after 3 weeks of storage and all were mannitol-nonfermenting. On the basis of microbiological and sensory quality, the shelf-life of the control sample was less than 1 week. All treated meat samples had a shelf-life of 3 weeks, but only combination treated samples were free from potentially pathogenic Staphylococcus spp .     

HIGH HYDROSTATIC PRESSURE EXTRACTION OF SALIDROSIDE FROM RHODIOLA SACHALINENSIS

A nonthermal process, high hydrostatic pressure (HHP), was used as a new extraction technique to obtain salidroside from Rhodiola sachalinensis. The leaching rates of salidroside were measured under different treatment conditions. When the material was treated under 500 MPa of hydrostatic pressure with 50 times of volume 60% (v/v) of ethanol solution as the extracting solvent, the leaching rate of salidroside was up to 0.401% in 3 min, while that of ultrasonic extraction was at 0.288% in 30 min and that of reflux extraction was at 0.302% in 120 min. The HHP extraction technique showed high efficiency in extracting salidroside at room temperature using moderate ethanol concentration.

PRACTICAL APPLICATIONS

Rhodiola is prevalently used as a safe and effective antifatigue drug and as an adaptogen around the world. But the common extraction methods used to yield commercial products are still traditional ones, which could easily lead some thermosensitive ingredients to lose their biologic activities. The nonthermal extraction method is the key point of the realization of herb modernization. The high hydrostatic pressure (HHP) technique can be operated at room temperature and can shorten processing time. The extract produced by the method has proved to be of higher extraction yield than traditional varieties. In addition, the HHP technique only consumes electric energy without exhaust emission, which is an environment-friendly method. In the future, we will cooperate with those far-seeing companies to achieve its industrialization. The annual sale of herbs in the whole world has reached 30 billion. We do not doubt the prospect of this technique.     

EFFECT OF pH AND ASCORBIC ACID ON HIGH HYDROSTATIC PRESSURE-PROCESSED MANGO PUREE

Mango puree containing ascorbic acid (AA) (500 ppm) standardized at low pH (3.5) with phosphoric acid and inoculated or noninoculated with Saccharomyces cerevisiae was treated at high pressure (207, 345, 483 and 552 MPa) for selected times. High hydrostatic pressure (HHP)‐processed (552 MPa/5 min) standardized mango puree (SMP) was stored at 3C for 1 month and periodically analyzed for color, residual polyphenoloxidase (PPO) activity and microbial load. The remaining PPO activity average in SMP, after HHP processing at 207, 345, 483 and 552 MPa, at all times, was 35.8 ± 6, 21.5 ± 13.2, 46.8 ± 53.2 and 61.8 ± 5.8% PPO activity units, respectively. The D₂₀₇ values of 8.5 and 7.2 min for total count and yeasts were observed, respectively, after 207 MPa of pressure. A log reduction of 1.62 and 1.35 was observed after applying 345 MPa of pressure (2 s) for total count and yeasts, respectively. However, no microbial growth (<10 cfu/g) was observed after applying 483 or 552 MPa at any time. The addition of AA and the standardization at pH 3.5 reduced the rate of browning during storage.     

THERMAL RESISTANCE OF BACILLUS STEAROTHERMOPHILUS SPORES IN DRIED PASTA AT DIFFERENT STAGES OF REHYDRATION

The thermal resistance of Bacillus stearothermophilus spores at different stages of rehydration was examined. Inoculation was achieved by rehydrating pasta samples in water containing 1‐mL spore crop (10⁶ spores/mL); the product was heat sealed in sterile pouches and processed in a laboratory retort. Thermal inactivation of spores was carried out using 10 time intervals at 121C for moisture contents, 70, 90, 105, 115, 125 and 145% dry basis. The death curves obtained consisted of two phases, an initial rapid decline (1.5 to 2.6 log reductions) followed by a slower, linear decrease of survivors from which the decimal reduction time (D value) was calculated. D₁₂₁ values were found to decrease with increasing moisture content and ranged from 4.6 to 6.5 min. The thermal resistance constant (z value) was examined to assess the effect of temperature (110–125C) on lethality. z values decreased with increasing moisture content and ranged from 10.7 to 15.6C. The results of this study demonstrate that temperature and moisture content influence the effectiveness of the intended heat process. For products that are rehydrated during heat treatment, the moisture content of the product is a critical factor. Therefore, if complete rehydration is not achieved, this may result in the heat process being less effective for the destruction of microorganisms, which may have implications in regard to food safety.     

EFFECT OF HIGH PRESSURE TREATMENTS ON PEROXIDASE AND POLYPHENOLOXIDASE ACTIVITIES

The effects of hydrostatic pressure of 1–9 kbar on peroxidase (POD) and polyphenoloxidase (PPO) activity were studied. Trials were carried out on crude enzymatic extracts obtained from carrots and apples as the sources of POD and PPO, respectively, with different pH values (from 4.5 to 7.0). In both cases pressurization caused a remarkable enzyme activation after 1-min treatments carried out at 3–5 kbar, while a complete enzyme inactivation was observed at 9 kbar. The pH values of the crude enzymatic extracts seemed to affect the degree of activation and/or inactivation due to different pressure levels. A slight recovery in enzyme activity was measured for POD samples during the first 24 h of storage after the treatments.     

INACTIVATION OF CLOSTRIDIUM BOTULINUM TYPE E SPORES BY HIGH PRESSURE PROCESSING

The effect of high pressure treatments at various temperature combinations on the inactivation of Clostridium botulinum type E spores of strains, Alaska and Beluga in phosphate buffer (0.067 M, pH 7.0) was investigated. No reduction of spores was observed at any pressurization (Maximum = 827 MPa) for temperatures below 35C. At pressurization of 827 MPa for 5 min, log unit reduction of spores increased as temperatures increased from 35 to 55C. An increase in the processing time from 5 to 10 min at a combination of high pressure (827 MPa) and low temperature (40C) resulted in a 5-log reduction of Alaska spores. About 5-log reductions for strains Alaska and Beluga occurred at 50 and 55C, respectively, after processing at pressures of 827 MPa for 5 min.     

EFFECTS OF HYDROSTATIC PRESSURE ON ALPHA-MACROGLOBULIN AND SELECTED PROTEASES

The enzymes, chymotrypsin, trypsin, collagenase and cathepsin C, as well as the protease inhibitor, α₂-macroglobulin, have been shown to be susceptible to hydrostatic pressure inactivation. a₂-Macroglobulin lost about 15% activity at a pressure of 2,000 atm and as much as 80% at 3,000 atm pressure applied for 1 h in both cases. Thermal stability studies also indicated the protein to be completely denatured at about 75C. The enzymes were also inactivated to various extents depending on the amount of pressure and the duration of application, The degree of susceptibility to pressures ranging between 2,000–4,000 atm applied for time periods ranging from 5–60 min was found to be as follows: chymotrypsin < cathepsin < trypsin < collagenase. The hydrostatic pressure effects were irreversible when pressure-treated enzymes were stored at room temperature (20–25C), but showed various levels of reactivation when stored at refrigerated temperatures (4–7C).     

EFFECT OF HIGH PRESSURE ON THE ACCUMULATION OF IMP AND ON THE STABILITY OF AMP DEAMINASE IN RABBIT SKELETAL MUSCLE

This study evaluated the effect of high pressure on rabbit skeletal muscle, specifically on the production of inosinic acid (IMP), one of “umami” components, and on the activity of adenosine triphosphate (AMP) deaminase, which plays a role in the conversion of AMP to IMP. By increasing the pressure (0.1 to 300 MPa), nucleotide analysis showed that IMP content in muscle increased instantly with a concomitant decrease in ATP content. The IMP content of muscle at 300 MPa was approximately 15% higher than with lower pressures (0.1–200 MPa) when stored for 1 week at 4C after pressurization. These results suggested that the metabolism of nucleotides in muscle was not significantly impaired by pressure treatment. At 300 MPa, AMP deaminase maintained approximately 70% of the activity at 0.1 MPa. In contrast, the activity of purified AMP deaminase was completely lost at 200 MPa, and irreversible conformational changes were observed by in situ fluorescence spectroscopy. These results indicated that purified AMP deaminase was irreversibly denatured under pressure as high as 300–400 MPa.     

HIGH HYDROSTATIC PRESSURE PROCESSING OF PEACH PUREE WITH AND WITHOUT ANTIBROWNING AGENTS

High hydrostatic pressure (HHP) (103 to 517 MPa) was applied for selected times (5 to 25 min) on peach puree with and without the addition of antibrowning agents (1000 ppm ascorbic acid or 300 ppm cysteine). Polyphenoloxidase (PPO) activity was assessed before and after HHP processing. Peach purees were stored at 3C for 13 days and then at 22C for 15 additional days. A z_p value of 224 MPa, in a range from 207 to 517 MPa of pressure, was obtained for peach puree without the addition of antibrowning agents. PPO activation at 103, 207 and 310 MPa or inhibition at 414 and 517 MPa was observed in peach purees containing added ascorbic acid. More than 50% inactivation of PPO was observed after adding cysteine to the peach puree and 99% inhibition was observed after 517 MPa of pressure at the experimental times. The higher the pressure treatment and the longer the treatment time for inactivating PPO, the less the discoloration of the peach puree with and without antibrowning agents during storage at low temperatures.