High pressure inactivation of Clostridium botulinum type E endospores in model emulsion systems

Clostridium botulinum type E is a cold-tolerant, neurotoxigenic, endospore-forming organism, primarily associated with aquatic environments. High pressure thermal (HPT) processing presents a promising tool to enhance food safety and stability. The effect of fat on HPT inactivation of C. botulinum type E spores was investigated using an emulsion model system. The distribution of spores in oil-in-water (O/W) emulsions and their HPT (300–750?MPa, 45–75?°C, 10?min) inactivation was determined as a function of emulsion fat content (30–70% (v/v) soybean oil in buffer). Approximately 26% and 74% of the spores were located at the oil–buffer interface and the continuous phase, respectively. Spore inactivation in emulsion systems decreased with increasing oil contents, which suggests that the fat content of food plays an important role in the protection of C. botulinum type E endospores against HPT treatments. These results can be helpful for future safety considerations.     

Effect of high hydrostatic pressure processing and squeezing pressure on some quality properties of pomegranate juice against thermal treatment

The aim of this study was to investigate the effect of high hydrostatic pressure (HHP) treatment (200, 300, 400?MPa; 5°C, 15°C and 25°C; 5 and 10 min) on some quality properties of pomegranate juice. Juice samples are obtained under industrial conditions at two different squeezing pressure levels (100 and 150?psi – 0.689 and 1.033?MPa, respectively). Results are compared against conventional thermal treatment (85°C/10 min) and raw sample. For all three processing temperature, HHP combinations at 400?MPa for 10 min were sufficient to decrease the microbial load around 4.0 log cycles for both squeeze levels. All HHP treatments showed no significant decrease at antioxidant activity, total phenolic content and monomeric anthocyanin pigment concentrations, while there was a significant decrease (p?≤?.05) in thermal-treated samples. Being the highest sugar alcohol in pomegranate juice, mannitol content must be considered for determining the authenticity, and mannitol content increased with squeezing pressure and thermal treatment.     

Comparison of microbiological loads and physicochemical properties of raw milk treated with single-/multiple-cycle high hydrostatic pressure and ultraviolet-C light

The effects of ultraviolet-C radiation (UV-C, 11.8?W/m²), single-cycle and multiple-cycle high hydrostatic pressure (HHP at 200, 400 or 600?MPa) on microbial load and physicochemical quality of raw milk were evaluated. Reductions of aerobic plate count (APC) and coliform count (CC) by HHP were more than 99.9% and 98.7%, respectively. Inactivation efficiency of microorganisms increased with pressure level. At the same pressure level, two-cycle treatments caused lower APC, but did not show CC differences compared with single-cycle treatments. Reductions of APC and CC by UV-C were somewhere between 200?MPa and 400/600?MPa. Both HHP and UV-C significantly decreased lightness and increased pH, but did not change soluble solids content and thiobarbituric acid-reactive substances’ values. Two 2.5?min cycles of HHP at 600?MPa caused minimum APC and CC, and maximum conductivity. Compared with HHP, UV-C markedly increased protein oxidation and reduced darkening.     

Moderate hydrostatic pressure–temperature combinations for inactivation of Bacillus subtilis spores

We report the effect of using moderate hydrostatic pressure, 40–140?MPa, at moderate temperature (38–58°C) to inactivate Bacillus subtilis spores in McIlvaine's citric phosphate buffer at pH 6. We have investigated several parameters: pressure applied, holding time, pressure cycling, and temperature. The kinetics of spore inactivation is reported. The results show that spore inactivation is exponentially proportional to the time the sample is exposed to pressure. Spore germination and inactivation occur at the hydrostatic pressures/temperature combinations we explored. Cycling the pressure while keeping the total time at high pressure constant does not significantly increase spore inactivation. We show that temperature increases spore inactivation at two different rates; a slow rate below 33°C, and at a more rapid rate at higher temperatures. Increasing pressure leads to an increase in spore inactivation below 95?MPa; however, further increases in pressure give a similar rate kill. The time dependence of the effect of pressure is consistent with the first-order model (R²?>?0.9). The thermal resistance values (Z_T) of B. subtilis spores are 30°C, 37°C, and 40°C at 60, 80, 100?MPa. The increase in Z_T value at higher pressures indicates lower temperature sensitivity. The pressure resistance values (Z_P) are 125, 125 and 143?MPa at 38°C, 48°C, and 58°C. These Z_P values are lower than those reported for B. subtilis spores in the literature, which indicates higher sensitivity at pressures less than about 140?MPa. We show that at temperatures <60°C, B. subtilis spores are inactivated at pressures below 100?MPa. This finding could have implications for the design of the sterilization equipment.     

The effect of high hydrostatic pressure on the physiological and biochemical properties of pepper (Capsicum annuum L.) seedlings

High hydrostatic pressure is a non-thermal food processing technology, which also has several successful applications in different areas besides food processing. In this study, Capsicum annuum L. (pepper) seeds are subjected to 50, 100, 200 and 300?MPa pressure for 5?min at 25°C and the seedlings of HHP processed seeds are used to compare percentage of seed germination and biochemical properties such as chlorophyll a, b and a/b, proline content, total protein, carotenoid, malondialdehyde, glucose, fructose and phenolic compounds concentrations. As a result of the study, it was observed that there are remarkable changes in terms of biochemical properties especially for seedlings, whose seeds were pressurized at 200 and 300?MPa. More detailed studies are needed to put forward the mechanism behind the changes in biochemical properties.     

Evaluation of high hydrostatic pressure effects on bovine red blood cells and platelets

The objective of this study was to investigate the effects of high hydrostatic pressure (HHP) on the stability of red blood cells (RBCs) and platelets. Bovine blood cells (n=5) were treated with the pressure of 55, 110, 154 and 220 MPa at 25 °C for 5 min. Light microscopy, atomic force microscopy and flow cytometry studies revealed that RBCs were morphologically stable up until the 220 MPa pressure treatments, at which surface modifications were observed. The platelets were found to be less stable than RBCs. HHP application did not cause any significant change in the signal intensity, band area and frequency values of the infrared bands with the exception that a significant variation was observed in the area of the cholesterol band. No statistically significant variations were observed in the secondary structure elements due to HHP treatment according to the artificial neural network study based on the FTIR data.     

High hydrostatic pressure pasteurization of a draft sake brewed using a Niigata-sake yeast

ABSTRACT

Draft (non-pasteurized) sake typically has a fresh flavor, but usually has to be consumed as soon as possible because of its short shelf life due to the potential for over-fermentation by residual yeast. In contrast, thermally pasteurized sake has a long shelf life, but the fresh flavor is lost during heat sterilization. High hydrostatic pressure (HHP) treatment can be used as a non-thermal pasteurization process while maintaining the characteristics of a draft sake. We evaluated the HHP inactivation behavior of sake yeast for the application of HHP pasteurization as an alternative to thermal pasteurization. The results showed complete pasteurization of sake yeast in the draft sake by HHP treatment at 400?MPa for 10?min. The viable cell count in HHP-pasteurized sake was below the detection limit during storage at 10°C for 3 months. Thus, we have established a HHP pasteurization technology to prevent over-fermentation and succeeded in producing a prototype of HHP-sparkling-cloudy sake, which we have designated AWANAMA.     

Use of pulsed-high hydrostatic pressure treatment to decrease patulin in apple juice

This study was aimed at reducing patulin content of apple juice using a non-thermal method, namely pulsed-high hydrostatic pressure (p-HHP). Commercially available clear apple juice was contaminated artificially with different concentrations of patulin (5, 50 and 100?ppb). Then, the samples were processed 5?min at different pressure treatments (300–500?MPa) in combination with different temperatures (30–50°C) and pulses (6 pulses?×?50?s and 2 pulses?×?150?s). To compare the impact of pulses, single pulse of high hydrostatic pressure (HHP) treatment was also applied with the same pressure/temperature combinations and holding time. Results indicated that pressure treatment in combination with mild heat and pulses reduced the levels of patulin in clear apple juice up to 62.11%. However, reduction rates did not follow a regular pattern. p-HHP was found to be more effective in low patulin concentrations, whereas HHP was more effective for high patulin concentrations. To the best of our knowledge, this is the first study using p-HHP to investigate the reduction of patulin content in apple juice.     

Enrichment of free amino acid content and reduction of astringent taste compounds in soybean by high hydrostatic pressure

ABSTRACT

We investigated the effect of high hydrostatic pressure (HHP) on the distribution of free amino acids and isoflavones in soybean immediately after pressure treatment (100–600?MPa, 10–60?min). HHP-treatment at 200 and 300?MPa resulted in high accumulation of free amino acids. Additionally, Gly, Val, and Pro levels increased even after HHP of 400?MPa. The application of HHP-treatment to soybean decreased sources of an astringent taste such as genistein-, daizein-, and glycitein-type molecules under all pressure conditions over 200?MPa. High accumulation of free amino acids with specific pressure–time conditions would be caused by the acceleration of proteolysis and specific amino acid metabolism. We concluded that HHP alone with no subsequent storage enabled the enrichment of specific amino acids such as GABA, Glu, Lys, and Pro in soybean. Soybean with enrichment of these amino acids and improved taste should be an innovative component of functional foods.     

The application of high pressure–mild temperature processing for prolonging the shelf-life of strawberry purée

The aim of this study was to monitor the shelf-life and quality of strawberry purée preserved using combined high pressure processing (HPP)–mild temperature processing at 300 and 600?MPa for 15 min during cold storage (6°C). Increasing the pressure resulted in a prolonged shelf-life of from 4 to 28 weeks for HPP-preserved purée at 300 and 600?MPa, respectively. The highest inactivation of peroxidases, pectinesterases and polygalacturonases was noted when a higher pressure was used, whereas a lower pressure was more efficient for polyphenoloxidases. The degradation of vitamin C and anthocyanins was 20% and 5% higher at 600?MPa than at 300?MPa, respectively. Significantly fewer changes in the colour coefficient, expressed as ΔE, and the browning index, were observed in purée preserved at 600?MPa. Oxidative and hydrolytic enzymes are highly pressure-resistant, which suggests other inhibitors should be used to increase the shelf-life of good-quality fruit products.     

Effects of high hydrostatic pressure on β-glucan content,swelling power,starch damage,and pasting properties of high-β-glucan barley flour

ABSTRACT

The effects of high hydrostatic pressure (HHP) on the physicochemical properties of high-β-glucan barley flour were investigated in the present study. Dough samples were made from two types of barley flour with low and high β-glucan content, respectively, and treated with HHP (200–600?MPa) for 10?min. Although the elevation of pasting properties for the samples treated at 600?MPa was reduced to the same extent as that in wheat flour at normal atmospheric pressure, β-glucan content was maintained regardless of the pressure applied. The significant increase in starch damage of the dough samples at 550 and 600?MPa was confirmed by the results of microscopic observation, which revealed that elliptical starch granules were cracked and damaged in samples with low β-glucan at 600?MPa, and in samples with high β-glucan content at 400?MPa or more. X-ray diffraction patterns of the samples treated at 600?MPa indicated the formation of amylose-lipid complexes that were considered to inhibit the elevation of viscosity.     

Fluorescence-based methods for the detection of pressure-induced spore germination and inactivation

The application of high pressure (HP) provides an opportunity for the non-thermal preservation of high-quality foods, whereas highly resistant bacterial endospores play an important role. It is known that the germination of spores can be initiated by the application of HP. Moreover, the resistance properties of spores are highly dependent on their physiological states, which are passed through during the germination. To distinguish between different physiological states and to detect the amount of germinated spores after HP treatments, two fluorescence-based methods were applied. A flow cytometric method using a double staining with SYTO 16 as an indicator for germination and propidium iodide as an indicator for membrane damage was used to detect different physiological states of the spores. During the first step of germination, the spore-specific dipicolinic acid (DPA) is released [P. Setlow, Spore germination, Curr. Opin. Microbiol. 6 (2003), pp. 550–556]. DPA reacts with added terbium to form a distinctive fluorescent complex. After measuring the fluorescence intensity at 270 nm excitation wavelength in a fluorescence spectrophotometer, the amount of germinated spores can be determined. Spores of Bacillus subtilis were treated at pressures from 150 to 600 MPa and temperatures from 37 °C to 60 °C in 0.05 M ACES buffer solution (pH 7) for dwell times of up to 2 h. During the HP treatments, inactivation up to 2log ₁₀ cycles and thermal sensitive populations up to 4log ₁₀ cycles could be detected by plate counts. With an increasing number of thermal sensitive spores, an increased proportion of spores in germinated states was detected by flow cytometry. Also the released amount of DPA increased during the dwell times. Moreover, a clear pressure-temperature-time-dependency was shown by screening different conditions. The fluorescence-based measurement of the released DPA can provide the opportunity of an online monitoring of the germination of spores under HP inside the HP vessel. Implementation can be done using diamond anvil cells, units with inspection glasses or by inserting an optical fiber into the HP vessel. The analytical methods used can help to understand the complex mechanism of germination and inactivation of bacterial spores. Due to its universal, process-independent character, the application of these methods is feasible for established and emerging technologies.     

Effect of plating and incubation conditions on the detection of lactic acid bacteria after high hydrostatic pressure treatment

ABSTRACT

Lactic acid bacteria (Leuconostoc mesenteroides, Enterococcus faecalis, and Lactobacillus fermentum) were subjected to high hydrostatic pressures (HHPs) of 400 and 600?MPa at 25°C for 10?min in phosphate-buffered saline. Differential plating methods were applied to evaluate HHP-treated cell populations, assuming that healthy and injured cells during plate incubation survived maximum and minimal stress, respectively. The stress was altered by using several selective media in combination with aerobic or anaerobic incubation at 25°C or 30°C. E. faecalis was detectable after 600?MPa treatment while L. mesenteroides and L. fermentum were nondetectable. Specific combinations of incubation conditions were suggested to determine maximum and minimum viable counts of L. mesenteroides and E. faecalis. The difference between the maximum and minimum counts can be used to evaluate HHP-injured population with reduced risks to overestimate healthy and/or underestimate HHP-injured cells.     

Effect of high hydrostatic pressure and reducing sodium chloride and phosphate on physicochemical properties of beef gels

ABSTRACT

The effect of high hydrostatic pressure (HHP) treatment (100–200?MPa, 10?min, 20°C) combined with sodium chloride and sodium phosphate on the physicochemical properties of beef gels was investigated. The water content, cooking losses, color, protein composition by SDS-PAGE analysis and texture parameters of beef gels were determined. The beef gels treated with high pressure at 150?MPa showed a synergistic effect in the increased water content and the decreased cooking losses compared with the unpressurized gels. The L*, a* and b* color values of beef gels were slightly decreased under HHP treatment at 100–200?MPa. In the SDS-PAGE analysis, the staining intensity of the α-actinin protein band was decreased in pressurized samples. The cohesiveness, adhesiveness, gel strength and modulus of elasticity were improved after HHP treatment. Application of high pressure treatment (150–200?MPa) before heat treatment would be beneficial for the manufacturing of low salt and/or low phosphate meat products for a healthy diet.     

Listeria monocytogenes cells injured by high hydrostatic pressure and their recovery in nutrient-rich or -free medium during cold storage

ABSTRACT

Cells of Listeria monocytogenes suspended in phosphate-buffered saline (PBS) were treated by high hydrostatic pressure (HHP; 500?MPa, 25°C, 10?min), diluted by ten folds using trypticase soy broth (TSB) or PBS, and stored at cold temperatures of 0–15°C. Viable cell count in TSB increased logarithmically close to the initial count at each storage temperature, while that in PBS increased temporarily and subsequently decreased to almost nondetectable level except the case at 15°C, where it showed logarithmic increase thereafter. Based on proliferation experiments where their healthy cells were inoculated to TSB or to PBS containing their heat-killed dead cells, it was suggested that increase in the viable count of HHP-treated cells in TSB and PBS could be ascribed to the recovery of colony forming ability and/or proliferation depending on the cold storage temperature.     

Effect of high hydrostatic pressure on enzyme activity in unpasteurized draft sake

ABSTRACT

Unpasteurized draft sake has a potentially high market value, due to its fresh flavor and fruity taste, compared with conventional thermal-pasteurized sake. However, the shelf life of draft sake is limited. To increase the shelf life of draft sake, it is necessary to suppress flavor and taste deterioration resulting from inactivation of enzymes produced by koji-mold. Draft sake was treated with high hydrostatic pressure (HHP) of 200 to 500?MPa at ?7 to 50°C to analyze the inactivation of α-amylases, glucose-forming enzymes, and acid carboxypeptidases. We found significant inactivation of enzymes produced by koji-mold in draft sake subjected to HHP treatment at both high and low temperature. However, HHP treatment at low temperature effectively inactivated enzymes while retaining the fresh flavor and fruity taste of draft sake.     

Inactivation of Staphylococcus aureus and native microflora in human milk by high pressure processing

The storage of unpreserved food, including breast milk, is associated with the growth of microorganisms, including pathogenic bacteria. It is therefore necessary to use suitable processes to eliminate pathogenic microorganisms and reduce the total microbial count in order to ensure product safety for consumers. In the present study, samples of milk obtained from volunteers donating to the human milk bank were artificially contaminated with Staphylococcus aureus ATCC 6538. This bacteria was the model microorganism of choice, being relatively resistant to high pressure as well as posing the most serious risk to infant health. The results obtained show that high pressure processing can reduce the count of S. aureus by about 5?log units at 4°C and about 8?log units at 50°C, and totally eliminate Enterobacteriaceae after 5?min of treatment, and result in a total microbial count reduction after 10?min treatment at 500?MPa at 20°C and 50°C. This suggests the possibility of this technology being applied to ensure the adequate safety and quality of human breast milk in human milk banks.     

Effects of high hydrostatic pressure on secondary structure and emulsifying behavior of sweet potato protein

In this study, secondary structures of sweet potato protein (SPP) after high hydrostatic pressure (HHP) treatment (200–600?MPa) were evaluated and emulsifying properties of emulsions with HHP-treated SPP solutions in different pH values (3, 6, and 9) were investigated. Circular dichroism analysis confirmed the modification of the SPP secondary structure. Surface hydrophobicity increased at pH 3 and decreased at 6 and 9. Emulsifying activity index at pH 6 increased with an increase in pressure, whereas emulsifying stability index increased at pH 6 and 9. Oil droplet sizes decreased, while volume frequency distribution of the smaller droplets increased at pH 3 and 6 with the HHP treatment. Emulsion viscosity increased at pH 6 and 9 and pseudo-plastic flow behaviors were not altered for all emulsions produced with HHP-treated SPP. These results suggested that HHP could modify the SPP structure for better emulsifying properties, which could increase the use of SPP emulsion in the food industry.     

The effect of high hydrostatic pressure on taste substance distribution in fresh green tea leaves

ABSTRACT

High hydrostatic pressure (HHP) can be an alternative method to steaming to inhibit enzymatic fermentation in green tea making process. However, the effect of HHP treatment on green tea taste is not clear. Thus, this study aimed to determine the effect of HHP on substances associated with green tea taste. Fresh green tea leaves were immediately treated with HHP at 300, 500, or 700?MPa for 10, 30, or 60 min at 25°C. The concentration of free amino acids, catechins, and caffeine in HHP-treated samples was quantified by LC-MS. The taste intensity of the samples was detected by taste sensors. HHP resulted in a high accumulation of free amino acids in green tea leaves, which was likely due to proteolysis. In particular, theanine synthesis may have been promoted by an increase in the concentration of substrates during HHP. Compared to steaming, HHP enhanced umami richness, and inhibited bitterness and astringency.     

Evaluation of the importance of germinative cycles for destruction of bacillus cereus spores in miniature cheeses

Our objective was to determine the effect of high pressure on inactivation of spores of Bacillus cereus ATCC 9139 inoculated into cheese made of raw cow's milk. Inoculated miniature cheeses were manufactured under controlled bacteriological conditions, vacuum packed and kept at 8 °C for 15 days after pressure treatment. Cheeses were submitted to pressures of 300, 400 or 500 MPa at 30 °C, during 15 min. Some of them were treated with a germination cycle of 60 MPa at 30 °C for 210 min. Lethality was calculated comparing surviving sample counts to control ones. Adding the germinative cycle resulted in higher efficiency, and when applied with 500 MPa, lethality reached 2.0 log cfu/mL. We saw that with both cycles initial counts of spores diminish, but all of them were not inactivated. However, considering that in raw milk mesophilic spore counts are 2.6-2.9 log cfu/mL, this treatment may be useful.