Model studies on the influence of high hydrostatic pressure on the formation of glycated arginine modifications at elevated temperatures

Aqueous solutions of Nalpha-acetylarginine and glucose were reacted for 2 h with pressure application from 0 to 600 MPa and varying temperatures between 90 and 120 degrees C. After enzymatic deacetylation of the reaction products, the glycated amino acids were separated by means of a self-assembled preparative ion exchange chromatography system using ninhydrin detection. On the basis of the use of eight synthesized reference compounds known in the literature as posttranslational arginine modifications, first, the presence of several glycated amino acids could be excluded. On the other hand, N5-[[(1-carboxyethyl)amino]iminomethyl]ornithine [N7-(1-carboxyethyl)arginine; N7-CEA; 12] was identified as a previously unknown arginine modification based on LC-MS, NMR measurements, and synthesis. In addition, N5-(5-hydro-5-methyl-4-imidazolon-2-yl)-L-ornithine (1) was identified as a further major reaction product. In further experiments, the formation of 1 and 12 was quantitatively followed at different pressures and/or temperatures. The results indicated that high hydrostatic pressure at elevated temperatures significantly increased the amounts of both arginine modifications. 2-Oxopropanal, known to form 1 in a reaction with arginine, was also quantified to explain the different yields observed after pressure application. A new formation mechanism leading to 12 by a reaction of the guanidine group or arginine with 2-oxopropanal is discussed.     

Changes in chymotrypsin hydrolysis of beta-lactoglobulin A induced by high hydrostatic pressure

Beta-lactoglobulin (beta-Lg) was subjected to high pressures up to 400 MPa and proteolysis with chymotrypsin. The hydrolysates were analyzed by SDS-PAGE and RP-HPLC, and the fragments obtained were identified by ESI-MS/MS. The results obtained showed that beta-Lg was hydrolyzed by chymotrypsin in a "progressive proteolysis" manner at either atmospheric or high pressure. Proteolysis during or after high-pressure treatment showed longer and more hydrophobic peptides than proteolysis at atmospheric pressure. Chymotrypsin showed a behavior similar to that of trypsin, with some differences, probably related to the orientation of the target residues specific for each enzyme. The similarities between proteolytic fragments produced by the two enzymes support that proteolysis enhancement under high pressure depends on the substrate changes rather than the enzyme. High pressure seemed to accelerate the first steps of proteolysis, probably through dimer dissociation, while leaving portions of the molecule more resistant to the enzyme.     

Investigation of binding behavior of alpha- and beta-ionones to beta-lactoglobulin at different pH values using a diffusion-based NOE pumping technique

Diffusion-based NMR techniques were employed to study effects of pH on beta-lactoglobulin (BLG) conformation and binding affinity to alpha- and beta-ionone. In the first part of the study, the influence of pH on the diffusion coefficient of BLG in D(2)O solution was investigated using a stimulated-echo NMR experiment. The diffusion coefficient of BLG decreased with increasing pH values. A significant decrease in the diffusion coefficient observed at pH 11 may be due to total unfolding (denaturation) of the protein, resulting in hydrophobically driven self-aggregation. A diffusion-based NOE pumping technique was then applied to determine the relative binding affinities between alpha- and beta-ionones and BLG at pH values varying from 3 to 11. An increase in signal intensities for beta-ionone with increasing molar concentration ratios between beta-ionone and BLG was observed at all pH ranges studied. The increased signal intensities reflect increased relative binding affinity. The greatest binding affinity occurred at pH 9 and the lowest at pH 11. alpha-Ionone showed binding evidence only at pH 9, and the binding was significantly weaker than that obtained for beta-ionone at the same pH. The high affinity observed for both aroma compounds at pH 9 may be due to a flexible conformation of BLG at this pH so that the flavor ligand accessibility increases. Conversely, alkaline denaturation occurring at pH 11 gives rise to relatively lower binding affinity compared to that observed at the other pH values.     

高静压杀菌对不同形状果块的黄桃罐头质地的影响

该文在保证黄桃罐头无菌的前提下,研究了高静压杀菌技术对不同形状果块的黄桃罐头质地的影响。首先通过测定不同的高静压条件下黄桃罐头细菌总数、霉菌和酵母数的变化,并结合热杀菌时间,确定出质地研究的杀菌条件为600 MPa/20 min。再将不同形状果块的黄桃罐头在600 MPa下处理20 min后,以90℃,20 min热杀菌的黄桃罐头为对照,通过测定2种杀菌方式前后及低温和常温贮藏过程中不同形状果块的黄桃罐头果肉硬度、汤汁黏度及果肉细胞结构的变化,得出了高静压杀菌结合低温贮藏技术在罐头质地保持方面的优越性。研究结果表明：高静压杀菌的3种形状果块的罐头果肉硬度分别为热杀菌的2.8倍（“条状”）,3.1倍（“丁状”）和2.9倍（“块状”）,高静压杀菌后“块状”黄桃罐头汤汁黏度较小,果肉细胞结构较完整;且高静压杀菌后低温贮藏（4±1）℃的黄桃罐头质地品质均优于常温贮藏（25±1）℃。在3种形状果块的黄桃罐头中,无论是低温贮藏（4±1）℃还是常温贮藏（25±1）℃,“块状”的黄桃罐头其果肉硬度最大,汤汁黏度最小,细胞结构破坏程度最小,是最适宜的黄桃罐头加工形状。研究结果为高静压技术取代热杀菌应用于黄桃罐头的加工提供了理论依据。     

Changes in conformation and subunit assembly of cod myosin at low and high pH and after subsequent refolding

Conformational and structural changes of cod myosin at pH 2.5 and 11 and after subsequent pH readjustment to pH 7.5 were studied. Results suggest that on acid unfolding, the myosin rod may fully dissociate due to electrostatic repulsion within the coiled coil, while it does not dissociate at alkaline pH. Both pHs led to significant conformational changes in the globular head fraction of the myosin heavy chains, suggesting that it takes on a molten globular configuration. A large part of the myosin light chains are lost on both pH treatments. On pH readjustment to neutrality, the heavy chains take on a structural form similar to the native state with the coiled-coil rod reassociating from acid pH while leaving the globular head less packed, more hydrophobic and structurally less stable. The irreversible change brought about in the globular head region leads to the failure of light chains to reassemble onto it, a drastic loss in ATPase activity, and more exposure of reactive thiol groups. The acid and alkali processes therefore lead to substantial changes in the globular part of the myosin molecule and perhaps more importantly to different molecular changes in myosin, depending on which pH treatment is employed.     

Soy glycinin: influence of pH and ionic strength on solubility and molecular structure at ambient temperatures

This study describes the relationship between the solubility of glycinin, a major soy protein, and its structural properties at a quaternary, tertiary, and secondary folding level under conditions representative for food products. When the ionic strength is lowered from 0.5 to 0.2 or 0.03, the basic polypeptides shift more to the exterior of the glycinin complex, as determined at pH 7.6 by labeling solvent-exposed lysines, supported by the study of the proteolytic action of clostripain on glycinin. This structural reorganization caused the pH of minimal solubility to shift to higher values. Ultracentrifugational analysis shows that at pH 7.6 and an ionic strength of 0.5 glycinin forms hexameric complexes (11S), whereas at pH 3.8 and at an ionic strength of 0.03 glycinin exists as trimers (7S). Intermediate situations are obtained by modulation of pH and ionic strength. The observed quaternary dissociation correlates with an increased amount of nonstructured protein at a secondary level and with changes in tertiary folding as determined using circular dichroism. Tryptophan fluorescence shows no significant structural changes for different ionic strengths but demonstrates a more tightly packed fluorophore environment when the pH is lowered from 7.6 to 3.8.     

The effect of high hydrostatic pressure on the strawberry anthocyanins

Color stability of fruit juice made from strawberries (Fragaria x ananassa, cv. Elsanta) that were subjected to high hydrostatic pressure was studied by measuring the anthocyanin content. High hysrostatic pressure is a method of preservation of food alternative to heat treatment. It is therefore essential to assess the impact of high pressure on color molecules. Samples were pressurized under 200, 400, 600, and 800 MPa for 15 min at a temperature controlled between 18 and 22 degrees C. After application of pressure, the anthocyanin content of the strawberry juice was analyzed by HPLC-UV using a novel isocratic elution system. The high-pressure treated samples were kept at refrigerator temperature (4 degrees C), room temperature (20 degrees C), and 30 degrees C. Two pigments were identified and quantified: pelargonidin 3-glucoside and pelargonidin 3-rutinoside. The highest stability of the anthocyanins was found when strawberries were stored at a temperature of 4 degrees C. High-pressure treatment at 800 MPa led to the lowest losses, at 4 degrees C.     

Modification of IgE binding to beta-lactoglobulin by fermentation and proteolysis of cow's milk

The effect of fermentation by Lactobacilli and of proteolytic hydrolysis of whole milk on the IgE binding ability of beta-lactoglobulin was studied using an ELISA inhibition assay. Sera from nine adult milk allergic patients were tested. The individual sera showed a similar inhibition pattern in the changes during fermentation and proteolysis. The degradation of beta-lactoglobulin was studied with liquid chromatography. In general, fermentation with Lactobacilli gave little effect on IgE binding, even though chromatography data showed a gradual degradation of beta-lactoglobulin. Proteolysis with trypsin, however, gave extensive degradation of beta-lactoglobulin and strongly decreased IgE binding. In addition, we measured the inhibition pattern of beta-lactoglobulin in various selected commercially available fermented milk products. These showed an IgE binding capacity similar to that of nonfermented high pasteurized milk.     

超高压对鲜榨西瓜汁杀菌效果和风味的影响

为了研究超高压（HHP）处理对鲜榨西瓜汁的杀菌效果及风味的影响,采用HHP技术对鲜榨西瓜汁进行处理。以95℃,1 min热处理作为对照,重点考察了600 MPa,60 min HHP处理对西瓜汁中菌落总数、霉菌和酵母菌数及典型风味化合物含量的影响;并探讨了2种处理西瓜汁在4℃,30 d贮藏过程中上述指标的变化情况。结果表明：95℃,1 min热处理和600 MPa,60 min HHP处理后西瓜汁中的微生物指标均符合《果、蔬汁饮料卫生标准》的要求;并且25 d的保质期满足鲜榨西瓜汁的消费要求（保质期≥7 d）。600 MPa,60 min HHP处理对西瓜汁的典型风味化合物含量影响较小;4℃,30 d贮藏过程中典型风味物质含量不发生变化(P＞0.05)。总体看来,HHP处理更适合于鲜榨西瓜汁的加工。     

Gelation of iota-carrageenan and micellar casein mixtures under high hydrostatic pressure

Effects of high-pressure treatment (HPT) on the rheological parameters and gelation of iota-carrageenan (iota-Car) and mixtures of micellar casein (MC) and iota-Car have been investigated under neutral pH conditions. It was established that HPT showed no significant effect, in the presence or absence of ionic calcium, with or without initial thermal processing, on the rheology or gelation/melting temperatures of the pure iota-Car solution. However, in mixed systems containing varying concentrations of iota-Car (up to 1 wt %) and MC (up to 8 wt %), considerable changes were detected. At the higher molar ratios of MC to iota-Car, and especially at the higher pressures, the dispersions were not thermoreversible in gelation, presumably due to the strong interactions of disrupted casein micelles with iota-Car molecules, as well as due to the formation of a dominant proteinaceous network at higher concentrations of MC. The associative protein-polysaccharide interactions in these systems are highly dependent on the ionic calcium content.     

Comparison of protein surface hydrophobicity measured at various pH values using three different fluorescent probes

The influence of type of fluorescent probe on the surface hydrophobicity values determined for three native and heated proteins was assessed using uncharged [6-propionyl-2-(N, N-dimethylamino)naphthalene or PRODAN] versus anionic aliphatic (cis-parinaric acid or CPA) and aromatic (1-anilinonaphthalene-8-sulfonic acid or ANS) probes. Surface hydrophobicities of whey protein isolate, beta-lactoglobulin, and bovine serum albumin under heated (80 degrees C for 30 min) and unheated conditions and at varying pH values (3.0, 5.0, 7.0, and 9. 0) were measured using ANS, CPA, and PRODAN. ANS and CPA yielded opposing results for the effects of pH and heating on protein hydrophobicity. Hydrophobicity was lower at pH 3.0 than at other pH values for all proteins measured by PRODAN, whereas the values measured by ANS and CPA at pH 3.0 were quite high compared to those at other pH values, suggesting the influence of electrostatic interactions on anionic probe-protein binding. These results suggest that the presence or absence of a permanent charge as well as the aromatic and aliphatic nature of fluorescent probes can affect protein hydrophobicity values measured under various pH conditions.     

高静压加工对黄桃罐头品质的影响（英）

为拓展非热加工技术在水果罐头加工中应用,以黄桃罐头为试材,研究了高静压加工对黄桃罐头品质的影响。分析了黄桃罐头果肉或汤汁的质地、显微结构、色泽、pH值、可滴定酸和可溶性固形物在加工前后变化,并与传统热加工产品进行比较。结果表明：高静压加工（600 MPa, >5 min）能有效地保持产品质地、显微结构、色泽、pH值、可滴定酸和可溶性固形物,而热加工（90±2°C, 20 min）导致果肉质地软化、显微结构破坏及可溶性固形物增加,严重影响产品品质。同时感官分析表明,与对照相比,高静压加工黄桃罐头除新鲜度及滋味稍差外,其他方面均无明显差别,而热处理黄桃罐头除保持较好色泽与香气外,其他方面均产生劣质的影响。所得研究结果将为高静压技术在水果罐头中工业化应用提供理论依据。     

食品工程用超高压设备的设计及性能试验

为了提高超高压设备的稳定性和处理效率,促进超高压技术应用于食品加工,该文阐述了HHP-700-400L型食品工程用大型超高压设备的工作原理、设计要求、主体结构和工作过程,分析并确定了双向水介质增压器、直推式容器堵头及其密封系统、承压容器材料和结构等相关参数。设备单体容积为100L,总容积为400L;最高工作压力为600MPa;处理能力达到1.68t/h。经性能试验表明:设备性能稳定、操作方便、安全可靠。研究结果为大型商业化超高压设备的设计和制造提供参考。     

超高压对鲜榨果蔬汁的杀菌效果

为防止鲜榨果蔬汁受微生物污染而腐败变质,通过超高压加工处理降低其污染的风险,延长产品货架期,提高产品的安全性。该文通过研究超高压处理鲜榨苹果汁和胡萝卜汁及其在贮藏期的菌落总数变化,确定超高压杀菌后的鲜榨苹果汁和胡萝卜汁的生物安全性。试验结果表明：超高压对低pH值的鲜榨苹果汁的杀菌效果优于pH值偏中性的鲜榨胡萝卜汁。随着处理压力的增大和处理时间的延长,鲜榨苹果汁和胡萝卜汁中的菌落总数显著降低（P＜0.05）。经过400 MPa、15 min处理的鲜榨苹果汁可在4℃下贮藏7 d仍保持食用安全性,而鲜榨胡萝卜汁经400 MPa、45 min处理,仅能在4℃下贮藏3d。     

超高压处理对绵羊肉嫩化机理的研究

实验研究了超高压处理条件下绵羊肌肉感官特性、显微结构、钙激活酶(Calpains)粗酶活性和剪切力值的变化，并探讨了超高压处理对绵羊肌肉的嫩化机理。超高压处理后绵羊肌肉的感官特性发生变化，随处理压力升高，绵羊肌肉颜色变淡，出现轻微的类似蒸煮的成熟风味。在压力为400 MPa，保压时间为10 min的处理条件下，绵羊肌肉显微组织结构变化明显：肌节收缩，肌原纤维的Z线断裂，M线降解，I带变白。当压力水平在100～400 MPa范围变动时，随压力升高，Calpains粗酶活性显著下降(P＜0.01)；当压力达到400 MPa时，Calpains粗酶活性几乎失活。超高压处理后绵羊肉的剪切力值显著下降(P＜0.05)。实验结果表明，超高压处理促进了绵羊肉的嫩化。     

Lipoxygenase inactivation in green beans (Phaseolus vulgaris L.) due to high pressure treatment at subzero and elevated temperatures

The kinetics of lipoxygenase (LOX) inactivation in green beans due to high-pressure treatment were studied in the pressure-temperature area of 0.1 up to 650 MPa and -10 up to 70 degrees C for systems with different levels of food complexity, i.e., in green bean juice and intact green beans (in situ study). For both systems, LOX was irreversibly inactivated by high-pressure treatment combined with subzero and elevated temperatures and the inactivation could be described as a first-order reaction. At ambient pressure, in situ LOX was less thermostable than in the juice at temperatures below 68 degrees C whereas the stability ranking was reverse at temperatures above 68 degrees C. At temperatures below 63 degrees C, sensitivity of the inactivation rate constants to temperature changes was on the same order of magnitude in the juice and in situ, while at higher temperature it was lower in situ. The pressure needed to obtain the same rate of LOX inactivation at a given temperature was lower in situ than in the juice. Application of high-pressure treatment at low/subzero temperature resulted in an antagonistic effect on LOX inactivation for both systems, whereas no such effect was found above room temperature. The pressure-temperature dependence of the LOX inactivation rate constants in green beans was successfully modeled.     

生物质炭对不同pH值土壤矿质氮含量的影响

为了揭示生物质炭作为土壤调理剂添加后对土壤矿质氮形态、含量等土壤性质的影响,该研究利用芒草分别在350和700℃裂解制得生物质炭,发现2个温度尤其是700℃制得的生物质炭,对NH4+有很强的吸附能力,但对NO3-的吸附能力很弱。将生物质炭分别加入到酸性(pH值为3.8)和碱性(pH值为7.6)土壤中,25℃下室内培养180d。结果表明,生物质炭提高了土壤全氮含量,酸性和碱性土壤分别平均提高了22%和17%;但使土壤铵态氮含量大幅降低至接近仪器检测限水平;生物质炭对土壤硝态氮含量的影响因生物质炭和土壤类型而异。生物质炭对土壤矿质氮形态和含量的影响,显然与生物质炭对铵的吸附作用、提高土壤pH值、增强氨挥发损失,以及形成微生物量氮等密切相关。该研究可为开展生物质炭基氮素新型肥料及制剂等方面的科学研究提供参考。     

Hydrophobic probe binding of beta-lactoglobulin in the native and molten globule state induced by high pressure as affected by pH,KIO(3) and N-ethylmaleimide

High hydrostatic pressure (HHP) at 500 MPa and 50 degrees C induces beta-LG into the molten globule state. Retinol, cis-parinaric acid (CPA), and 1-anilino-naphthalene-8-sulfonate (ANS) fluorescence from pH 2.5 to 10.5 in the presence of the native and molten globule states of beta-LG indicate that retinol binds to beta-LG in the calyx, CPA at the surface hydrophobic site, and ANS in multiple hydrophobic sites. HHP treatment results in a decrease of beta-LG affinity for retinol and CPA, suggesting conformational changes in the calyx and surface hydrophobic site of beta-LG during HHP treatment. beta-LG treated by HHP in the presence of N-ethylmaleimide (NEM) retains retinol affinity, suggesting that NEM protects the calyx conformation of beta-LG during HHP treatment. HHP treatment of beta-LG in the presence of KIO(3) exhibits a great decrease of CPA affinity compared to HHP-treated beta-LG in the absence of KIO(3), suggesting the formation of non-native disulfide bonding at the CPA binding site.     

Effect of high hydrostatic pressure on the volatile components of a glucose-lysine model system.

An aqueous glucose-lysine model system (initial pH 10.1) was incubated at 60 degrees C and atmospheric pressure (system A) or 600 MPa (system B) to the same absorbance value at 420 nm. Volatile reaction products were isolated by solvent extraction and analyzed by gas chromatography/mass spectrometry. Thirty-two compounds were identified; most contained nitrogen, and pyrazines predominated. Yields of all compounds were suppressed at 600 MPa. Further incubation, at either atmospheric pressure (system C) or 600 MPa (system D), of system A, resulted in lower yields of many compounds at 600 MPa, compared to prolonged incubation at atmospheric pressure. Many of the compounds reported may be formed by, or subsequently react via, aldol condensation. The observed differences among the systems in the profiles and yields of volatile compounds suggest that aldol condensations increase in rate in the systems under pressure.