beta-Lactoglobulin A with N-ethylmaleimide-modified sulfhydryl residue, polymerized through intermolecular disulfide bridge on heating in the presence of dithiothreitol

Roles of sulfhydryl groups on thermal aggregation of beta-lactoglobulin A (betaLG A) at pH 7.5 were investigated. It is known that betaLG A modified at Cys(121) with N-ethylmaleimide (NEM-betaLG A) does not form an aggregate by heating and that dithiothreitol (DTT) reduces cystine residues and induces the intermolecular sulfhydryl/disulfide interchange reaction and/or oxidation. NEM-betaLG A was heated in the presence of DTT. The molecules were linked together with an intermolecular disulfide bridge, and the polymer formed increased with increase in DTT concentration. The largest portion of polymer was formed when DTT was added at around the same molar concentration as that of NEM-betaLG A. Then, polymer formation decreased with further increase in DTT concentration. The results suggest that sulfhydryl/disulfide residues other than Cys(121), generated from cysteine residues, can induce intermolecular sulfhydryl/disulfide interchange reactions to polymer and that thiol compounds, for example, added DTT, are capable of starting such reactions.     

Cholesterol oxidation in meat products and its regulation by supplementation of sodium nitrite and apple polyphenol before processing

The levels of cholesterol oxidation derivatives (OxChol) in eight commercial species of meat products were examined. These products contained more than 1 mg/100 g of OxChol, and 7beta-hydroxycholesterol + 5beta-epoxycholesterol (111-1092 microg/100 g), 5alpha-epoxycholesterol (80-712 microg/100 g), cholestanetriol (0-368 microg/100 g), and 7-ketocholesterol (708-1204 microg/100 g) were detected. To know the interaction of sodium nitrite supplementation against cholesterol oxidation in meat products, sausage was produced with or without varying levels of sodium nitrite and stored in the refrigerator for 15 days. As a result, cholesterol oxidation in sausage was inhibited by addition of sodium nitrite in a dose-dependent manner. This observation may be associated with inactivation of O(2)(-) radical and stabilization of polyunsaturated fatty acids (PUFAs). In fact, the levels of OxChol in sausage increased, accompanying the decrease of coexisting linoleic acid when sodium nitrite was not added to sausage meat. Thus, cholesterol oxidation in meat products seems to be considarably promoted by the oxidation of coexisting PUFAs. On the other hand, additive apple polyphenol also inhibited linoleic acid oxidation in sausage and then suppressed cholesterol oxidation through its radical scavenging effects. Therefore, apple polyphenol, having a large amount of an oligomer of catechin, may interfere with cholesterol oxidation in meat processing or storage of meat products through its antioxidative action and be useful as a new antioxitant for meat products when it is added to the original meat before processing.     

超声辅助变压滚揉对鸡肉蛋白质结构及含水量的影响

为了探讨超声辅助变压滚揉对原料肉腌制过程蛋白质结构及水分含量的影响,该研究以鸡胸肉为原料分别进行静置腌制(对照)、真空滚揉(vacuum tumbling,VT)腌制和超声辅助变压滚揉(pressure-transform tumbling curing assisted by ultrasound,PTU)腌制处理100 min,采用拉曼光谱、低场核磁共振(low-field nuclear magnetic resonance,LF-NMR)、扫描电镜(scanning electron microscopy,SEM)和透射电镜(transmission electron microscope,TEM)测定不同腌制处理后鸡肉肌原纤维蛋白的二级结构、肌肉中水分含量以及肌肉组织微观结构的变化.结果显示,经过VT和PTU腌制处理的鸡肉其各项指标较对照组均发生显著变化(P＜0.05),其中拉曼光谱分析显示PTU处理的鸡肉肌原纤维蛋白质二级结构组成发生更显著变化,α-螺旋含量降低(P＜0.05),β-折叠和-转角的含量显著增加(P＜0.05);LF-NMR结果显示PTU处理的鸡肉的结合水横向弛豫时间和不易流动水横向弛豫时间较VT处理显著减小(P＜0.05),而对应的结合水峰面积百分比和不易流动水峰面积百分比都较其他处理组增加(P＜0.05).扫描电镜和透射电镜研究结果也显示经PTU处理的鸡肉组织结构破坏最严重,Z线以及肌原纤维膜降解最为明显.综上,PTU处理能够通过改变蛋白质结构、水分存在状态分布以及微观结构的变化来加速原料鸡肉腌制效率,改善嫩度和提高保水性;研究结果为PTU技术在肉制品加工中的应用及相关设备的开发提供理论依据.     

Biogenic amine formation and nitrite reactions in meat batter as affected by high-pressure processing and chilled storage

Changes in biogenic amine formation and nitrite depletion in meat batters as affected by pressure-temperature combinations (300 MPa/30 min/7, 20, and 40 degrees C), cooking process (70 degrees C/30 min), and storage (54 days/2 degrees C) were studied. Changes in residual nitrite concentration in raw meat batters were conditioned by the temperature and not by the pressure applied. Cooking process decreased (P < 0.05) the residual nitrite concentration in all samples. High-pressure processing and cooking treatment increased (P < 0.05) the nitrate content. Whereas protein-bound nitrite concentration decreased with pressure processing, no effect was observed with the heating process of meat batters. High-pressure processing conditions had no effect on the rate of residual nitrite loss throughout the storage. The application of high pressure decreased (P < 0.05) the concentration of some biogenic amines (tyramine, agmatine, and spermine). Irrespective of the high processing conditions, generally, throughout storage biogenic amine levels did not change or increased, although quantitatively this effect was not very important.     

Myoglobin oxidation in a model system as affected by nonheme iron and iron chelating agents

A model system was used to study the effect of nonheme iron on myoglobin oxidation at pH 5.6 and pH 7.2 at 23 degrees C. The addition of ferrous iron significantly (p < 0.05) increased the rate of myoglobin oxidation in the absence of lipid, demonstrating that iron promoted myoglobin oxidation independent of the effect of lipid oxidation. The addition of the type II, iron chelating antioxidants sodium tripolyphosphate (at pH 7.2) or milk mineral (at pH 5.6) negated the effect of added iron, slowing oxidation of myoglobin. A clear concentration dependence was seen for iron-stimulated myoglobin oxidation, based on both spectral and visual evidence. Further investigation is needed to determine the possible role for nonheme ferrous iron on myoglobin oxidation in vivo or in meat.     

发色剂对中式香肠营养、风味成分的影响

为寻求应用于肉制品加工的亚硝酸盐替代物,以中式香肠为研究对象,从感官评价、营养成分及挥发性风味物质组成探究不同发色剂(试验组1:0.5%甜菜粉+50 mg·kg^-1亚硝酸钠、试验组2:0.1%甜菜粉+0.06%改性血红蛋白、试验组3:100 mg·kg^-1亚硝酸钠)对产品营养和风味品质的影响。结果表明,试验组1和2香肠的外观、色泽和口感与试验组3差异不显著,且均与对照组差异显著(P<0.05);试验组1氨基酸总含量显著高于其他组(P<0.05);试验组2脂肪酸含量相对最高,其中十八碳二烯酸含量显著高于其他各组(P<0.05);通过风味分析,试验组1、试验组2分别检出27、26种挥发性成分,风味物质种类多于试验组3,其中试验组2醛类、酯类化合物含量相对较高,风味更好。本研究结果为中式香肠加工过程中亚硝酸盐的有效替代,提升中式香肠品质提供了一定的技术指导。     

Investigation on the possible formation of N-nitroso-N-methylurea by nitrosation of creatinine in model systems and in cured meats at gastric pH

N-Nitroso-N-methylurea (NMU) is a highly potent direct-acting carcinogen that has been shown to induce cancer in a number of animal species. Although previous research has indicated that nitrosation of creatinine (CRN), a common constituent of meats, dried fish, and seafoods, can form traces of NMU, there is uncertainty as to (1) the yield of NMU and (2) whether detectable amounts of NMU can be formed from cured meats following nitrosation under acidic conditions given the low residual levels of nitrite found in cured meats at the present time. Lack of sensitive and specific analytical methods most likely has hindered progress in research in these areas. An HPLC postcolumn denitrosation-thermal energy analyzer technique and a GC-MS confirmation technique were developed for the determination of NMU in cured meats. Both techniques are highly sensitive (0.5 and 0.03 ppb, respectively) and specific. The optimum pH for NMU formation from CRN ranged between pH 1 and pH 3, and the yields of NMU under variable reactant concentrations ranged between 0.00004 and 0.0046%. When 27 samples of various cured meats (10 g aliquots each) were acidified with HCl (final pH values of 0.8-2.5) and incubated at room temperature for 2 h, without any additional nitrite, 24 gave results below detectable levels but 3 formed 2-26 ng of NMU/10 g of meat. Incubation of the negative meats with additional nitrite (50-500 microg/g of meat) formed 0.6-176 ng of NMU/10 g of sample. Although the amounts of NMU formed were extremely small, this seems to be the first reported formation of NMU from cured meats with and without additional nitrite.     

超声联合滚揉腌制对鸡胸肉品质的影响研究

为了改善淘汰蛋鸡鸡胸肉的品质,探明超声联合滚揉腌制对鸡胸肉保水性、嫩度和挥发性风味成分的影响。本研究基于不同腌制工艺下鸡胸肉腌制液的吸收率和肉品质(蒸煮损失、离心损失、剪切力)的差异,通过低场核磁共振、组织切片和顶空固相微萃取气相质谱联用(HS-SPME-GC-MS)技术分析了鸡胸肉品质发生变化的原因。结果表明,滚揉腌制和超声联合滚揉腌制均加快了腌制速率,提高了鸡胸肉的保水性和嫩度,原因可能是滚揉和超声波的机械作用促进了水分的迁移和均匀分布,破坏了肌纤维结构,并且超声和滚揉具有协同效果。此外,采用滚揉腌制和超声联合滚揉腌制对鸡胸肉挥发性风味成分的种类及含量都有一定影响。本研究结果为改良鸡肉品质、开发低磷酸盐含量的鸡肉制品提供了理论参考。     

羟自由基氧化对牦牛肉肌原纤维蛋白生化特性的影响

为探究羟自由基(·OH)氧化处理对宰后牦牛肉成熟过程中肌原纤维蛋白氧化特性的影响,以经不同浓度(0、0.5、1、5、10、20 mmol·L^-1)羟自由基氧化体系氧化不同时间(0、0.5、1、12、24、48 h)的牦牛肉为研究对象,以未经处理的牦牛肉为空白对照,测定不同浓度不同氧化时间下肉样肌原纤维蛋白的羰基含量、巯基含量、二硫键含量、表面疏水性、Ca/K-ATPase活性等指标的变化规律。结果表明,随着H₂O₂浓度的增加及氧化时间的延长,不同处理组间牦牛肉肌原纤维蛋白均表现出不同程度的氧化水平,其中羰基含量、二硫键含量和Ca-ATPase活性均呈显著增加趋势(P<0.01),而表面疏水性呈先上升后下降的趋势,巯基含量和K-ATPase活性则显著降低(P<0.01)。H₂O₂浓度增加至20 mmol·L^-1时,与空白对照组相比,羟自由基处理组羰基含量升高46.03%,而巯基含量下降79.79%。综上,羟自由基促使宰后牦牛肉成熟过程中肌原纤维蛋白发生了显著氧化,增加了蛋白的氧化程度。本研究结果为牦牛肉生产加工过程中蛋白氧化控制提供了一定的参考。     

解冻方式对调理猪肉饼品质的影响

为探索不同解冻方式对猪肉饼品质和蛋白质氧化程度的影响,本试验将-18℃冻藏的调理猪肉饼分别采用室温解冻、冷藏解冻、流水解冻以及盐水解冻4种方式进行解冻,通过测定解冻后肉饼的保水性(解冻损失率、离心损失率)、pH值、TBARS值、TVB-N值,并测定猪肉饼肌原纤维蛋白羰基含量、巯基含量、蛋白溶解等,分析不同解冻方式对其蛋白氧化程度的影响。结果表明,解冻对调理猪肉饼保水性有较大影响,其中空气解冻与冷藏解冻组的保水性最好,解冻损失维持在5.03%~5.75%之间,离心损失率在31.02%~32.19%之间;同时与新鲜肉饼组相比,空气解冻、盐水解冻和流水解冻组猪肉饼TVB-N值显著上升(P<0.05),盐水解冻组TVB-N值上升至3.17 mg·100g^-1;但解冻对调理肉饼pH值、TBARS值、色泽的影响均不显著(P>0.05);通过对肌原纤维蛋白氧化程度的测定发现,解冻会引起肌原纤维蛋白羰基含量上升、巯基含量和溶解度下降,SDS-PAGE试验也证明解冻会引起蛋白质发生不同程度的降解。本研究为实际生产中冻结肉制品解冻工艺的选择与优化提供了一定的理论依据。     

Determination of total N-nitroso compounds and their precursors in frankfurters, fresh meat, dried salted fish, sauces, tobacco, and tobacco smoke particulates.

Total N-nitroso compounds (NOC) and NOC precursors (NOCP) were determined in extracts of food and tobacco products. Following Walters' method, NOC were decomposed to NO with refluxing HBr/HCl/HOAc/EtOAc and NO was measured by chemiluminescence. NOC were determined after sulfamic acid treatment to destroy nitrite, and NOCP were determined after treatment with 110 mM nitrite and then sulfamic acid. Analysis without HBr gave results < or =20% of those with HBr. This NOC method was efficient for nitrosamines but not nitrosoureas. The standard nitrosation for determining NOCP gave high yields for readily nitrosated amines, including 1-deoxy-1-fructosylvaline, but not for simple amines, dipeptides, and alkylureas. Mean NOC and NOCP results were (respectively, in micromol/kg of product) 5.5 and 2700 for frankfurters, 0.5 and 660 for fresh meat, 5.8 and 5800 for salted, dried fish, and 660 and 2900 for chewing tobacco (all for aqueous extracts) and 220 and 20000 nmol/cigarette for MeCN extracts of cigarette smoke filter pads.     

Intake and food sources of nitrites and N-nitrosodimethylamine in Spain

OBJECTIVE: To conduct a comprehensive assessment of dietary intakes of nitrites and N-nitrosodimethylamine (NDMA). SUBJECTS AND SETTING: A study was conducted within the Spanish cohort of the European Prospective Investigation in Cancer and Nutrition (EPIC) to assess the intake and food sources of these compounds in Spanish adults. The study included 41,446 health volunteers, aged 29-69 years, from Northern and Southern regions. Usual food intake was estimated by in-person interviews using a computerised dietary questionnaire. RESULTS: The estimated geometric mean was 0.994 mg day(-1) for nitrites and 0.114 microg day(-1) for NDMA. For both compounds a positive trend in consumption with increasing energy intake was observed. Dietary NDMA was related to age and sex after energy adjustment, while nitrite consumption increased with higher intakes of vitamin C (P < 0.001). The food groups that contributed most to intakes were meat products, cereals, vegetables and fruits for nitrites, and processed meat, beer, cheese and broiled fish for NDMA. Current and past smokers, who had high levels of NDMA from tobacco exposure, were also identified as the highest consumers of dietary NDMA. Furthermore, smokers had low intakes of vitamin C (an inhibitor of endogenous nitrosation). CONCLUSIONS: Intake levels of NDMA and nitrites in a Mediterranean cohort are currently relatively lower than those previously reported, although processed meat, beer and cured cheese still are the most important contributors to NDMA intake.     

Interaction of soluble peptides and proteins from skeletal muscle with volatile compounds in model systems as affected by curing agents

The effect of curing agents (salt, glucose, nitrate, nitrite, and ascorbic acid) on the binding of skeletal peptides (carnosine and anserine) and a sarcoplasmic protein (myoglobin) with key flavor compounds (hexanal, octanal, 2-pentanone, 2-methylbutanal, and 3-methylbutanal) has been studied by solid-phase microextraction (SPME). Curing agents had an effect on the interaction process between carnosine and volatile compounds, which was higher than the interactions observed with anserine and myoglobin. Sodium chloride decreased the interaction of volatiles with carnosine except for octanal, which was increased, and 2-pentanone, which was unaltered. Ascorbic acid exerted the highest effect by decreasing the interaction of carnosine with all of the volatile compounds except for octanal and 2-pentanone. The interaction with anserine was affected by sodium chloride, nitrate, and nitrite, producing a decrease in the interaction with hexanal, octanal, and methional. Finally, sodium chloride, glucose, and nitrite increased the interaction of myoglobin with hexanal, octanal, and methional. The effect of simulated stages of the curing process on the binding was also studied. A combined effect of the curing agents resulted in a change in the relative proportions of volatile compounds that can lead to different flavor perceptions of dry-cured meat products.     

Effect of glutathione on oxymyoglobin oxidation

The oxidation of oxymyoglobin (OxyMb) to metmyoglobin (MetMb) is responsible for fresh meat discoloration. Glutathione (GSH) is an important tripeptide reductant that can protect lipid and protein from oxidation. The objective of this research was to investigate the effect of GSH on MetMb formation in vitro and in bovine skeletal muscle cytosol. Equine MetMb formation was greater in the presence of GSH than controls at pH 5.6 or 7.2 and 25 or 37 degrees C (p < 0.05); GSH addition to purified bovine OxyMb solution also resulted in more MetMb formation at pH 7.2 and 25 or 37 degrees C (p < 0.05). This effect on MetMb formation was partly or completely inhibited by EDTA or catalase in the GSH-equine OxyMb system (p < 0.05). The addition of GSH to bovine muscle cytosol inhibited MetMb formation at pH 5.6 or 7.2 and 4 or 25 degrees C (p < 0.05); the effect was concentration-dependent. The inhibitory effect was observed in a high molecular weight (HMW) but not low molecular weight fraction of cytosol at pH 7.2 and 25 degrees C (p < 0.05); there was no effect when HMW was heated at 90 degrees C for 15 min. These results suggest the antioxidant effect of GSH on bovine OxyMb is dependent on heat-sensitive HMW cytosolic component(s).     

Determination of nitrite in cured meats by ion-exclusion chromatography with electrochemical detection

A rapid liquid chromatographic (LC) method was developed for a sensitive determination of nitrite in cured meats, using ion-exclusion chromatographic separation and electrochemical detection (IEC-EC). The current AOAC colorimetric method requires 2 h shaking in a steam bath to eliminate interference from reducing compounds such as ascorbic acid. In the present method, nitrite was analyzed in the presence of ascorbic acid without interference, and the extraction time was reduced to 1 min. The extracted nitrite was determined by ion chromatography using anion-exclusion/HS column and amperometric detector equipped with platinum or glassy carbon electrode operating at +1.0 V vs Ag/AgCl reference electrode. The detection limit was 1 ppb as NO2-. The recoveries of 50 ppm nitrite added to frankfurter and meat stick were 103 and 99.6%, respectively, with relative standard deviations less than 4%. The high speed, sensitivity, and selectivity make the new method a useful alternative to the AOAC colorimetric method.     

S-sulfonate determination and formation in meat products

A treatment with cyanide for the analysis of S-sulfonates in meat and meat derivatives, after a study of the effectiveness of this agent and that of dithiothreitol (DTT), is proposed. Once the protein-bound sulfite has been released, it is determined by HPLC ion exclusion with electrochemical detection. In the assay on the reproducibility of the method, standard deviations were 7.4, 9.2, and 11.4 for mean S-sulfonate values of 69, 107, and 130 microg of SO(2)/g, respectively. Mean recovery was 91.2% for different amounts (56, 111, and 223 microg of SO(2)/g) of S-sulfocysteine added. A study was made of the formation of S-sulfonates in model systems and in meat from different species-chicken and beef-with different fat contents. In the assays with meat, two different levels of sulfite addition were used: 600 and 1200 microg of SO(2)/g. From the assays carried out in model systems with sulfite and cystine it may be concluded that one factor limiting the interaction is the accessibility to disulfide groups. The proportion of S-sulfonates in sulfited meat remains relatively constant and does not seem to be governed by the meat component, the level of sulfite addition, or the fat content. However, the latter two factors are inversely correlated with the retention of sulfite in the foods analyzed.     

三种方法对细胞周期诱导效率的比较

比较两种处理方法对细胞周期调控的影响,本研究用饥饿法,胸腺嘧啶处理法和饥饿法结合胸腺嘧啶法诱导胎儿成纤维细胞进入G1/G0期。诱导两周后,收集3种方法来源的细胞用流式细胞技术、细胞活力检测、real-time PCR和核型分析检查细胞诱导效率、细胞活力、成纤维细胞特异基因的表达水平和核型情况。结果表明,实验组细胞G1/G0期比率显著高于对照组（p〈0.05）;实验组中,饥饿联合药物诱导法比饥饿法和药物法稍高,但未达显著水准。实验组与对照组细胞核型正常,均为40条染色体,性染色体为XX。细胞活力和显示,对照组比试验组稍高,但差异不显著（p〉0.05）;FGF4、FGF2及Sox2的基因表达水平在各组间差异不显著（p〉0.05）。这些结果表明,实验组3种方法对细胞特性没有显著影响,饥饿结合药物诱导可适当提高细胞诱导效率,为相关研究提供参考。     

Formation of N-nitroso-N-methylurea in various samples of smoked/dried fish, fish sauce, seafoods, and ethnic fermented/pickled vegetables following incubation with nitrite under acidic conditions

In continuation of our previous studies on N-nitroso-N-methylurea (NMU) formation in cured meats following incubation with nitrite at gastric pH, we extended the investigation to other foods mentioned in the title of this paper. The main objective was to determine whether these foods have the potential to form NMU at pH's that can be found in the human stomach. This was done by nitrosating an aliquot (5 g for fish sauce, 10 g for the others) of each with 7.25 microM to 1.59 mM levels of sodium nitrite for 2 h at room temperature at pH 0.8--1.5 and measuring the amounts of NMU formed. Of the samples tested, fish sauce formed 2--712 ng of NMU, followed in decreasing order by herring (<0.3--688 ng); dried anchovy, shrimp, and other fishes (<0.3--134 ng); crab and lobster paté (<0.3--342 ng); sardines (6--59 ng); oysters and mussels (11--31 ng); dried squid (3--47 ng); kimchi (7--107 ng); and Japanese pickled radish (<0.3--72 ng). Incorporation of 200-2000 ppm of ascorbic acid in the fish sauce and other foods, prior to nitrosation, appreciably inhibited such NMU formation. Although previous researchers in China reported NMU formation in nitrosated samples of fish sauce, this is the first reported formation of NMU upon nitrosation of the other foods mentioned above, and the first reported inhibition of such formation by added ascorbic acid.     

Comparison of chemiluminescent and AOAC methods for determining nitrite in commercial cured meat products

A chemiluminescent detector was used to measure nitrite in a variety of commercial cured meat products, using the AOAC sample preparation procedure, 24.041. A comparison of the NaNO2 values obtained by using sulfanilamide/N-(1-naphthyl)ethylenediamine (SAN/NED) reagent, sulfanilamide/1-naphthylamine reagent, and chemiluminescent detection (CLD) revealed no significant differences between the latter 2 detection methods. The AOAC SAN/NED reagent combination gave an average of 24.6% lower NaNO2 results than CLD. Examination of the sample preparation extraction and heating steps indicated that the procedure could not be made more rapid because of the need to destroy residual reductants and release "bound-complexed-reacted" nitrite from the meat samples.     

海绵铁缓解污水厌氧氨氧化反应器中硝酸盐积累的效果

该文旨在通过向厌氧氨氧化反应器中投加海绵铁来减轻厌氧SBR(sequencingbatchreactoractivatedsludge process)反应器中的硝酸盐积累,试验研究了海绵铁与硝酸盐和亚硝酸盐在静态条件下的反应。在静态条件下,部分硝酸盐和亚硝酸盐被海绵铁还原成了氨。对比动态试验表明投加海绵铁可以将SBR出水硝酸盐质量浓度控制在25～30 mg/L左右。相同条件下不投加海绵铁出水硝酸盐质量浓度不断累积,直至超过55 mg/L。这可能是由于铁将硝酸盐还原为亚硝酸盐并与厌氧氨氧化进行了耦合。采用高通量测序发现投加海绵铁的反应器中厌氧氨氧化菌在微生物群落中所占的比例(22.55%)约为不投加反应器(8.85%)的3倍,表明投加海绵铁有利于反应器中厌氧氨氧化(ANAMMOX)菌的生长和厌氧氨氧化反应器的启动。