低温贮藏中华管鞭虾肌肉品质及组织蛋白酶H活性变化

为探究不同贮藏条件下中华管鞭虾组织蛋白酶H活性及肌肉品质的变化情况,以中华管鞭虾为对象,分别在冷藏（4 ℃,0~6 d）和冻藏（?18 ℃,0~120 d）条件下,比较分析完整虾组和去头虾组肌肉pH、持水力、硬度、弹性、肌原纤维蛋白含量、水分含量及各亚细胞分级中组织蛋白酶H活性等指标的变化。结果表明,随着贮藏时间的延长,整虾和去头虾肌肉pH均不断升高,但去头虾组pH升高幅度略低于完整虾组;虾肉中肌原纤维蛋白含量随贮藏时间延长而逐渐降低,完整虾组和去头虾组在冷藏至6 d时分别降低了38.32%和30.88%,在冻藏至120 d时分别减低了61.67%和52.09%;在整个冻藏过程中,两组虾的硬度和弹性均出现先上升后下降的趋势,而冷藏贮藏过程中则始终呈现下降趋势;组织蛋白酶H在不同亚细胞分级中活性变化不同,冻藏条件下两组虾组织蛋白酶H酶活整体低于冷藏条件下酶活,且完整虾组组织蛋白酶H活性整体高于去头虾组。综上,在低温贮藏过程中,相比于完整虾形式,以去头虾的贮藏方式更有利于对虾肌肉品质的保障,且组织蛋白酶H活性受贮藏温度影响较大。研究结果为不同贮藏方式下中华管鞭虾理化特性及组织蛋白酶活性变化提供了理论基础。     

罗氏沼虾冻藏过程中质构与理化特性研究

以剪切力、肌原纤维小片化指数、肌原纤维表观直径、TCA-溶解肽、总蛋白酶活和微生物为指标,比较分析了整只罗氏沼虾冻品和去头罗氏沼虾冻品品质的差异,并探讨了其质构劣化的机理。结果表明:随着冻藏时间的延长,虾肉的剪切力均呈下降趋势,整只罗氏沼虾冻品的下降速度明显高于去头罗氏沼虾冻品,冻藏16周后剪切力分别下降了36.81%,25.40%;随着冻藏时间延长整只罗氏沼虾冻品的肌原纤维小片化指数增加,肌原纤维表观直径明显下降,TCA-溶解肽含量增加,说明罗氏沼虾冻藏期间肌原纤维内部结构被破坏,骨架蛋白出现了降解,整只罗氏沼虾冻品在贮藏过程中蛋白发生了降解;虾肉中蛋白酶活性呈上升趋势而虾头中蛋白酶活性则略微下降,说明虾头内的蛋白酶可能向虾肉发生了迁移。菌落总数则随着冻藏时间的延长而下降。相关性分析表明:罗氏沼虾在冻藏期间蛋白变化的各项指标与蛋白酶活性变化和质构劣化之间存在显著相关性(P0.01)。     

冷藏三疣梭子蟹腹部和螯足肌肉品质特性变化

目的：探究冷藏过程中三疣梭子蟹腹部和螯足肌肉品质特性变化情况。方法：以三疣梭子蟹为对象，在4℃冷藏过程中，对蟹腹部和螯足肌肉进行定量描述分析，同时测定肌肉持水力、水分含量、水分活度、pH、挥发性盐基氮、三甲胺、TCA-可溶性肽、肌原纤维蛋白含量及其小片化指数等理化指标。结果：随着冷藏时间延长，三疣梭子蟹腹部和螯足肌肉特性出现相似的变化趋势，其中肌肉品质感官特性、持水力、水分含量和水分活度均呈下降趋势；肌肉pH呈先下降后上升趋势，腹部肌肉pH在冷藏第5 d时为7.7，而螯足肌肉pH在冷藏第4 d时已达7.99；挥发性盐基氮、三甲胺、TCA-可溶性肽含量和肌原纤维小片化指数则呈不断上升趋势；肌原纤维蛋白含量呈显著下降趋势(P<0.05)，腹部和螯足肌肉在冷藏5 d后分别下降了38.11%和49.51%。在整个冷藏周期中，三疣梭子蟹腹部肌肉品质特性均显著优于螯足肌肉。结论：随着贮藏时间延长，三疣梭子蟹肌肉品质逐渐发生劣变，其腹部肌肉品质特性相比螯足部分更加稳定。本研究为三疣梭子蟹冷藏品质劣变及调控技术提供理论参考。     

凡纳滨对虾内源蛋白酶对肌原纤维蛋白的降解作用

为确定引起凡纳滨对虾自溶的主要酶类及分布部位,本文测定了凡纳滨对虾各内源蛋白酶的活性,分析主要内源蛋白酶贮藏期间的变化,并比较主要内源蛋白酶对虾肌原纤维蛋白的降解作用。结果显示,虾头蛋白酶活达90.17U/mg,是虾肉的300倍。虾头中的胰蛋白酶活性最高,达13.59U/mg。全虾贮藏4d后虾肉总酶活和胰蛋白酶活增加,于第8d虾肉第一腹节总酶活增至1.66U/mg,胰蛋白酰胺酶和酯酶活分别达到0.97U/mg和1.84U/mg。SDSPAGE结果显示,随贮藏温度和时间的增大,内源酶对肌原纤维蛋白的肌球蛋白重链(MHC)和肌动蛋白(Actin)降解程度越大,其中虾头中胰蛋白酶对肌原纤维蛋白的降解作用最强。因此凡纳滨对虾胰蛋白酶很可能是肌肉软化的主要作用酶。     

虾夷扇贝肌肉丝氨酸蛋白酶的研究

为了探究虾夷扇贝肌肉冷藏过程中内源蛋白酶对其质构的影响,本文对虾夷扇贝肌肉在4 ℃下冷藏7 d过程中肌肉质构及蛋白变化进行测定,结果表明,整个冷藏过程中扇贝肌肉硬度、弹性、咀嚼性和粘性整体呈下降趋势。SDS-PAGE分析显示肌肉蛋白在第3 d开始出现明显降解。在内源酶活力方面,丝氨酸蛋白酶（Serine proteinase,SP）活力在冷藏2 d后开始急剧下降。通过硫酸铵盐析、离子交换、凝胶过滤和疏水柱层析等从虾夷扇贝肌肉中获得高度纯化的丝氨酸蛋白酶并对其酶学性质进行研究。SDS-PAGE和明胶酶谱结果表明,SP在天然状态下主要以分子量约为52 kDa的二聚体形式存在。SP的最适pH为9.0,最适温度为37 ℃。SP特异性水解羧基侧P₁位含有精氨酸或赖氨酸残基的底物。丝氨酸蛋白酶特异性抑制剂PMSF、Leupeptin、Pefabloc SC、Benzamidine分别能抑制其97%、98%、90%和85%的酶活力;金属离子Fe2+、Zn2+、Cu2+也能明显抑制SP的酶活力。37 ℃下SP可有效降解扇贝肌原纤维蛋白,为揭示SP对扇贝肌肉蛋白的品质影响提供参考。     

UV-C辐照对凡纳滨对虾虾头主要内源酶酶学性质的影响

以凡纳滨对虾虾头为研究对象,探究UV-C辐照对其主要内源酶的影响。用功率为30 W的UV-C紫外灯以20 cm高度、辐照20 min,处理虾头均浆液,检测内源酶酶活,分析比较辐照前后温度、pH对其内源酶酶活的影响规律。从虾头中检测出酸性及碱性蛋白酶、脂肪酶、几丁质酶和多酚氧化酶等酶活性; UV-C辐照前后,脂肪酶、几丁质酶的最适pH有所变化,脂肪酶最适pH由10变为6～10,几丁质酶最适pH由5变为6左右;各主要内源酶最适温度无明显变化; UV-C辐照后,在最适条件下酸性蛋白酶酶活升高23. 99%,碱性蛋白酶升高37. 49%,脂肪酶升高21. 37%,几丁质酶升高37. 70%,多酚氧化酶升高46. 52%。结果显示,UV-C辐照对凡纳滨对虾虾头酸性及碱性蛋白酶、脂肪酶、几丁质酶和多酚氧化酶等主要内源酶均有激活作用。     

罗非鱼冷藏期间内源蛋白酶和热休克蛋白70对肌原纤维解离的影响

为研究罗非鱼冷藏期间内源蛋白酶和热休克蛋白70与肌原纤维解离的关系，实验测定了罗非鱼肉在4℃真空包装冷藏48 h内肌原纤维小片化指数、肌原纤维解离、内源蛋白酶活性和热休克蛋白70含量的变化，并对其进行相关性分析。结果表明鱼肉冷藏期间肌原纤维小片化指数变化整体呈增加的趋势，最终为初始值的3.52倍；解离的肌球蛋白重链和肌动蛋白的含量呈升高的趋势，最终分别较初始值提高了70%和65%;pH变化为先减少后略微回升，变化范围为6.5～7.0;组织蛋白酶B、L活性持续增加，组织蛋白酶D和钙激活蛋白酶活性呈先增加后减少的趋势，且分别在8、12 h到达最大值；热休克蛋白70含量为先增加后减少，在4 h时达到最大值，最终相比初始值下降了22.27%;肌原纤维小片化指数与组织蛋白酶B、L活性呈极显著正相关(P<0.01),与肌原纤维解离的肌球蛋白重链和肌动蛋白含量均呈极显著正相关(P<0.01),与热休克蛋白70含量呈显著负相关(P<0.05);热休克蛋白70含量与与组织蛋白酶L呈显著负相关(P<0.05)。组织蛋白酶B、L是影响肌原纤维完整性的关键蛋白酶，热休克蛋白70能与肌原...     

包装方式对冷藏过程中牦牛肉蛋白质生化特性的影响

为研究不同包装方式对冷藏过程中牦牛肉蛋白质生化特性的影响。取牦牛背最长肌,采用真空和非真空2种包装方式,在0~4℃下分别冷藏0、2、4、6、8、10、12 d,分析其在冷藏过程中蛋白质生化特性变化。结果显示,随着冷藏时间的延长,真空和非真空包装的牦牛肉肌原纤维蛋白和肌浆蛋白均表现出不同程度的降解,但2种包装方式间差异不显著。真空组和非真空组肌原纤维蛋白小片化指数和表面疏水性均呈显著(P0.05)上升趋势,肌动球蛋白盐溶性、巯基含量、Ca~(2+)-ATPase酶活性则均呈显著(P0.05)下降趋势。2种包装方式下的肌动球蛋白盐溶性、Ca~(2+)-ATPase酶活性和肌原纤维蛋白表面疏水性、肌原纤维蛋白小片化指数在0~8 d存在显著(P0.05)差异,10 d以后差异不显著。研究表明,牦牛肉在冷藏期间蛋白质发生了显著降解和变性,含氧包装促进了肌原纤维蛋白小片化和表面疏水性的增加,加重了肌动球蛋白盐溶性、巯基含量、Ca~(2+)-ATPase酶活性的降低。     

凡纳滨对虾虾头协同水解工艺的响应面优化

利用风味蛋白酶协同虾头内源酶水解凡纳滨对虾虾头,制备水解度高、苦味值低的新型虾风味食品基料.以苦味感官为指标,采用响应面法研究pH、温度、底物浓度对产物水解度和苦味的影响.在pH值6.92,温度52.5 ℃,底物浓度为22.25 g/100 mL的条件下,每克虾头添加144 U风味蛋白酶水解3 h,与自溶水解液相比,水解度增加了6.4%,苦味值降低了48.5%,游离氨基酸总量增加了16.8%,疏水性氨基酸总量增加了4.5%.风味蛋白酶可以协同虾头内源酶水解凡纳滨对虾虾头,制备出水解度高、苦味值低的水解液.     

真空低温慢煮加工对中华管鞭虾肌肉品质的影响

探究真空低温慢煮（sous vide,SV）加工对中华管鞭虾肌肉品质特性的影响。以去头去壳（虾仁组）和去头带壳（带壳虾组）中华管鞭虾为对象,经真空密封后在70℃水浴下分别加热5、8、10和12 min(SV加工),以传统100℃沸水加热2 min为对照（CK）,测定加工后虾仁肌肉感官特性、色差、水分含量、蒸煮损失率、质构特性、蛋白质含量及提取率、肌原纤维蛋白总巯基含量、Ca2+-ATPase活性和组织微观结构等指标。结果发现,随着煮制时间延长,SV加工中华管鞭虾肌肉L*值和b*值不断上升,a*值先上升后下降,肌肉中水分含量逐渐下降,而肌肉蒸煮损失率、硬度和咀嚼性不断升高。煮制加热过程中,肌肉中盐溶性蛋白质、肌原纤维蛋白总巯基含量、Ca2+-ATPase活性和肌原纤维蛋白溶解度不断下降。HE染色和SEM结果表明,煮制过程中中华管鞭虾肌纤维受热逐渐发生变性,肌纤维间距增大、肌丝断裂。与CK处理相比,SV处理可增加虾感官评分和水分含量,有效减少蒸煮损失率、硬度和咀嚼性,更好保留水溶性蛋白质、盐溶性蛋白质、肌原纤维蛋白总巯基含量、Ca2+-ATPase活性等,并减少不溶性蛋白质的形成。此外,与...     

Retardation of post-mortem changes of freshwater prawn (Macrobrachium rosenbergii) stored in ice by legume seed extracts

Meat quality of freshwater prawn (Macrobrachium rosenbergii) treated with soybean and bambara groundnut extracts at different concentrations was monitored during 10 days of iced storage. During storage, the control sample (without treatment) had a higher pH, TCA-soluble peptide content, heat soluble collagen content, proteolytic activities and psychrophilic bacterial count than did samples treated with soybean and bambara groundnut extracts. Conversely, shear force value and likeness scores of the control sample decreased (p<0.05), more likely associated with softening of muscle. The decrease in myosin heavy chain in the control sample was found after 6 days of storage. However, no changes in protein patterns of samples treated with soybean extracts at 2.5 mg/mL were found after 10 days of storage. Therefore, the injections of legume seed extracts, especially soybean extract, at a sufficient concentration, could be a means to retard muscle softening and maintain the qualities of freshwater prawn during iced storage.     

Characterisation of proteolytic enzymes from muscle and hepatopancreas of fresh water prawn (Macrobrachium rosenbergii)

BACKGROUND: Fresh water prawn in Thailand is widely consumed due to its delicacy. During postmortem handling and storage, prawn meat becomes soft and mushy, probably as a result of indigenous proteases. Therefore, an understanding of prawn proteases associated with the degradation of muscle proteins from fresh water prawn could pave the way for prevention of such a phenomenon during extended storage. RESULTS: Proteolytic enzymes in the crude extract (CE) from muscle and hepatopancreas of fresh water prawn (Macrobrachium rosenbergii) were characterised. CE from muscle exhibited the highest hydrolytic activities towards haemoglobin at pH 5 and 50 °C, while that from hepatopancreas had the highest activity on casein at pH 7 and 60 °C. Based on inhibitor study, cysteine protease and serine protease were dominant in CE from muscle and hepatopancreas, respectively. CE from muscle rarely hydrolysed natural actomyosin (NAM), but could not degrade pepsin‐soluble collagen (PSC). Conversely, NAM and PSC were susceptible to hydrolysis by CE from hepatopancreas as evidenced by the marked decreases in band intensity. Activity staining using haemoglobin, casein and gelatin as substrates revealed that no proteolytic or gelatinolytic activity was observed in CE from prawn muscle, while CE from hepatopancreas exhibited pronounced hydrolytic activities towards all substrates. CE from muscle showed calpain and cathepsin L activities but CE from hepatopancreas mainly exhibited tryptic and chymotryptic activities. CONCLUSION: Serine proteases, mainly trypsin‐like or chymotrypsin‐like, from hepatopancreas were probably responsible for the softening of prawn meat during postmortem storage via the degradation of both muscle and connective tissues. Copyright © 2010 Society of Chemical Industry     

Investigation of the activity of cathepsin B in red shrimp (Solenocera crassicornis) and its relation to the quality of muscle proteins during chilled and frozen storage

Cathepsin B is a cysteine protease that has important effects on the quality of muscle products. In this study, the changes of cathepsin B activity and its relation to muscle proteins were investigated in intact and beheaded shrimp during chilled and frozen storage. The obtained results indicated that the water holding capacity (WHC), shear force, hardness, and myofibrillar protein (MP) content all significantly decreased in both the intact and beheaded shrimp samples with increasing storage period (p < 0.05). Specifically, beheading shrimp exhibited much more stable characteristics than intact shrimp samples during both chilled and frozen storage. The enzyme activity results suggested that cold temperature and storage induced the release of cathepsin B from the lysosomes to the mitochondria, sarcoplasm, and myofibrils in the muscle tissues. Furthermore, SDS-PAGE and transmission electron microscopy (TEM) analysis revealed that beheading the shrimp greatly inhibited the dissociation of shrimp muscle proteins during storage. The current findings suggest that cathepsin B located in the head of shrimp was likely transferred to the muscle through the first abdominal segment during storage, accelerating the dissociation of the muscle proteins. Therefore, beheading the shrimp was conducive to prolonging the shelf-life of stored shrimp products.     

Collagenolytic serine protease in fresh water prawn (Macrobrachiumrosenbergii): Characteristics and its impact on muscle during iced storage

Proteolytic activity of crude protease extract (CPE) from the hepatopancreas of fresh water prawn (Macrobrachium rosenbergii) was studied. Optimal activity of CPE was found at pH 7 and 60 °C when casein was used as a substrate. The activity was strongly inhibited by 10 mM N-ρ-tosyl-L-lysine chloromethylketone (TLCK), suggesting that trypsin-like protease was dominant. CPE also showed the collagenolytic activity toward pepsin soluble collagen extracted from prawn muscle. During extended iced storage of 4 days, proteolytic and trypsin activities were found in the first segment of prawn abdomen. These activities were detected in the second segment after 4 days of storage. Heat soluble collagen content was continuously increased during the storage. Nevertheless, no changes in proteolytic activity and heat soluble collagen content were obtained in the abdomen of prawn with the removal of hepatopancreas. Therefore, the release of trypsin-like collagenase from hepatopancreas was most likely responsible for the softening of prawn meat during iced storage.     

Changes in the ultrastructure and texture of prawn muscle (Macrobrachuim rosenbergii) during cold storage

Changes in the ultrastructure and proteolytic activity of prawn muscle were determined during storage at 5 °C, in order to better understand changes in physical and sensory texture measurements. Progressive deterioration of myofibril structure was observed during refrigeration of prawn for 14 d. The loss of density and order in Z-line alignment was first detected after 3 d of storage. Progressive disruption of Z-line, I-bands and M-lines was observed after 4-6 d of storage. Muscle degradation included pronounced disruption of the mitochondria as revealed by swollen cristae, loss of cristae material, and membrane breakage. Along with ultrastructural changes, decreased shear force values and mean textures scores were measured. An initial shear force value of 18.21 N/g decreased to 14.50, 12.46, and 10.79 N/g on days 3, 6, and 14, respectively. Mean texture scores indicated that prawn muscle maintained firm texture during 0-3 d of storage, and became soft during 4-6 d of storage. After 6 d of storage, the prawn texture was very soft. Increased deterioration of the muscle ultrastructure coincided with the increase of proteolytic microorganisms and salt soluble muscle proteins found by Sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It is concluded that the weakening of the ultrastructure is related to proteolytic activity and results in a more soft texture in prawns.     

Endogenous proteases in giant freshwater prawn (Macrobrachium rosenbergii): changes and its impacts on texture deterioration during frozen storage

Effects of endogenous enzymes on texture deterioration of Macrobrachium rosenbergii during frozen storage (−18 °C) were investigated. The prawn was pretreated with iodoacetic acid (IAA) solution to inhibit cysteine proteases and/or phenylmethylsulfonyl fluoride (PMSF) solution to inhibit trypsin before frozen storage, and sterile distilled water was used as the control. After 16 weeks of cryopreservation, the shear force of the groups treated with enzyme inhibitors was significantly higher than that of sterile distilled water-treated group (control; P < 0.05), while significantly fewer TCA-dissolved peptides, α-amino nitrogen and Myofibril fragmentation index (MFI) value were observed in all inhibitor-treated groups (P < 0.05). Compared with PMSF treatment, IAA or IAA + PMSF treatment effectively alleviated softening and remarkably reduced the accumulation of TCA-soluble peptides and AAN (α-amino nitrogen) during frozen storage, with a lower MFI value and a higher sodium dodecyl sulphate-polyacrylamide gel electrophoresis band intensity. Therefore, endogenous cysteine proteases in muscle play a crucial part in texture deterioration of giant freshwater prawn during cryopreservation.     

Effect of protein oxidation and degradation on texture deterioration of ready-to-eat shrimps during storage

The impact of protein oxidation and degradation on texture deterioration of ready-to-eat (RTE) shrimps during storage was investigated. The deterioration in texture during storage was manifested by decreased instrumental hardness, elasticity, chewiness, and recoverability. The occurrence of protein oxidation was revealed by a significant increase in the contents of free radicals and carbonyls. The increases in trichloroacetic acid-soluble peptide (TCA-soluble peptide) content and myofibril fragmentation index (MFI) were also observed, suggesting the degradation of protein. Pearson correlation analysis showed that the decreased instrumental texture parameters were negatively correlated with the increased carbonyl content, TCA-soluble peptide, MFI, porosity, and pore size as well as the decreased water-holding capacity (WHC), thus, it was hypothesized that protein oxidation and degradation were responsible for changes in the microstructure and reduction of WHC, which ultimately resulted in texture deterioration of RTE shrimps.     

分散固相萃取-GC-MS法测定海产品中9种有机锡化合物

本实验建立了一种同时检测海产品中9种有机锡化合物的新方法。样品依次经过超声波提取,冷冻脱脂,N-丙基乙二胺（PSA）分散固相萃取、四乙基硼酸钠衍生、石墨化炭黑（GCB）分散固相萃取,最后采用气相色谱-质谱法（GC-MS）进行测定。在优化条件下,9种有机锡化合物的线性范围为10~1000 μg/L,相关系数为0.9952~0.9988,检出限（3S/N）为0.4~1.0 μg/kg dw （干重）,以Sn计。在2.00、20.0和200 μg/kg 3个添加水平的回收率为81.2%~115.0%,相对标准偏差为4.1%~7.5%（n=5）。对龙头鱼、梅童鱼、小黄鱼、大黄鱼、带鱼、鲳鱼、海鳗、哈氏仿对虾、口虾蛄、梭子蟹、缢蛏、牡蛎等海产品中9种有机锡化合物进行检测,结果表明梅童鱼、小黄鱼、大黄鱼、海鳗中均未检出9种有机锡化合物。在龙头鱼、带鱼、鲳鱼、哈氏仿对虾、口虾蛄、梭子蟹、缢蛏、牡蛎中9种有机锡化合物总量为ND~162.0 μg/kg ww （湿重）,以Sn计。该方法简捷高效,基体干扰小,可用于海产品样品中的9种有机锡化合物的同时检测。     

Autolysis of Pacific white shrimp (Litopenaeus vannamei) meat: characterization and the effects of protein additives

ABSTRACT:  Autolytic activity of Pacific white shrimp ( Litopenaeus vannamei ) mince in the absence and in the presence of 2.5%NaCl was investigated. Pacific white shrimp mince exhibited the maximum autolytic activity at 35 and 40 °C in the absence and in the presence of 2.5%NaCl, respectively, as evidenced by the highest TCA-soluble peptide content and the greatest disappearance of myosin heavy chain (MHC). The autolysis was more pronounced in the acidic pH values, followed by alkaline pH ranges. Pepstatin A showed the highest inhibition toward autolysis in the acidic condition, revealing that aspartic proteinase was dominant in shrimp muscle. Nevertheless, soybean trypsin inhibitor effectively inhibited the autolysis at neutral and alkaline pH values, suggesting that serine proteinase was present in shrimp mince but contributed to autolysis at a lower extent in shrimp meat. Autolysis in shrimp meat could be inhibited partially by all protein additives, including bovine plasma protein (BPP), egg white (EW), and whey protein concentrate (WPC). The inhibition of autolysis increased when the level of protein additives increased with the concomitant increase in band intensity of MHC retained. WPC and BPP in the range of 2% to 3% exhibited the highest inhibition toward autolysis of shrimp mince.