乳酸脱氢酶在冷却牛肉肉色稳定性和高铁肌红蛋白还原中的作用机理研究

发布时间：2018-03-26 04:11

本文选题：乳酸脱氢酶　切入点：MetMbR活性　出处：《甘肃农业大学》2017年硕士论文

【摘要】：肉色是消费者判断肉新鲜程度和货架期的重要指标,冷却肉表面的褐变被消费者认为是品质劣变的表征。本实验以有氧储藏条件下的冷却牛肉为试验材料,探究了宰后冷藏过程中影响肉色稳定性的主要生化因子变化规律以及LDH活力与MRA活性表达的相关性,从稳定肌红蛋白氧化还原形式的生化机制入手,明确了乳酸脱氢酶和高铁肌红蛋白还原与肉色稳定性的关系,并结合体外孵化揭示乳酸脱氢酶在冷却牛肉肉色稳定及高铁肌红蛋白还原作用机理。结果如下:1.宰后冷藏期间三组冷却肉肉色L*和b*值都呈现出先增大后减小的变化趋势,在1~3 d内无明显变化。抑制组L*值和b*值相对初始值降低了2.26%和6.79%,而对照组和激活组L*值分别升高了3.47%和4.46%(P0.05),b*值分别降低了2.89%和0.74%。对照组、抑制组和激活组a*值在冷藏过程中相对初始值降低了17.5%、25.23%和2.35%(P0.05)。2.冷藏初期冷却肉中肌红蛋白主要以OMb的形式存在(约占56%),成熟7d后OMb降低至10.93%,MMb则升高至68.90%(P0.05),另外,肉的冷藏期间LDH活性、NADH的浓度、MetMbR的活力也显著下降,相比于初始状态降幅分别为45.96%27.81%和84.64%(P0.05),导致MMb大量积累。3.草氨酸钠抑制剂处理过的肌肉中LDH活性、NADH浓度显著降低(P0.05),而甜菜碱处理肌肉后LDH活性,NADH浓度及显著升高(P0.05),肉色更稳定。说明草氨酸钠在肌肉组织中能够有效的抑制乳酸脱氢酶的活性从而导致NADH浓度及MetMbR活性降低,影响MetMb还原因而降低肉色的稳定性,甜菜碱作用与之相反。4.SDS-PAGE电泳图谱(乳酸脱氢酶同工酶)分析发现,对照组的LDH1同种型表达显著高于处理组1,显著低于处理组2(P0.01),表明草氨酸钠在肌肉组织中能够有效抑制LDH-B活性,阻碍了LDH1蛋白表达,降低了NADH的生成速率。从而导致MetMbR活性降低,影响MetMb还原因而降低肉色的稳定性,甜菜碱作用与之相反。5.乳酸-LDH-NAD~+体系在缺乏牛肌肉提取物(LDH)、NAD~+或乳酸盐的反应混合物中,高铁肌红蛋白还原酶活力不明显。将L-乳酸盐为替换D-乳酸盐或者增加草酸酸钠到混合物中导致没有高铁肌红蛋白还原,增加混合物中牛肌肉提取物的量、NAD~+或乳酸盐浓度、适当提高乳酸盐 NAD~+ 肌肉提取物(LDH)系统的pH和反应温都会明显提高高铁肌红蛋白还原力(P0.05)。用乳酸钾代替混合物中的乳酸钠时高铁肌红蛋白还原力没有差异(P0.05)。离体实验验证了乳酸脱氢酶参与MetMb还原,同时是L-乳酸盐和NAD~+的相互作用。
[Abstract]:Meat color is an important index for consumers to judge the freshness and shelf life of meat. The browning of cooled meat surface is regarded by consumers as a sign of poor quality. The changes of main biochemical factors affecting the stability of meat color during postmortem refrigeration and the correlation between the activity of LDH and the expression of MRA activity were investigated. The biochemical mechanism of stabilizing the redox form of myoglobin was discussed. The relationship between the reduction of lactate dehydrogenase and high iron myoglobin and the stability of meat color was clarified. Combined with in vitro incubation, lactate dehydrogenase was found to be stable in the color of chilled beef and the mechanism of the reduction of high iron myoglobin. The results were as follows: 1. During the period of postmortem refrigeration, the values of L* and b* of the three groups of chilled meat showed a tendency of increasing first and then decreasing. The values of L * and B * decreased by 2.26% and 6.79%, while those of control group and activation group increased by 3.47% and 4.46%, respectively, and decreased by 2.89% and 0.74%, respectively. The relative initial values of inhibition group and activation group decreased by 17.525% and 2.35% respectively during cold storage. The myoglobin in the chilled meat was mainly in the form of OMb (about 56.5%). After 7 days of maturity, OMb decreased to 10.93% and increased to 68.90%, P0.05%. The activity of LDH activity nadh and MetMbR also decreased significantly during cold storage of meat. Compared with the initial state, the decreases were 45.96% and 84.64%, respectively, which resulted in a large accumulation of MMb in the muscle treated with sodium oxalate inhibitor. The concentration of LDH activity and nadh in the muscle treated with sodium oxalate inhibitor decreased significantly, while the concentration of nadh in muscle treated with betaine significantly increased the concentration of P0.05. The results show that sodium oxalate can effectively inhibit the activity of lactate dehydrogenase in muscle tissue, resulting in the decrease of NADH concentration and MetMbR activity. The effect of betaine on the stability of meat color was found by SDS-PAGE electrophoresis (lactate dehydrogenase isozyme) analysis. The expression of LDH1 isotype in the control group was significantly higher than that in the treatment group 1 and significantly lower than that in the treatment group 2 (P 0.01), which indicated that sodium oxalate could effectively inhibit the activity of LDH-B in muscle tissue, hinder the expression of LDH1 protein, and decrease the rate of NADH production, which resulted in the decrease of MetMbR activity. The effect of betaine on the stability of meat color was also affected by the effect of betaine. 5. Lactic acid-LDH-NAD ~ system was used in the reaction mixture of lack of bovine muscle extract (MetMb) or lactate. The activity of ferric myoglobin reductase was not obvious. When L- lactate was substituted for D- lactate or sodium oxalate was added to the mixture, there was no reduction of ferric myoglobin, and the amount of bovine muscle extract in the mixture was increased, and the concentration of NAD- or lactate in the mixture was increased. When the pH and reaction temperature of the lactate NAD ~ + muscle extract (LDH) system were properly increased, the reductive capacity of ferric myoglobin was significantly increased (P0.05). There was no difference in the reductive capacity of ferric myoglobin with potassium lactate instead of sodium lactate in the mixture (P0.05N). The results showed that lactate dehydrogenase was involved in MetMb reduction. It is also the interaction of L-lactate and NAD ~.
【学位授予单位】：甘肃农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TS251.52

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