【作者】 孙月梅；

【导师】 江连洲；

【作者基本信息】 东北农业大学 ， 农产品加工与贮藏， 2008， 硕士

【摘要】 大豆活性肽由于其高效的营养价值及功能活性,是目前营养界研究得比较多的一个热门课题,而且对这方面的研究也具有很实际的经济和利用价值。本试验通过比较六种不同的蛋白酶对大豆分离蛋白的水解效果,筛选出最适合生产抗氧化肽的Alcalase碱性蛋白酶,用正交实验法确定了其水解大豆分离蛋白的最佳工艺条件为:底物浓度7.0%、酶用量9000U/g底物、温度52.5℃、pH值8.5、反应时间5h。在上述条件下,蛋白质水解度为24.28%、氮溶指数95.7%。蛋白质含量43.5mg/mL,蛋白回收率71.2%。分别从羟基自由基（OH·）清除能力、超氧阴离子自由基（O₂^-·）清除能力、抑制脂质过氧化（ROO·）能力这三方面研究水解产物的抗氧化活性,水解5h的产物OH·抑制率为36.43%、清除O₂^-·活性为317.73U/g、ROO·抑制率为46.24%。采用超滤法对得到的水解产物分离,用截留分子量5000D、2000D的超滤膜将水解产物分成分子量大于5000D、5000D²000D和小于2000D三种组分,然后比较这三种组分的抗氧化活性。结果显示,分子量小于2000D的组分抗氧化活性最强。因此试验选择截流分子量为2000D的超滤膜对大豆分离蛋白的水解产物进行分离纯化。在单因素试验的基础上,以膜渗透通量为响应值设计了4因素（压力、pH值、温度、时间）3水平的响应面实验,通过响应面实验得到最佳超滤条件为:压力0.24MPa、pH值7.47、温度44.8℃、时间109.6min。在此条件下平均膜渗透通量为28.62L/（m2h）。超滤后其抗氧化活性得到了提高,清除O₂^-·活性由超滤前317.73U/g升高至超滤后387.9U/g。得到的活性较高的大豆抗氧化肽,用微胶囊化的方法对其活性进行保护。试验采用了复凝聚法和乳化-凝胶法制备大豆抗氧化肽微球。复凝聚法的最佳制备工艺为:壳聚糖浓度0.8%,阿拉伯胶与芯材肽的比8:1,成囊温度45℃,成囊pH4.75。得到的微囊包埋率为64.0%,载药量11.78%。乳化-凝胶法制备微球最佳工艺是:2.5%海藻酸钠,8.0%的氯化钙,乳化速度8000rpm,乳化剂浓度8.0%。得到的抗氧化肽微胶囊包埋率73.65%,载药量13.2%。体外试验证明:两种微球在胃液中的释放率均较低,在肠液中可继续释放,达到了缓释的目的。包埋后的微囊抗氧化活性与未包埋时活性比较,只有少量损失,说明微胶囊化使抗氧化肽的活性得到了保护。更多还原

【Abstract】 The research on bioactive peptide is a popular subject in nutrition nowdays because it has high nutritional value, functional activities and economic value .This experiment contrasted the hydrolytic effect of SPI among the six different proteolytic ferments and choosed the Alcalase basic protein which can produce the best antioxygen peptide, the optimization technology condition of hydrolytic SPI was determined by orthogonality experimentation: concentration of substrate 7.0%, the dosis of enzyme 9000U/g substrate, temperature 52.5℃, pH 8.5,the reaction time 5h..In the optimum condition,the degree of hydrolysis of protein was 24.28% ,NSI 95.7% ,the protein content was 43.5mg/mL and the recovery rate of protein was 71.2%. The antioxygen activity of hydrolysis product was researched from the capability of cleaning OH·, the capability of cleaning O₂^-·and the capability of inhibiting ROO·.After hydrolyzing of 5h ,the OH·inhibition ratio of the production was 36.43%, the activity of cleaning O₂^-·of the production was 317.73U/g, the ROO·inhibition ratio of the production was 46.24%.The hydrolysis products abstracted by the ultrafiltration were classified into three parts: above2000D,between 2000D and 5000D ,below 2000D ,which were cut off by 5000 D and 2000D ultrafiltration membrane .Then comparing the antioxdative activity of the three compositions,the result showed that the antioxidative peptide of the composition of which molecular weight was less than 2000D was the most powerful. So 2000D ultrafiltration membrane was choosed in the abstraction. Above monofactorial found, the response experiment of 4 factors （pressure, pH, temperature, time） and 3 levels was designed by the response value of the film percolation flux. The best ultrafiltrate condition gained by the response experimentation was that : pressure 0.24MPa ,pH 6.47, temperature 44.8℃, time 109.6min. Under the optimal condition ,the membrance flux was 28.62 L/（m2h）. The antioxdative activity had enhanced after ultrafiltration, the activity of cleaning O₂^-·increased from 317.73U/g to 387.9U/g.The activity of antioxygen bean peptide that was gained and has higher activity was protected by microencapsulation. Microballoons of bean antioxygen peptide were prepared by complex coacervation and multi-phase emulsion in this experiment. The optimal technology condition of complex coacervation : concentration of chitosan 0.8%,the ratio of acacia and peptide of core material 8:1, temperature of sacculation45℃, pH 4.75. The ratio of embedment of microcapsule gained was 64.0%, dosage was 11.78%.The best technology condition of multi-phase emulsion: Sodium Alginate 2.5%, Calcium chloride 8.0%, the velocity of emulsification 8000rpm, concentration of emulsifier 8.0%, The ratio of embedment of microcapsule of soybean antioxidative peptide gained was 73.65%, dosage was 13.2%.The vitro test certified: the release rate of two kinds of microballoons was both low in the gastric fluid. These microballoons continue to release for the purpose of delayed release in the intestinal juice. Comparing the activity of antioxygen embedment with not embedment, embedded activity had a little loss. The test showed that the activity of antioxygen peptide was protected by microencapsulation..更多还原