小麦蛋白低苦味肽的制备及其脱苦机理研究

发布时间：2018-03-06 00:05

  本文选题：小麦面筋蛋白　切入点：酶法水解　出处：《江南大学》2017年博士论文　论文类型：学位论文

【摘要】：小麦面筋蛋白作为小麦淀粉加工过程中的副产物,主要用于面制品和饲料工业,是一种物美价廉、食用安全的植物蛋白。小麦面筋蛋白复杂的分子组成和结构特性,且含有高比例的疏水性氨基酸,造成该蛋白的水溶性较差,极大地限制了其在食品工业中的应用。采用蛋白酶酶解小麦面筋蛋白制备生物活性肽的研究与开发,将有助于小麦产业链向高附加值产品延伸。但传统酶解工艺不够成熟,尚存在苦味较重、小肽含量较低等问题,制约了产品的开发利用。本文以小麦面筋蛋白为原料,围绕酶解技术在制备低苦味肽粉中的应用及其脱苦原理进行研究。首先分析了小麦面筋蛋白酶解产物的苦味特性,设计内切酶和端解酶连续水解,制备小麦面筋蛋白低苦味肽粉,并采用脱酰胺改性改善小麦面筋蛋白酶解产物的风味。为针对性优化小麦面筋蛋白-Proteax酶水解体系,探讨了小麦面筋蛋白酶解产物中苦味肽的释放机制,最终确定了分步酶解小麦面筋蛋白制备低苦味小肽的工艺。内切酶和端解酶连续水解,比单一的内切酶和端解酶有更强的水解能力,因此连续水解可以更有效的制备小肽。采用中性蛋白酶、木瓜蛋白酶、风味蛋白酶、碱性蛋白酶、胰蛋白酶、复合蛋白酶和Proteax酶酶解小麦面筋蛋白,通过滋味稀释分析评估单酶酶解产物的苦味强度,构建内切酶和端解酶连续水解方案。对比不同方案中酶解产物的肽氮含量、氨基酸组成、分子量分布和氨基酸序列对感官特性和电子舌分析的影响。研究结果表明连续水解制备的酶解产物,在苦味肽含量上的差异不大。氨基酸序列和分子量分布影响多肽的苦味阈值,决定酶解产物的苦味强度。与其它方案制备的酶解产物相比,Pro-m酶解产物具有最高的肽氮含量(61.54%)、小肽含量(53.26%)和最低的苦味值(1.33),并且其可溶性氮含量也高达82.82%。因此,确定以小麦面筋蛋白-Proteax酶水解体系为基本框架,开展后续工作。采用Glutaminase SD-C100S、Glutaminase和盐酸(HCl)对Pro-m酶解产物进行脱酰胺改性,促使酶解产物中谷氨酰胺转化为谷氨酸,发挥鲜味肽和游离谷氨酸增鲜抑苦的特性,改善酶解液的风味。对比分析脱酰胺改性酶解产物的风味特征、分子量分布和氨基酸组成,探讨脱酰胺改性酶解产物中呈鲜物质的主体成分以及其对苦味的抑制效果。研究结果表明,脱酰胺度增大,鲜味物质的含量上升,引起酶解产物鲜味增强、苦味降低。谷氨酸钠亲水性强,比鲜味肽有更强的唾液溶解性,能够更好的抑制苦味信号的传导。鲜味肽比谷氨酸钠有更久的受体作用时间,增强了鲜味的持续时间,会表现出一种更为柔和、协调的浓厚感。为针对性优化小麦面筋蛋白-Proteax酶水解体系,研究了小麦面筋蛋白酶解产物中苦味肽的释放机制。采用异丁醇萃取苦味释放特征发生明显变化的酶解产物,有效萃取酶解产物中苦味极强的苦味肽。系统地阐述了Proteax酶水解过程中苦味肽氨基酸序列、分子量分布、氨基酸组成的变化规律,并利用偏最小二乘回归法分析了异丁醇萃取物、感官属性、分子量分布与氨基酸组成之间的相关性。研究表明,相对于氨基酸序列的变化,苦味肽的氨基酸组成、分子量分布对苦味肽的苦味强度有一个更显著的影响。酶解反应在0-120 min时,苦味氨基酸在苦味肽(分子量处于180-500 Da、500-1000 Da和1000-3000 Da区间)中所占的比例逐渐增大。酶解反应在120-300 min时,苦味肽中苦味氨基酸的含量不再提高。萃取物中高分子量苦味肽的含量降低,尤其是苦味最强,相对分子量处于500-1000 Da区间的苦味肽。因此,小麦面筋蛋白-Proteax酶水解体系中苦味肽的苦味强度呈先升高后降低的趋势。采用Proteax酶、碱性蛋白酶、复合蛋白酶和胰蛋白酶分别水解小麦面筋蛋白,生成可溶性酶解产物和水不溶性聚集体。以水不溶性聚集体为原料,重复上述操作,收集聚集体酶解产物。对可溶性酶解产物和聚集体酶解产物进行苦味评定,发现采用同一种蛋白酶,在水解度大小一致的前提下,聚集体酶解产物比可溶性酶解产物产生更高的苦味强度。根据上述结论提出分步酶解技术,优化了小麦面筋蛋白-Proteax酶水解体系。即采用Proteax酶水解小麦面筋蛋白35 min,离心去除沉淀,加入Glutaminase SD-C100S,采用Design-Expert 8.0.6软件优化复合酶法水解工艺。酶解工艺以低苦味值为目标,复合酶法的最佳工艺参数为:酶解时间280.79 min,酶解温度53.09°C,pH6.94,苦味预测值为0.41。对结果进行验证,调整酶解参数为酶解时间280 min,酶解温度53°C,pH6.9。制备出苦味值仅为0.43的小麦面筋蛋白低苦味肽粉。
[Abstract]:Wheat gluten as a by-product of wheat starch processing, mainly used for flour products and feed industry, is a kind of high quality and inexpensive, safe to eat the vegetable protein. Molecular wheat gluten protein complex composition and structure characteristics of hydrophobic amino acids and contains a high proportion of the protein, resulting in poor water solubility, which greatly limits its application in the food industry. The research and development of protease preparation of bioactive peptide of wheat gluten system solutions, will contribute to the wheat industry chain to high value-added product extension. But the traditional enzymolysis technology is not mature, there are still bitter heavy, small peptide content is low, restricted the development and utilization of products. In this paper, wheat gluten was used as raw material, around the low bitter peptide powder in the system and the principle of debittering enzyme technology. First analysis of wheat gluten hydrolysate. Bitter taste characteristics, design enzymes and exopeptidase continuous hydrolysis, preparation of wheat gluten powder with low bitter peptide, deamidation of wheat gluten hydrolysate flavor. For the optimization of enzymatic hydrolysis of wheat gluten -Proteax system, discusses the release mechanism of bitter peptide products of wheat gluten protein by enzymatic hydrolysis finally, determine the process by hydrolysis of Wheat gluten to produce low bitter peptide. Exopeptidase enzymes and continuous hydrolysis, enzyme hydrolysis ability has better solution than the single enzyme and end, therefore continuous hydrolysis preparation of small peptides can be more effective. Using neutral protease, papain, flavourzyme, alkaline protease, trypsin, wheat gluten, composite protease and Proteax enzyme, through analysis and evaluation bitter taste dilution strength products of single enzyme hydrolysis, enzyme and construction of exopeptidase hydrolysis continuous square Comparison of different schemes in the case. The amino acid hydrolysate peptide nitrogen content, composition, influence analysis of the sensory characteristics of electronic tongue and the molecular weight distribution and amino acid sequence. The results showed that the product of continuous hydrolysis preparation of enzyme solution, differences in the content of the bitter peptides. Little amino acid sequence and molecular weight distribution of bitter threshold polypeptide the decision of bitterness intensity hydrolysates. With other solutions prepared hydrolysates compared to Pro-m hydrolysate peptide has the highest nitrogen content (61.54%), the content of small peptide (53.26%) and the lowest value (1.33), and the bitter taste of the soluble nitrogen content as high as 82.82%. so determined to wheat gluten protein -Proteax enzyme hydrolysis system as the basic framework, to carry out follow-up work. By Glutaminase SD-C100S, Glutaminase and hydrochloric acid (HCl) of Pro-m hydrolysis products were deamidation of glutamine, prompted the enzymolysis products into the valley Ammonia acid, umami peptide and free glutamate play by fresh suppression and bitter characteristics, improve hydrolysate flavor. Comparative analysis of flavor characteristics of the deamidation of hydrolysates, molecular weight distribution and amino acid composition of a main component of fresh matter and the bitter taste of the inhibitory effect of deamidation of hydrolysates. The results show that the deamidation degree increases, the rising content of umami substances, causing hydrolysate flavor enhancement, reduce the bitter taste. Glutamate strong hydrophilicity, saliva solubility stronger than umami peptides, can inhibit the bitter better signal conduction. Umami peptides receptors longer than the role of glutamate increased duration of flavor, will show a more gentle, strong sense of coordination. In order to optimize the enzymatic hydrolysis of wheat gluten -Proteax system of wheat gluten protein enzymolysis bitter peptide product of release The isobutanol extraction mechanism. Bitter product release features changed by enzymatic hydrolysis, bitter peptide product bitter strong effective extraction enzyme. Systematically bitter peptide amino acid sequence of Proteax hydrolysis process, molecular weight distribution, variation of amino acid composition, and analysis of isobutanol extract, using partial least squares regression of sensory attributes correlation weight distribution and amino acids composition. The results show that, compared to the changes in the amino acid sequence, amino acid composition of bitter peptides, the molecular weight distribution of bitterness intensity of bitter peptides have a more significant effect. The enzymatic reaction in 0-120 min, in the bitter bitter amino acid peptide (molecular weight at 180-500 Da, 500-1000 Da and 1000-3000 Da interval) in the proportion increased gradually. The enzymatic reaction at 120-300 min, the content of amino acid in the bitterness of bitter peptides and improving extraction. 鐗╀腑楂樺垎瀛愰噺鑻﹀懗鑲界殑鍚�閲忛檷浣�,灏ゅ叾鏄�鑻﹀懗鏈�寮，

本文编号：1572462