【作者】 刘亮亮；

【导师】 韩玲；

【作者基本信息】 甘肃农业大学 ， 营养与食品卫生学， 2010， 硕士

【摘要】 目前国内生产的胰酶主要是猪胰酶,羊胰酶还是空白;羊胰酶属清真产品,为广大穆斯林消费者专用。本研究以羊胰脏为试验材料,采用有机溶剂法进行羊胰酶的提取,优选异丙醇为提取剂和沉淀剂;通过单因素试验和正交设计,对主要影响因素和工艺参数进行了研究,确定了胰酶提取的主要工艺流程和参数,优选了保护剂、激活剂和脱脂剂;参照中华人民共和国药典,测定了羊胰酶活力和理化指标;并研究了羊胰酶水解酪蛋白动力学特性。研究结果如下:1.工艺流程:采用有机溶剂法,以冻藏的羊胰脏为原料,经过胰脏的破碎、激活、提取、沉淀、脱脂、真空干燥等工艺过程后得到羊胰酶。2.提取工艺参数:优选提取剂浓度、激活时间、激活温度、激活pH值、提取温度和提取时间的最佳提取条件及参数。通过单因素试验,选出影响羊胰酶提取过程的主要因素及其参数范围,进一步通过L₁₈（3⁷）正交试验,以胰蛋白酶活力为依据,筛选出的最佳工艺参数:提取剂异丙醇浓度10%、提取温度15℃、提取时间5h、激活pH6、激活温度15℃、激活时间5h。3.优选保护剂和激活剂:筛选麦芽糖、淀粉和氯化钠为保护剂,Ca²⁺-胰酶粉为激活剂,均得到高活力的羊胰酶。4.沉淀工艺参数:优选沉淀剂浓度、沉淀温度、沉淀pH和静置时间的最佳沉淀条件及参数。通过单因素试验,选出影响胰酶沉淀的主要因素及其参数范围,进一步通过L₉（3⁴）正交试验,以胰蛋白酶活力为依据,筛选出的最佳工艺参数:沉淀剂浓度60%、沉淀温度5℃、沉淀pH值6、静置时间1h。5.优选丙酮为最佳脱脂剂,按上述最佳工艺提取的羊胰酶,通过胰酶活力的测定,胰蛋白酶的活力为2654U/g,胰淀粉酶的活力13800U/g,胰脂肪酶的活力为10400U/g,羊胰酶的收率为11.2%,脂肪残留为17.1mg/g,干燥失重为3.2%,均符合药典的规定。6.羊胰酶水解酪蛋白的动力学特征:最适温度40℃、最适pH值8;通过热稳定性的研究,酶活力随时间延长逐渐降低,温度越高,胰酶失活越快,热稳定性越差;通过pH稳定性的研究,酶活力随时间延长逐渐降低,最适pH时,胰酶失活最快,pH稳定性越差;Ca²⁺激活羊胰酶,并提高胰酶的稳定性,Na⁺对胰酶活力无影响,EDTA可以抑制胰酶活性。综上所述,采用异丙醇提取羊胰酶,工艺简单、提取效率高、产品酶活高、成本低、生产周期短。提高了生产效率和原料利用率,节约资源,减少污染。利用先进的生物分离工程技术进行羊胰酶的生产工艺研究,提高动物副产品综合利用中的回收率和保存率,在生化制药业中以及动物副产品综合利用中具有积极的意义。更多还原

【Abstract】 Now production of pancreatin is mainly pig pancreatin,Ovine pancreatin in domestic blank, Ovine pancreatin is Muslim product for the Muslim consumers,The raw materials of the experiment is ovine pancreas, pancreatin extracting was studied by the methods of organic solvents, isopropyl alcohol as optimum extraction agent and precipitation agent.Through the single factor and the orthogonal test, the major effect factor and the process parameter of the pancreatin extracting method were studied, the main technical process and the parameter of pancreatin’s product has determined; protectant,activator and degreaser were studied;Through the pancreatin activities and physics chemistry Indicators were determined by the chinese pharmacopoeia,Kinetics of Ovine pancreatin hydrolyse casein were studied, the research results are as follows:1. Technical process: During the experiment which product ovine pancreatin by organic solvents method, take the frozen ovine pancreas as the raw material, during the pancreas broken,activation,extract,precipitation,remove fat,vacuum drying to get the pancreatin.2.Parameters of Extracting process: Optimizing activating time, activating temperature, activating pH value, extraction agent concentration, extracting temperature and extracting time of the best condition and parameters. Through the single factor test, determined the main impact factors and parameters, then using L₁₈（3⁷）orthogonal test, take the activities of trypsin as the index, screened out the best process parameter were: extraction agent concentration 10%,extracting temperature 15℃, extracting time 5h, activating pH6,activating temperature15℃and activating time 5h.3. Optimizing protectants and activators :Starch, maltose and sodium chloride as protectants and select the CaCl₂ -Pancreatin for the best activator.4.Parameters of precipitating process : Optimizing Precipitation agent concentration, Precipitation temperature, Precipitation pH value and Static placed time of the best condition and parameters. Through the single factor test,Determined the main impact factors and parameters, then using L₉（3⁴）orthogonal test, take the activities of trypsin as the index, screened out the best process parameter were: Precipitation agent concentration 60%,Precipitation temperature 5℃,Precipitation pH value 6 , Static placed time1h.5. Optimizing degreaser : Acetone as the best degreaser . According to the confirmation test of above process parameter, the result indicated that the yield of pancreatin was 11.2%,the activities of trypsin,amylopsin,lipase respective were 2654 U/g,13800U/g,10400 U/g,Loss on drying 3.2%,Fat content 17.1mg/g, the results fit for the chinese pharmacopoeia.6. Kinetics of Ovine pancreatin hydrolyse casein were studied: The optimum temperature was measured 40℃, the optimum pH was measured 8.Through the research of Thermal stability, pancreatin activity decreased with the time gradually increased, The higher the temperature, the faster inactivation of trypsin, The poor thermal stability;Through the research of pH stability, pancreatin activity decreased with the time gradually increased, At the optimum pH, the fastest inactivation of trypsin, The poor pH stability. According to research the affect of trypsin activity by Ca²⁺,Na⁺and EDTA: Ca²⁺ can activate ovine pancreatin,and to improve the stability of the ovine pancreatin,Na⁺ does not affect the activity of trypsin, but EDTA has Strong Inhibition effect to the activity of trypsin.In summary,Using the isopropyl alcohol extraction ovine pancreatin powder,Simple process,higher yield ,higher activity of three enzymes, Lower cost and Short production cycle. Improve the production efficiency and the utilization rate of raw materials, conservation of resources, reduce pollution.Use advanced bioseparation engineering to produce pancreatin, increase the yield and enzyme activity which has positive significance in biochemical pharmaceutical industry and comprehensive utilization of animal by-products. Key words: Ovine Pancreas; Pancreatin; Isopropyl alcohol; Extract;更多还原