胰蛋白酶提取香菇可溶性含氮化合物的研究

[目的]研究各种蛋白酶水解提取香菇可溶性含氮化合物,优化筛选胰蛋白酶酶解条件。[方法]采用各种食品级蛋白酶对香菇进行酶解,选取较优蛋白酶酶解条件,并结合螺杆挤压预处理提取香菇可溶性氮。[结果]胰蛋白酶提取香菇可溶性含氮化合物的最佳酶解条件为料液比1∶11 g/ml,酶解时间1.5 h,酶解温度50℃,酶量0.5%。在最优酶解条件结合螺杆挤压物理方法,可使香菇可溶性氮释放率及氨基态氮含量提高94.94%、82.94%。[结论]研究可为食用菌鲜味物质的提取利用提供参考。     

不同热处理对黑豆干物质消化率,蛋白质消化率的影响

为了研究热处理的温度和时间对黑豆干物质消化率(DMD)、蛋白质消化率(PD)及脲酶活性的影响规律,对经过110℃和121℃下分别处理3,9和15min的饲用黑豆进行了离体干物质消化率和离体蛋白质消化率及脲酶活性的测定。结果表明:在121℃处理15min的样本干物质消化率和蛋白质消化率最高,分别为75.13%,90.91%,脲酶活性最低,为0.04mg/min·g。作者认为:在本研究范围内,湿热处理最适宜的温度是121℃,时间是15min。     

Ontogenetic development of digestive enzymes and effect of starvation in miiuy croaker <Emphasis Type="Italic">Miichthys miiuy</Emphasis> larvae

The ontogenetic development of the digestive enzymes amylase, lipase, trypsin, and alkaline phosphatase and the effect of starvation in miiuy croaker Miichthys miiuy larvae were studied. The activities of these enzymes were detected prior to exogenous feeding, but their developmental patterns differed remarkably. Trypsin activity continuously increased from 2 days after hatching (dah), peaked on 20 dah, and decreased to 25 dah at weaning. Alkaline phosphatase activity oscillated at low levels within a small range after the first feeding on 3 dah. In contrast, amylase and lipase activities followed the general developmental pattern that has been characterized in fish larvae, with a succession of increases or decreases. Amylase, lipase, and trypsin activities generally started to increase or decrease at transitions from endogenous to exogenous feeding or diet changes, suggesting that these enzymatic activities can be modulated by feeding modes. The activities of all the enzymes remained stable from 25 dah onwards, coinciding with the formation of gastric glands and pyloric caecum. These results imply that specific activities of these enzymes underwent changes due to morphological and physiological modifications or diet shift during larval development but that they became stable after the development of the digestive organs and associated glands was fully completed and the organs/glands functioned. Trypsin and alkaline phosphatase were more sensitive to starvation than amylase and lipase because delayed feeding up to 2 days after mouth opening was able to adversely affect their activities. Enzyme activities did not significantly differ among feeding groups during endogenous feeding; however, all activities were remarkably reduced when delayed feeding was within 3 days after mouth opening. Initiation of larvae feeding should occur within 2 days after mouth opening so that good growth and survival can be obtained in the culture.     

The changes in digestive enzymes and hormones of gilthead seabream larvae (Sparus aurata, L 1758) fed on Artemia nauplii enriched with free methionine

Variations in digestive enzymes and hormones during the larval development of gilthead seabream (Sparus aurata) fed on live prey (Artemia nauplii) enriched with free methionine were investigated for 16 days (from day 24 to day 40). Prior to initiation of the experiment, newly hatched larvae were transferred from incubators to fiber glass tanks (300 l) with black walls and fed with same diets until day 24. Each experiment was performed in triplicate. In the experimental group, the content of the free methionine in the Artemia nauplii was increased by adding a 5.3 mM free methionine solution to the culture water during a 16-h enrichment period. The larvae of both the control and enriched-methionine groups were sampled four times, with 4-day intervals between samplings, during a 16-day period. The larvae in the control group had a significantly lower growth than those of the methionine group at the end of the study (P < 0.05). The highest trypsin activity and leucine aminopeptidase N/leucine–alanine peptidase ratios were observed in the control group. A significant difference between bombesin activities in the treatment groups was not found at 5th minute after the initiation of feeding (P > 0.05), but they were significant at 15th minute post-initiation of feeding (P < 0.05). A significant difference between the cholecystokinin levels of the treatment groups was found (P < 0.05).     

缺失SBTi—A2大豆杂交后代品系籽粒化学成分分析

对一个杂交组合后代１０个稳定缺失胰蛋白酶抑制剂（ＳＢＴｉ－Ａ２）品系及其亲本籽粒化学成分组成进行的分析比较表明，大豆籽粒中缺失ＳＢＴｉ－Ａ２对其蛋白质含量及天门冬氨酸、甘氨酸和丙氨酸含量有一定影响，表现为缺失ＳＢＴｉ－Ａ２品系显著低于高亲和双亲平均值；而对其他氨基酸和脂肪含量及脂肪酸组成基本没有显著的影响。     

胰蛋白酶基因的系统发育分析

下载NCBI数据库中的动物胰蛋白酶基因的氨基酸序列,用CLUSTAL X2程序进行序列比对,用MEGA3.1软件中的Neighbor-joining法构建系统发育树.序列分析表明,胰蛋白酶氨基酸有多处保守基序,且保守性较高,最典型的保守序列有催化三联体结构:His(组氨酸)-Asp(天冬氨酸)-Ser(丝氨酸)以及金有6个保守的半胱氨酸形成3对肽内二硫键.另外,此序列还具有特异性的底物结合位点,这就决定了胰蛋白酶的专一性.系统发育分析结果显示,脊椎动物与无脊椎动物被分成2个大的分支,而脊椎动物中trypsinX3物种被单独分为一支,这表明typsinX3物种与脊椎动物的同源性较低.     

紫花芸豆胰蛋白酶抑制剂的分离和纯化

[目的]从紫花芸豆中分离纯化胰蛋白酶抑制剂。[方法]以紫花芸豆为材料,采用脱脂、酸抽提、热变性、硫酸铵分级沉淀及离子交换层析和凝胶过滤层析等方法,分离纯化胰蛋白酶抑制剂。[结果]胰蛋白酶抑制剂经PAGE和IEF电泳显示单一条带。凝胶过滤法测定其表观分子量约为59kD。SDS电泳结果显示它有3个亚基,分子量分别为34、16、15kD。等电聚焦法测定其等电点为5．25。动力学的方法检测PVTT与胰蛋白酶发生不可逆抑制作用。[结论]离子交换层析和凝胶过滤层析法能快速地分离纯化紫花芸豆中的胰蛋白酶抑制剂。     

人胰岛素B、C、A链基因定点突变及表达载体的构建

采用定点突变技术,将组成人胰岛素原基因的C、A两条肽链进行了PCR突变连接,构建了人胰岛素C、A突变基因。采用T-A法将PCR产物B、CA克隆到T载体,分别构建了中间体pHB和pHCA,其测序结果与GenBank公布的一致。应用BamH I和Sse8387 I对pHB和pHCA进行双酶切,然后将回收纯化的片段进行定向连接,构建了重组质粒pHBCA,酶切分析证实重组质粒pHBCA构建成功。     

壳聚糖固定化胰蛋白酶研究

以自制的壳聚糖为载体,以戊二醛为交联剂,制备了壳聚糖固定化胰蛋白酶。经测定该酶活性回收率达到72.7%,操作半衰期为12 d,表观米氏常数为5.88 mg/ml,最适pH值为8.0,最适温度为40℃;与游离胰蛋白酶相比,酶的耐热性和操作半衰期有明显提高,但表观米氏常数也略有增大。     

Capacity for digestive hydrolysis and amino acid absorption in Atlantic salmon (Salmo salar) fed diets with soybean meal or inulin with or without addition of antibiotics

This experiment was done to study the effects of dietary soybean meal (SBM) and inulin (a prebiotic) on the capacity for digestive hydrolysis and amino acid absorption by Atlantic salmon, and how a dietary supplement of the broad-spectrum antibiotic oxytetracycline (OTC) modulated these responses. A control diet (FM) was made from fish meal, fish oil and extruded wheat. Two similar diets were made with 250 g soybean meal (SBM) or 75 g inulin kg^− 1. Each diet was made with or without a supplement of 3 g OTC kg^− 1. All six diets contained yttrium oxide for estimation of apparent nutrient absorption. Each diet was fed to two groups of 172 g salmon kept in 1 m² tanks with 9 °C saltwater for 3 weeks. Intestinal organs were then sampled and weighed. Gastrointestinal tracts (GIT) were sectioned for analyses of brush border alkaline phosphatase (ALP) and leucine aminopeptidase (LAP) activities. Tissue from the distal intestine (DI) was also fixed for histological examination. Digesta from the different sections were freeze dried for estimation of trypsin and amylase activities, and of apparent absorption of amino acids (AA), nitrogen (N), and sulphur (S). About 85% of the trypsin activity, 70% of the amylase activity, 85% of the ALP activity, and 82% of the LAP activity were found in the proximal (PI) and mid (MI) intestine of fish with functional DI, and the absorption of AA, N, and S was quantitatively completed in the MI. Dietary OTC resulted in lower relative liver weight, but apart from increased ALP and LAP activities in DI when feeding OTC in combination with inulin, OTC did not modify the responses to dietary SBM or inulin. Dietary SBM resulted in lower relative liver weight, and induced pathomorphological changes in the DI mucosa, thus lower the ALP and LAP activities in the DI. SBM also stimulated absorption of AA, N, and S in the PI, but at the same time increased the activities of trypsin and amylase in the DI, indicating reduced re-absorption and increased faecal losses of these endogenous enzymes. Dietary inulin did not damage the DI, and stimulated intestinal growth and higher relative mass of the GIT. Inulin without OTC did not affect the hydrolytic and absorptive capacity of the salmon GIT.