脂肪酶水解壳聚糖作用研究

通过还原端基含量测定、粘度测定研究了脂肪酶非专一性对壳聚糖的降解作用,探索了酶反应过程中温度、pH、加酶量、底物浓度、反应时间、金属离子等因素对脂肪酶降解壳聚糖的影响,以及不同脱乙酰度和不同分子量的壳聚糖与脂肪酶降解反应的关系。结果表明,以壳聚糖为底物的脂肪酶的一些催化特性为最适温度50～55℃,最适pH5～5.5,反应时间在3～4h范围内;降解作用随着酶用量和底物浓度的增加而增加,水解反应不符合Michaelis-Menten方程;3mmol/L的Cu2+,10mmol/L的Mg2、Ca2+对脂肪酶有一定的激活作用,10mmol/L的Ba2+、Zn2+、Fe3+对该酶有一定的抑制作用,随着底物壳聚糖脱乙酰度的提高,降解速度降低。      

脂肪酶水解壳聚糖作用的研究

壳聚糖被水解生成的甲壳低聚糖(chitooligosaccharides)是水溶性产物，可被人体、动物和植物吸收，有很高的免疫调节功能及抗肿瘤、抗菌等生理活性。甲壳低聚糖的制备中酶法水解由于其条件温和，易控制而受到广泛关注。据报道，甲壳素和壳聚糖除被甲壳素酶、壳聚糖酶和溶菌酶降解外，还能被葡萄糖酶、蛋白酶、脂肪酶、纤维素酶、淀粉酶、半纤维素酶和果胶酶等非专一性酶有效地水解，非专一性酶由于价格便宜、易制得等优点而倍受关注，而且有些非专一性酶甚至比专一性酶更加有效，例如脂酶。Pantaleone等研究了脂酶的非专一性作用，发现来自猪胰腺的脂肪酶对壳聚糖有强烈的水解作用。国内夏文水教授等     

根霉脂肪酶降解壳聚糖条件研究

以德氏根霉(Rhizopus delemar)为菌种,采用固态发酵法生产脂肪酶,并用此酶对壳聚糖进行非专一性水解.研究了脂肪酶降解壳聚糖的多种影响因素,包括温度、pH、底物浓度、酶浓度、反应时间和常见金属离子等.结果表明,在45℃、pH5.0务件下,该脂肪酶能显著地降解壳聚糖,反应6h后,壳聚糖溶液的粘度即下降为原溶液的8%.适当增加酶浓度可使降解速度加快,但此酶促反应不遵循Michealis-Menten动力学方程.     

根莓脂肪酶降解壳聚糖条件研究

以德氏根霉（Rhizopus delemar）为菌种,采用固态发酵法生产脂肪酶,并用此酶对壳聚糖进行非专一性水解。研究了脂肪酶降解壳聚糖的多种影响因素,包括温度、pH、底物浓度、酶浓度、反应时间和常见金属离子等。结果表明,在45℃、pH5.0条件下,该脂肪酶能显著地降解壳聚糖,反应6h后,壳聚糖溶液的粘度即下降为原溶液的8%。适当增加酶浓度可使降解速度加快,但此酶促反应不遵循Michealis-Menten动力学方程。      

脂肪酶解聚壳聚糖及其衍生物的研究

壳聚糖及其衍生物能很容易被脂肪酶解聚。当壳聚糖乳酸盐溶液与脂肪酶在２５℃下作用１０ｍｉｎ后,其溶液粘度被降低到初始值的３５％,分子量从约７００ｋＤａ减少到１３ｋＤａ（还原糖滴定法测定）。脂肪酶浓度在４．５×１０－３～９×１０－１ｇ／Ｌ范围内,解聚初速度与酶浓度成对数线性关系。Ｎ－羧甲基壳聚糖比壳聚糖更容易被脂肪酶解聚。     

壳聚糖涂层法固定化脂肪酶的研究

研究了壳聚糖涂层在纤维素滤纸上成膜后再固定化猪胰脂肪酶的最佳条件。结果表明,当戊二醛浓度为５％,活化１２ｈ,与ｐＨ７．６的酶的磷酸盐缓冲溶液于室温（１５℃）交联１２ｈ,获得的固定化酶活最高,为０．２６Ｕ／ｃｍ２。固定化酶最适温度４０℃,比游离酶提高了５℃;最适ｐＨ８．５,与游离酶相比,向碱性偏移了０．５个ｐＨ     

纤维素酶水解壳聚糖条件的研究

利用绿色木霉产生的纤维素酶水解壳聚糖,并对其水解条件进行了研究.在50℃、pH5.0条件下,按照8 U/g底物的酶量,将纤维素酶添加到3%壳聚糖溶液中,水解20 h可得到平均聚合度为4的低聚壳聚糖.     

磁性壳聚糖微球固定化脂肪酶研究

以磁性壳聚糖微球为载体,通过戊二醛交联进行脂肪酶固定化,对影响脂肪酶固定化各种因素进行考察,确定最佳条件,并比较游离酶与固定化酶pH和热稳定性。结果表明,固定化适宜条件为:脂肪酶加入量5.0 mg/100 mg载体、温度40℃、时间5 h、pH 8.04、戊二醛浓度10%、最高固载率可达90.56%,酶活4034 U/g载体;与游离酶相比,固定化酶pH和热稳定性都有较宽适用范围。     

壳聚糖微球的制备及其对脂肪酶的固定化研究

采用反相悬浮法制备壳聚糖微球,并以此作为载体固定了脂肪酶。对壳聚糖微球的制备条件、微球的性能及其固定化脂肪酶的条件进行了探讨,结果表明,壳聚糖微球成球效果最好的制备条件是壳聚糖溶液与分散相液体石蜡体积比为1∶2,吐温-80使用量为15mL,壳聚糖浓度为4%,所制得的壳聚糖微球具有良好的热稳定性、耐酸碱性和抗氧化性;壳聚糖微球固定化脂肪酶的最佳条件为戊二醛用量0.6mL,交联时间60min,加酶量1mg/g载体,pH值为7。采用壳聚糖微球固定化脂肪酶具有较高的酶活回收率,为60%     

酶法制备低聚壳聚糖的研究

通过对α-淀粉酶、纤维素酶、脂肪酶降解壳聚糖产物的黏均分子量的测定,确定了这三种酶降解壳聚糖的最佳工艺条件。结果表明：α-淀粉酶降解的最传条件为时间120min,温度45℃,酶量1400U／g;纤维素酶降解的最佳条件为时间75min,温度45℃,酶量400U／g;脂肪酶降解的最佳条件为时间90min,温度45℃,酶量800U／g。同时得到了分子量小于2000的低聚壳聚糖,为进一步研究低聚壳聚糖与金属离子络合提供了条件。     

脂肪酶水解乳清中混合油脂的研究

研究了乳清中脂肪及其添加脂肪的水解作用。单因素试验详细比较了酶添加量、pH值、温度、水解时间、底物浓度等对乳清中油脂水解度的影响。利用脂解率和水解产物中游离脂肪酸（FFA，以油酸计）质量分数分析结合响应面分析（SAS）确定最佳水解条件：加酶量为300U／g乳脂肪；温度为45℃；质量分数40％；转速150r／min。     

磁性壳聚糖微球固定化脂肪酶的研究

目的:制备具有高活性的固定化脂肪酶;方法:以磁性壳聚糖微为载体,用物理吸附法固定化脂肪酶,对影响固定化的各种因素进行考察,确定最优条件,并比较游离酶和固定化酶的pH和热稳定性,研究固定化酶的使用稳定性;结果:固定化的适宜条件为采用加酶量600 U/g,温度5℃.pH 7.0,固定时间2 h,固定化酶的pH和热稳定性都优于游离酶,固定化酶连续使用5次,其相对酶活仍为使用前的57.8%,具有较好的操作稳定性;结论:磁性壳聚糖微球是固定脂肪酶的良好载体.     

Hydrolysis of bovine milk fat globules by lipoprotein lipase: inhibition by proteins extracted from milk fat globule membrane

Extraction of membrane proteins from milk fat globules by GuHCl or by MgCl2 made the lipids more accessible to lipolysis by added lipoprotein lipase. The increase in lipolysis paralleled the loss of membrane proteins and was continuous up to 2.5 M GuHCl, which was the highest concentration used. About twice as much protein was extracted with 2.5 M GuHCl as with buffer only. The amount of protein lost was about 50% of total milk fat globule protein. Lipolysis of milk fat globules was inhibited by addition of the extracted protein. The extracted proteins also reduced lipolysis when added to whole milk. More protein was needed to inhibit lipolysis of milk fat globules treated with GuHCl compared with globules treated with buffer only. The inhibition by a given amount of protein decreased if more milk fat globules were used. Protein extracted with MgCl2 had similar effects as those extracted with GuHCl. The major components extracted with MgCl2 migrated in the 40 to 50-kdalton region on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. By gel filtration chromatography, two protein fractions were obtained, which inhibited lipolysis more efficiently than the total extract. As has previously been found for inhibition of lipolysis by skim milk, the amount of extracted protein needed to inhibit lipolysis varied between preparations of milk fat globules. Milk with propensity to cold-induced ("spontaneous") lipolysis was normalized by addition of extracted proteins.     

Influence of chitosan on stability and lipase digestibility of lecithin-stabilized tuna oil-in-water emulsions

The influence of chitosan concentration (0–0.3 wt%) and molecular weight (120, 250 and 342.5 kDa) on the physical stability and lipase digestibility of lecithin-stabilized tuna oil-in-water emulsions was studied. The ζ-potential, droplet size, creaming stability, free fatty acids and glucosamine released was measured for the emulsions when they were subjected to an in vitro digestion model. The ζ-potential of the oil droplets in lecithin-chitosan stabilized emulsions changed from positive (≈+53 mV) to negative and the emulsions were unstable to droplet aggregation for all chitosan concentrations and molecular weights used after being subjected to the digestion model. The amount of free fatty acid and glucosamine released per unit amount of emulsion was higher when pancreatic lipase was included in the digestion model. These results suggest that lecithin-chitosan coated droplets can be degraded by lipase under simulated gastrointestinal conditions. Consequently, chitosan coated lipid droplets may serve as useful carriers for the delivery of bioactive lipophilic nutraceuticals.     

Alcoholysis of salicornia oil using free and covalently bound lipase onto chitosan beads

Porcine pancreatic lipase was immobilized on chitosan by covalent binding and retention of its activity was examined. The activities of free and immobilized lipase were determined using olive oil as substrate. The free and immobilized enzymes showed pH 9 as optimum and retained 50% of activity after five cycles. When the substrate concentration was kept constant and enzyme concentration was varied, the K_m and V_max were observed to be 4.0 × 10⁻⁷ and 0.32, and 3.32 × 10⁻⁷ and 0.32, respectively, for free and covalently bound enzyme. This indicates that there is no possibility of conformational change during immobilization. Immobilized enzyme showed improved thermal and storage stability. Alcoholysis of salicornia oil, mediated by free and immobilized lipase, was carried out at 25 °C using methanol in hexane and acetone. Free and immobilized enzyme in hexane produced, respectively, 45% and 55% of fatty acid methyl ester after 12 h.     

酸性酶对纤维素的水解性能

用酸性纤维素酶分别处理棉、粘胶和天丝织物,通过测定处理液中还原糖含量,研究了影响酶水解的各种因素.结果表明,酸性酶对粘胶的水解活力最高,纯棉次之,对天丝织物的活力最低;织物所受前处理条件、酸性酶浓度、机械搅拌条件和不同缓冲体系等对酸性纤维素酶水解性能也会产生影响.经退浆、煮练、漂白后的织物,酶水解能力较高;适当地增加机械搅动,采用HAc-NaAc缓冲体系,酶活力也较高.     

纤维素酶水解玉米芯的研究

研究了影响纤维素酶水解玉米芯的各个因素(pH、温度、酶添加量、底物浓度)，得到纤维素酶水解玉米芯的最佳条件——pH4．5，温度35℃，酶添加量10U／g玉米芯，底物浓度100g／L。在最佳条件下纤维素酶水解玉米芯的葡萄糖产率为27．1％。     

通过酶促反应制备壳寡糖

用酶降解法对壳聚糖进行降解 ,研究了温度、pH值、金属离子等对酶促反应的影响 ,降解的最佳温度和pH值分别为 5 0℃和 5 0 ,Mg2 + 和Ca2 + 对酶降解具有一定的促进作用 ,而重金属离子Cu2 + ,Ni2 + 和Zn2 + 对酶降解有强烈的抑制作用。通过降解过程中壳聚糖溶液粘度的变化可以得出 ,该壳聚糖酶是以内切方式作用于壳聚糖。采用离子交换色谱柱对降解产物进行分离 ,得到了聚合度为 2～ 9的壳寡聚糖。该酶促反应符合米氏动力学方程 ,米氏常数Km 值为 2 60 1g/L ,Vmax 值为 3 5 79× 10 - 2 g/min·L。