Isolation and identification of lipase producing thermophilic Geobacillus sp. SBS-4S: cloning and characterization of the lipase

A thermophilic microorganism, SBS-4S, was isolated from a hot spring located in Gilgit, Northern Areas of Pakistan. It was found to be an aerobic, gram-positive, rod-shaped, thermophilic bacterium that grew on various sugars, carboxylic acids and hydrocarbons at temperatures between 45°C and 75°C. Complete 16S rRNA gene sequence of the microorganism exhibited homology to various species of genus Geobacillus. A highest homology of 99.8% was found with Geobacillus kaustophilus. A partial (0.7 kbp) chaperonin gene sequence also showed a highest homology of 99.4% to that of G. kaustophilus whereas biochemical characteristics of the microorganism were similar to Geobacillus uzenensis. Based on biochemical characterization, 16S rRNA and chaperonin gene sequences, we identified SBS-4S as a strain of genus Geobacillus. Strain SBS-4S produced several extracellular enzymes including amylase, protease and lipase. The lipase encoding gene was cloned, expressed in Escherichia coli and the gene product was characterized. The recombinant lipase was optimally active at 60°C with stability at wide pH range (6-12). The enzyme activity was enhanced remarkably in the presence of Ca(+2). The K(m) and the V(max) for the hydrolysis of p-nitrophenyl acetate were 3.8mM and 2273 μmol min(-1)mg(-1), respectively. The ability of the recombinant enzyme to be stable at a wide pH range makes it a potential candidate for use in industry.     

无溶剂催化体系酶法合成低热结构脂的研究

采用无溶剂体系,以三丁酸甘油酯及硬脂酸甲酯为原料,1,3-专一性脂肪酶Lipozyme RMIM为催化剂,研究了含短、长碳链脂肪酸的低热结构脂的酶法合成.探讨了反应器种类等单因素对反应体系的影响.结果表明:在减压旋转蒸发反应器中,在三丁酸甘油酯与硬脂酸甲酯摩尔比1∶2,反应温度60℃,反应时间6.0h,加酶量10%时,产物中SSL、LLS、LLL含量分别为54.15%、42.10%和5.16%.     

动态高压微射流对猪胰脂肪酶的影响

以猪胰脂肪酶(PPL)为原料,研究动态高压微射流(DHPM)对酶活性和构象的影响.结果显示:DHPM处理对PPL具有钝化作用,处理压力越大,钝化作用越小;4℃下贮存24 h,相对酶活有所回升.通过紫外光谱和圆二色谱分析DHPM对PPL构象的影响,发现经100 MPa DHPM处理后,PPL的紫外吸收强度降低,β-折叠含量减少,但随着处理压力增大,紫外吸收强度和β-折叠含量逐渐增大;4℃放置24 h后,紫外吸收强度和β-折叠含量进一步上升.PPL的相对酶活与紫外吸收强度、β-折叠含量呈一定的正相关.     

Cooling causes changes in the distribution of lipoprotein lipase and milk fat globule membrane proteins between the skim milk and cream phase

Lipoprotein lipase (LPL) activity and free fatty acid levels were studied in freshly milked, uncooled milk from individual Danish Holstein or Jersey cows, or after storage for up to 24 h at either a cooling temperature (4°C) or at the milking temperature (31°C). Upon cooling for up to 24 h, LPL activity increased in the cream phase, whereas the activity in the skim milk was steady, as observed for Jersey cows, or increased, as seen for the Holsteins. Storage at 31°C decreased the LPL activity in both the cream phase and the skim milk phase. The increase in free fatty acid levels was found to depend on LPL activity, incubation temperature, substrate availability, and incubation time. Furthermore, the migration of milk proteins between the skim milk phase and the cream phase upon cooling of milk from Jersey cows or from Danish Holstein cows was studied using proteomic methods involving 2-dimensional gel electrophoresis and mass spectrometry. Proteins associated with the milk fat globules were isolated from all milk fractions and analyzed. Major changes in the distributions of proteins between the skim milk phase and the cream phase were observed after cooling at 4°C for 4 h, where a total of 29 proteins between the 2 breeds was found to change their association with the milk fat globule membrane (MFGM) significantly. Among these, the MFGM proteins adipophilin, fatty acid-binding protein, and lactadherin, as well as the non-MFGM proteins β-casein, lactoferrin, and heat shock protein-71, were identified. Adipophilin, lactadherin, and lactoferrin were quantitatively more associated with the MFGM upon cold storage at 4°C, whereas β-casein, fatty acid-binding protein, and heat shock protein-71 were found to be less associated with the MFGM upon cold storage.     

纸浆和造纸白水中DCS的生物酶控制

随着造纸白水的循环回用,化学机械浆中的溶解与胶体物质(DCS)在造纸系统中不断积累,会给纸机运行和产品质量造成不利影响。以机械浆为原料的纸厂白水中积累的DCS类物质主要来源于原材料中的亲脂性抽出物、制浆过程引入的大量胶体状木素、半纤维素和果胶物质等。DCS中的树脂酸和脂肪酸组分会沉积在纸机设备表面或留在白水系统中,进而形成沉积物并产生障碍,利用脂肪酶处理DCS能够有效消除树脂障碍。DCS中的聚半乳糖醛酸对阳离子需求量贡献较大,利用果胶酶处理DCS能够有效地消除DCS中聚半乳糖醛酸所带来的危害。     

酶催化法拆分共轭亚油酸异构体

在Candida rugosa脂肪酶的催化下,采用正辛醇与共轭亚油酸(CLA)进行酯化反应,对CLA异构体进行拆分,并通过单因素实验优化酶法酯化的工艺条件.结果表明最优条件为:AY-30脂肪酶加量40.44 U/g,CLA与正辛醇摩尔比1:1,pH 6.5,温度30℃,反应时间12 h.当酯化率为41.92%时,c9,t11-CLA在两种异构体中的相对含量达到88.13%.     

酶法分离共轭亚油酸两种主要异构体的工业生产研究

应用脂肪酶酯化法研究了共轭亚油酸两种主要异构体分离纯化的生产条件。当在CLA和无水乙醇比为1:1、1%的脂肪酶LipaseAY30、100%的水、恒温30℃的反应条件下反应9h,再经分子蒸馏和碱化脱酸,可以得到酸值0.5,CLA总含量81.84%,c9,t11CLA:t10,c12CLA为3.02的共轭亚油酸乙酯产品。     

潜力工业催化剂——白地霉脂肪酶

白地霉脂肪酶因具有独特的底物特异性,在油脂改性等领域有着广泛应用。文中概述了白地霉脂肪酶的发酵生产、分离纯化、克隆与表达、底物特异性分子机理及应用等,并展望其未来发展方向,为该工业催化剂的研发与广泛应用提供参考。     

糙米微波稳定化技术研究

对微波处理糙米的稳定效果进行研究。以米糠作为对照组,探讨比较二者过氧化物酶、脂肪酶的酶灭活情况,同时考察大米品质的变化。结果表明：微波对糙米稳定化效果非常显著,而大米品质也受到影响,因此为获得大米品质损失最小、灭酶效果最好的工艺条件,通过三因素三水平响应面分析,对糙米微波稳定进行优化得最佳工艺参数为微波功率700W、微波时间100s、水分含量21%,理论脂肪酶活、碎米率分别为15.66μmol/(g ·h)、18.92%,实际脂肪酶活、碎米率分别为16.25μmol/(g ·h)、17.33%,强化储藏4周后的游离脂肪酸值较未处理糙米减少13.7%,说明所得的糙米微波稳定工艺可用于糙米稳定的实践。     

江米酒脂肪酶对米酒奶脂解风味的影响

利用传统江米酒制备凝乳剂来制作米酒奶、其中江米酒脂肪酶活力为1.0～1.5LU/mL。通过酸度值(ADV)分析发现,脂肪酸的积累主要发生在凝乳期间。采用气相色谱- 质谱联用仪(GC-MS)检测各游离脂肪酸的含量,棕榈酸、油酸、豆蔻酸和硬脂酸占有较高的比例。