有机溶剂对脂肪酶ipozyme^IM催化活性的影响

脂肪酶Ｌｉｐｏｚｙｍｅ＾ＩＭ经有机溶剂浸泡处理后活性显著提高。该文系统地探讨了有机溶剂特性,有机溶剂水含量以及有机溶剂包温度和浸泡时间对Ｌｉｐｏｚｙｍｅ＾ＩＭ催化活性的影响。发现Ｌｉｐｏｚｙｍｅ＾ＩＭ经疏水性较强的有机沉沦 泡处理后活性成倍增长;浸泡的最佳温度为６０℃左右,浸泡时间对酶活影响不大;随着有机溶剂水含量的增加,酶活性急剧下降。     

耐有机溶剂脂肪酶的研究进展

脂肪酶是一种重要的酶,在有机介质中脂肪酶能催化某些特定的化学反应。获得具有较高催化活性的耐有机溶剂脂肪酶具有重要的价值。该文概述了一些具有代表性的耐有机溶剂脂肪酶,介绍了耐有机溶剂脂肪酶的特性和应用,并对耐有机溶剂脂肪酶的筛选途径和分子改造方法进行了分析。今后有机溶剂耐受性脂肪酶的研究将集中在酶的利用、微生物筛选、脂肪酶表征和基因克隆、酶的分子改造等方面。     

甲醇溶液对木瓜蛋白酶催化活性的影响

试验结果表明,木瓜蛋白酶在一定浓度甲醇溶液中水解酪蛋白的活性显著上升;动力学测定表明,该酶在甲醇溶液中Km下降;在甲醇溶液中木瓜蛋白酶催化酪蛋白水解反应的最适pH为8.5,最适温度为70℃;紫外差示光谱研究表明,在甲醇溶液作用下木瓜蛋白酶的二级结构发生了变化;荧光发射光谱研究表明,木瓜蛋白酶在甲醇溶液中发射峰位向低波长移动,荧光峰值明显增高。     

反应介质对脂肪酶稳定性的影响

脂肪酶Ｌｉｐｏｚｙｍｅ~（ＩＭ）在有机溶剂中的热稳定性显著提高。在水溶液中,热处理温度高于５０℃后,其催化甘油三酯水解的活力迅速下降。温度升到６０℃时,水解活力仅残存１３．６％。温度高于７０℃该酶完全失活。而在有机溶剂正庚烷中,温度高达８５℃仍表现出较高的活性。反应介质的疏水性越强,酶抗超声辐射变性作用的能力越强。经同样的超声辐射处理后,正己烷中Ｌｉｐｏｚｙｍｅ~（ＩＭ）的酯水解活力仅损失ｌｌ．８％,而在水溶液中其活力却损失了９５．９％。     

有机溶剂中固定化脂肪酶催化硅醇的酯化反应

固定化Mucor miehei脂肪酶可催化有机硅醇和脂肪酸的酯化反应．对固定化酶用量、脂肪酸链长、不同有机硅醇底物、有机溶剂极性和水含量等影响因素进行了初步研究．     

耐受有机溶剂洋葱伯克霍尔德菌ZYB002全细胞脂肪酶酶学性质

一株对多种有机溶剂具有良好耐受能力的产脂肪酶菌株ZYB002经分子鉴定为洋葱伯克霍尔德菌。其产生的细胞结合脂肪酶最适温度为65°C,最适pH为8.0,在低于70°C和pH3-8.5的范围内,全细胞脂肪酶保持稳定。Ca2+、K+、Na+和NO3-等离子对脂肪酶活性有激活作用,而Zn2+有抑制效应。全细胞脂肪酶对正丁醇有较强的耐受能力,但曲拉通X-100对脂肪酶活性有强烈的抑制效应。洋葱伯克霍尔德菌ZYB002全细胞脂肪酶良好的碱稳定性、热稳定性和有机溶剂耐受性,表明该全细胞脂肪酶具有重要的工业应用潜力。     

响应面法优化耐有机溶剂脂肪酶营养条件

目的：通过优化蜡状芽孢杆菌SWWL6产耐有机溶剂脂肪酶营养条件,产酶量有较大提高。方法与结果：通过单因素实验确定了产酶的最佳碳源、氮源及无机盐分别为可溶性淀粉、酵母膏、NH4NO3、MgSO4·7H2O和NaCl。部分因子实验结果表明初始培养基中酵母膏、NH4NO3的质量浓度对产酶的影响显著。通过最陡爬坡实验逼近最大响应区域。以中心组合设计和响应面分析法确定了最优培养基。结论：优化的培养基为酵母膏0.64%、NH4NO30.384%、可溶性淀粉1%、MgSO4·7H2O0.1%、NaCl0.25%、甲苯20%。优化后脂肪酶相对酶活为348.44%,比优化前提高了3.48倍。     

超声波对猪胰脂肪酶催化活性影响的机理研究

猪胰脂肪酶(porcine pancreas Lipase,PPL)反应系统经适宜参数的超声波处理,催化活性提高了11.22%～24.42%。PPL经超声波处理后,其动力学参数Km和Vmax都变小,酶分子的紫外吸收光谱不变,荧光发射光谱不变。而酶分子经超声波处理后,其紫外差示光谱出现了明显的正峰或负峰。探讨了超声波影响PPL活性的作用机理。     

反应介质对脂肪酶稳定性的影响

脂肪酶Ｌｉｐｏｚｙｍｅ＾ＩＭ在有机溶剂中的热稳定性显著提高,在水溶液中,热处理温度高于５０℃后,其催化甘油三酯水解的活力迅速下降,温度升到６０℃时,水解活力仅残存１３．６％,温度高于７０℃该酶完全失活,而在有机溶剂正庚烷中,温度高达８５℃仍表现出较高的活性,反应介质的疏水性越强,酶抗超声辐射变性作用的能力越强,经同样的超声辐射处理后,正已烷中Ｌｉｐｏｚｙｍｅ＾ＩＭ的酯水解活力仅损失１１．８％,而在     

PARTIAL PURIFICATION AND CHARACTERIZATION OF THE ORGANIC SOLVENT-TOLERANT LIPASE PRODUCED BY Pseudomonas fluorescens RB02-3 ISOLATED FROM MILK

Eighty-five putative Pseudomonas isolates were obtained from various raw milk and pasteurized milk samples using Pseudomonas CFC agar. Among them, 36 isolates were identified as Pseudomonas fluorescens, and one isolate was identified as Pseudomonas putida. Lipase activity of the strains was quantitatively measured by the spectrophotometric method using p-nitrophenyl palmitate (p-NPP) as substrate. Detected lipase activity of the strains was between 10.03 U/mL and 22.16 U/mL. Pseudomonas fluorescens RB02-3 possessed the highest lipase activity. The extracellular lipase of P. fluorescens RB02-3 strain was homogeneously purified using a combination of ammonium sulfate precipitation, dialysis, and gel filtration column chromatography. This purification procedure resulted in 2.97-fold purification with 20.3% recovery. The enzyme was characterized, and exhibited maximum activity at pH 7.0 and 50°C; after it was incubated for 1 h it was activated in the presence of hexane, ethyl acetate, isopropanol, and ethanol and remained stable after the incubation was extended for 2 hr. The lipase was slightly inhibited in the presence of Zn²⁺, Co²⁺, Cu²⁺, Ni²⁺ salts, and ethylenediamine tetraacetic acid (EDTA), whereas Cd²⁺, sodium dodecyl sulfate (SDS), and Tween-80 had no effect on its activity.     

脂肪酶催化有机硅化合物的生物转化

C.cylindracea脂肪酶可催化有机介质中有机硅醇与脂肪酸的酯化反应。微水有机介质比水-水不溶有机介质更有利于酶的反应,有机硅醇是比其碳结构类似物更好的酰基受体。对不同有机硅醇底物,当其空间障碍大时,不利于酶催化酯化反应,对不同脂肪酸底物,有机硅醇未影响该脂肪酶的脂肪酸底物特异性。     

卡门柏青霉-PG3脂肪酶的研究

从242株青霉属菌株中筛选出脂肪酶产生菌青霉-PG3。经鉴定,定名为卡门柏青霉(Penicillium camembertii Thom)。卡门柏青霉-PG3在由4％豆饼粉,0.5％糊精,0.75％橄榄油,0.5％K_2HPO_4,0.1％(NH_4)_2SO_4组成的液体培养基中,28℃,振荡培养96小时,发酵液脂肪酶活力(39℃,pH7.0)达60U/ml。PG3脂肪酶以橄榄油为底物,水解反应最适温度为48℃,最适pH为8.0。pH稳定范围6.0—11.0。Cu~(2+),Ca~(2+),Fe~(2+),Pb~(2+)等金属离子对酶活力有抑制作用。PG3脂肪酶对椰子油、菜籽油、亚麻油等油脂的水解率分别达到96％,94％和90％。     

菌种保存方法对冰核细菌冰核活性的影响

本文测定了5种保存方法及3种保存温度,对4株代表3个属4个种(变种)的冰核细菌冰棱活性的影响。结果表明,不同保存方法对冰核细菌的存活力和冰核活性存在着不同程度的影响,保存温度越高,对冰核活性的影响越大,不同菌种对保存的敏感性不同。所适合的保存方法也不同。真空冷冻干燥和灭菌水于-20℃冷冻保存对各菌株的存活力和冰核活性的影响都较小,适用于一般冰棱细菌的长期保存。而10%甘油冷冻保存则不适用于冰核细菌的保存。     

EFFECT OF LIGHT ON THE DEVELOPMENT OF THE CIRCADIAN RHYTHM OF MOTOR ACTIVITY IN THE MOUSE

In previous experiments, we found that rats raised in constant light (LL) manifested a more robust circadian rhythm of motor activity in LL and showed longer phase shifts after a light pulse in constant darkness (DD) than those raised under constant darkness. In addition, we observed that the effects produced by constant light differed depending on the time of postnatal development in which it was given. These results suggest that both sensitivity to light and the functioning of the circadian pacemaker of the rat could be affected by the environmental conditions experienced during postembryonic development. Thus, the present experiment aimed to study whether postnatal exposure to light could also affect the circadian system of the mouse. Three groups of mice were formed: One group was raised under constant darkness during lactation (DD group), the second under constant light (LL group), and the third under light-dark cycles (LD group). After lactation, the three groups were submitted first to constant light of high intensity, then to LD cycles, and finally to constant darkness. In the DD stage, a light pulse was given. Finally, mice were submitted to constant light of low intensity. We observed that the circadian rhythm of the DD group was more disturbed under constant light than the rhythm of the LL group, and that, when light intensity increased, the period of the rhythm of the DD group lengthened more than that of the LL group. No significant differences among the groups were found in the phase shift induced by the light pulse. Therefore, it appears that DD mice are more sensitive to light than their LL counterparts. However, at present there is no evidence to affirm that the light environment experienced by the mouse during postnatal development affects the circadian pacemaker. (Chronobiology International, 18(4), 683–696, 2001)     

电刺激延髓最后区对血浆肾素活性及肾交感神经活动的影响

实验在68只家兔上进行。乌拉坦麻醉。用RM-6000型多道仪同步记录呼吸(Respir)、血压(BP)、心率(HR)及肾神经放电(RSNA)。用放免法测定血浆肾素活性(PRA)。应用外径0.3mm的双芯同心电极置于延髓最后区(AP)处进行电刺激,每30s刺激4s。实验分为三组。第一组仅刺激AP观察到PRA增加了91％,RSNA出现以兴奋为主的三种放电形式,BP升高,HR减慢,呼吸无显著变化。第二组去除双侧肾神经后再刺激AP,PRA增加甚微,RSNA及血流动力学反应与第一组刺激前后的变化相似。第三组注射心得安后,刺激AP,这时除PRA明显抑制外,RSNA、BP、HR及呼吸亦与第一、第二组刺激前后的变化相似。上述结果表明电刺激兔AP能引起肾素释放增加,肾交感神经放电及血流动力学活动增强。     

碱性脂肪酶产生菌扩展青霉W—1382的诱变育种

以扩展青毒（Ｐｅｎｉｃｉｌｕｍｅｘｐａｎｓｕｍ）Ｓ－１４作为出发菌株,分别比较ＵＶ、Ｃｏ６０－γ射线、ＮＴＧ单因子的诱变效应和以ＮＴＧ、Ｃｏ６０－γ射线、ＵＶ＋ＮＴＧ作为诱变剂及自然分离在内四代的诱变育种。单因子诱变剂最适剂量分别为ＵＶ１５０ｓ,Ｃｏ６０－γ射线４８０００ｒａｄ,ＮＴＧ３００μｇ／ｍｌ８０ｍｉｎ,比较诱变效果表明：ＮＴＧ最佳,Ｃｏ６０－γ射线次之,ＵＶ较差;连续三代诱变各代诱变剂及剂量分别为ＮＴＧ３００μｇ／ｍｌ１００ｍｉｎ,Ｃｏ６０－γ射线２００００ｒａｄ,ＵＶ３０ｓ＋ＮＴＧ１５０μｇ／ｍｌ,４０ｍｉｎ。复合处理比单因子处理效果好。实验中选育成功高产酶变株Ｗ－１３８２,结合自然分离和诱变育种,其酶活力较出发菌株提高了５３．６％。     

溶剂极性及盐酸胍对碳酸酐酶构象的影响

以紫外荧光光谱为检测手段,比较研究了溶剂极性及盐酸胍对碳酸酐酶构象的影响,结果表明,当盐酸胍浓度小于１ｍｏｌ／Ｌ时酶呈天然构象,１．５ｍｏｌ／Ｌ－２．０ｍｏｌ／Ｌ的盐酸胍导致该酶呈伸展构象;而乙醇不以导致该酶产生稳定的中间态构象,１ｍｏｌ／Ｌ和３ｍｏｌ／Ｌ的甲醇以及３ｍｏｌ／Ｌ的乙醇对盐酸胍致碳酸酐酶中间态的影响不大,甲醇略使致中间态的盐酸胍浓度下降,乙醇略使之上升。     

有机物质对紫球藻生长的影响

研究了有机碳、氮源扩B族维生素对紫球藻生长的影响。葡萄糖促生长作用最佳,添加2％（W／V）葡萄糖时,藻细胞生长速度比对照组明显提高,培养10d收获的生物量增加92.6％,培养液中的溶解氧含量和藻体叶绿素a含量也有变化,有机氮源的利用率低,仅蛋白胨、酵母汁可被利用。维生素B2和B12也有促长作用。