超临界酶法酯交换合成结构脂质中酶活力的研究

超临界酯交换合成结构脂质的反应中使用脂肪酶Lipozyme RM IM,不同的工艺条件对脂肪酶的催化活性会有影响.当反应温度为45℃,底物摩尔比(辛酸乙酯:鱼油)为15:1,酶用量为底物总量的10%,添加水量为酶量的30%,反应时间为11h,超临界CO2压力为12.5 MPa时,脂肪酶Lipozyme RM IM的催化效果最好,辛酸结合率在反应产物甘三酯中达最大值.肪脂酶Lipozyme RMIM在以上反应体系中重复使用17次,其酯交换活力降低50%.     

脂肪酶催化玉米胚芽油与辛酸制备MLM型结构脂质

为研究无溶剂系统脂肪酶催化玉米胚芽油与辛酸酸解制备功能性脂质的可能性及反应条件,以玉米胚芽油为原料,比较6种脂肪酶(Lipozyme RM IM、Lipozyme TL IM、Novozym 435、Lipase AK、Lipase AY和Newlase F)对无溶剂体系中固定化脂肪酶催化玉米胚芽油与辛酸酸解制备MLM结构脂质的影响。研究结果表明:所选6种脂肪酶中,来自米黑根毛霉的固定化脂肪酶Lipozyme RM IM的催化效果最好。当玉米胚芽油与辛酸的比率为1∶4(物质的量比)、脂肪酶量为12%(底物质量分数)、反应时间16 h、反应温度50℃时,玉米胚芽油中辛酸插入率最高。在无溶剂系统,利用Lipozyme RM IM脂肪酶催化玉米胚芽油与辛酸进行酸解反应,可制备高品质的MLM型功能性脂质。     

Lipozyme TL IM催化合成中长链脂肪酸甘油三酯的工艺优化

为探索高含量中长链脂肪酸甘油三酯(MLCT)和低成本的酶法催化合成MLCT工艺,以一级菜籽油为原料,采用脂肪酶Lipozyme TL IM酶法催化酯交换合成MLCT,采用单因素试验研究了底物配比(菜籽油与中链甘油三酯质量比)、反应时间、反应温度、酶添加量对酯交换反应的影响,在此基础上,通过正交试验对MLCT合成工艺条件进行优化。结果表明,最佳的MLCT合成工艺条件为底物配比3∶1、反应时间4 h、反应温度50℃、酶添加量8%(基于底物的质量),在此条件下MLCT含量达到87.50%。优化的MLCT合成工艺具有MLCT含量高,反应时间短、反应温度低的优势,可降低能耗及减少酶失活,从而降低了生产成本。     

超临界体系酶催化制备甘油二酯及其纯化

以二氧化碳为流体,在超临界体系中用脂肪酶催化大豆油脂与甘油反应制备甘油二酯及其纯化研究。选取Lipozyme RMIM、Novozyme 435、Lipozyme TLIM 三种固定化脂肪酶为试验酶进行酯化反应,通过单因素试验,分别确定3种酶的最佳工艺条件,3种酶的最佳添加量分别为2.5%、3%、8%;反应温度分别为65、70、65℃,反应时间分别为7、8、9h,底物比均为2:1,得甘油二酯含量分别为68.6%、67.7%、64.8%。采用二级分子蒸馏工艺对超临界体系生产的甘油二酯混合物进行纯化,甘油二酯产品纯度从68.6%提高到90.4%,产品得率为60.0%。综合考虑,在工业生产中,建议用脂肪酶Lipozyme RMIM。     

酶催化酸解棉籽油分提产物合成结构脂质研究

以棉籽油分提产物和辛酸为原料,Sn1,3-位定向固定化脂肪酶为催化剂,在无溶剂体系中催化酸解棉籽油分提产物合成一类具有供能低、易消化、熔点低等优点的功能性结构脂质。对3种不同碱液进行脱酸能力比较,结果表明KOH-乙醇水溶液脱酸效果最好;对3种不同来源的脂肪酶脱酸能力进行筛选,结果表明Lipozyme RM IM脂肪酶催化活性高;在单因素试验基础上,以辛酸含量为响应值,采用响应面试验确定最优工艺条件为:反应温度68℃,反应时间7.88 h,酶添加量11.8%,底物比2.98∶1。在最佳反应条件下,产物中辛酸质量分数达35.25%。     

酶催化桑蚕蛹油酯交换制备中长链脂肪酸甘油酯

以桑蚕蛹油为底物,通过酶法催化其与三辛酸甘油酯进行酯交换反应,制备富含α-亚麻酸的中长链脂肪酸甘油三酯(MLCTs)。通过单因素实验考察了酶种类、底物质量比、反应温度、加酶量以及反应时间对酯交换反应的影响,对酯交换条件进行了优化。结果表明,最佳反应条件为：采用Lipozyme TL IM脂肪酶,三辛酸甘油三酯与桑蚕蛹油质量比1∶ 4,加酶量为底物质量的8%,反应温度45 ℃,反应时间10 h。在最佳反应条件下,酯交换反应的转化率为98.42%,酯交换产物中中长链脂肪酸甘油三酯含量为98.73%,辛酸含量为20.00%,α-亚麻酸含量为30.09%。     

Lipozyme RM IM脂肪酶催化酸解制备MLM型结构脂质

采用脂肪酶Lipozyme RM IM催化辛酸、癸酸与大豆油进行酸解反应以制备MLM型结构脂质。通过单因素实验研究底物摩尔比、辛癸酸摩尔比、酶添加量、反应时间、反应温度和初始水分含量对酸解反应的影响。得到适合的反应条件为:底物摩尔比3∶1(总脂肪酸/大豆油),辛酸与癸酸摩尔比(辛酸/癸酸)2.5∶1,酶添加量7.5 wt%(基于底物总重),反应时间5 h,反应温度65℃,加水量1.0 wt%(基于底物总重),得到MLM结构脂脂肪酸组成中辛酸含量为20.0 wt%,癸酸含量为10.5 wt%,两者质量比为1.92。     

脂肪酶改良猪油制备功能性脂的研究

我国有丰富廉价的动植物油资源，但这一资源并末得到有效的利用，为了充分利用这些资源，开展了脂肪酶催化猪油与辛酸酸解制备功能性脂的研究工作。脂肪酶筛选实验表明，在所选用的五种脂肪酶中，来自T.languginosa的同定化脂肪酶Liopzyme TL IM的催化效果最好。以Lipozyme TL IM为催化剂，进一步研究了酶量、有机溶剂、底物比率、反心时间和反应温度对猪油中辛酸插入率的影响。反应产物通过高效液相色谱法（HPLC）进行分析。研究结果表明，在正己烷介质中，脂肪酶量为15％（底物重量厅分比），底物比率为1：2（猪油：辛酸）反应时间24h，反应温度为55～60℃时，辛酸插入率最高。     

MLM型结构脂的酶法合成及其理化性质分析

以醇解反应制得单甘酯和辛酸为底物,1,3-特异性固定化脂肪酶为催化用酶,在单因素试验的基础上,通过正交试验获得酯化反应制备MLM型结构脂质的最适条件：Lipozyme RM IM酶加入量9%(以底物质量计)、反应时间14h、反应温度40℃、单甘酯与辛酸(底物)物质的量比1:6;该条件下辛酸插入率达60.48%,其中92.84%的辛酸分布在甘油结构的1,3位上。经二级分子蒸馏纯化后得到的MLM型结构脂质,辛酸插入率达到73.34%;对其进行理化指标检测分析表明,该产品最低可达国家四级菜籽油的标准。     

单辛酸甘油酯的酶法合成

以Novo435固定化脂肪酶为催化剂,在无溶剂条件下催化甘油和辛酸合成单辛酸甘油酯。通过单因素试验确定加酶量、反应温度、反应时间三个因素的取值范围,并用响应面实验设计和分析方法对合成反应条件进行了优化。结果表明,在甘油辛酸摩尔比1:1、反应温度66.8℃、脂肪酶与反应底物质量比0.88%、无水的条件下反应11.5h,辛酸转化率达93.53%,单辛酸甘油酯含量为47.69%。     

酶催化菜籽油酸解制备结构脂质工艺

以菜籽油和辛酸为原料,用来自Thermomyces lanuginosa固定化脂肪酶TL IM作为催化剂,采用单因素试验结合响应曲面(Box-Behnken设计)优化得到酶法制备结构脂质的工艺条件,最佳制备条件为底物(菜籽油:辛酸)物质的量比1:3.57、加酶量10.57%(以底物质量计)、加水量5.2%(以酶质量计)、反应温度48.6℃、反应时间12h。该条件下辛酸插入率可达23.19%。对提高油菜籽油附加值、延长油菜加工产业链具有重要意义。     

无溶剂体系酶法催化酸解合成共轭亚油酸甘油酯

采用商业化固定化酶Novozym 435作为生物催化剂,催化共轭亚油酸(CLA)和葵花籽油的酸解反应合成富含CLA的结构脂质(CLA-SL).研究了在无溶剂体系中,底物摩尔比、酶用量、体系含水量、反应温度和反应时间对产物中CLA含量和Sn-2位CLA含量的影响.结果表明,最佳反应条件为:CLA与葵花籽油摩尔比3 :1,酶用量10%,体系含水量1%,反应温度55 ℃,反应时间36 h.在最佳反应条件下,产物中的CLA含量和Sn-2位CLA含量分别为15.7%和2.73%.     

LIPASE-CATALYZED PRODUCTION OF STRUCTURED LIPIDS VIA ACIDOLYSIS OF FISH OIL WITH CAPRYLIC ACID

Structured lipids containing eicosapentaenoic and docosahexaenoic acids were manufactured in a batch reactor by lipase-catalyzed acidolysis of fish oil with caprylic acid. The following free lipases (Lipase AP, Aspergillus niger ; Lipase P, Pseudomonus sp. ; Lipase AY, Candida rugosa ; Lipase AK, Pseudomonas fluoresescens ; Lipase F, Rhizopus oryzae ; Lipase D, Rhizopus delemar ) were screened under selected reaction conditions. The conditions were enzyme load 5%, substrate mole ratio 1:6 (fish oil: caprylic acid), and reaction temperature of 50C. Lipase AK had the highest activity and was suitable for production of structured lipids from fish oil. The optimal mole substrate ratio of fish oil to caprylic acid for Lipase AK was 1:6 to 1:8. The time course of the reaction at different enzyme loads demonstrated that 40% incorporation of caprylic acid could be obtained for Lipase AK in 5 h with 10% enzyme load. Addition of water had little effect on the activity of the lipase. Lipase AK and Lipozyme IM were further compared under the same conditions, in which Lipase AK had a slightly higher incorporation of caprylic acid, similar acyl migration of caprylic acid from sn-1,3 positions to the sn-2 position, and a slightly lower selectivity towards docosahexaenoic acid.     

Lipase-catalysed acidolysis of lard with caprylic acid to produce structured lipid

Enzymatic acidolysis of lard with caprylic acid was investigated. Of the five lipases that were tested in the initial screening, immobilised lipase TL IM from Thermomyces lanuginosus resulted in the highest incorporation of caprylic acid into lard. This enzyme was further studied for the effect of enzyme load, organic solvent, substrate ratio, reaction time and temperature. HPLC was used to analyse the products from the acidolysis reaction. The highest incorporation was attained at 15% enzyme load. Among the solvents tested, n‐hexane was the best reaction medium for the acidolysis of lard with caprylic acid. Time course studied suggests that the incorporation of caprylic acid into lard was increased up to 37.7 mol% after 24 h. Desirable mole ratio of lard to caprylic acid was 1:2, caprylic acid incorporation up to 34.2 mol%. Temperature had no significant effect on enzyme activity in the range of 40–80 °C.     

酶法甘油解制备甘油二酯的研究

以精炼菜籽油为底物,通过酶法甘油解制备甘油二酯,比较了3种常用固定化脂肪酶甘油解制备甘油二酯的能力。结果表明:LipozymeRM IM具有较好的甘油耐受性,采用甘油预吸附的方式进行甘油解反应,可明显减少反应中酶活损失,显著提高酶的重复使用寿命。在菜籽油与甘油摩尔比1∶1,酶添加量为油质量的5%,硅胶与甘油质量比1∶1,反应温度60℃的优化条件下,甘油解反应8 h后,产物中的甘油二酯含量达到57.5%。通过硅胶预吸附甘油可以使LipozymeRM IM酶的多批次操作稳定性得到很大提高,半衰期达到22次,有应用于工业生产的潜力。     

PREPARATION OF STRUCTURED LIPIDS WITH SPECIAL FUNCTIONAL PROPERTIES

Enzymatic acidolysis of rapeseed oil with capric acid was carried out to obtain structured lipids. The reaction was catalyzed by Lipozyme IM lipase from Rhizomucor miehei. The enzyme preparations contained 2.8 and 10% water. The reaction conditions were enzyme load of 8% (w/w total substrates), substrate mole ratio of 1:6 (rapeseed oil:capric acid), and reaction temperature of 65C. The results showed that triacylglycerols (TAG) after transesterification contained mainly oleic, linoleic and linolenic acids (about 90%) in the internal sn-2 position, whereas capric acid was mostly in the external sn-1,3 positions (approximately 40%). The quantity of water in the reaction medium had a significant influence on the yield and quality of the TAG fraction.     

Production of olive oil enriched with medium chain fatty acids catalysed by commercial immobilised lipases

Structured triacylglycerols, containing medium chain fatty acids, were produced by acidolysis of virgin olive oil with caprylic or capric acid, at a molar ratio of olive oil:fatty acid of 1:2, at 45 °C for 24 h, in solvent-free media or in n-hexane, catalysed by Thermomyces lanuginosa (Lipozyme TL IM), Rhizomucor miehei (Lipozyme RM IM) and Candida antarctica (Novozym 435) immobilised lipases. Incorporations were always greater for capric than for caprylic acid. For both acids, higher incorporations were always attained in solvent-free media: the highest caprylic acid incorporations were obtained with Novozym 435 (25.5 mol%) and Lipozyme RM IM (25.7 mol%), while similar capric acid incorporations were obtained with all biocatalysts (27.1–30.4 mol%).     

Lipase-catalyzed acidolysis of olive oil and caprylic acid in a bench-scale packed bed bioreactor

Lipase-catalyzed acidolysis of olive oil and caprylic acid was performed in a bench-scale packed bed bioreactor to produce structured lipids (SL). A 1,3-specific lipase, IM 60 from Rhizomucor miehei was used as the biocatalyst. Reaction products were analyzed by reversed phase high-performance liquid chromatography, with an evaporative light scattering detector. Olive oil is characterized by four major clusters of triacylglycerol species with equivalent carbon number (ECN), C44, C46, C48, and C50. Three monosubstituted products and two disubstituted products were detected after the reaction. Monosubstituted products had ECN of C36, C38, and C40, and disubstituted products had ECN of C30 and C32. The effect of solvent, temperature, substrate mol ratio, and flow rate/residence time were studied. Optimal solvent-free production of SL was obtained at a substrate flow rate of 1 ml/min, residence time 2.7 h, temperature 60°C, and mol ratio 1:5 (olive oil/caprylic acid). Fatty acid distribution at the sn-2 position of olive oil was determined by pancreatic lipase hydrolysis as 74.8% oleic acid and 25.2% linoleic acid. SL produced at optimal conditions had 7.2% caprylic acid, 69.6% oleic acid, 21.7% linoleic acid and 1.5% palmitic acid at the sn-2 position.     

固定化脂酶催化合成生物柴油的研究

探讨了酶法制备生物柴油的过程,通过脂肪酶酯化和醇解两种工艺路线合成生物柴油的试验研究,考察了反应条件如醇油比、催化剂用量、反应温度、反应时间等对酯化率和产品纯度的影响.试验表明,采用分批加入甲醇的酯化工艺,酯化率可以达到95%以上;采用醇解工艺菜籽油酯化率达95%以上,产品经GC分析其纯度可达98%以上,固定化酶的半衰期至少达100 d.