重组白地霉脂肪酶催化菜籽油水解制备α-亚麻酸工艺的研究

The Preparation Procedure of α-Lionlenic Acid by Hydrolysis of Rapeseed Oil with Recombinant Lipase from Geotrichum Candidum

利用毕赤酵母GS115表达白地霉的脂肪酶基因,以离子交换法纯化的重组脂肪酶催化菜籽油水解,确定了最佳水解条件为温度42℃,水油比为1.5∶1,p H 8.0,酶用量为250 U/g,反应18 h水解率可达到40.1%。以水解得到的混合脂肪酸为原料,建立一种尿素包合法富集α-亚麻酸的工艺。通过单因素试验对影响尿素包合富集效果的包合温度、时间、溶剂配比进行研究,由正交试验得到包合过程的最佳工艺条件为m(脂肪酸)∶m(尿素)∶V(乙醇)为1∶4∶20,包合温度-20℃、包合时间14 h,此条件可使其中α-亚麻酸的含量由8.03%提高至19.24%。

The gene coding the lipase of Geotrichum candidum was expressed in Pichia pastoris GS115. The hy- drolysis of rapeseed oil was catalyzed with recombinant lipase purified by one single step of ion - exchange. The opti- mum enzymatic hydrolysis conditions were determined as follows ： temperature is 42 ℃, ratio of water to oil is 1.5： 1, with pH 8.0 and lipase dosage 250 U/g,the reaction time is 18 h and the hydrolysis ratio is 40.1%. The mix-fattyacids obtained by the hydrolysis were used as raw material and a new technology to purify α -linolenic acid is eatab- lished through urea inclusion method. The single factor experiment determined factors affecting the urea complex puri- fication of temperature, the reaction time, urea and fatty acids ratio and urea and ethanol ratio were studied. An or- thogonal experiment was applied to optimize the parameters in purifying α-linolenic acid by urea complexation. The optimal purification process conditions as follows ： fatty acid and urea ratio was 1 ： d （ m/m）, urea and ethanol ratio was 1 ： 5 （ m/V）, the reaction temperature is - 20 ℃, the reaction time was 14 h, and the relative content of et - linolenic acid could be raised from 8.03% to 19.24% under the conditions mentioned above.