Enzymatic Synthesis And Purifieation Of Dicaprylin Glycerol

Diacylglycerol (DAG) has been widely used in food, cosmetics and pharmaceuticals. Recently, it has been found that DAG have beneficial effects on decreasing postprandial blood lipid level, preventing the accumulation of visceral fat and body weight gain. However, the diacylglycerol content is very low in natural oils, which limits its wide application. Thus, the artificial synthesis of diacylglycerol has attracted increasingly attention at home and abroad. Two strategies are mainly employed to synthesis diacylglycerol, such as chemical and enzymatic method. Compared to chemical catalysts, enzymes have many advantages, of relatively mild reaction conditions, low energy consumption, low pollution dischargement, and positional and enantio selectivity. Moreover, the enzymatic synthsis produced less by-products, which results in higher purity DAG. The main topics of this work are as follows:1. From three different kinds of immobilized lipases, Novozyme 435, Lipozyme RM IM and Lipozyme TL IM, Novozyme 435 was selected as the best one to catalyze esterification reaction of glycerol and caprylic acid.2. Enzymatic esterification of glycerol and caprylic acid for diacylglycerol prodution was investigated by Novozyme 435 in the free system. In the single factorial experients, severoy crucial paramerers were examined, such as dehydration methods, substrate molar ratio, reaction temperature, the enzyme loading, moosture content on caprylic acid conversion rate and deglyceride yield. Based on the results of single factorial experiments, Response Surface Method (RSM) was employed to further optimize the reaction conditions and the quadric regression equation on the yield of deglyceride was established. The optimal conditions were: the molar ratio of caprylic acid to glycerol 1.99:1, reaction temperature 51.5℃, enzyme loading 3.05 % and rection time 6.29 h. Under the optimal conditions, the measured DAG yield was up to 71.9 %, which coincides with the predicted yield of 71.3±0.88 %. It is revealed that the quadric regression equation is reasonable and reliable.3. Silica gel column chromatography was used to purify dicaprylin glycerol. Six eluant systerms(ether chloroform and hexyl acetate, chloroform methanol, hexane and acetic acid, petroleum ether and butanol, petroleum ether and isobutyl alcohol, petroleum ether and absolute ether) were screened for purification. The result showed that petroleum ether and absolute ether was preliminarily selected as the best eluant system. Subsequently, the effects of eluant volume ratio, flow rate, sample load on the purity and the recovery of dicaprylin glycerol were optimized. The optimal conditions were: flow rate 2.5 ml/min, sample loading 4 g, eluted successively by 4.5:1, and 0.5:1(V/V) mixture of petroleum ether and absolute ether solution with discrete method of gradient elution. Under the above optimal conditions, the purity of dicaprylin glycerol and the recovery of dicaprylin glycerol were 94 % and 90 %, respectively. The silica gel column chromatography show high operational stability. After 5 cycles repeated usage, the purity and the recovery of the product respetively remained above 90 % and 85 %, respectively.