Preparation Of Diacylglycerol By Hydrolysis Of Soybean Oil Catalyzed By Phospholipase A1

Diacylglycerols(DAGs) are esters of glycerol in which two of the hydroxyl group are esterified with fatty acids(FFA).They can exist in two different isomeric forms which are 1,2-DAG and 1,3-DAG..It occurs as a natural minor component of plant oils and the endogenetic intermediate metabolite of lipid.In 2000 DAG oil has received the status "Generally Recognized as Safe"(GRAS) by the U.S.Food and Drug Adminstration.In recent years,many researchers have shown that diacylglycerols can reduce visceral fats and flood lipid and suppress the increase of body weight.The triacylglycerol of the soybean oil hydrolyzed by the catalysis of Phospholipase A1 was studied.Parameters of the hydrolysis,such as reaction time, reaction temperature,the load of enzyme and the water amount were investigated.The optimal conditon of hydrolysis of soybean oil was obtained by the orthogonal experiment as the following:reaction temperature of 35℃,load of enzyme of 26u/g, water amount of 40%(wt%,of the oil),reaction time of 8h.Under this condition,the acid value of the oil layer of the hydrolyzed mixture was 58.07mg KOH/g.Soybean oil was hydrolyzed by catalysis of Phospholipase A1 to release diacylglycerol and free fatty acid.The molecular distillation was introduced to remove the free fatty acid from the crude diacylglycerol at the different temperatures. The results showed that the purified diacylglycerol with acid value of 0.77 mgKOH/g and iodine number of 138.20 gI₂/100g was obtained after molecular distillation twice of the hydrolysis mixture.The light phase of molecular distillation had the lower iodine number and higher content of saturated acids compared to the soybean oil.High-performance liquid chromatography(HPLC)was found to determine DAG.The content of glyceride in the soybean oil and its product obtained by molecular distillation after phospholipase A1 catalyzed hydrolysis was analyzed by high performance liquid chromatography(HPLC).The content of diacyglycerol in the hydrolyzed product was 42.64%detected by the Refractive Index detector.The composition of glyceide in the soybean oil and its hydrolyzed product was analyzed by HPLC-ESI(Electrospray ionization)-MS(mass spectrometry).The fatty acid composition of soybean oil,the light phase of molecular distillation and its hydrolyzed product by molecular distillation was determined by the gas chromatography.The results showed that,compared to the soybean oil,the content of palm acid was higher in the released fatty acids which was from hydrolysis of soybean oil catalyzed by phospholipase A1.The hydrolyzed product by molecular distillation contains undersaturated fatty acids,which are easy to suffer from oxidation when exposure to oxygen.The antioxidants of TBHQ and ascorbyl palmitate were used to test their preservative effect on the product by Rancimat method.In Rancimat method,50ppm,100ppm 150ppm and 200ppm with DAG product were kept at 110℃with air flux 10L/h,their inducing time were determined by a 743 Rancimat.The results showed that the optimum consentration of TBHQ was 200ppm and the inducing time of the DAG product was 9.62h.