Enzymatic Synthesis And Surfactant Performance Characterization Of Different Fatty Acids Phosphatidylglycerol

Phosphatidylglycerol(PG)as an important component of pulmonary surfactant,which can reduce pulmonary surface tension and maintain the structure and function of alveolar stability,it has a great potential application in the diagnosis and treatment of pulmonary diseases,for instance,neonatal respiratory distress syndrome.As a natural surfactant,PG also has a potential application in the field of tissue antifreeze and skin care.The natural source of PG is the same as other phospholipid components,their structures are characterized with different fatty acids.In this paper,transphosphatidylation was carried out for synthesizing different fatty acids phosphatidylglycerol(DFA-PG)from easily obtained soybean phosphatidylcholine(PC)with the help of phospholipase D(PLD)in the presence of glycerol(GI).The reaction conditions were optimized to improve the yield of the product,and the process of separation and purification of DFA-PG by silica gel column chromatography were researched;The fatty acid composition of DFA-PG was characterized,the surfactant performance and aggregation behavior were also characterized,which provide a reference for industrial production and application.A single factor experiment was used to optimize the transphosphatidylation for the preparation of DFA-PG in the bi-liquid system catalyzed by PLD,and the appropriate reaction conditions were obtained.The organic solvent was dichloromethane,the reaction temperature 28?,the molar ratio of glycerol to PC was 190:1,pH 5.5,the volume ratio of organic phase to aqueous phase was 2.5:1.Under these conditions,the yield of DFA-PG was 85.59%.On this basis,uniform design method was used to design the experiment to investigate the interaction between the factors,and the reaction conditions were further optimized by multiple nonlinear regression and response surface analysis:temperature 30.5?,the molar ratio of glycerol to PC 190:1,pH 5.5,the volume ratio of organic phase to aqueous phase 1.5:1.Under these conditions,the yield of DFA-PG reached 97.54%.Enzymatic synthesis of crude DFA-PG was purified by column chromatography with 100?200 silica gel as stationary phase.Optimization of column chromatography conditions at room temperature and atmospheric pressure,results showed that in the eluent composition of chloroform:methanol 2:1(v/v),the loading volume 0.03 g/g-silica gel,flow rate 3 mL/min,water content in silica gel 9%(wt),DFA-PG purity was obtained 97.6%,recovery was reached 90.5%.At the same time,PC product with a purity of 98.2%can be obtained.The fatty acids of DFA-PG and PC were analyzed by gas chromatography mass spectrometry(GC-MS).The results showed that the products and raw materials all contained 5 kinds of fatty acids,which with the same type and content.Surfactant performance characterization showed that the critical micelle concentrations(CMC)of DFA-PG and the purified raw material PC were 0.4904 mg/mL and 1.4673 mg/mL respectively,which were measured by Wilhelmy plate method,their surface tension of aqueous solution were 35.3 mN/m and 40.1 mN/m respectively,all these showed that DFA-PG has better capability and efficiency than PC in reducing water surface tension;Transmission electron microscopy(TEM)test showed that the aggregation state of DFA-PG changed with pH,and when pH was 7.4,which was in a stable vesicular structure with an average diameter of 131.19 nm;when pH was 6.9,the aggregation state was a rod-shaped micelle structure with an average diameter of 88.23 nm;when pH was 6.4,the aggregation state was spherical micelles with an average diameter of 25.98 nm.