Enzymatic Preparation Of Human Milk Fat Substitutes Rich In Medium And Long Chain Triacylglycerols

Medium and long chain triacylglycerols(MLCT)are a class of structured lipids containing both medium and long chain fatty acids on one glycerol backbone,accounting for about 20% of total triacylglycerols in human milk fat.MLCT have beneficial effects on health and nutrition,however,the molecular structure of which in human milk fat is different from vegetable oils or mammalian milk fat.In addition,the infant formulas almost do not contain MLCT similar to human milk fat,and there were fews studies on the synthesis of MLCT resemble human milk fat.Therefore,it is possible to synthesize MLCT mimic human milk fat to make infant formulas more similar to human milk.However,it is difficult to prepare MLCT by one-step method for its composition is complicated in human milk fat.Therefore,the objective of this study was to prepare human milk fat substitutes rich in medium and long chain triacylglycerols by two-step method.The major contents were as follows:Firstly,58oC palm stearin was fractioned in solvent to obtain palm stearin with total and sn-2 palmitic acid content of 91.57% and 91.96%,respectively.Then,three sn-1,3 position specific lipases were screened for enzymatic acidolysis between fractionated palm stearin,oleic acid and linoleic acid in a solvent-free system.The results showed that there was a significant difference in the total yields of OPO and OPL in the products when catalyzed by different lipases.The catalytic effect of lipase was as follows: NS40086>Lipozyme RM IM >Lipozyme TL IM.In order to obtain high total yields of OPO and OPL,lipase NS40086 was selected for the subsequent reaction.Subsequently,the effects of substrate molar ratio,enzyme load,temperature,and reaction time on the yields of OPO and OPL,palmitic acid content at the sn-2 position,and the relative percentage of palmitic acid at the sn-2 position in relation to total palmitic acid content were investigated.Under optimal conditions(palm stearin/oleic acid/linoleic acid molar ratio of 1:8:4,NS40086 load of 8%,60oC,4 h),the yield of OPO was 39.20%,the yield of OPL was 30.06%,the total yields of OPO and OPL were 69.26%,palmitic acid content at the sn-2 position was 87.75%,and the relative percentage of palmitic acid at the sn-2 position was 71.07%.Next,lipase NS40086 was used for the interesterification of coconut oil and structured lipids rich in OPO and OPL.The effects of reaction system,substrate molar ratio(OPO+OPL/coconut oil),reaction temperature,enzyme load on the yield of MLCT were investigated.The optimal reaction conditions were as follows: substrate molar ratio,0.8:1;reaction temperature,65oC;enzyme load,8%;reaction time,6 h.The content of MLCT in the product was 67.37% when the reaction was magnified 20 times under optimal conditions.The composition of the triacylglycerols of interesterified product analyzed by UPLC-Q-TOF-MS showed significant difference when compared with raw oils.MLCT similar to human milk fat were produced by interesterification,in which OPLa,LPLa,OLaL,OLaO,OPCa,OMLa,LLLa,OLaLa,LLaLa were all high-content in human mik fat.Finally,the products were purified by molecular distillation and the effect of the evaporation surface temperature on the composition of heavy phase was investigated.The optimal conditions were as follows: preheating temperature,65oC;evaporation surface temperature,210oC;condenser temperature,30oC;pressure,1Pa;scraper speed,120 r/min;feed rate,2.5 mL/min.After molecular distillation,the content of triacylglycerols in the product could reach 97.22%,the content of MLCT was increased to 74.82%.UPLC-Q-TOF-MS was used to analyze the composition of triacylglycerols and the results showed that the content of OPLa,LPLa,OLaL,OLaO,OMLa,LLLa,OLaLa and LLaLa were all increased in different degree.Human milk fat substitutes rich in medium and long chain triacylglycerols were obtained by two-step enzymatic method and purified by molecular distillation in this study,in which OPLa,LPLa,OLaL,OLaO,OPCa,OMLa,LLLa,OLaLa,LLaLa were all high-content in human mik fat.The lipids imitated the composition of MLCT in human milk fat,which may have potential to be applied in infant formulas to mimic the chemical composition of human milk fat.