Modification Of Structural Lipids With Long Chain Unsaturated Fatty Acids As Sn-2 Acyl-site Donors And Their Effects On Oxidation Stability

The absorption effect of triglyceride with n-3 long chain polyunsaturated fatty acids?n-3 LC-PUFA?as acyl-site donors was better than that in sn-1,3 position and random distribution for whole natural triglyceride components.How to keep n-3LC-PUFA in the digestible acyl-site?sn-2?of triglycerides to play a better physical and chemical properties and physiological functions was not only a research focus of structural lipids,but also one of the key and difficult problems in the construction of long-chain polyunsaturated fatty acid structural lipids.Due to the high degree of unsaturation of the n-3 LC-PUFA linked to the sn-2 acyl-site of the structural lipids,it was easy to oxidize and deteriorate which resulting in the decline of its stability or the loss of nutrition.This one-way irreversible change has a significant impact on the sensory and safety of the food,and it will directly affect the stability of the physical and chemical properties of the structural lipid and its physiological function.Therefore,it was necessary to study the modification and oxidation characteristics of structural lipid using long-chain polyunsaturated fatty acids as sn-2 acyl-site donors.In this study,two-steps method was used to synthesize monoglyceride using long chain polyunsaturated fatty acid as sn-2 acyl-site donors?2D-MAG?and MLM structural triglyceride?1,3C-2D-TAG?.The synthesis conditions and influencing factors of monoglycerides and MLM structured lipids were also explored.The oxidation mechanism of gel made of 2D-MAG was further studied.Meanwhile,the intrinsic relationship and interaction mechanism between the physico-chemical properties,oxidative stability and structural characteristics of MLM structural lipid were also studied in this work.The six lipases were used to synthesize 2D-MAG by alcoholysis in this study.The sequence of enzyme activity was:lipozyme RM IM>Novozym 435>lipozyme TL IM>Newlase F>lipase AK>lipase AY.Compared with Novozym 435 and lipozyme TL IM,lipozyme RM IM has lower price and the strongest ability to catalyze 2D-MAG.The results of TLC and UPLC/MS showed that the purity of 2D-MAG was 88.5%,and¹³C NMR showed that 2D-MAG accorded with the structural characteristics of2-docosahexaenyl monoglyceride.When the molar ratio of algal oil to ethanol with 1:40,added 10%enzyme,3 hours reaction time,and 35?reaction temperature,the use of lipozyme RM IM to catalyze the alcoholysis of algal oil was more conducive to the accumulation of MAG synthesis products.In the mixed lipids with ratio of SFA,PUFA and MUFA approximate 1:1:1,the ratio of LC-PUFA as sn-2 acyl-site donors was significantly higher than that of other fatty acids in monoglyceride.The effects of different concentrations of monoglyceride on the thermodynamic properties,crystal morphology and oscillatory rheology of algae oleogel prepared with MAG were compared.The results showed that the prepared MAG could be used as a semi-solid gel for algae oleogel.The algae oleogel formed by high concentration of MAG had higher peak melting temperature?Tm,71.48??and melting enthalpy?Hm,190.2 J/g?.The oleogel melting temperature,melting enthalpy and gel strength decreased significantly with the decrease of MAG concentration.At lower MAG concentration?5%,10%?,the oleogel network structure was unstable,and free algal oil can not be effectively captured in oleogel network,which resulting in the separation of algae oil and the phenomenon of two-phase separation.In addition,the oleogel network adding 5%MAG present weaker intensity and larger crystallization.After increasing the concentration of MAG to 10 or 20%,the oleogel network was composed of smaller MAG crystals and algal oil.At this time,algal oil can be suspended in the network well,and the physical barrier formed outside can prevent the attack of oxygen molecules,which resulting in better oxidation stability of the gel.This mechanism played an important role in the application of MAG to high unsaturated fatty acid oleogel system.On the basis of 2D-MAG synthesis,the 1,3C-2D-TAG structured lipids with MLM type were further synthesized by two-steps enzymatic method with octanoic acid.The results of TLC and UPLC/MS showed that MLM type was the main structural lipids,which contained 93.82%octanoic acid at sn-1,3.The iodine value of the modified1,3C-2D-TAG is significantly lower than that of the algal oil and has higher oxidation stability.The calorific value of the modified 1,3C-2D-TAG?22.41%?lower than that of the algal oil.The esterified 1,3C-2D-TAG had not only the physical properties of the traditional oil but also the calorific value of the oil.Compared with 1,2,3C-TAG,the viscosity of 1,3C-2D-TAG increased with the increase of hydrogenation degree.In order to evaluate the effect of ultrasound/non-ultrasound on the synthesis rate of1,3C-2D-TAG structured lipids,the differences of synthesis rate of MLM structured lipids synthesized by enzymatic hydrolysis of sn-2 containing four fatty acids?C16:0,C18:0,C18:1 and C22:6?were also studied,and the effects of ultrasound conditions,reaction temperature,reaction time and substrate molar ratio on the synthesis rate of MLM structured lipids were also evaluated.Compared with the conventional enzymatic hydrolysis,the synthesis rate of structural lipids under ultrasound was higher than that under non-ultrasound conditions,and the proportion of sn-2 binding to C18:1 and C22:6increased by 33.2%and 42.4%respectively,which significantly improved the synthesis rate of sn-2 long chain polyunsaturated fatty acid monoglycerides to MLM type structural lipids.With the increase of ultrasound intensity,the synthesis rate of sn-2unsaturated fatty acid MLM type structural lipid increased significantly.The results showed that the enzyme substrate activity can be increased and the reaction rate can be accelerated with the increase of ultrasound intensity.When the lipase loading increased from 1:1 to 1:3,the C16:0 and C18:0 as sn-2 acyl-site donors increased from 66.4%and68.2%to 76.1%and 73.6%respectively.Further increasing the lipase loading,the synthesis rate of each structural lipid decreased,which indicating that the higher lipase ratio did not significantly improve the synthesis rate of MLM structural lipid.In general,ultrasound can effectively improve the formation rate of MLM type structured lipids synthesized by monoglycerides with monounsaturated fatty acids and polyunsaturated fatty acids as sn-2 acyl-site donors.Under the condition of accelerated oxidation at 50?,2D-MAG and 1,3C-DAG can significantly prolong the oxidation induction time of stripped soybean oil?SSO?.Compared with 2D-MAG,the oxidation rate of DHA at sn-2 position in 1,3C-2D-TAG was significantly faster than that in 2D-MAG,which indicating that the position of sn-2acyl group was not the main reason for the decrease of the oxidation stability of structured lipids.Compared with 1,2,3C-TAG,1,3C-2D-TAG can obviously induce the oxidation of structural lipids and promote the oxidation of SSO,which showed that the type of sn-2 acyl fatty acids?DHA?in structural triacylates had a great influence on the degree and speed of lipid oxidation.The oxidation induction period of 1,3C-2D-TAG was shorter in pure SSO system,which showed that 1,3C-2D-TAG can accelerate SSO oxidation and reduce its stability.Adding antioxidants to 1,3C-2D-TAG significantly prolonged the induction period of SSO oxidation.Compared with the pure SSO system,1,3C-2D-TAG needs to add antioxidants to avoid oxidation.In order to further discover the relationship between antioxidant and the content of DHA at sn-2,the content of DHA at sn-2 was compared during the oxidation period.The results showed that compared with 2D-MAG group,the oxidation rate of DHA as sn-2 acyl-site donors of1,3C-2D-TAG was significantly faster than that of 2D-MAG during the oxidation process.There are fatty acids on the three acyl-sites of 1,3C-2D-TAG,and DHA at sn-2acyl-site was not be transesterified,which affects the antioxidant's direct inhibition on the oxidation of 1,3C-2D-TAG.After adding 1,3C-2D-TAG to commercial coconut oil,5%level of 1,3C-2D-TAG can significantly shorten the induction period of coconut oil oxidation to 10.1 h.When the content of 1,3C-2D-TAG in coconut oil increases,the oxidation stability of commercial coconut oil continues to decline.The results showed that 1,3C-2D-TAG still needs antioxidant protection in the practical application.