Study On The Enzymatic Synthesis Of Ethyl Oleate By Lipex Lipase In CTAB Reverse Micellar System

As one of the important types of intermediate chemical, ethyl oleate is used as to be lubricant, water-resisting admixture, colophony intenerater, insecticide assistant. As Lipase catalyzed biotransformations are gaining importance because of their substrate specificity, their milder reaction conditions and the relatively lower energy requirement. The Lipases have been employed for direct esterification and transesterification reactions in reverse micellar system. It has been confirmed that lipase in the reverse micellar sysetem exihibited more active than that in organic solvents and water. Thus the synthesis of ethyl oleate by Lipex in CTAB reverse micellar was investigated.Firstly, ethyl oleate was synthesized by oleic acid and ethanol, It was confirmed that Lipex has a good activation in the CTAB reverse micellar system, and the best solvent is isooctane. The influences of organic solvent, ratio of substrates, pH value, water content W0, concentration of oleic, acidreaction time and additive were investigated. As a result, a more stable reverse micellar system was obtained, which led to a better activity of Lipex lipase. The optimization of the reaction was studied in detail, when the temperature is 25℃, the molar ratio of oleic acid to alcohol is 1:4, buffer pH is 7.0, W0 is 10, oleic acid concentration was 0.01 mol·L-1, PEG400 0.05g, agitation speed was 150 r·min-1 and reaction time was 36h, which brought the yield of 93%.Secondly, the kinetic model of the reverse micellar system influenced by double substrates was established for the first time. And the simulated result agreed well with the experimental result. The kinetic model eaquation is as followed:Thirdly, It shows that the activity of Lipex was effected by the particle size of reverse micellar, the suitable article size of CTAB reverse micellar is 150 nm. And there is a repulsion function between the cation of CTAB and the active of Lipex. The system added PEG400 is analasised by infrared, it shows that, when PEG400 was added to the system, the surfactant cation has a microscopic changes, stretching weakened. It's maybe the reason why the yield of ethyl oleate is improved. Lastly, Lipex was analysed by fluorescence and the software of ExPASy. It is summarized that the accession of PEG400 induce a structure changing of Lipex, and it is the conclusion that the activity of Lipex was effected by the structure of Lipex.