Study On Entrapment Of Candida Antarctic Lipase B In Sol-Gel Glass Matrix

In this work, the sol–gel method was used to immobilize CALB. We present how the mixed solution of liposome, made up of sugar as additive, lecithin and cholesterol, affects both the activity and stability of lipase CALB entrapped into tetraethyl orthosilicate (TEOS) derived silica.Firstly, a simple and rapid test used for the selection of active hydrolases. The hydrolytic activity of free and immobilized lipase were usually investigated by the olive oil emulsion method and the liberated fatty acids were titrated with alkaline solution at the presence of phenolphthalein as an indicator. The phenolphthalein in emulsion turns blurry red and the result was accordingly imprecise. Herein, the application of the pH-stat titration on the determination of lipase activity, which can determines the oleic acid released in the enzymatic hydrolyzation of the ethyl oleate emulsion for the tests of lipase.Secondly,the conditions effecting lipase CALB active, sach as: water content in lipase solution, pH value of buffer, PEG400 content in sol-gel solution and lipase content in immobilized lipase, were studied in this work. Two ways for protecting CALB molecule during sol-gel process were applied. One, different sugars were selected to be protecting additives in lipase solution. Another, liposomes could be as protective capsules for active silica sol-gel biocomposite. The lipase solution was dispersed among decane solution with lecithin and cholesterol, and then were entrapped in sol-gel matrixes together. Furthermore, the xylose content, water content, ratio of lecithin and cholesterol, cholesterol content and lipase solution content were determined with cross-over method. And the best result is xylose=10mg,water=60μL,ratio=1:4,cholesterol=1.5% and lipase=60μL. The mixture was kept under static conditions for 48h to complete the chemical condensation and obtained immobilized lipase about in 0.2g.Thirdly, the character of CALB entrapped in sol-gel was analysed by fourier transform infrared spectroscopy, differential scanning calorimetry, scanning electron microscopy and laser granularity scanning. Furthermore, the compare between immobilized CALB with N435 in stability was carried. As a result, the immobilized CALB was better.Finally, the thorough study was carried by mathematic method. A kinetics of immobilized lipase-catalyzed esterification of lactic acid and ethanol have been investigated with Ping-Pong Bi-Bi mechanism. And the result was well validated by experiments.