The Studies On Liquid-state Fermentation, Immobilization And Application Of Fungal Lipase

The properties,production and immobilization of alkaline lipase,and its applications in production of biodiesel were discussed in this paper.1.The main factors affecting the alkaline lipase production by marine fungus H-19 were studied.The results showed that the optimal fermentation medium for lipase production was:Glucose 1.0%,soybean oil 1.0%,peptone1.0%,(NH₄)₂SO₄ 0.5%,NaCl 2%,KCl0.1%,MgCl₂ 0.5%,MgSO₄·7H₂O 0.2%,CaCl₂ 0.05%.The optimal initial pH were 8.0.The centigrade was 28℃and the speed was 180rpm.Under the optimised conditions,the yield of lipase activity reached 17.43U/mL after 48h.The crude enzyme of lipase from H-19 was purified using saluration of ammonium sulfate,dialyzing,freeze concentrating and sephadex G-100.The purification fold was 10,and the specific activity was 1.1U/mg.The optimum reaction temperature and pH of the Alkaline lipase was 36℃and 9.4 respectively.The temperature to lose 50%activity of the enzyme was 60℃.2.Two methods of immobilization were used for lipase physical adsorption on unmodified copolymers and covalent attachment on carriers with amino groups.The maximum yield of immobilized enzyme was 90%by covalent attachment,and 73%by physical adsorption.And immobilization by covalent bonding on silica and adsorption on macroporous resin all afforded 25-30u/g.The immobilized enzyme displayed excellent stability in the pH range of 8.6-10 and even at higher temperature,i.e,50-60℃,compared to free enzyme,which is unstable under extreme conditions.However,physically immobilized lipase retained only 15%of the initial activity after 10-baths feedback.Covalent attachment gave stable preparations,retaining 50%of the initial activity after 10-baths feedback.3.The effects of factors on production of biodiesel have been studied by immobilized Penicilium lipase in solvent,such as the amounts of lipase added,solvents, substrate concentration ration,water et.al.After 9h,esterification by immobilized Penicilium lipase was 80%in 20mL n-hexane with 60U of lipase,4%water,and 1:1.2 molar ration of methanol to fatty acids,and lipase inactivation is avoided by 3-step addition of methanol.