Screening And Immobilization Of Lipase-producing Strain, And Its Catalytic Activity In Ionic Liquid

Biodiesel, as a renewable and green fuel, is made from animal fats and vegetable oils. Enzymatic process for biodiesel producing, with advantage of mild reaction conditions, easy recovery of products and low chemical wastes, has a promising perspective and become a hot topic at home and abroad. However, there are several problems in the enzymatic reaction, such as high cost of biocatalysts, low activity and stability of enzyme, and poor solubility of methanol in oil. To solve these problems, screening of lipase-producing strain and optimizing of fermentation conditions had been down. The cell catalyst immobilized was used to catalyze the transesterification of soybean oil and methanol to produce biodiesel in the presence of ionic liquids. Main contents and results of this thesis are as follows:(1) Screening and identifieation of lipase-produeing strains. A lipase-producing strain is isolated from oil soil on olein assimilation plates, which contained bromcresol purple as indicator. According to the microbial taxonomy, the strain isolated was identified as Penicillium and named Penicillium camembertii Thom NS27. The initial activities of hydrolytic lipase and synthetic lipase was 76 U/g dry cell and 0.41 U/g dry cell, respectively.(2) Fermentation conditions optimization and immobilization. Through optimizating the culture media and fermentation conditions, the optimum lipase-producing media of Penicillium camembertii Thom NS27 was:1.0% tragantine,0.5% oleic acid,1.5% peptone,0.6% ammonium chloride,0.1% K2HP04 and 0.1% MgSO4·7H2O. The lipase activity could reach 335 U/g dry cell under the optimum conditions. Oleic acid can be used as carbon source and inducer. The strain cells were immobilized in polyurethane foam particles. From the SEM pictures, it could be seen that the growth and immobilization of strain Penicillium camembertii Thom NS27 in supporter was very well and stability. After been washing five times, the polyurethane foam particles reached constant weight, and there were 57.9% of strain cells stabilized in it.(3) Synthesis and screening of ionic liquids. Nine kinds of target ionic liquids were synthesised by microwave radiation, and the suitable ionic liquid [Bmim]PF6 for transesterification of soybean oil and methanol to produce biodiese has been abtained. The results of optimization of [Bmim]Br preparation conditions showed that the yield of intermediate [Bmim]Br reached 95.16% under the amount ratio of 1-butyl bromide to N-methylimidazole of 1.1, the microwave power of 200W and the intermission irradiation time of 100 s. The tolerance of immobilized cell in [Bmim]PF6 were investigated, the results showed that the remaining activity of cell catalyst reached 60% after being cultured at 35℃for 24 h.(4) The immobilized cell lipase catalyst of Penicillium camembertii Thom NS27 used for catalyzing the transesterification of soybean oil and methanol to produce biodiesel was studied in the presence of ionic liquids [Bmim]PF6. The conversion ratio of soybean oil could reach 62% under the conditions that the molar ratio of methanol/oil was 4, reaction time was 40 h, temperature was 40℃, water content was 4% and shaking table was 160rpm. When the immobilized cell particles and [Bmim]PF6 were used four times repetitivly, the conversion ratio of soybean oil and the remaining enzyme activity were 43% and 63%, respectively. It demonstrated that the experimental system of catalysis reaction was stability.