Continuous production of biodiesel fuel from vegetable oil using immobilized Candida antarctica lipase

Candida antarctica lipase is inactivated in a mixture of vegetable oil and more than 1:2 molar equivalent of methanol against the total fatty acids. W e have revealed that the inactivation was eliminated by three successive ad ditions of 1:3 molar equivalent of methanol and have developed a three-step methanolysis by which over 95% of the oil triacylglycerols (TAG) were conv erted to their corresponding methyl esters (ME). In this study, the lipase was-not inactivated even though 2:3 molar equivalent of methanol was presen t in a mixture of acylglycerols (AG) and 33% ME (AG/ME33). This finding led to a two-step methanolysis of the oil TAG: The first-step was conducted at 30 degrees C for 12 h with shaking ina mixture of the oil, 1:3 molar equiv alent of methanol, and 4% immobilized lipase; the second-step reaction was done for 24 h after adding 2:3 molar equivalent of methanol (36 h in total) . The two-step methanolysis achieved more than 95% of conversion. When two- step reaction was repeated by transferring the immobilized lipase to a fres h substrate mixture, the enzyme could be used 70 cycles (105 d) without any decrease in the conversion. From the viewpoint of the industrial productio n of biodiesel fuel production, the two-step reaction was conducted using a reactor with impeller. However, the enzyme carrier was easily destroyed, a nd the lipase could be used only several times. Thus, we attempted flow rea ction using a column packed with immobilized Candida lipase. Because the li pase packed in the column was drastically inactivated by feeding a mixture of AG/ME33 and 2:3 molar equivalent of methanol, three-step flow reaction w as performed using three columns packed with 3.0 g immobilized lipase. A mi xture of vegetable oil and 1:3 molar equivalent of methanol was fed into th e first column at a constant flow rate of 6.0 mL/h. The eluate and 1:3 mola r equivalent of methanol were mixed and then fed into the second column at the same flow rate. The final step reaction was done by feeding a mixture o f eluate from the second column and 1:3 molar equivalent of methanol at the same flow rate. The ME content in the final-step eluate reached 93%, and t he lipase could be used for 100 d without any decrease in the conversion.