Dynamic analysis of medium chain glycerides synthesis by using immobilized lipase in a solvent-free system

Abstract

Enzymatic synthesis of medium chain glycerides (MCGs) was studied by using capric acid (decanoic acid) and glycerol as the substrates for an immobilized lipase (Lipozyme). Quantitative analysis of the reaction mixture was conducted by using a reverse phase HPLC, which enabled the exact tracing of the glyceride synthesis. The esterification activity of Lipozyme was determined at 40℃ in an open batch reactor. The synthetic activity for the capric glyceride was about 400 units per gram of Lipozyme where one unit is defined as one unit is defined as one μ Imole of fatty acid consumed (transformed into glycerides) per minute, while the activity for the oleic glyceride was about 200 units per gram of Lipozyme. Maximum initial reaction rate was obtained at 50℃ for capric and 60℃ for oleic glyceride synthesis. The time courses of the capric glyceride synthesis were analyzed in terms of different molar ratios, from which we infer that this enzyme is 1,3-specific, but not absolute, in this esterification reaction. The final conversion was greatly influenced by various water removing methods, among which the cold trap method resulted in a noticeable improvement. Dehydrated Lipozyme was prepared by storing it in the $P_2O_5$ desuccator for 4 weeks at room temperature. When we compared the esterification activity of dehydrated Lipozyme with that of intact one, the dehydrated Lipozyme did not show any activity during the first 4 hrs, but after this period, reaction proceeded at a similar rate compared to intact one, which implies that the dehydrated one can retain original activity but a certain amount of water is required for its catalytic activity. For the study of reaction kinetics, dynamic analysis was conducted. As this lipase shows 1,3specificity, a simple consecutive reaction model was suggested and a set of linear differential equations was developed. From the time course of the closed batch reaction, the kinetic constants were estimated by solving this simultaneous ...