Abstract
In this study, we make the favorable applications of chitosan in immobilization techniques. In enzymatic study, a method for immobilization of Candida rugosa lipase to two types of chitosan beads by activating the hydroxyl groups of chitosan using 1-Ethyl-3-(3-dimethyl-aminopropyl) carbodiimide hydrochloride (EDC) coupling agent has been successfully developed. The ability of EDC to activate the hydroxyl groups of chitosan was confirmed using the electron spectroscopy for chemical analysis (ESCA) technique. The properties of lipase immobilized using dry and wet chitosan beads were also investigated and compared. Immobilization enhanced the enzyme stability against changes of pH and temperature. High storage stability of 30 days and an increased enzyme activity of 2,110% were observed in wet immobilized lipase, which indicated that the enzyme had a high affinity for chitosan supports. Immobilized lipase using dry and wet chitosan beads retained 78 and 85 % of its initial activity after 10 batch hydrolytic cycles. It was found that the swelling of dry chitosan beads in aqueous medium lead to a decrease in lipase loading as well as stability. The kinetic parameters such as Michaelis-Menten constant (Km) and maximum velocity (Vm) were determined for the free and immobilized lipase. The activation energy (Ea) was found to decrease for immobilization of lipase on chitosan beads. On the other hand, immobilization of bata-cyclodextrin to chitosan (Ch-BCD) by chemical binding has been successfully developed and demonstrated that it possessed excellent capacity for cholesterol adsorption. The applicability of Langmuir and Freundlich equations to interpret the behavior of adsorption was studies. The experimental data on cholesterol adsorption fitted well in the Langmuir isotherm equation as compared with using Freundlich isotherm equation. The maximum amount of cholesterol adsorption determined using the Langmuir isotherm equation was 0.33 gram of cholesterol per gram of Ch-BCD which was the same as the experimental data. Cholesterol adsorption from yolk was also carried out with the Ch-BCD. Results indicated that 92% of cholesterol could be removed using 1% (w/v) Ch-BCD in 2 hours at 25oC. On the other hand, 96% of adsorbed cholesterol could be dissociated from the Ch-BCD by using 95% ethanol at 50oC. The adsorption of plasma cholesterol from human blood was also investigated. About 55% of total cholesterol from 20 ml of plasma could be removed in 30 minutes by using 0.5 gram of Ch-BCD.