Abstract
Ascorbic acid (AA) plays an important role in physiological reactions and serves as a biological antioxidant. It possesses various functions in immune system modulation, regulation of collagen synthesis, and nitrous acid neutralization. Recently, it is confirmed that AA also plays a beneficial role against skin aging and shows anticancer activity. Therefore, it has the potential for uses in medicine and in cosmetic industry. However, ascorbic acid easily undergoes oxidation to an inactive compound under oxidative conditions such as heat, light, presence of transition metals and enzymes such as oxidases and consequently loses its anti-oxidative activity. Ascorbic acid can be converted into stable derivatives by structurally modifying it. Enzyme-mediated structural modifications are prefered over chemical processes.Several studies conducted in the past revealed that glycosyltransferase, an enzyme that modifies the structure of ascorbic acid to a more stable ascorbic acid glucoside, can be obtained from several microorganisms. A mold, Aspergillus niger, is used as a biocatalyst in this study for the transglucosylation of ascorbic acid to ascorbic acid glucoside.Through the design of medium by Response Surface Methodology (RSM), enzyme activity was increased by 230 U/ g-pellet compared to that using the unoptimized medium, using pellets from 2 days culture at 30oC and 200 rpm. It was observed that by using yeast extract, tryptone or soytone as nitrogen sources resulted in compact pellets than by using rice bran or corn steep liquor. Optimal cultivation medium contained: rice bran (1.8 %), starch (4.0 %) and K2HPO4 (0.05 %).Glucosyltransferase and amyloglucosidase were identified to be the two key enzymes for the biosynthesis of ascorbic acid glucoside. 89%-93% conversion was obtained at 30oC and 200 rpm after 6 h of reaction using maltose (0.55 M) as sugar donor and ascorbic acid (0.14 M). The maximum velocity (Vm) and Michaelis constant (Km) for the transglucosylation were 0.99 U and 0.19 M, respectively. The activation energy (Ea) was 1.97 cal /g-mole