Immobilization of Lactobacillus acidophilus β-galactosidase on chitosan obtained from the shells of the African giant snail, Achatina achatina

High enzyme activity and reusability are the major factors that limit enzyme application in the industry. This study explored the properties of Lactobacillus acidophilus β-galactosidase immobilized on Achatina chitosan, for improved enzyme reusability in industry. β-Galactosidase was produced from environmentally well-adapted Lactobacillus acidophilus. The enzyme was purified by ion exchange chromatography using DEAE-cellulose and had a molecular weight of 43kDa. Mg2+ was a major positive effector of the β-galactosidase activity. Chitin was extracted from Achatina shells by demineralization and deproteination, and deacetylated to chitosan. The chitin and chitosan yields were 74.64% and 58.60%. However, a hypochlorite-decolorized chitin, deacetylated to chitosan, gave a yield of 71%. FTIR spectra of chitin showed major bands at 711 cm-1, 855 cm-1, 1082 cm-1, and 1438 cm-1 for chitin and for chitosan at 6778 cm-1, 711 cm-1, 851 cm-1, 1082 cm-1 and 1436 cm-1. The β-galactosidase was immobilized on chitosan beads by adsorption and covalent linkage using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS), and glutaraldehyde, separately. The enzyme had optimal temperature and pH of 70°C and 5.5. The Michaelis-Menten constant (KM) and maximal velocity (Vmax) of the free and immobilized β-galactosidase were 0.262/0.251mM, and 270.27/290 μmol/min using p-NPG as substrate, and 0.53/10.02 mM and 250/275 μmol/min for lactose as substrate, respectively. Covalent immobilization by glutaraldehyde improved the β-galactosidase activity more than adsorption, in comparison to EDC/NHS. The results show that extracellular β-galactosidase from Lactobacillus acidophilus, isolated from dairy wastewater, can be immobilized on chitosan support produced using cheaply available Achatina shell chitosan for greater reusability.