Separation, Characteristics And Immobilization Application Of Alliumin

The unique flavor of shallot was generated by the enzymatic reaction whose catalyst and substrate is alliinase and alliin. The study was using buffer extraction, disruption and filtration, ammonium sulfate fractionation, Sephadex G-75 separation to purified alliinase; the sodium alginate was used to immobilize alliinase to improve the stability and utilization of alliinase. The immobilized alliinase was used in the flavor precursors from shallot. The results show:1 ? The alliinase in shallot was purified by extraction with phosphate buffer(pH=7), broken, high speed refrigerated centrifugation, 30%~50% ammonium sulfate precipitation, using UF to desalinate, Sephadex G-75 chromatography. SDS-PAGE was used to verify the purity alliinase and the molecular weight of alliinase was 50.23 kDa.2?The pyruvic acid method was used to survey the enzyme kinetics and Characterization with natural alliin. The result show that the optimum reactive temperature of alliinase was 40? and the optimum reactive pH was 8. The relatively stable temperature of alliinase was 30?, and the pH=6~8 was more conducived to keep the alliinase activity. Alliinase optimum reaction time was 4.5min. Michaelis-Menten equation was y=1.85056x+0.9204, and the values of Km and Vmax were 2.01mmol/L and 1.09?mol. PLP, glycerol, Ca2+?Mg2+?K+?Na+ and Fe3+ were activated alliinase activity and the activation of PLP?Na+?Fe3+?K+ were more obvious3?Purificed alliinase was immobilized by embedded-crosslinking method with sodium alginate which was as immobilized carrier, adding glutaraldehyde as cross-linking agent. The best single factor was determined by single factor experiments in immobilization efficiency and relative activity as an index, and the optimum conditions of immobilization was determined by orthogonal experimental validation. The optimum conditions was that the concentration of sodium alginate was 2.5%, calcium chloride concentration of 4.0%, 0.75% glutaraldehyde, sodium alginate-alliinase volume ratio of 4:3. The immobilization efficiency was 94.05%. The inactivation of the enzyme was lower than the free state. 4?The volatile components of flavor precursors, flavor and flavor precursors adding with alliinase were analyzed by GC-MS. The result show that the flavor precursors was contained 32 kinds of volatile components and the relative content of the sulfide-containing accounted for 59.49% of the total; the flavor was contained 30 kinds of volatile components and the relative content of the sulfide-containing accounted for 69.98% of the total; the flavor precursors adding with alliinase was contained 31 kinds of volatile components and the relative content of the sulfide-containing accounted for 61.40% of the total. The types of sulfide-containing from flavor were the same as the sulfide-containing flavor precursors adding with alliinase...