关键词: 云南莴笋尖, 酪氨酸酶, 分离纯化, 酶学性质

Abstract: To find a new cheap and easily available source of tyrosinase and to provide an experimental basis for further research on inhibitors of tyrosinase from lettuce tip, electrophoretically pure tyrosinase was obtained from Yunnan-grown lettuce tip through consecutive homogenization, buffer extraction, ammonium sulfate fractionation, DEAE-Sepharose fast flow chromatography and Superdex-200 prep grade chromatography. The results showed that the specificity activity of the purified enzyme was 20.62 U/mg. The relative molecular weight of the tyrosinase was approximately 995.40 kD, in which the subunit molecular mass was roughly 65.23 kD. The enzymatic properties showed that the optimum temperature and pH for the tyrosinase were 60 ℃ and 8.0, respectively. The enzyme was stable in the pH range of 6.0–8.0 and in the temperature range of 20–40 ℃. Its apparent Km and vmax were 1.567 mmol/L and 0.037 6 μmol/(min·L), respectively. The tyrosinase activity was slightly affected by EDTA, urea, K+, Mg2+and Li+. The enzyme activity could be inhibited by ethanol, isopropanol, n-butanol, but activated by methanol, Co2+, Mn2+, Pb2+, Ag+, Cd2+, Ba2+, and Ca2+, and it could be obviously activated by Co2+, Mn2+. Ascorbic acid, Zn2+, Cu2+, and oxalic acid enhanced the enzyme activity at low concentrations but inhibited it at high concentrations.

Key words: Yunnan-grown lettuce tip, tyrosinase, isolation and purification, enzymatic properties