Inhibition of mushroom tyrosinase by flavonoid from Sorbus tianschanica Ruper in Xinjiang

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1 Chinese Journal of Bioprocess Engineering Vol. 13 No. 4 Jul doi /j. issn L- 50% IC mg /ml K I - K IS mg /ml DPPH DPPH 50% IC mg /ml Q A Inhibition of mushroom tyrosinase by flavonoid from Sorbus tianschanica Ruper in Xinjiang FU Jianhong 1 QI Rui 2 ZHENG Jing 2 ZHOU Chenjie 2 CHEN Qingxi 2 1. Xinjiang Key Laboratory of Special Species Conservation and Regulatory Biology College of Life Science Xinjiang Normal University Urumqi China 2. School of Life Sciences Xiamen University Xiamen China Abstract Flavonoid was extracted from powder of Sorbus tianschanica Ruper stems under assistance of ultrasonic wave. Adopting enzymatic kinetic method inhibitory action of flavonoid extract on the tyrosinase activity was studied using L-DOPA as substrate. Results showed that flavonoid extract from Sorbus tianschanica Ruper inhibited tyrosinase. Its IC 50 was estimated as mg /ml. The inhibition mechanism was studied and it was found that the reaction was reversible and type of the inhibition belonged to the mixed-type. The inhibition equilibrium constants for inhibitor binding with free enzyme K I and with enzyme-substrate complex K IS were determined as and mg /ml respectively. The DPPH 1 1-diphenyl-2-picrylhydrazyl radical test results showed that flavonoid extract from Sorbus tianschanica Ruper had the antioxidant activity to a certain extent and the inhibitor concentration leading to 50% DPPH free radical activity loss IC 50 was determined to be mg /ml. The above study provides a basis for further development of the fruits and vegetables preservatives and whitening ingredient in the Sorbus tianschanica Ruper XJEDU2012I30 XJSD fjh_719@ 163. com

2 64 13 Keywords Sorbus tianschanica Ruper flavonoid tyrosinase inhibitory mechanism EC tyrosinase U /mg L- L- 3 4 DOPA DMSO Sigma 1. 2 KQ5200E SHB-Ⅲ 1 EYELA DU800 DKB-501A g 300 ml 60% 6 Kubo W min Xie 8 5% DMSO mg /ml μl Sorbus tianshanica Ruprer 3. 0 ml mg /ml A C mg 60% mg /ml ml 10 ml ml 60% 5% NaNO ml 6 min 10% Al NO ml 6 min

3 4 65 4% NaOH 4. 0 ml 60% ml 1. 0 ml 15 min510 nm 0. 1 mol /L ph ml A % 0. 1 ml 3 mmol /L DPPH ml 0 ~ mg /ml min 517 nm 12 V c DPPH 0. 5 mmol /L L-DOPA mol /L 14 ph = ml ml 2. 8 ml 0. 1 ml A 475 A ρ A = ε ε = L / mol cm ρ R = U L-DOPA ~ mg /ml 1 μmol 1 = V s /V c 100% 1 V c OD 475 min - 1 V s 6 g OD 475 min % 300 ml W L-DOPA min 0. 5 mmol /L 60% 510 nm 2. 75% 2. 2 L-DOPA L-DOPA 1 1 = 1 000A 475 /3. 7 Lineweaver- IC mg /ml Burk K m v max 2. 3 L-DOPA K I - K IS DPPH 2 2

4 66 13 Fig. 1 2 Fig L-DOPA Lineweaver-Burk Effect of flavonoid extract of Sorbus tianschanica 3 3 Ruper stems on activity of diphenolase of tyrosinase K m v max E - ES 3 b 3 c K I mg /ml - K IS mg /ml K I < K IS Determination of the inhibitory mechanism of - flavonoid extract from Sorbus tianschanica Ruper on diphenolase of tyrosinase Fig. 3 1 ~ mg /ml 3 Detemination of inhibition type and inhibition constant of flavonoid extract from Sorbus tianschanica Ruper on diphenolase of tyrosinase 2. 5 DPPH DPPH

5 4 67 DPPH EIS DPPH 4 4 DPPH IC mg /ml V c IC mg /ml 5 DPPH Fig. 4 Fig Effect of flavonoid extract from Sorbus tianschanica 3 Sánchez Ferrer A Rodríguez López J N García Cánovas F et al. Ruper on activity of antioxidation Tyrosinase a comprehensive review of its mechanism J. Biochim Biophysica Acta V c Effect of vitamin C on activity of antioxidation 1 10 J 60% W J min % IC mg /ml L-DOPA J. Bioorg Med Chem J Osvaldo Z Antonella B Patrizia C et al. Truffle-thio-flavours reversibly inhibit truffle tyrosinase J. FEMS Microbiol Lett 4 Martinez M V Whitaker J R. The biochemistry and control of enzymatic browning J. Trends Food Sci Technol J J Kubo I Kinst Hori I Isiguro K et al. Flavonols from H. inuloides. tyrosinase inhibitory activity and structural criteria J. Bioorg Med Chem Xie L P Chen Q X Huang H et al. Inhibitory effects of some flavonoids on the activity of mushroom tyrosinase J. Biochemistry Moscow J Chen Q X Song K K Wang Q et al. Inhibitory effects of mushroom tyrosinase by some alkylbenzaldehydes J. Enzyme Inhibition Med Chem J Khatib S Nerya O Musa R. Chalcones as potent tyrosinase inhibitors the importance of a 2 4-substituted resorcinol moiety