葡萄籽原花青素及其柔性纳米脂质体的抗氧化及对酪氨酸酶活性的抑制作用

Antioxidation and Inhibition of Tyrosinase Activity of Grape Seed Proanthocyanidins and Their Flexible Nano-Liposomes

目的探讨葡萄籽原花青素(GSP)及其柔性纳米脂质体(GSP-FL)的抗氧化活性,以及对酪氨酸酶活性的影响。方法体外测定GSP及GSP-FL对1,1-二苯基-2-三硝基苯肼(DPPH)、2,2’-联氮-双-3-乙基苯并噻唑啉-6-磺酸(ABTS)、2-苯基-4,4,5,5-四甲基咪唑啉-1-氧基-3-氧化物(PTIO)自由基的清除率,以及对三价铁离子(Fe3+)的还原能力。采用四甲基偶氮唑盐(MTT)法检测GSP对过氧化氢(HO)诱导的细胞氧化损伤的保护作用。体外测定GSP及GSP-FL对酪氨酸单酚酶、酪氨酸二酚酶活性的影响。以3,4-二羟基苯丙氨酸(L-DOPA)为底物,测定GSP及GSP-FL对Melan-a细胞胞内酪氨酸酶活性的影响;采用氢氧化钠裂解法测定GSP-FL对Melan-a细胞胞内黑色素含量的影响。结果GSP及GSP-FL均具有良好的抗氧化活性。与GSP比较,GSP-FL可显著降低对Melan-a细胞的毒性(P<0.05);与模型组(250μmol/L HO)比较,GSP质量浓度为12.5~50.0 mg/mL时可显著提高氧化损伤细胞的存活率(P<0.05)。GSP及GSP-FL均可抑制细胞内酪氨酸酶的活性,并减少细胞黑色素的生成。结论GSP-FL具有抗氧化活性,能降低药物的细胞毒性,且可通过抑制酪氨酸酶的活性来抑制黑色素的生成,有望进一步开发成具有美白功效的化妆品制剂。

Objective To investigate the antioxidation activity of grape seed proanthocyanidins(GSP),their flexible nano-liposomes(GSP-FL),and their effects on the activity of tyrosinase.Methods The scavenging rates of GSP and GSP-FL on the free radicals such as 1,1-diphenyl-2-picrylhydrazyl(DPPH),2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid),that was ABTS,2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide(PTIO)and the reduction ability of GSP and GSP-FL on the ferric ion(Fe)were measured in vitro.The protective effect of GSP on the oxidative damage of cells induced by the hydrogen peroxide(HO)was tested by the methyl thiazolyl tetrazolium(MTT)method.The effects of GSP and GSP-FL on the monophenolase and diphenolase activities of tyrosinase were measured in vitro.The effects of GSP and GSP-FL on the activity of intracellular tyrosinase of Melan-a cells were measured with the L-3,4-dihydroxyphenylalanine(L-DOPA)as the substrate.The effect of GSP-FL on the intracellular melanin content of Melan-a cells was measured by the sodium hydroxide lysis method.Results Both GSP and GSP-FL had good antioxidant activities.Compared with GSP,GSP-FL can significantly reduce the toxicity to Melan-a cells(P<0.05).Compared with those in the model group(250μmol/L HO),the survival rate of oxidatively damaged cells could significantly improved when the mass concentration of GSP was in the range of 12.5-50.0 mg/mL(P<0.05).Both GSP and GSP-FL could inhibit the activity of intracellular tyrosinase and reduce the synthesis of melanin.Conclusion GSP-FL has antioxidation activity,can reduce the cytotoxicity of drugs,and can inhibit the synthesis of melanin by inhibiting the activity of tyrosinase,which is expected to be further developed into a cosmetic preparation with the whitening effect.