Anti-tumor-promoting activity of a polyphenolic fraction isolated from grape seeds in the mouse skin two-stage initiation-promotion protocol and identification of procyanidin B5-3'-gallate as the most effective antioxidant constituent.
Anti-tumor-promoting activity of a polyphenolic fraction isolated from grape seeds in the mouse skin two-stage initiation-promotion protocol and identification of procyanidin B5-3'-gallate as the most effective antioxidant constituent.
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DOI：
10.1093/carcin/20.9.1737
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发表时间：
1999-09
期刊：
影响因子：
4.7
通讯作者：
Jifu Zhao;Jiannong Wang;Yingjie Chen;R. Agarwal
中科院分区：
文献类型：
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作者：
Jifu Zhao;Jiannong Wang;Yingjie Chen;R. Agarwal
葡萄籽中的原花青素具有抗炎、抗关节炎和抗过敏的作用，防止皮肤老化，抑制氧自由基，抑制紫外线辐射引起的过氧化作用。由于这些事件中的大多数与肿瘤促进阶段的致癌作用，这些研究表明，葡萄籽多酚和原花青素存在于其中可能是抗癌和/或抗肿瘤促进剂。因此，我们评估了从葡萄籽中分离的多酚组分（GSP）的抗肿瘤促进作用，采用7，12-二甲基苯并[a]蒽（DMBA）引发和12-O-十四烷酰基佛波醇13-乙酸酯（TPA）促进的SENCAR小鼠皮肤两阶段致癌方案作为模型系统。在用DMBA引发肿瘤后，以0.5和1.5 mg/小鼠/应用的剂量局部应用GSP至背部引发的小鼠皮肤导致TPA肿瘤促进的高度显著抑制。观察到的GSP的抗肿瘤促进作用具有剂量依赖性，并且在0.5和1.5 mg GSP下分别在肿瘤发生率（35%和60%抑制）、肿瘤多样性（61%和83%抑制）和肿瘤体积（67%和87%抑制）降低方面是明显的。基于这些结果，我们致力于分离和鉴定GSP中存在的单个多酚，并评估其在抑制表皮脂质过氧化方面的抗氧化活性。采用高效液相色谱法，然后与真实标准品的保留时间进行比较，在HPLC图谱，理化性质和光谱分析，9个单独的多酚被鉴定为儿茶素，表儿茶素，原花青素B1-B5和C1和原花青素B5- 3 '-没食子酸酯。对这些具有明显结构差异的多酚中的五种，即儿茶素、原花青素B2、原花青素B5、原花青素C1和原花青素B5- 3 '-没食子酸酯，进行了抗氧化活性评估。所有这些都显着抑制表皮脂质过氧化反应，虽然到不同的水平。构效关系研究表明，随着多酚结构聚合度的增加，对脂质过氧化的抑制潜力增加。此外，黄烷单元间的连接位置也影响脂质过氧化活性，具有4-6键的原花青素异构体比具有4-8键的异构体表现出更强的抑制活性。当没食子酸酯基团连接在原花青素二聚体的3 '-羟基位置时，对表皮脂质过氧化的抑制作用也明显增加。原花青素B5- 3 '-没食子酸酯在表皮脂质过氧化测定中显示出最有效的抗氧化活性，IC（50）为20 μ M。总之，这些结果首次表明，葡萄籽多酚具有高的抗肿瘤促进活性，由于其中存在的原花青素的强抗氧化作用。总之，葡萄籽多酚，特别是原花青素B5- 3 '-没食子酸酯，应该更详细地研究，以开发为癌症化学预防和/或抗癌剂。
Procyanidins present in grape seeds are known to exert anti-inflammatory, anti-arthritic and anti-allergic activities, prevent skin aging, scavenge oxygen free radicals and inhibit UV radiation-induced peroxidation activity. Since most of these events are associated with the tumor promotion stage of carcinogenesis, these studies suggest that grape seed polyphenols and the procyanidins present therein could be anticarcinogenic and/or anti-tumor-promoting agents. Therefore, we assessed the anti-tumor-promoting effect of a polyphenolic fraction isolated from grape seeds (GSP) employing the 7,12-dimethylbenz[a]anthracene (DMBA)-initiated and 12-O-tetradecanoylphorbol 13-acetate (TPA)-promoted SENCAR mouse skin two-stage carcinogenesis protocol as a model system. Following tumor initiation with DMBA, topical application of GSP at doses of 0.5 and 1.5 mg/mouse/application to the dorsal initiated mouse skin resulted in a highly significant inhibition of TPA tumor promotion. The observed anti-tumor-promoting effects of GSP were dose dependent and were evident in terms of a reduction in tumor incidence (35 and 60% inhibition), tumor multiplicity (61 and 83% inhibition) and tumor volume (67 and 87% inhibition) at both 0.5 and 1.5 mg GSP, respectively. Based on these results, we directed our efforts to separate and identify the individual polyphenols present in GSP and assess their antioxidant activity in terms of inhibition of epidermal lipid peroxidation. Employing HPLC followed by comparison with authentic standards for retention times in HPLC profiles, physiochemical properties and spectral analysis, nine individual polyphenols were identified as catechin, epicatechin, procyanidins B1-B5 and C1 and procyanidin B5-3'-gallate. Five of these individual polyphenols with evident structural differences, namely catechin, procyanidin B2, procyanidin B5, procyanidin C1 and procyanidin B5-3'-gallate, were assessed for antioxidant activity. All of them significantly inhibited epidermal lipid peroxidation, albeit to different levels. A structure-activity relationship study showed that with an increase in the degree of polymerization in polyphenol structure, the inhibitory potential towards lipid peroxidation increased. In addition, the position of linkage between inter-flavan units also influences lipid peroxidation activity; procyanidin isomers with a 4-6 linkage showed stronger inhibitory activity than isomers with a 4-8 linkage. A sharp increase in the inhibition of epidermal lipid peroxidation was also evident when a gallate group was linked at the 3'-hydroxy position of a procyanidin dimer. Procyanidin B5-3'-gallate showed the most potent antioxidant activity with an IC(50) of 20 microM in an epidermal lipid peroxidation assay. Taken together, for the first time these results show that grape seed polyphenols possess high anti-tumor-promoting activity due to the strong antioxidant effect of procyanidins present therein. In summary, grape seed polyphenols in general, and procyanidin B5-3'-gallate in particular, should be studied in more detail to be developed as cancer chemopreventive and/or anticarcinogenic agents.