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.