Potent suppressive activity of pheophytin a and b from the non-polyphenolic fraction of green tea (Camellia sinensis) against tumor promotion in mouse skin
Introduction
In Japan, various kinds of green tea have been traditionally consumed as a daily beverage and some epidemiological studies have indicated that the ubiquitous consumption of green tea might be a protective factor against certain types of cancer 1, 2. Furthermore, many experimental studies have shown that the hot water-soluble extract of green tea or tea-derived polyphenols have significant suppressive activities against carcinogenesis 3, 4. The major polyphenols derived from green tea have been known as catechins, such as (−)epicatechin, (−)epicatechin gallate, (−)epigallocatechin and (−)epigallocatechin gallate, and they are detected in the water-soluble fraction of green tea [5]. However, the anticarcinogenic activity in the water-insoluble fraction has been poorly analyzed. To study this problem, we analyzed the effects of the non-polyphenolic fraction of green tea on tumor initiation and promotion using in vitro and in vivo experiments and detected the suppressive activities of the non-polyphenolic fraction against umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002) caused by genotoxins in a tester bacteria, tumor promoter-induced ornithine decarboxylase (ODC) induction of 3T3 fibroblasts and chemically-induced skin tumor initiation and promotion in BALB/c mice [6]. Furthermore, we have recently identified pheophytin a and b as antigenotoxic substances in the non-polyphenolic fraction of green tea [7].
In the present paper, we describe the effects of pheophytin a and b on tumor promotion in mouse skin using in vivo and in vitro experiments.
Section snippets
Chemicals
7,12-Dimethylbenz[a]anthracene (DMBA) and 12-O-tetradecanoylphorbol-13-acetate (TPA) were purchased from Sigma (St. Louis, MO).
Purification of pheophytin a and b in the non-polyphenolic fraction of green tea
The preparation of the non-polyphenolic fraction of green tea was carried out using our previous method [7]. Briefly, green tea leaves (Sansui-En, Kumamoto, Japan) were brewed with hot water (25 ml/g tea leaves) and tea extract was decanted. After the same treatment was repeated three times, the residual tea leaves were dried and minced extensively by an electric mill
Suppressive effects of pheophytin a and b from the non-polyphenolic fraction of green tea against tumor promotion in BALB/c mouse skin
We examined the effects of pheophytin a and b from the non-polyphenolic fraction of green tea on the promotion phase of chemically-induced mouse skin tumorigenesis. As shown in Table 1, as a positive control, the combined treatment with 50 nmol DMBA (initiator) and 2 nmol TPA (promoter) induced remarkable tumors. Namely, all tested mice developed skin tumors and a high incidence of tumors was observed (19.2±3.6 tumors per mouse). However, the application of 100 or 500 μg pheophytin a or b prior
Discussion
Previous experimental studies have indicated that green tea extract or tea-derived polyphenols known as catechins have antitumor promoting activities 3, 4. However, the effect of the non-polyphenolic fraction of green tea against tumor promotion has been poorly analyzed. Therefore, the authors analyzed the effect of the non-polyphenolic fraction of green tea against tumor promotion using in vitro and in vivo experiments [6]. On the other hand, the authors have recently identified
Acknowledgements
The authors would like to thank Dr Y. Nakamura (Faculty of Agriculture, Kyoto University) for the useful information on tumor promotion in mouse skin and Dr Y. Yano (Department of Second Biochemistry, Osaka City University, Medical School) for his kind suggestion for the ODC assay.
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