An isocratic HPLC procedure was developed for simultaneous determination of six catechins, gallic acid, and three methylxanthines in tea water extract. A baseline separation was achieved on a Cosmosil C18-MS packed column with a solvent mixture of methanol/doubly distilled water/formic acid (19.5:80.2:0.3, v/v/v) as mobile phase. A gradient HPLC procedure was also provided for the separation of these tea components. The contents of catechins, gallic acid, and methylxanthines have been measured in infusions of a range of green tea, oolong tea, and pu-erh tea products sold and consumed in the China, Japan, and Taiwan. When 15 Chinese green tea and 13 Japanese green tea products were analyzed by the HPLC method, the mean levels of the total catechins, (-)-epigallocatechin 3-gallate, (+)-catechin, and caffeine were found to be very similar in these two groups, but other minor catechins such as (-)-epigallocatechin, (-)-epicatechin, and (-)-gallocatechin 3-gallate were found to be higher in Japanese green tea products, whereas (-)-epicatechin 3-gallate, gallic acid, theophylline, and theobromine were found to be higher in Chinese green tea products. Oolong tea products possessed lower levels of catechins, whereas pu-erh tea products contained negligible amounts of these constituents. The new HPLC method is rapid, reliable, and reproducible and should be highly recommended to tea industries for routine analysis of commercial tea samples.

The polyphenols in various parts of the tea plant used in manufacture of tea products (young leaves, old leaves, and stem) were analyzed by HPLC. The young leaves (apical bud and the two youngest leaves) were found to be richer (2.7-fold) in polyphenols than old leaves (from the tenth to the fifth leaf). Also, the tea polyphenols were found to be higher (1.4-fold) in summer than in spring. Ten different types of commercial tea (manufactured tea), including unfermented, semifermented, and fermented tea, were analyzed for their polyphenol compounds, and it was found that both yields of solids in tea water extracts (TWEs) and the amount of (-)-epigallocatechin 3-gallate (EGCG) in these products varied with different tea leaves and processing methods. Longjing tea (unfermented green tea) contained the highest concentration of EGCG and polyphenols, whereas Assam black tea (most fermented) contained the least. Longjing TWEs showed strong inhibitory effect on DNA synthesis in A-431 tumor cells. We also found that green tea polyphenols (GTPs) and EGCG strongly inhibited tumor cell DNA synthesis and peroxyl-radical generation. The evidence suggests that the strong antioxidative and antiproliferative activities of Longjing TWEs and GTPs are mainly due to the higher contents of tea polyphenols. EGCG (the major component of GTPs) showed strong inhibitory action on the growth of immortalized cells (NIH3T3) and tumor cells (S-180II) but almost no effect on the growth of normal cells (C3H10T1/2).

Peroxisome proliferator-activated receptor (PPAR)Q Q transcription factor has been implicated in anti-inflammatory response. Of the compounds tested, apigenin, chrysin, and kaempferol significantly stimulated PPARQ Q transcriptional activity in a transient reporter assay. In addition, these three flavonoids strongly enhanced the inhibition of inducible cyclooxygenase and inducible nitric oxide synthase promoter activities in lipopolysaccharide-activated macrophages which contain the PPARQ Q expression plasmids. However, these three flavonoids exhibited weak PPARQ Q agonist activities in an in vitro competitive binding assay. Limited protease digestion of PPARQ Q suggested these three flavonoids produced a conformational change in PPARQ Q and the conformation differs in the receptor bound to BRL49653 versus these three flavonoids. These results suggested that these three flavonoids might act as allosteric effectors and were able to bind to PPARQ Q and activate it, but its binding site might be different from the natural ligand BRL49653. ß 2001 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.

(-)-Epigallocatechin-3-gallate (EGCG), a major polyphenol in green tea, was shown to have cancer chemopreventive activity. In this study, we examined the antimetastatic effects of EGCG or the combination of EGCG and dacarbazine on B16-F3m melanoma cells in vitro and in vivo. First, the antimetastatic potentials of five green tea catechins were examined by soft agar colony formation assay, and the results show that EGCG was more effective than the other catechins in inhibiting soft agar colony formation. Second, EGCG dose-dependently inhibited B16-F3m cell migration and invasion by in vitro Transwell assay. Third, EGCG significantly inhibited the spread of B16-F3m cells on fibronectin, laminin, collagen, and Matrigel in a dose-dependent manner. In addition, EGCG significantly inhibited the tyrosine phosphorylation of focal adhesion kinase (FAK) and the activity of matrix metalloproteinase-9 (MMP-9). In animal experiments, EGCG alone reduced lung metastases in mice bearing B16-F3m melanomas. However, a combination of EGCG and dacarbazine was more effective than EGCG alone in reducing the number of pulmonary metastases and primary tumor growths, and increased the survival rate of melanoma-bearing mice. These results demonstrate that combination treatment with EGCG and dacarbazine strongly inhibits melanoma growth and metastasis, and the action mechanisms of EGCG are associated with the inhibition of cell spreading, cell-extracellular matrix and cell-cell interactions, MMP-9 and FAK activities.