Functional analysis of flavonoid 3',5'-hydroxylase from tea plant (Camellia sinensis): critical role in the accumulation of catechins.

BACKGROUND: Flavonoid 3',5'-hydroxylase (F3'5'H), an important branch point enzyme in tea plant flavan-3-ol synthesis, belongs to the CYP75A subfamily and catalyzes the conversion of flavones, flavanones, dihydroflavonols and flavonols into 3',4',5'-hydroxylated derivatives. However, whether B-ring hydroxylation occurs at the level of flavanones and/or dihydroflavonols, in vivo remains unknown. RESULTS: The Camellia sinensis F3'5'H (CsF3'5'H) gene was isolated from tea cDNA library. Expression pattern analysis revealed that CsF3'5'H expression was tissue specific, very high in the buds and extremely low in the roots. CsF3'5'H expression was enhanced by light and sucrose. Over-expression of CsF3'5'H produced new-delphinidin derivatives, and increased the cyanidin derivative content of corollas of transgenic tobacco plants, resulting in the deeper transgenic plant flower color. Heterologous expressions of CsF3'5'H in yeast were carried out to demonstrate the function of CsF3'5'H enzyme in vitro. Heterologous expression of the modified CsF3'5'H (CsF3'5'H gene fused with Vitis vinifera signal peptide, FSI) revealed that 4'-hydroxylated flavanone (naringenin, N) is the optimum substrate for CsF3'5'H, and was efficiently converted into both 3'4'- and 3'4'5'-forms. The ratio of 3'4'5'- to 3'4'-hydroxylated products in FSI transgenic cells was significantly higher than VvF3'5'H cells. CONCLUSIONS: CsF3'5'H is a key controller of tri-hydroxyl flavan-3-ol synthesis in tea plants, which can effectively convert 4'-hydroxylated flavanone into 3'4'5'- and/or 3'4'-hydroxylated products. These findings provide animportant basis for further studies of flavonoid biosynthesis in tea plants. Such studies would help accelerate flavonoid metabolic engineering in order to increase B-ring tri-hydroxyl product yields.

[Controlled study of auricular point taping and pressing therapy for treatment of vascular dementia].

OBJECTIVE: To assess the therapeutic effect of auricular point taping and pressing therapy on vascular dementia (VD). METHODS: One hundred and eighty cases of vascular dementia were randomly divided into an auricular point taping and pressing group and a western medicine group, 90 cases in each group. The auricular point taping and pressing group was treated by auricular point taping and pressing at auricular points Shennen, Nao (brain), Shen (kidney), Zhen (pulvinal), and the western medicine group by oral administration of Nimodipine, thrice each day, 30 mg each time. They were treated for 12 weeks. The scores of mini-mental state examination (MMSE) and activities of daily living (ADL), and adverse reaction were observed. RESULTS: The scores of MMSE before treatment and 12 weeks after treatment were 18.00 +/- 3.88 and 20.83 +/- 3.74 in the auricular point taping and pressing group, and 17.80 +/- 3.82 and 20.70 +/- 3.53 in the western medicine group, respectively, with significant increases in scores of MMSE after treatment in the two groups (both P < 0.01) and with no significant difference between the two groups (P > 0.05). The scores of ADL before treatment and 12 weeks after treatment were 44.90 +/- 14.84 and 39.60 +/- 12.45 in the auricular point taping and pressing group, and 45.70 +/- 14.86 and 39.10 +/- 13.44 in the western medicine group, respectively, with significant decreases after treatment in the two groups (both P < 0.01) and with no significant difference between the two groups (P > 0.05). In the auricular point taping and pressing group, 2 cases withdrew from the test because adhesive plaster allergic reaction induced severe itch of the auricle. In the western medicine group, one case had mild dizziness and another case had diarrhea respectively. CONCLUSION: Auricular point taping and pressing and Nimodipine have similar therapeutic effect on vascular dementia, and have no obvious adverse reaction, except adhesive plaster allergic reaction.