No abstract

Gambir, an official pharmacopoeic medicine in Japan, is the aqueous extract of the leaves and young twigs of Uncaria gambir ROXB. (Rubiaceae).2) The extract has been used for the treatment of diarrhea and sore throat as an astringent medicine in Southeast Asia, 3) and also for hoarseness, as an ingredient of the kampo medicine "Kyoseihatekigan."Flavan monomers, (ϩ)-catechin (1) and (ϩ)-epicatechin (2), 4) several other dimeric compounds 5) related to 1, as well as alkaloids, 6,7) have been reported to be the constituents. Our investigation on the polyphenolic contents of a manufactured gambir product led to the isolation of 1, 2, and seven dimeric flavans 3-9. Although the seven dimeric flavans were known compounds, the present study revealed that the stereostructures of four of the chalcane-flavan dimers should be revised to 6-9. Their 1 H-and 13 C-NMR spectral assignments are also shown here, since those were not reported in detail previously. 5,8) Results and DiscussionA manufactured product of gambir purchased in Japan was extracted with MeOH, and the extract was fractionated by column chromatography on Dia-ion HP-20 using aqueous MeOH. (ϩ)-Catechin (1) was crystallized from the eluate with 20% MeOH, and the mother liquor was subjected to column chromatography on a Toyopearl HW-40 to give a fraction containing both monomeric and dimeric flavans. The fraction was further chromatographed on octadecylsilyl (ODS) silica gel, Sephadex LH-20, and MCI-gel CHP-20P columns to give (ϩ)-catechin (1), (ϩ)-epicatechin (2), procyanidin B1 (3), 9,10) procyanidin B3 (4), 11) gambiriin C (5), 5) and gambiriin A1 (6). The eluate, with 40% MeOH from the column of Dia-ion HP-20, was separated in an analogous way to give gambiriin A2 (7), gambiriin B1 (8), and gambiriin B2 (9). Gambiriin A1 (6) was obtained as an amorphous powder. The dimeric molecular formula, C 30 H 28 O 12 , was indicated by the [MϩH] ϩ ion peak at m/z 581 in the electrospray-ionization (ESI)-MS. The showed a large coupling constant (7.5 Hz), indicating that the lower unit was 2,3-trans catechin. The 2,3-trans structure was also validated by the 13 C-NMR spectrum, which showed the C-2 L signal at d 82.5. The remaining set of the methine-methine-methylene protons [d: 2.56 (H-ga U ), 2.95 (Hgb U ), 4.68 (H-b U ), 4.75 (H-a U )] was attributable to the substituted propyl group in the chalcane structure. The molecular formula of 6, which was indicated by the ESI-MS, also substantiates this open-chain chalcane structure.The rotating frame nuclear Overhauser effect spectroscopy (ROESY) of 6 showed correlations of H-a U /H-2Ј L and Hb U /H-2Ј L , indicating the presence of an interflavonoid link between position C-8 L and the upper chalcane unit (C-a U ) (Fig. 2). These correlations would not be observed if the linkage occurred at C-6 L . The aliphatic protons of 6 appeared as broad signals, due to the restricted rotation around the interflavonoid linkage, which confirmed the location of the linkage at C-8 L .The assigned structure of 6 based on these data suggest...

We have recently reported that inhibition of tumor necrosis factor-a (TNF-a) release is a useful method for screening of cancer preventive agents, 1) because TNF-a and other inflammatory cytokines stimulate tumor promotion and progression of initiated cells as well as premalignant cells. 2,3) Since TNFa and interleukin-1 (IL-1), as endogenous tumor promoters, activate nuclear factor kB (NF-kB), 4,5) inhibitors of NF-kB activation are also assumed to be cancer preventive agents. 6) However, in a practical sense, it is quantitatively easier to measure inhibition of TNF-a release from cells treated with a tumor promoter such as okadaic acid than inhibition of NFkB activation using a gel mobility shift assay. 7) Based on the study of green tea as cancer preventive agent in the United States 8) and as cancer preventive beverage in Japan, 9) we decided to take a closer look at Japanese herbal medicine for other likely cancer preventives. One classical Japanese herb, Geranium thunbergii SIEB. et ZUCC. (Gennoshoko in Japanese), is used as an antidiarrhetic 10) ; it contains geraniin, which has been intensively studied as a representative tannin. 11) And the bark of Acer nikoense MAXIM (Megusurino-ki in Japanese), which contains diarylheptanoids, is sold as a folk medicine for eye and liver diseases in Japan. 12) The Forestry Experiment Station of Saitama Prefecture succeeded in culturing young plants of A. nikoense collaborated with us on the scientific utilization of the plant. We received four water extracts-leaves, bark, small branches, and timber-from the Station and subjected them to inhibition of TNF-a release assay using BALB/3T3 cells treated with okadaic acid 13) : The leaves showed the strongest activity among the four extracts. Each extract was subjected to highperformance liquid chromatography (HPLC) with CAP-CELL PAK C18, and geraniin and corilagin were identified in the active leaf extract (Fig. 1).In The success of green tea as a cancer preventive is based on evidence that green tea contains tannins and antioxidants, does not show toxicity in humans and has long traditional use in Asia. In the light of this, herbal medicines are now also attracting attention as potential sources of cancer preventive agents. Using the inhibition of TNF-a a release assay, we studied Acer nikoense (Megusurino-ki in Japanese), one of the herbal medicines. The inhibitory activity of TNF-a a release was found in the leaf extract rather than the bark extract, and the main active constituents were identified as geraniin and corilagin, which are present in another Japanese traditional herb, Geranium thunbergii (Genno-shoko). The IC 50 values of TNF-a a release inhibition were 43 m mM for geraniin and 76 m mM for corilagin, whereas that for (؊)-epigallocatechin gallate (EGCG) was 26 m mM. Treatment with geraniin prior to application of okadaic acid, a tumor promoter on mouse skin initiated with 7,12-dimethylbenz(a)anthracene, reduced the percentage of tumor-bearing mice from 80.0 to 40.0% and the average numbers of tumor per mouse ...

Three new hellinoyl-type ellagitannins, nilotinins M4 (7), D7 (8), and D8 (9), and a new macrocyclic-type, nilotinin D9 (10), together with eight known tannins, hirtellins B (2), C (11), and F (12), isohirtellin C (13), tamarixinin A (3), tellimagrandins I and II, and 1,2,6-tri-O-galloyl-beta-d-glucose (14), were isolated from an aqueous acetone extract of Tamarix nilotica dried leaves. Nilotinin M4 (7) is a monomeric tannin possessing a hellinoyl moiety. The structure of 8 demonstrated replacement of one of the HHDP groups at the glucose core O-4/O-6 in ordinary dimeric tannins with a galloyl moiety at O-6. This is a new structural feature among the tamaricaceous ellagitannins. On the basis of the results, reported spectroscopic assignments for 2, 3, and the macrocyclic tannins 11-13 were revised. Unusual shifts in the NMR spectra of these macrocyclic tannins are also discussed in relation to their conformations. Several tannins isolated from T. nilotica were assessed for possible cytotoxic activity against four human tumor cell lines, and nilotinin D8 (9) and hirtellin A (1) showed high cytotoxic effects.