Partial agonistic effect of yokukansan on human recombinant serotonin 1A receptors expressed in the membranes of Chinese hamster ovary cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Yokukansan (YKS) is a traditional Japanese medicine consisted of seven medicinal herbs and has been used for treatment of neurosis, insomnia, and behavioral and psychological symptoms of dementia (BPSD) in Japan. AIM OF THE STUDY: The aim of the present study is to clarify the intrinsic activity of YKS on serotonin (5-HT)1A and 5-HT2A receptors and also to determine the constituent herbs which are responsible for the effect of YKS. MATERIALS AND METHODS: The dry powdered extracts of YKS, seven constituent herbs, and YKS-analogues which were produced by eliminating one of the constituent herbs from YKS in the manufacturing process, were used for the evaluation. Competitive binding assays for 5-HT receptors and [(35)S]GTPgammaS binding assays for the evaluation of agonistic/antagonistic activity were performed using Chinese hamster ovary cell membranes stably expressing human recombinant 5-HT1A or 5-HT2A receptors. RESULTS: YKS (6.25-400 microg/ml) concentration-dependently inhibited the binding of [(3)H]8-OH-DPAT to 5-HT1A receptors. The IC(50) value was estimated to be 61.2 microg/ml. In contrast, YKS failed to inhibit the binding of [(3)H]ketanserin to 5-HT2A receptors. Only Uncaria hook (3.13-50 microg/ml), of the seven constituent herbal extracts, inhibited the [(3)H]8-OH-DPAT binding to 5-HT1A receptors in a concentration-dependent manner, and the IC(50) value was estimated to be 7.42 microg/ml. The extracts of YKS or Uncaria hook increased [(35)S]GTPgammaS binding to 5-HT1A receptors to approximately 50% of that of a full agonist, 5-HT. Both the competitive binding and [(35)S]GTPgammaS binding of YKS to 5-HT1A receptors were remarkably attenuated by eliminating Uncaria hook from YKS, but it was almost unchanged when one of the other constituent herbs was eliminated from YKS. CONCLUSION: These results suggest that YKS has a partial agonistic effect on 5-HT1A receptors, which is mainly attributed to Uncaria hook.

The herbal medicines Saireito and Boiogito improve the hypertension of pre-eclamptic rats induced by Nomega-Nitro-L-arginine methyl ester.

The chronic inhibition of nitric oxide (NO) synthesis with N(omega)-Nitro-L-arginine methyl ester (L-NAME) induces a pre-eclampsia-like syndrome including hypertension in pregnant rats. We tested the traditional herbal medicines Saireito (SR) and Boiogito (BO), which have been used clinically for the treatment of pre-eclampsia, in this model. L-NAME was infused subcutaneously into pregnant rats from gestational day 14 (G14). SR and BO (both at 1, 2g/kg) were administered by gavage from G14 to G20. Systolic blood pressure was measured on G19. SR and BO (both at 1, 2g/kg) inhibited L-NAME-induced hypertension. SR was effective in both pregnant and non-pregnant rats while BO was effective only in pregnant rats. BO increased blood levels of CGRP and decreased levels of endothelin-1; both are known to play important roles in regulation of blood pressure in pre-eclampsia. SR and BO may be beneficial for the treatment and prevention of hypertension in pre-eclampsia.

Effects of Saireito, a Japanese herbal medicine, on edema via antagonistic actions against aldosterone in anti-GBM nephritic rats.

BACKGROUND: In order to clarify the diuretic mechanisms of Saireito, a Japanese herbal medicine, the mineralcorticoid receptor antagonistic action of Saireito was evaluated in anti-glomerular basement membrane (GBM) nephritic rats. METHODS: Anti-GBM nephritis was induced in rats by the intravenous, injection of anti-GBM serum, and test drugs were administered 5 days after the induction of nephritis. In addition, we also investigated aldosterone-loaded mice to clarify the effects of test drugs on aldosterone signal transduction. In an in vitro study, a mineralocorticoid receptor binding assay of the components of Saireito was performed. RESULTS: Saireito and spironolactone inhibited the development of proteinuria and abdominal ascites in anti-GBM nephritic rats. Saireito and spironolactone increased the urine volume and decreased the abdominal saline content in aldosterone-loaded mice. Saikosaponin H, a component of Saireito, inhibited the receptor binding of aldosterone in the in vitro assay 50% inhibitory concentration ([IC(50)], 22 micromol/l). Saikosaponin H also inihibited the decrease in urine volume in aldosterone-loaded mice. CONCLUSIONS: These results suggest that the diuretic action of Saireito may be partly due to an antagonistic action on the mineralocorticoid receptor, exerted by saikosaponin H.

A new acetylenic compound from the rhizomes of Atractylodes chinensis and its absolute configuration.

A new acetylenic compound "atractyloyne", (3S,4E,6E,12E)-1-isovaleryloxy-tetradeca-4,6,12-triene-8,10-diyne-3,14-diol (1) was isolated from the rhizomes of Atractylodes chinensis (Compositae) together with a known compound (4E,6E,12E)-3-isovaleryloxy-tetradeca-4,6,12-triene-8,10-diyne-1,14-diol (2). These structures were determined on the basis of the spectroscopic data and chemical evidence, and the absolute configuration of 1 was established by the modified 2-methoxy-2-trifluoromethylphenylacetic acid (MTPA) method.

Effects of rikkunshi-to, a traditional Japanese medicine, on the delay of gastric emptying induced by N(G)-nitro-L-arginine.

We evaluated the effects of Rikkunshi-to and several of its ingredients on the delay of gastric emptying induced by a nitric oxide (NO) synthase inhibitor, N(G)-nitro-L-arginine (L-NNA). After oral administration of L-NNA to rats, the gastric emptying rate at 24 h was decreased from 82.8 +/- 2.4% to 53.3 +/- 5.7%. The decrease of the gastric emptying rate induced by L-NNA treatment was markedly ameliorated by administration of Rikkunshi-to (250 and 500 mg/kg, p.o.) in a dose-dependent manner. To identify the active ingredient of Rikkunshi-to, the components were separated according to polarity, and the effects of the respective fractions on gastric emptying were evaluated. Significant efficacy was found in the water and methanol fractions, but not in the 50% aqueous-methanol fraction. Furthermore, hesperidin (1 - 4.29 mg/kg, p.o.) contained in the methanol fraction and L-arginine (4.5 mg/kg, p.o.) contained in the water fraction ameliorated the decrease in the gastric emptying rate induced by L-NNA treatment. These results suggest that Rikkunshi-to ameliorated abnormalities of NO-mediated gastric functions such as delayed gastric emptying, and hesperidin and L-arginine were identified as two of the active ingredients contributing to the ability of Rikkunshi-to to facilitate gastric emptying.

Inhibitory effects of triterpenes isolated from Chuling (Polyporus umbellatus Fries) on free radical-induced lysis of red blood cells.

Chuling, sclerotia of Polyporus umbellatus FRIES, has long been used for urological disorders in traditional medicine. In this study, we demonstrated that Chuling in vitro protects red blood cells from 2,2-azo-bis(2-amidinopropane)dihydrochloride (AAPH)-induced hemolysis. The inhibitory effect was dose-dependent at concentrations of 50 to 1000 microg/ml. Moreover, tests were carried out to identify the main ingredient of Chuling with scavenging effect on free radicals. Triterpene carboxylic acids isolated from the methanol extract of Chuling, namely, polyporusterone A and polyporusterone B, were found to have inhibitory activities against AAPH-induced lysis of red blood cells. The anti-hemolytic effect was significantly stronger in polyporusterone B compared with polyporusterone A. Furthermore, the ingestion of 150 mg of Chuling was associated with a significant increase in free-radical scavenging effect of plasma in rats.

Separation and isolation methods for analysis of the active principles of Sho-saiko-to (SST) oriental medicine.

Sho-saiko-to (SST) was introduced into Japan as an oriental classical medicine from China approximately 1500 years ago, and it is currently the most representative Kampo medicine (traditional Japanese medicine). SST is manufactured in Japan as an ethical drug on a modern industrial scale in which the quality of ingredients is standardized with Good Manufacturing Practices (GMP) regulation. SST is widely used for the treatment of chronic hepatitis. Experimental and clinical studies including multi-center, placebo-controlled, double-blind studies have demonstrated the various pharmacological effects of SST. SST is prepared from the hot water extraction of seven raw materials, therefore many kinds of constituents are included. Three-dimensional (3D) HPLC analysis is useful for obtaining many kinds of constituents, especially low molecular ultraviolet (UV) quenching compounds, contained in SST as well as its fractions. Fingerprint pattern provided by 3D HPLC analysis makes possible to identify the overall-viewing of SST. Databases of UV spectra of the components of medicinal herbs obtained by reversed-phase (RP) HPLC using a photodiode array (PDA) and fingerprint patterns of crude drugs made by 3D HPLC analysis facilitate the identification, analysis and quality of herbal drugs. Studies using both PDA HPLC and an amino acid analysis with a fluorometric detector have found that SST contains fifteen major low molecular compounds (i.e. baicalin, wogonin-7-O-glucuronide, liquiritin, their three aglycons, liquiritin apioside, glycyrrhizin, saikosaponin b1, saikosaponin b2, ginsenoside Rg1, ginsenoside Rb1, (6)-gingerol, (6)-shogaol and arginine). These compounds have various pharmacological actions, and are assumed to be responsible, at least partly, for the pharmacological effects of SST. Although there have only been a few investigations on high molecular compounds with pharmacological actions contained in SST, several kinds of polysaccharides have been isolated from constituent herbs of SST. This review paper summarizes analytical methods of separation, isolation and identification of compounds with biological activities from SST, which is a mixture drug of medicinal herbs. Accordingly, this paper would not focus on methods of separation, isolation and analysis of particular compounds from each constituent herb of SST.

Effect of the rhizomes of Atractylodes lancea and its constituents on the delay of gastric emptying.

The effect of rhizomes of Atractylodes lancea on gastric disorders, in particular, the delay in gastric emptying induced by N(G)-nitro-L-arginine in rats, was investigated. Intragastric treatment with an aqueous extract (250 mg/kg) and its lipophilic fractions (4 mg/kg) significantly improved delayed gastric emptying. The major constituents of the lipophilic fraction were two sesquiterpens, hinesol and beta-eudesmol, and four known polyacetylenic compounds, atractylodin, atractylodinol, acetylatractylodinol and 4,6,12-tetradecatriene-8,10-diyne-1,3,14-triol. The activity was found in the four polyacetylenic compounds at a similar potency, but not in the two sesquiterpens. To clarify the effect of the four polyacetylenic compounds in this extract, we attempted to evaluate the activity of atractylodin, as representative, at a dose of 0.2 mg/kg based on the total amounts (0.2 mg/250 mg aqueous extract) of the four polyacetylenic compounds. In addition, atractylodin improved the delay in gastric emptying at between 0.1 and 0.3 mg/kg in a dose-dependent manner. These results suggest that the aqueous extract improved the delayed gastric emptying, and polyacetylenic compounds contributed to its activity.

Pharmacological evaluation of Shokyo and Kankyo (1).

Zingiberis Rhizoma (Shokyo, [Japanese characters: see text]) showed significant ameliorative effect on the BaCl2-induced delay of gastric emptying in rat. Bioassay-guided fractionation of the aqueous extract of Shokyo resulted in isolation of 6-gingesulfonic acid (1) and shogasulfonic acid A (3). These compounds significantly improved the delay of gastric emptying on both BaCl2-induced and N(G)-nitro-L-arginine (L-NNA)-induced model in rat. Zingiberis Siccatum Rhizoma (Kankyo, [Japanese characters: see text]) had significant efficacy against castor oil-induced diarrhea. In addition, Kankyo showed the activity increasing intestinal blood flow in normal rat.

Effects of Sho-Saiko-to on hepatocarcinogenesis and 8-hydroxy-2'-deoxyguanosine formation.

Oxidative stress plays an important role in hepatocarcinogenesis. Although Sho-saiko-to (TJ-9), a Japanese herbal medicine which has been recently administered to patients with chronic liver disease in Japan, prevents hepatocarcinogenesis, the mechanism by which TJ-9 protects against cancer development is not fully understood. 8-Hydroxy-2'-deoxyguanosine (8-OHdG), a DNA adduct by reactive oxygen species, is known as a parameter of genetic risk for hepatocarcinogenesis. To clarify whether the preventive effect on hepatocarcinogenesis by TJ-9 is dependent on 8-OHdG, the effect on 8-OHdG levels by TJ-9 was examined by using high-performance liquid chromatography-mass spectrometry (LC-MS) in a diethylnitrosamine (DEN)-induced hepatocarcinogenesis model of male Fisher rats. TJ-9 reduced the number of preneoplastic cells, detected as the glutathione S transferase P (GST-P)-positive hepatocytes, and inhibited the development of liver tumors. TJ-9 also significantly decreased the formation of 8-OHdG, as indicated by LC-MS and immunohistochemical analysis. In addition, ornithine decarboxylase (ODC) activity and the number of proliferating cell nuclear antigen (PCNA)-positive cells were not altered. An electron paramagnetic resonance spin-trapping technique showed that TJ-9 scavenges hydroxyl radicals in a dose-dependent manner. In conclusion, the results of the present study suggest that TJ-9 prevents hepatocarcinogenesis in association with inhibition of 8-OHdG formation.