A simultaneous determination of principal compounds in tokishakuyakusan by high-performance liquid chromatography with diode array detector.

We developed a simultaneous analysis method using high-performance liquid chromatography coupled with diode-array detector (HPLC-DAD) for six principal compounds (atractylenolide III, alisol A, alisol B, paeoniflorin, ferulic acid and (Z)-ligustilide) in a traditional Japanese (Kampo) medicine, tokishakuyakusan (TSS). The HPLC separation was conducted on a reversed-phase TSK-gel ODS-80TS column (4.6 i.d. × 250 mm, 5 µm) at 40°C with a 0.1% phosphoric acid-acetonitrile gradient system. The DAD detection wavelength was set at 205, 232 and 330 nm. Calibration curves for the compounds showed linear regressions with correlation coefficients of >0.999. The intra- and inter-day precision (i.e., the relative standard deviation) were in the range of 0.50-1.55 and 0.70-1.80%, respectively. The average recovery yields of the compounds ranged from 98.3 to 103%. The present results will contribute to shorter analysis times with less organic solvent compared with the individual analysis of each compound for the evaluation of TSS. The application of the established method to TSS will also provide helpful information for the further pharmacological and clinical studies.

Extraction efficiency of shosaikoto (xiaochaihu tang) and investigation of the major constituents in the residual crude drugs.

Shosaikoto (Xiaochaihu Tang) is one of the frequently used traditional herbal medicines (Kampo medicines, Japan). To realize the effective use of precious crude drugs, we investigated the major constituents in the residual crude drugs after decoction and aimed for extraction efficiency of shosaikoto with regard to the extract, tannin content, and major constituents. We found that the residual crude drugs had large amounts of compounds, especially saikosaponin b(2), which had a 78.3% yield compared to that in the first decoction. The extraction efficiency increased when decoction time and volume of water increased. Both increases had an additive effect on the yield of the extract and saikosaponin b(2) in particular. We also found that the size of crude drug pieces that are available in Japanese markets is suitable for decoction because of quick permeation of water. From our study, the second decoction may be a valuable contribution to medical treatment and of effective use of crude drugs. Moreover, time and volume of water should be increased when patients have trouble in preparing a decoction. Our study revealing the factors that influence the extraction efficiency of shosaikoto will be the basis for empirical evidence about decocting Kampo medicine.

Efficient preparation of Hangekobokuto (Banxia-Houpo-Tang) decoction by adding perilla herb before decoction is finished.

Hangekobokuto (banxia-houpo-tang) is a Kampo (traditional Japanese) medicine used mostly for anxiety disorder and consists of Pinellia Tuber, Poria Sclerotium, Magnolia Bark, Perilla Herb and Ginger. Perilla Herb contains an essential oil rich in perillaldehyde, which has a pleasant flavor, but this is easily lost due to heating. We therefore investigated how the major constituents of hangekobokuto, namely perillaldehyde, rosmarinic acid, magnolol and [6]-gingerol, varied with time during decoction and approached an optimal condition for decoction. Rosmarinic acid at 15 min after boiling, and magnolol and [6]-gingerol at 30 min were mostly extracted, while 0.09 mg of perillaldehyde remained at 15 min, but was not detected at 30 min. From these results, a decoction was prepared by adding Perilla Herb 1, 2, 5 and 10 min prior to the termination time of decoction at 30 min. When Perilla Herb was added 1 min beforehand, perillaldehyde was abundant (1.58 mg) and the amount of rosmarinic acid was already the same as that in the conventional decoction at 30 min, but the amount of total extract was inadequate. When Perilla Herb was added 5 min beforehand, perillaldehyde remained to some extent (0.61 mg) and rosmarinic acid was significantly increased compared with that in the conventional decoction at 30 min. From these results, we can conclude that the decoction should be prepared by boiling for not more than 30 min and, if possible, Perilla Herb should be added 5 min prior to the termination time. Considering the antidepressant-like activity of perillaldehyde and rosmarinic acid, the suggested method will not only achieve better treatment for anxiety, but also provide an effective use of crude drugs in the resource-limited environment.

A new lignan from Balanophora abbreviata and inhibition of lipopolysaccharide (LPS)-induced inducible nitric oxide synthase (iNOS) expression.

Six lignans including a new lignan (1), beta-sitosterol glucopyranoside and phenylpropanoids were isolated from the whole plants of Balanophora abbreviata Bl. (Balanophoraceae). Their structures were determined by NMR, MS analysis and other spectroscopic methods. Lignans (1, 2 and 4) showed potent inhibitory activities on the lipopolysaccharide (LPS)-induced inducible nitric oxide synthase (iNOS) expression in RAW 264.7 cells.

Immunomodulatory constituents from an Ascomycete, Chaetomium seminudum.

A screening study focusing on immunomodulatory activity of the EtOAc extract of an Ascomycete, Chaetomium seminudum, has afforded a known epipolythiodioxopiperazine, chetomin (1), together with three new chetomin-related metabolites named chetoseminudins A (2), B (3), and C (4). Among these four metabolites, 1 and 2 have been deduced as the immunosuppressive features of this fungus.

A new ent-clerodane diterpene from the aerial parts of Baccharis gaudichaudiana.

A new ent-clerodane diterpene, named bacchariol (1) was isolated from the aerial parts of Baccharis gaudichaudiana DC. (Compositae), together with known ent-clerodane diterpenes (2, 3), eight known flavonoids (4-11) and 3, 5-dicaffeoylquinic acid (12). Their structures were determined by spectroscopic analyses. Flavonoids (7, 8, 11) and 12 showed moderate scavenging activities toward 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radicals.

Ardisiphenols and other antioxidant principles from the fruits of Ardisia colorata.

Novel alkylphenols, ardisiphenols A-C (1-3) and a novel bergenin derivative, demethoxybergenin (10) were isolated from the fruits of Ardisia colorata (Myrsinaceae), together with known alkylresorcinols (4-6), embelin (7), myricetin (8), quercetin (9), bergenin (11), norbergenin (12), kaempferol (13), quercetin-3-O-beta-D-glucopyranoside (14) and gallic acid (15). Their structures were determined by NMR, MS(/MS) analyses and other spectroscopic methods. Ardisiphenols showed moderate scavenging activities toward 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals and showed cytotoxicity against the murine breast cancer cell line, FM3A.