Flavonoids rather than alkaloids as the diagnostic constituents to distinguish Sophorae Flavescentis Radix from Sophorae Tonkinensis Radix et Rhizoma: an HPLC fingerprint study.

Sophorae Flavescentis Radix (Sophora flavescens Ait., SFR) and Sophorae Tonkinensis Radix et Rhizoma (S. tonkinensis Gapnep., STR) are two commonly used traditional Chinese medicines from Sophora (Leguminosae) plants, which are believed to possess similar bioactive components with entirely different clinical applications. In order to find out the characteristic chemical constituents potentially leading to the unique medicinal properties claimed for each of the two closely related TCMs, an HPLC fingerprint method was developed for analyses of the alkaloid and flavonoid constituents of SFR and STR, respectively, which were further evaluated and compared through similarity calculation and hierarchical clustering analysis (HCA). The results from the present study showed that the alkaloid fingerprints of the two herbs were similar, with many components co-existing in both drugs and various batches of samples from different species being mixed together in the HCA dendrogram. However, their flavonoid constituents were totally different with specific fingerprints being yielded for each herb, and further HCA analysis showed that the tested samples could almost be clearly divided into two groups based on their origins of species. The results from the present study indicated that the flavonoid constituents could serve as the differentially diagnostic constituents of SFR and STR and might potentially attributed to their distinct therapeutic effects.

Preparative separation and quantitative determination of two kaurenoic acid isomers in root barks of Acanthopanax gracilistylus.

The kaurenoic acid-type diterpenoids in Acanthopanacis Cortex have been reported to be the major active components. However, the diterpenoids are present as position isomers that exacerbate the challenges in obtaining standards compounds. Little work has been done on the quantitative analysis of the diterpenoids in the herb. In the present study, two diterpenoid isomers ent-16ßH,17-isovalerate-kauran-19-oic acid (1) and ent-16ßH,17-methyl butanoate-kauran-19-oic acid (2) with high purity were separated by analytical HPLC, followed by recrystallization in acetone. Furthermore, an HPLC-ELSD method was developed and validated for simultaneous determination of 1 and 2 in 9 batches of Acanthopanacis Cortex samples. The HPLC separation and quantification was achieved in 40 min using an Agela Promosil C18 column eluted with a gradient of water and acetonitrile. The calibration curves showed good linearity (r2 ≥ 0.999 9) within the test ranges. The LOD ranged from 0.407 2 to 0.518 0 µg and LOQ ranged from 1.018 0 to 1.295 0 µg. The precisions (%RSD) were within 1.47% for the two isomers. The recovery of the assay was in the range of 98.78%-99.11% with RSD values less than 2.76%. It is the first time to establish a quantitative HPLC method for the analysis of the bioactive kaurenoic acid isomers in the herb.

[Anti-complement alkaloids from whole plants of Viola yedoensis].

Fifteen alkaloids were isolated from the 95% ethanol extract of the whole plants of Viola yedoensis by various column chromatographic techniques such as silica gel and Sephadex LH-20. Their structures were identified as neoechinulin A（1）,N-benzoyl-L-p-hydroxy-phenylalaninol（2）,aurantiamide acetate（3）,aurantiamide（4）,anabellamide（5）,trichosanatine（6）,indole-3-carboxylic acid methyl ester（7）,3-carboxyindole（8）,N-trans-feruloyl-tyramine（9）,paprazine（10）,7'-（3', 4'-dihydroxyphenyl）-N-[(4-methoxyphenyl)ethyl]propenamide（11）,cannabisin F（12）,N-(4-hydroxyphenethyl)octacosanamide（13）,N-(4-hydroxyphenethyl)hexacosanamide（14）and N-benzoyl-L-phenylalaninol（15）. All the compounds except 3 and 4 were isolated from this plant for the first time. These alkaloids exhibited anti-complement activity against the classical pathway(CP)and the alternative pathway(AP)with the CH50 and AP50 values ranging from 0.12 to 0.33 g•L⁻¹ and 0.22 to 0.50 g•L⁻¹, respectively. Preliminary mechanism study using complement-depleted sera showed that these alkaloids acted on different complement components in the complement activation cascade.

Anti-Complementary Components of Helicteres angustifolia.

A first phenalenon derivative with an acetyl side chain at C-8, 8-acetyl-9-hydroxy-3-methoxy-7-methyl-1-phenalenon (compound 1), and a pair of new sesquilignan epimers at C-7″ of hedyotol C and hedyotol D analogs, hedyotol C 7″-O-ß-d-glucopyranoside (compound 2) and hedyotol D 7″-O-ß-d-glucopyranoside (compound 3) were isolated from the aerial parts of Helicteres angustifolia together with nine known compounds (4-12). Their structures were elucidated on the basis of spectroscopic methods, including mass spectroscopy, and 1D and 2D nuclear magnetic resonance. Eleven isolates exhibited anti-complementary activity. In particular, compounds 4 and 5 exhibited potent anti-complementary activities against the classical and alternative pathways with CH50 values of 0.040 ± 0.009 and 0.009 ± 0.002 mM, and AP50 values of 0.105 ± 0.015 and 0.021 ± 0.003 mM, respectively. The targets of compounds 4 and 5 in the complement activation cascade were also identified. In conclusion, the anti-complementary components of H. angustifolia possessed chemical diversity and consisted mostly of flavonoids and lignans in this study.

Regio- and stereo-selective hydroxylations of ingenane diterpenoids by Mortierella ramanniana and Gibberella fujikuroi.

The regio- and stereo-selective hydroxylations of two ingenane diterpenoids, 20-deoxyingenol (1) and 13-oxyingenol dodecanoat (2), by the filamentous fungi Mortierella ramanniana and Gibberella fujikuroi were investigated in the present study. Four undescribed metabolites (3-6) of substrate 1 and two undescribed metabolites (7 and 8) of substrate 2 were isolated. All the metabolites were identified as hydroxylated ingenane derivatives by extensive NMR and HR-ESI-MS data analyses. All the biotransformed compounds and the substrates were evaluated for their cytotoxicities against three human cancer cell lines, including human colon cancer Caco-2, breast cancer MCF-7, and adriamycin (ADM)-resistant MCF-7/ADM cell lines. All ingenane alcohols (1, and 3-6) displayed no significant cytotoxic activities. The substrate 13-oxyingenol dodecanoat (2) showed moderate cytotoxicity with IC50 values being 35.59 ± 5.37 µmol·L-1 (Caco-2), 24.04 ± 4.70 µmol·L-1 (MCF-7), and 22.24 ± 5.19 µmol·L-1 (MCF-7/ADM). However, metabolites 7 and 8 displayed no significant cytotoxicity. These results indicated that the hydroxylation at the C-13 aliphatic acid ester of substrate 2 can significantly reduce the cytotoxic activity.

Anticomplement monoterpenoid glucosides from the root bark of Paeonia suffruticosa.

Six new (1-6) and 19 known monoterpenoid glucosides were isolated from the root bark of Paeonia suffruticosa. The monoterpenoid glucosides 1, 2, 7, 10-19, and 22 exhibited anticomplement effects with CH50 and AP50 values ranging from 0.14 to 2.67 mM and 0.25 to 3.67 mM, respectively. In a mechanistic study, suffrupaeoniflorin A (1) interacted with C1q, C3, C5, and C9, while galloylpaeoniflorin (12) and galloyloxypaeoniflorin (19) acted on C1q, C3, and C5 components in the complement activation cascade.

Qualitative and quantitative analysis of flavonoids in Sophora tonkinensis by LC/MS and HPLC.

AIM: To develop analytical methods for the identification and determination of the flavonoids in Sophora tonkinensis for comprehensive quality evaluation. METHODS: An HPLC-DAD-ESI-MS method was used for the separation and characterization of the flavonoids in S. tonkinensis, and a liquid chromatographic method was employed to simultaneously determine five major active flavonoids in this crude drug. RESULTS: Seventeen flavonoids were identified, among which, seven were unambiguously identified as trifolirhizin, quercetin, formononetin, macckiain, kurarinone, sophoranone, and sophoranochromene by comparing their retention times, and UV and MS spectra with those of the authentic compounds, and the other ten flavonoids were tentatively identified by comparing their UV and MS/MS spectra with those of literature data. Furthermore, five major active flavonoids, including trifolirhizin, quercetin, maackiain, sophoranone, and sophoranochromene were determined in S. tonkinensis. All calibration curves expressed good linearity (r > 0.999 8) within the test ranges, and the recovery from this method was 96.40%-104.43%. The developed HPLC method was successfully applied to quantitatively determine the five flavonoids in seventeen samples of S. tonkinensis. CONCLUSION: The developed method rapidly characterized the bioactive flavonoids of S. tonkinensis, and could be readily utilized to enhance the quality assurance approaches for this traditional Chinese medicine.

Viola yedoensis liposoluble fraction ameliorates lipopolysaccharide-induced acute lung injury in mice.

Viola yedoensis is a component of traditional Chinese herb medicine for inflammatory diseases. Chemical constituents of V. yedoensis have been shown to possess antibacterial, anti-HIV, and anticoagulant effects in experimental research; however, their anti-inflammatory properties remain to be demonstrated. In this study, a mouse model of lipopolysaccharide (LPS)-induced acute lung injury was used to investigate the effect of petroleum ether fraction of V. yedoensis (PEVY) on inflammation in vivo. After being shown to have anti-complementary activity in vitro, PEVY was orally administered to the mice at doses of 2, 4, and 8 mg/kg. Treatment with PEVY significantly decreased the wet-to-dry weight ratio of the lung, total cells, red blood cells, protein concentration, and myeloperoxidase activity in bronchoalveolar lavage fluid. PEVY markedly attenuated lung injury with improved lung morphology and reduced complement deposition. In addition, PEVY suppressed the expression of pro-inflammatory cytokines, TNF-α, IL-1ß, and IL-6. Taken together, PEVY protects the lung from acute injury, potentially via inhibiting the activation of the complement system and excessive production of proinflammatory mediators.

[A study on quality standard for Herba Siegesbeckidae].

OBJECTIVE: To establish the qualitative and quantitative methods of Herba Siegesbeckiae. METHOD: A TLC method was used for qualitative identification and a HPLC analysis was applied for quantitative determination of Herba Siegesbeckiae with kirenol as the reference substances. RESULT: Chloroform-methanol-formic acid (25:5:1) as a mobile phase of TLC, the spot of kirenol can be easily detected; Methanol extracts of Herba Siegebeckiae were separated on a Polaris C18 column with acetonitrile-water (25:75) as mobile phase and kirenol was separated well. The average content of kirenol in Herba Siegebeckiae was 0.14%. A good linear relationship between the peak areas and injected amounts of kirenol in the range of 0.19-14.9 microg and the average recovery was 100.0% (RSD = 2.4%). CONCLUSION: The method can be used for qualitative identification and quantitation determination of Herba Siegesbeckiae.

[Identification of the fumatory Radix Bupleuri with sulfur by static headspace GC-MS].

OBJECTIVE: To identify whether Radix Bupleuri (Bupleurum chinense) was fumed with sulfur. METHOD: A static headspace GC-MS method was used to detect sulfur in the fumatory Radix Bupleuri, the authentic samples free of sulfur was detected as reference. RESULT: Sulfur was detected in six samples from nine samples collected in different locations. CONCLUSION: The method can be used to detect sulfur rapidly in the fumatory Radix Bupleuri with sulfur.

cis-Eudesmane sesquiterpene glycosides from Liriope muscari and Ophiopogon japonicus.

Two new cis-eudesmane sesquiterpene glycosides, liriopeoside A (1) and ophiopogonoside A (2), were extracted and purified from tubers of Liriope muscari and Ophiopogon japonicus, respectively, along with three known compounds. Their structures were elucidated as 1beta,6beta-dihydroxy-cis-eudesm-3-ene-6-O-beta-D-glucopyranoside (1) and 1beta,4beta,6beta-trihydroxy-cis-eudesmane-6-O-beta-D-glucopyranoside (2) by spectral data analysis. The structure and the relative configuration of compound 1 were confirmed by X-ray crystallographic analysis. This is the first time that cis-eudesmane-type sesquiterpenes have been reported from the genera Ophiopogon and Liriope.

[Triterpenoid saponins from bark Mitragyna inermis].

OBJECTIVE: To study the constituents of Mitragyna inermis that have an anti-tumor activity on Bel-7402 of hepatic carcinoma. METHOD: The compounds were isolated by column chromatography on silica gel, ODS, Sephadex LH-20 separately and their structures were elucidated by chemical and spectral technology. RESULT: Three quinovic acid glycosides, quinovic acid 3 beta-beta-D-glucopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-28-O-beta- D-glucopyranoside(I), quinovic acid 3 beta-O-beta-D-quinovopyranoside(II), quinovic acid 3 beta-O-beta-D-glucopyranoside(III), were obtained. CONCLUSION: I, II were isolated from Mitragyna for the first time.