Résumé
Objective: To establish an HPLC fingerprint of Cnidii Fructus formula granule analysis method for simultaneous determination of six main coumarin components, including osthol, xanthotoxin, xanthotol, bergapten, imperatorin and isopimpinellin, in order to provide reference for the study of the material basis of Cnidii Fructus formula granule. Methods: The method was performed by high performance liquid chromatography with a Waters XBridge C18 (250 mm × 4.6 mm, 5 μm) column and methanol (A)-0.1% acetic acid (B) as the mobile phase for gradient elution. The flow rate was 0.5 mL/min, the injection volume was 10 μL and the column temperature was 40 ℃. The detection wavelength was set at 320 nm. The chromatographic fingerprint evaluation system published by the State Pharmacopoeia Commission (2012 Edition) was used to establish the fingerprint of Cnidii Fructus formula granule, and the content of six main coumarin components was simultaneously determined. Results: The research on the 18 batches of Cnidii Fructus formula granule showed that the fingerprint similarity was greater than 0.992 and 19 common peaks were calibrated with satisfied peak resolution. The content determination results showed that the content of both xanthotoxin and osthol were the main coumarin components in Cnidii Fructus formula granule. According to the methodological investigation, the precision RSD values were all less than 1.6%. The sample was stable within 48 h and this method had good repeatability. The average recovery rates of xanthotol, xanthotoxin, imperatorin, isopimpinellin, bergapten and osthol were 100.69%, 101.03%, 99.48%, 100.88%, 101.27% and 100.35%, respectively. All of these coumarin components’ RSD were less than 2.5%. The six components showed a good linear relationship within a certain concentration range. The results of the content determination of xanthotol, xanthotoxin, isopimpinellin, bergapten, imperatorin and osthol respectively were 8.01-8.29, 2.37-2.63, 4.30-4.61, 4.04-4.40, 3.45-3.90 and 6.02-6.80 mg/g among the 18 batches of the Cnidii Fructus formula granule. Conclusion: The fingerprint method and the determination method of six main coumarin components in the Cnidii Fructus formula granule established in this study are simple, stable, accurate and reliable. This method can be used for the quality control of the Cnidii Fructus formula granule.
Résumé
Objective To study the chemical constituents from the fresh fruits of Physalis pubescens. Methods Compounds were isolated and purified by repeated column chromatography on silica gel column, Sephadex LH-20 gel column, and preparative HPLC methods. Physicochemical properties and spectroscopic methods were employed for the structure elucidation. Results Twenty compounds were obtained and identified as osthol (1), xanthotoxin (2), isopimpinellin (3), imperatorin (4), (-)-meranzin hydrate (5), auraptenol (6), 1-(4-hydroxy-3,5-dimethoxyphenyl) ethanone (7), 2,5-dimethoxybenzoquinone (8), 2-(4-hydroxyphenyl) acetic acid (9), (S)-methyl 2-hydroxy-2-(4-hydroxyphenyl) acetate (10), pyrocatechol 1-O-β-D-glucopyranoside (11), benzyl β-D-glucopyranosyl (1→6)-β-D-glucopyranoside (12), 2-phenylethyl-O-β-D-glucopyranoside (13), p-hydroxybenzene propanoic acid (14), 3,4-dihydroxybenzene propionic acid (15), (1-O-p-coumaroyl)-(6-O-β-glucosyl)-β-glucoside (16), 1-O-trans-cinnamoyl-β-D- glucopyranosyl-(1→6)-β-D-glucopyranoside (17), N-trans-feruloyltyramine (18), kaempferol 3-O-β-D-glucopyranoside (19), and bergapten (20), respectively. Conclusion Compounds 1-16 are obtained from Physalis genus for the first time.
Résumé
Objective To study the chemical constituents in 95% ethanol aqueous extract of Trigonostemon lutescens. Methods The open silica gel, MCI, Sephadex LH-20 column chromatography, and the semi-preparative HPLC were used to isolate and purify the chemical constituents from the EtOAc fraction of T. lutescens. The structures of the isolates were elucidated by their physiochemical properties, NMR, and MS spectroscopic data, as well as comparison with literature data. Results Eleven compounds were isolated from the EtOAc fraction of the 95% aqueous EtOH extract of T. lutescens, and their structures were identified as seven coumarins, auraptenol (1), meranzin hydrate (2), xanthotoxin (3), bergapten (4), isoimpinellin (5), alloisoimperatorin (6), and isodemethylfuropinarine (7); Two phenylalanine glycosides, 3,4-dihydroxy allylbenzene-4-O-β-D-glucopyranoside (8) and 1-O-β-D-glucopyranosyl-4- allylbenzene (9); One phenylethanol, 1-(2-ethylphenyl)-1,2-ethanediol (10); One alkaloid, 8-hydroxy-3-methoxy-5H-pyrido [2,1-c] pyrazin-5-one (11). Conclusion All these compounds are isolated from the genus Trigonostemon for the first time.
Résumé
Objective: To investigate the effects of xanthotoxin from Apiaceae medicinal plants on cell proliferation and apoptosis, and explore its mechanism of action against human gastric carcinoma SGC-7901 cells in vitro. Methods: SGC-7901, HepG-2, MCF-7, and A549 cells were treated with different concentrations of xanthotoxin (10, 20, 60, 80, 100, 120, 140, and 160 µg/mL) for 48 h, and the cell viability (IC50) was determined by MTT assay; Xanthotoxin-induced apoptosis in cells was observed by using Hoechst 33258 Staining Kit and Annexin V-FITC Apoptosis Detection Kit; Flow cytometry was used to detect apoptosis related proteins of Fas/FasL, Bid, and DR5/TRAIL proteins in human gastric carcinoma SGC-7901 cells after being treated by xanthotoxin; The influence of xanthotoxin on Caspase-8 protein expression in the cells was determined by Flouormetric Assay Kit. Results: Xanthotoxin obviously inhibited SGC-7901, HepG-2, MCF-7, and A549 cells proliferation, and its inhibition was in a concentration-dependent manner; flow cytometry results showed that in a certain concentration range, xanthotoxin can increase the expression levels of Fas/FasL and DR5/TRAIL proteins in a concentration-dependence manner. The content of Bid protein in cells was increased, and it showed concentration-dependence. Conclusion: Xanthotoxin may induce SGC-7901 cells apoptosis in a certain concentration range through the Fas/FasL protein mediated death receptor pathway, or by DR5/TRAIL mediated death receptor pathway, and increase the expression level of death receptor protein, activation Caspase-8, activating downstream effect factor, inducing cell apoptosis, or activate Caspase-8 cutting activate protein Bid, and then enter the mitochondrial pathway, induction of apoptosis.
Résumé
Objective: The chemical constituents were isolated and identified in order to find the bioactive natural products from Gelsemium elegans. Methods: Silica gel, sephadex LH-20 and HPLC column chromatographic techniques were used for separation and purification of the compounds and extensive spectral analysis spectrum were employed for structural elucidation. Results: Fifteen compounds were separated from G. elegans, identified by physicochemical properties, spectral analysis and other means. They are koumine (1), gelsemine (2), gelsevirine (3), gelsemicine (4), xanthotoxin (5), bergapten (6), isopimpinellin (7), imperatorin (8), osthol (9), p-hydroxybenzoic acid (10), vanilla acid (11), β-sitosterol (12), β-daucosterol (13), gelsemamide (14), and 16-epivoacarpine (15). Conclusion: Compounds 5-9 are first isolated from the plants in genus Gelsemium Juss.
Résumé
Objective: To study the chemical constituents of lipophilic parts from the roots of Angelica dahurica. Methods: The compounds were separated and purified by repeated column chromatography on silica gel and HPLC, as well as the chemical structures of isolated compounds were determined by spectral data analyses. Results: Forty compounds were obtained and identified as isoimperatorin (1), psoralen (2), bergapten (3), β-sitosterol (4), imperatorin (5), phellopterin (6), xanthotoxin (7), isopimpinellin (8), dehydrogeijerin (9), isooxypeucedanin (10), oxypeucedanin (11), dibutylphthalate (12), umbelliferone (13), xanthotoxol (14), 5-hydroxy-8-methoxypsoralen (15), p-hydroxyphenethyl-trans-ferulate (16), pabularinone (17), isobyakangelicol (18), heraclenin (19), columbianetin (20), isogosferol (21), pabulenol (22), byakangelicin ethoxide (23), neobyakangelicol (24), oxypeucedanin methanolate (25), isoscopletin (26), 1′, 2′-dehydromarmesin (27), angelol A (28), angelol E (29), tert-O-methylheraclenol (30), (-)-marmesin (31), dahurianol {8-[(2′R)-2′, 3′-dihydroxy-3′-methylbutoxy] bergaptol; 32}, oxypeucedanin hydrate (33), heraclenol (34), byakangelicin (35), angelol L (36), angelol G (37), trans-coniferylaldehyde (38), vanillic acid (39), and trans-ferulic acid (40), respectively. Conclusion: Compound 32 is a new natural product named dahurianol.
Résumé
Objective: To develop an HPLC-DAD method for the simultaneous determination and comparative analysis on the contents of prim-O-glucosylcimifugin, 4′-O-β-D-glucosyl-5-O-methylvisamminol, cimifugin, sec-O-glucosylhamaudol, psoralen, imperatorin, bergapten, and xanthotoxin in the roots of Saposhnikovia divaricata from Longxi areas. Methods: The extracts were obtained by methanol reflux method and the contents of the eight compounds were determined by HPLC-DAD. The mobile phase, consisting of acetonitrile-water, was programmed for a gradient elution. The flow rate was 1.0 mL/min, the detection wavelength was 254 nm, and the column temperature was 25°C. Results: Excellent linearity with correlation coefficents (r) of 0.9992-0.9999 was obtained. The average recoveries of the eight compounds were 96.2%-104.1% and all RSD values were less than 3%. The contents of the four coumarins in the roots of S. divaricata were much less than those four chromones. The contents of the four coumarins in the roots of Carum carvi and Peucedahum ledebourielloides were more than those in S. divaricta, while no chromones were detected except sec-O-glucosylhamaudol in P. ledebourielloides. Conclusion: The method appears to be simple, accurate, and well reproducible, which could be used for the simultaneous determination of the above-mentioned eight compounds in S. divaricata. According to the above analysis, C. carvi and P. ledebourielloides could not be used as the succedaneum of S. divaricata on the basis of the eight compound contents.
Résumé
Objective: To study the absorption and transportation characteristic of xanthotoxol (1), xanthotoxin (2), imperatorin (3), isoimperatorin (4), cnidilin (5), and isopimpinellin (6), which were classified as the linear type furocoumarins, in a model of Caco-2 cell monolayers in human intestinal epithelium. Methods: Caco-2 (the human colon adenocarcinoma cell lines) cell monolayer was used as an intestinal epithelial cell model. The permeability of the six coumarins from apical side (AP side) to basolateral side (BL side) or from BL side to AP side was evaluated. The concentration of the six coumarins was measured by HPLC coupled with UV detector. Transportation parameters and permeability coefficients (Papp) were then calculated, and P app values were compared with the reported values for model compounds, Propranolol and Atenolol. Based on the absorption and transportation characteristic of coumarins 1-6, and psoralen (7), bergaptol (8), bergaptol-O-β-D-glucopyranoside (9), bergapten (10), nodakenin (11), nodakenetin (12), decuroside V (13), umbelliferone (14), osthole (15), angelol-A (16), and angelol-B (17) in a model of Caco-2 cell monolayer, the relationship of absorption and transportation with diversed chemical structures and lipophilicity was reviewed. Results: In the Caco-2 cell monolayer model, the Papp magnitudes of the linear furocoumarins 1-6 were 10-5 cm/s in the bi-directional transport, which was identical with Propranolol. And the permeability of Caco-2 cell monolayer is mainly via passive absorption. Conclusion: The above-mentioned linear furocoumarins 1-6 are well-absorbed compounds. The results show that a significant Sigmoid dependence of permeability on 1g Papp AP→BL and 1g D at pH 7.35 of all 1-17 furocoumarins can be absorbed across intestinal epithelial cells by passive diffusion mechanism.