Simultaneous Determination of Seven Constituents in Qinggan Lidan Mixture by HPLC / 中国实验方剂学杂志

Objective: To establish an HPLC method for simultaneous determination of geniposide,chlorogenic acid,neochlorogenic acid,cryptochlorogenic acid,isochlorogenic acid A,B and C in Qinggan Lidan mixture,in order to provide references for its quality control. Method: The analysis of methanol extract of this drug was performed on a 35℃ Luna C18 column (4.6 mm×250 mm,5μm),with the mobile phase comprised of acetonitrile-0.4% phosphoric acid flowing at 1.0 mL·min-1 in a gradient elution mode (0-10 min,8%-12%A;10-30 min,12%A;30-60 min,12%-35%A),and the detection wavelengths were set at 238 and 327 nm. Result:Geniposide,chlorogenic acid,neochlorogenic acid,cryptochlorogenic acid,isochlorogenic acid A,B and C were completely separated,and well separated from other constituents. The linear ranges of geniposide,chlorogenic acid,neochlorogenic acid,cryptochlorogenic acid,isochlorogenic acid A,B and C were 0.188-2.355,0.083-1.040,0.074-0.920,0.075-0.940,0.064-0.800,0.076-0.955,0.071-0.888 μg (r ≥ 0.999 0),respectively. The average recoveries were 99.45%,98.45%,99.06%,98.50%,98.16%,101.01%,96.93%,with the RSDs of 0.5%,1.8%,1.3%,2.4%,2.3%,1.6%,1.6%,respectively.The contents of geniposide,chlorogenic acid,neochlorogenic acid,cryptochlorogenic acid,isochlorogenic acid A,B and C were 3.420-3.794,0.835-0.890,1.222-1.275,1.064-1.210,0.377-0.398,0.419-0.464 and 0.362-0.405 g·L-1,respectively. Conclusion:This method can be used for simultaneous determination of muti-ingredients in Qinggan Lidan mixture,and the established method is simple and accurate,with a good reproducibility and high sensitivity. It can be used for the quality control of Qinggan Lidan mixture.

Selection of quality markers of Jasminum amplexicaule based on its anti-diarrheal and anti-inflammatory activities: Effect-target affiliation-traceability-pharmacokinetics strategy / 中草药·英文版

Objective: To investigate therapeutic mechanism in Jasminum amplexicaule (Oleaceae) and verify its main active component as quality control markers Methods: Established mouse models of diarrhea, intestinal angina, and inflammation were firstly used to select herb fractions with optimum efficacy, followed by an in vitro experiment to determine key targets associated with effects of J. amplexicaule extract. The selected fractions were isolated and purified, its components were identified, and the obtained compounds were verified for their effects on NF-κB and iNOS. Finally, effective compounds were administered to rats, their plasma pharmacokinetic parameters were calculated, and quality markers (QMs) reflecting therapeutic activities of J. amplexicaule were confirmed. Results: Trichloromethane and ethyl acetate fractions had significant anti-diarrheal, anti-inflammatory, and analgesic effects. The trichloromethane fraction also reduced BDNF, p38 MAPK, p-p38 MAPK, NF-κB p65, and p-NF-κB p65 levels in the ileum in a rhubarb-induced diarrhea mouse model. Additionally, it inhibited LPS-induced NF-κB transcription and nitric oxide (NO) production in RAW264.7 macrophages, which suppressed iNOS expression. Therefore, the trichloromethane fraction was further investigated. QMs candidate selection identified 17 compounds, and results of in-vitro therapeutic validation indicated that methyl caffeate and isochlorogenic acid B had the strongest anti-diarrheal, anti-inflammatory, and analgesic activities. After being validated by a UHPLC–MS-MS method, concentrations of these target compounds were accurately determined in the rat plasma and pharmacokinetic parameters were calculated. Cmax, tmax, and t1/2 were respectively 575.35 ng/mL (2.963 nmol/mL), 0.5 h, and 0.45 h for methyl caffeate and 262.03 ng/mL (0.5034 nmol/mL), 0.25 h, and 2.03 h for isochlorogenic acid B. Because these candidate compounds exhibited favorable pharmacokinetics, they were considered as QMs of J. amplexicaule. Conclusions: The present study accurately and effectively identified QMs of J. amplexicaule that act as indicators of efficacy and quality.

Simultaneous determination of ten components in Lonicerae Japonicae Flos from different habitats by UPLC / 中草药

Objective: To establish a method for the determination of seven organic acids, one flavone and two iridoid terpenoids in Lonicerae Japonicae Flos from different habitats. Methods: The content of 10 components in Lonicerae Japonicae Flos from different habitats were determined by ultra-performance liquid chromatography (UPLC). The mobile phase for gradient elution was 0.1% phosphoric acid solution (A)- methanol (B); The flow rate was 0.3 mL/min, and the column temperature was 30 ℃. Results: A UPLC method for simultaneous determination of seven organic acids, one flavone and two iridoid terpenoids in Lonicerae Japonicae Flos was established. Principal component analysis (PCA) and partial least squares method (PLS-DA) were used to analyze the distribution and characteristics of 10 constituents of Lonicerae Japonicae Flos collected from different habitats; Three production areas of Shandong, Jiangsu and Shaanxi are respectively grouped into one group. Conclusion: The method is stable and feasible, which could be used as a reference for evaluating the quality of Lonicerae Japonicae Flos in a more comprehensive way.

Establishment of UPLC fingerprints of Jasminum elongatum and determination of two constituents / 中成药

AIM To establish the UPLC fingerprints of Jasminum elongatum (Bergium) Wild.and to determine the contents of isochlorogenic acid B and ethylcaffeate.METHODS The analysis of methanol extract of J.elongatum was developed on a 25 ℃ thermostatic Agilent Ecplise XDB-C18 column (2.1 mm × 100 mm,1.7 μm),with the mobile phase comprising of acetonitrile-0.1％ methanoic acid flowing at 0.5 mL/min in a gradient elution manner,and the detection wavelength was set at 260 nm.RESULTS There were eighteen common peaks in the fingerprints of ten batches of samples,with the similarities of more than 0.85.Isochlorogenic acid B and ethylcaffeate showed good linear relationships within the ranges of 7.67-38.35 μg/mL (R2 =0.999 4),9.60-96.0 μg/mL (R2 =0.999 7),whose average recoveries were 98.61％,99.09％ with the RSDs of 0.84％,1.25％,respectively.CONCLUSION This stable and reliable method can be used for the quality control of J.elongatum.

Accumulation rules of effective constituents in Farfarae Flos of different growth stage based on HPLC characteristic fingerprint / 中草药

Objective To analyze the accumulation patterns of active constituents in Farfarae Flos with different flower bud colors (yellow, purple, and deep purple) at different growth stages, and provide theoretical guidance for the production and quality control of Farfarae Flos. Methods The medicinal materials of Farfarae Flos of different growth stages with different colors were determinated by HPLC method. The Similarity Evaluation System for Chromatographic Fingerprint of TCM (2012 A edition) was used to evaluate the similarity of the samples. The differences among samples were identified by chemical pattern recognition methods including hierarchical principal component analysis (PCA) and partial least squares discriminate analysis (PLS-DA). Results The HPLC fingerprint of different flower bud colors of Farfarae Flos at different growth stages was obtained, 27 common peaks were found in the chromatography, and 11 of them were identified. Similarities of samples of all batches with reference fingerprint were among 0.901-0.995. There were differences in the accumulation characteristics of Farfarae Flos at different growth stages according to the peak area, and showing significant differences among different flower bud colors. PCA and PLS-DA results demonstrated obvious distinction among different flower bud colors. Twelve constituents, such as gallic acid, chlorogenic acid, rutin, hyperin, isochlorogenic acid B and quercetin, were screened as biomarkers, representing major differences among colors. The quality evaluation demonstrated that deep purple buds was the best, followed by purple buds and yellow buds for the worst. Conclusion The HPLC fingerprint can reflect the accumulation characteristics of the active constituents of Farfarae Flos in different growth stages and the differences among different flower bud colors. Combining chemical pattern recognition can provide reference for the production and quality evaluation of Farfarae Flos.

Protective effect of isochlorogenic acid B on CCl4 induced acute liver injury in mice / 国际药学研究杂志

Objective To investigate the effect of isochlorogenic acid B(ICAB)on CCl4-induced acute liver injury(ALI)in mice. Methods The animal model of CCl4-induced ALI in mice was established and then the protective effect of ICAB was evaluated using this model. Serum levels of alanine transaminase(ALT)and aspartate aminotransferase(AST),hepatic levels of malondialdehyde (MDA)and the activities of superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px)were measured by spectrophotometric methods. Liver cell morphology was observed by hematoxylin and eosin staining method and the effects of ICAB on the protein expres-sion of nuclear factor erythroid-2-related factor 2(Nrf2),heme oxygenase-1(HO-1)and quinone oxidoreductase 1(NQO1)in mice he-patocyte were determined by Western blot method. Results ICAB(5,10 and 20 mg/kg)significantly protected against CCl4-induced liver injury by reducing the elevated levels of serum aminotransferases and hepatic MDA and remarkably restored the impaired antioxi-dants. Meanwhile,the histopathological changes were also attenuated in mice. In addition,ICAB could induce the protein expression of Nrf2 and promote its nuclear translocation,and further increase the protein expression of HO-1 and NQO1. Conclusion ICAB has protective effect against CCl4-induced ALI in mice,which is mainly due to its ability to promote the nuclear translocation of Nrf2 and decrease the oxidative stress.

Simultaneous determination of chlorogenic acid, aesculetin, isochlorogenic acid B, and isochlorogenic acid A in Cichorii Herba by QAMS / 中草药

Objective: To establish the quantitative analysis of multi-components by single marker (QAMS) method to determine the content of chlorogenic acid, aesculetin, isochlorogenic acid B, and isochlorogenic acid A in Cichorii Herba (chicory). Methods: Selecting aesculetin as the internal reference substance, the relative correction factors (RCF) of chlorogenic acid, isochlorogenic acid B and isochlorogenic acid A were established respectively, and then the contents of the three constituents were calculated by RCF, to achieve the quality of chicory through QAMS. At the same time, the external standard method was used to determine the content of four constituents in chicory and compare the difference between calculated values and measured values, so as to verify the construction method for accuracy, applicability, and repeatability. Results: No significant difference was observed in chlorogenic acid, isochlorogenic acid B, and isochlorogenic acid A from eight batches of chicory in the quantitative results by these two methods. The validated QAMS method had good precision, reproducibility, and reliability. Conclusion: The established QAMS method is suitable and feasible for the quality control of chicory.

Chemical constituents in stems of Ilex cornuta / 中草药

Objective: To investigate the chemical constituents in the stems of Ilex cornuta and the ability of scavenging free radicals of compounds 1-9. Methods: The compounds were isolated and purified by silica gel, medium pressure column chromatography, and semi-preparative liquid chromatography, and their structures were elucidated by chemical properties and spectroscopic analyses. The antifreeradical efficiency of compounds 1-9 was evaluated by DPPH radical scavenging assay. Results: Fifteen compounds were isolated and their structures were identified as isochlorogenic acid B (1), 3,4,5-tricaffeoylquinic acid (2), 4,5-di-O-caffeoyl quinic acid methyl ester (3), 3,4-di-O-caffeoyl quinicacid methyl ester (4), 3,5-di-O-caffeoyl quinicacid methyl ester (5), 3,4,5-tri-O-caffeoyl quinic acid methyl ester (6), 3,5-dimethoxy-4-hydroxybenzaldehyde (7), ethyl gallate (8), dihydrosyringenin (9), 2,6-dimethoxy-1,4-benzoquinone (10), arctigenin (11), 1-O-(vanillic acid)-6-O-(3″, 5″-dimethoxy-galloyl)-β-D-glycoside (12), (+)-1-hydroxypinoresinol-1-O-β-D-glucopyranoside (13), (+)-(7S,8S)-syringylglycerol 8-O-β-D-glucopyranoside (14), and schaftoside (15). Compounds 1-7 had good antifreeradical efficiency. Conclusion: Compounds 6,8-10,14, and 15 are obtained from the plants of Ilex L. the first time, and compounds 2,7,11, and 12 are obtained from this plant for the first time. Compounds 1-6 have good antifreeradical efficiency.

Preparation technology of isochlorogenic acids A, B, and C / 中草药

Objective: To establish a method for separation of isochlorogenic acids A, B, and C from Lonicerae Flos. Methods: Isochlorogenic acids A, B, and C in Lonicerae Flos were isolated and purified by macroporous resin and medium-low-pressure preparative chromatography. Their structures were identified on the basis of the spectral data and physicochemical property. Results: The contents of prepared isochlorogenic acids A, B and C were 98.7%, 99.2%, and 97.6%, respectively. Conclusion: This method is economic, simple, rapid, and effective for the preparation of isochlorogenic acids A, B, and C with high purity.

Simultaneous determination of the contents and fingerprint of multi-components in honeysuckle extract by HPLC / 中国药学杂志

OBJECTIVE: To establish a HPLC method for the simultaneous determination of the contents and fingerprint of 7 active constituents (neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, caffeic acid, isochlorogenic acid B, isochlorogenic acid A, and isochlorogenic acid C) in honeysuckle extract. METHODS: The chromatographic separation was achieved on a AkzoNobel Kromasil C18 column(4.6 mm × 250 mm, 5 μm), the mobile phase was acetonitrile-0.1% phosphoric acid aqueous solution with gradient elution at a flow rate of 1.0 mL · min-1, the detection wavelength was set at 326 nm, and the column temperature was 30°C. RESULTS: All the 7 compounds showed good linearity in the ranges of the test concentrations. The RSDs of the precision, stability and reproducibility tests were less than 3%. The average recoveries were in the range of 97.98%-99.29%. CONCLUSION: This method is simple, sensitive, accurate, and can be used for quality control of honeysuckle extract. Copyright 2012 by the Chinese Pharmaceutical Association.

HPLC fingerprint and pattern recognition of Deng's Herbal Tea Granula / 中草药

Objective: To establish a method for the quality control of Deng's Herbal Tea Granula. Methods: An HPLC method was developed to establish the fingerprint of Deng's Herbal Tea Granula methanol extract and 11 samples from various batches were analyzed. Furthermore, principal component analysis (PCA) was used to investigate pattern recognition. Results: The similarity among 11 batches of samples was no less than 0.95. The PCA showed that 16 common peaks were integrated into three principal components and their accumulation contributing rate amounted to 91.89%. The relative peak area of isochlorogenic acid B was the most effective index in fingerprint of Deng's Herbal Tea Granula. Conclusion: This method is available for the quality evaluation and quality control of Deng's Herbal Tea Granula.