Exploration of nanofiltration conditions for separation of alcohol precipitation liquid during preparation of Reduning Injection by Box-Behnken design combined with flux attenuation / 中国中药杂志

This study employed Box-Behnken design combined with flux attenuation to explore the nanofiltration conditions for separation of alcohol precipitation liquid during the preparation of Reduning Injection and discussed the applicability of nanofiltration in the separation of the liquid with high-concentration ethanol. The effects of nanofiltration molecular weight cut-off(MWCO) and pH on the rejection of chlorogenic acid, 3,5-dicaffeoylquinic acid, and 4,5-dicaffeoylquinic acid were consistent with the principles of pore size sieving and charge effect, respectively. The rejection of the three phenolic acids was reduced by concentration polarization effect caused by trans-membrane pressure(TMP). The swelling of membrane surface decreased the pore size and membrane flux for effective separation. Chlorogenic acid and 4,5-dicaffeoylquinic acid were more sensitive to pH and ethanol concentration than 3,5-dicaffeoylquinic acid. A certain correlation existed between the compound structure and the separation factors of nanofiltration, and the separation rules were associated with the comprehensive effect of charge effect, pore size sieving, concentration polarization, steric hindrance and so on.

Mechanism of Farfarae Flos in Qingfei Paidu Decoction against COVID-19 based on network pharmacology and molecular docking / 中草药

Objective: To investigate the mechanism of Farfarae Flos (FF) in Qingfei Paidu Decoction against coronavirus disease 2019 (COVID-19) based on network pharmacology and molecular docking. Methods: Based on our previous study, the main compounds in FF were selected. The potential targets of FF were searched by Swiss Target Prediction and BATMAN-TCM database. GenCLiP 3 and GeneCard were used to predict and screen the therapeutic targets of COVID-19, and then Cytoscape 3.7.1 was used to build the compound-target-disease network. The String database was used to build the target PPI network. Gene ontology (GO) function enrichment analysis and KEGG pathway enrichment analysis were performed in the DAVID database. Molecular docking was performed based on the above compounds and the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) 3CL hydrolase and angiotensin converting enzyme II (ACE2). Results: The compound-target-disease network contained 14 compounds, 104 targets and four diseases. GO function enrichment analysis revealed 444 GO items (P < 0.05), including 325 biological process (BP) items, 44 cell composition (CC) items and 75 molecular function (MF) items. A total of 94 signal pathways (P < 0.05) were screened out by KEGG pathway enrichment analysis. The results of molecular docking showed that the affinity of 3,4-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid with proteins were better than Remdesivir. Conclusion: The compounds in FF can bind with SARS-CoV-2 3CL hydrolase and ACE2, and then act on many targets to regulate multiple signaling pathways, thus exerting the therapeutic effect on COVID-19.

Analysis of metabolites of 4,5-dicaffeoylquinic acid in rat plasma and urine based on LC-MS / 中国中药杂志

Ultra high performance liquid chromatography tandem high field orbital trap mass spectrometry(UPLC-Orbitrap Elite-MS/MS) method was applied in this paper to analyze the metabolites of 4,5-dicaffeoylquinic acid in rat plasma and urine after oral administration. A gradient elution was performed by using Thermo C_(18) column(2.1 mm×100 mm, 1.9 μm), with 0.1% formic acid solution-acetonitrile as the mobile phase. Mass spectral data of biological samples were collected in negative ion mode. The data were extracted by Compound Discovery 2.1 software. Then the blank group samples and the drug samples were compared for exact molecular weight and the mass fragmentation information, and the secondary fragment fitting ratio was calculated to finally attribute the metabolites. As a result, 15 metabolites were detected in rat plasma, and 16 metabolites were detected in urine. The involving metabolic reactions included methylation, hydration, dehydration, reduction, glucuronide conjugation, and sulfation reaction. The metabolites in plasma and urine complemented each other and initially revealed the migration and excretion patterns of this compound in the body. A method for pre-processing biological samples, high-resolution LC-MS instrumentation data, and qualitative software was established in this study to identify metabolite structures, laying the foundation for the study of the active ingredients and in vivo pharmacodynamics forms of Chinese medicines.

Preliminary prediction and analysis of Q-marker of multi-original Tibetan medicine meiduoluomi / 中草药

Objective Based on the concept of quality marker (Q-marker), a chemical method was developed to evaluate the quality of the multi-original medicinal material “Meiduoluomi” and predict its Q-marker. Methods Firstly, the Q-marker were preliminary predicted based on the relationship between genetic relationship and biosynthesis pathway, pharmacological activity and chemical composition. Secondly, the main active components were determined by UHPLC. The analysis was performed using ACQUITY UHPLC HSS C18 column (100 mm × 2.1 mm, 1.8 μm); Mobile phase was acetonitrile (A)-0.3% acetic acid water (B) with gradient elution; The flow rate was 0.2 mL/min; The column temperature was 35 ℃; The detection wavelength was 320 nm. Results A new method for simultaneous determining five compounds of meiduoluomi was established, including chlorogenic acid, caffeic acid, 3,4-dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid, and 4,5-dicaffeoylquinic acid. The linear relationship of them was good at the range of 9.60-480.00, 10.27-513.50, 10.08-504.03, 9.64-482.80, and 3.82-380.24 mg/L, respectively; And the average recovery rates were 102.43%, 99.78%, 99.39%, 103.12%, and 101.83%, respectively. The five components were detected in each sample and the total content of the five components was more than 10% in the herbal medicine. It was suggested that the five components can be used as a Q-marker of meiduoluomi for quality control. Conclusion The coffeic acid components are scientific and reasonable to be considered as a Q-marker of meiduoluomi. The developed UHPLC method can be used for the quality control of meiduoluomi. This study provides new idea for the rational evaluation of multi-original medicinal material based on Q-marker.

Chemical constituents from seeds of Cuminum cyminum / 中草药

Objective To investigate the chemical constituents from the seeds of Cuminum cyminum. Methods The chemical constituents were isolated and purified by repeated silica gel column chromatography, Sephadex LH-20 gel column chromatography, medium pressure column chromatography, high pressure flash chromatography, and semi-preparative HPLC, and their structures were elucidated on the basis of physico-chemical constants and spectral analysis. Results Fifteen compounds were identified as astragaline (1), 6-β-hydroxy glycyrrhetinic acid (2), ursolic acid (3), quercetin (4), protocatechuic acid (5), (6R,7E,9R)-9-hydroxy-4,7- megastigma-dien-3-one (6), 4,5-dicaffeoylquinic acid methyl ester (7), 3,5-dicaffeoylquinic acid methyl ester (8), rosin (9), kaempferol 3-O-β-D-glucopyranosyl (1→2)-β-D-glucopyranoside-7-O-α-L-rhamnopyranoside (10), cuminacid (11), cuminoside A (12), luteolin (13), apigenin-7-O-β-D-(6″-O-acetyl)-glucopyranoside (14), and 5,7,4’-trihydroxyflavanone (15). Conclusion Compounds 2, 3, 6-10, and 14 are obtained from this genus for the first time.

Dynamic changes of phenolic acids and anthraquinones in aerial parts of Xanthium sibiricum from different collection periods / 中草药

Objective: To analyze the dynamic changes of phenolic acids and anthraquinones from the aerial parts of Xanthium sibiricum (Xanthii Herba) in different collection periods. Methods: To establish an HPLC method for simultaneous determination of chlorogenic acid, neochlorogenic acid, protocatechualdehyde, protocatechuic acid, cryptochlorogenic acid, caffeic acid, 1,3-dicaffeoylquinic acid, ferulic acid, 3,4- dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid, 4,5-dicaffeoylquinic acid, aloeemodin, emodin, and chrysophanol in the aerial parts of X. sibiricum. Results: The contents of phenolic acids and anthraquinones in different harvest time showed dynamic changes. In mid July, the content of total phenolic acids was higher. The higher content of total anthraquinones was in late July. The contents of five total phenolic acids were higher in mid July, such as chlorogenic acid, protocatechuic aldehyde, ferulic acid, 3,5-dicaffeoylquinic acid, and 1,3-dicaffeoylquinic acid. The content of chlorogenic acid was higher in late June, and the rest of five phenolic acids were higher in mid July. Conclusion: This can provide the basis for determining the suitable harvest time of X. sibiricum.

Chemical constituents from whole plant of Gynura procumbens (I) / 中草药

Objective: To investigate the chemical constituents from the whole plants of Gynura procumbens. Methods: The chemical constituents were isolated and purified by repeated silica gel column chromatography, Sephadex LH-20 gel column chromatography, medium pressure column chromatography, and semi-preparative HPLC, and their structures were elucidated by chemical properties and spectroscopic analyses. Results: Sixteen compounds were identified to be quercetin (1), apigenin (2), luteolin (3), kaempferol (4), astragaline (5), kaempferol-5-O-(6″-O-acetyl)-β-D-glucopyranoside (6), negletein (7), 4-methoxycinnamic acid (8), benzyl-O-β-D-glucopyranoside (9), 2-phenylethyl-O-β-D-glucopyranoside (10), 3,5-dicaffeoylquinic acid methyl ester (11), 3,5-dicaffeoylquinic acid ethyl ester (12), 3,4-dicaffeoylquinic acid methyl ester (13), 4,5-dicaffeoylquinic acid methyl ester (14), protocatechuic acid (15), and eugenol glucoside (16). Conclusion: Compounds 7, 8, 12, 14, and 16 are obtained from the plants in Gynura Cass. for the first time, and compounds 3, 6, 9-11 and 13 are obtained from this plant for the first time.

Improvement of different processing technologies of Xanthii Fructus / 中草药

Objective: To compare the different processed products of Xanthii Fructus and improve the processing technology of Xanthii Fructus. Methods: Using 2015 version of Chinese Pharmacopoeia to determine the contents of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, caffeic acid, 3,5-dicaffeoylquinic acid, and 4,5-dicaffeoylquinic acid as well as toxic ingredients, such as carboxyatractyloside and atractyloside in raw products, stir-fried and sand fried active ingredients of Xanthii Fructus; The fingerprint of Xanthii Fructus samples was established, and the Chinese Pharmacopoeia Commission promulgated the chromatographic fingerprint similarity evaluation system software (version: 2004A) for the similarity evaluation; SPSS 19.0 software analyzed cluster analysis and SIMCAI 13.0 software analyzed partial least squares-discriminant analysis (PLS-DA). Results: The results showed that the active ingredient content of improved processing technology was high and toxic constituents content was low; The similarity of the 19 samples was greater than 0.98; Cluster analysis showed that the samples might be roughly classified into four groups; PLS-DA analysis showed that different samples can be distinguished. Conclusion: The established analysis method of Xanthii Fructus has a good reproducibility verifies the feasibility of improved processing technology, and provides a scientific basis for quality control of Xanthii Fructus.

Simultaneous Determination of Schaftoside, Isoschaftoside, Deoxyelephantopin and 4, 5-Dicaffeoylquinic Acid in Shennongcha Granules by HPLC / 中国药师

Objective:To establish an HPLC method for determining four constituents ( schaftoside, isoschaftoside, deoxyelephan-topin and 4,5-dicaffeoylquinic acid) in Shennongcha granules. Methods:An HPLC method was performed on a Hypersil C18 column (200 mm × 4. 6 mm,5 μm) with the mobile phase of acetonitrile as the phase A and 0. 025 mol·L-1 phosphoric acid solution as the phase B with gradient elution. The flow rate was 1. 3 ml·min-1 . The detection wavelength was set at 270 nm for schaftoside and isos-chaftoside, 208 nm for deoxyelephantopin and 327 nm for 4, 5-dicaffeoylquinic acid. The column temperature was room temperature. Results:The calibration curve was linear over the concentration range of 5. 850-117. 000 μg·ml-1 for schaftoside, 4. 650-93. 000 μg ·ml-1 for isoschaftoside, 4. 160-83. 200 μg · ml-1 for deoxyelephantopin and 5. 470-109. 400 μg · ml-1 for 4, 5-dicaffeoylquinic acid. The correlation coefficient of all curves was more than 0.999. The average recoverywas 97.70% (RSD=1.40%), 96.87%(RSD=1.13%), 97.53%(RSD =1.69%) and 99.29%(RSD =1.01%) (n =6) , respectively. Conclusion: The developed HPLC method is simple,accurate,and can be used in the content determination of Shennongcha granules.

Research on quality control method for Juhuang Oral Liquid / 中草药

Objective: To establish the quality control method for Juhuang Oral Liquid (JOL) based on the qualitative and quantitative analysis methods. Methods: TLC was used to identify Lycii Fructus and Polygonati Rhizoma in JOL. Phenol-sulfuric acid method was used for the detemination of total polysaccharide, HPLC was used to simultaneously determine the contents of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, coffeic acid, galuteolin, 3,4-dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid, and 4,5-dicaffeoylquinic acid. The Odyssil C18 (250 mm × 4.6 mm, 5 μm) column was used. The mobile phase was acetonitrile-0.05% phosphoric acid solution in gradient elution, at the flow rate of 1.0 mL/min, the detection wavelength was set at 325 nm. Results: The identified characteristics of Lycii Fructus and Polygonati Rhizoma by TLC were distinct and the spots were clear, without interference and easy to recognize. When phenol-sulfuric acid method was used to determinate the total polysaccharide content, the regression equation was A=0.057 8 C + 0.032 5 (r=0.999 1, n=8). The linear with its absorbance in the range of 26.15-196.13 mg/mL, the average recoveries was 99.82%. The average content of the total polysaccharide in JOL from six batches is 157.74-166.49 mg/mL. When RP-HPLC was used to determine the eight chemical components, the compounds showed a good linearity(r ≥ 0.999 8) in the determination range. The average recoveries were between 99.18%-101.06% with RSDs between 0.79%-1.91%. The average content from the six batches showed neochlorogenic acid 102.9-142.6 μg/mL, chlorogenic acid 488.6-563.8 μg/mL, cryptochlorogenic acid 58.9-71.6 μg/mL, coffeic acid 240.7-326.1 μg/mL, galuteolin 228.6-302.7 μg/mL, 3,4-dicaffeoylquinic acid 398.0-485.4 μg/mL, 3,5-dicaffeoylquinic acid 184.1-203.1 μg/mL, and 4,5-dicaffeoylquinic acid 476.2-561.8 μg/mL. Conclusion: TLC identification method is simple. The determination method of total polysaccharide and chemical components is simple, accurate, and with a good repeatability. It can be used for the quality control of JOL.

Quality comparison on Farfarae Flos from various origins by UPLC based on multi-components content determination method / 中草药

Objective: To establish a UPLC method for simultaneously determining 12 components and evaluating the quality of Farfarae Flos from various origins. Methods: The UPLC method was achieved by BEH C18 column (100 mm × 2.1 mm, 1.7 μm) using a mobile phase made up of acetonitrile (A)-0.03% trifluoroacetic acid in water (B). The flow rate and detection wavelength were 0.5 mL/min and 254 nm, respectively.The column temperature was 40 ℃. Coefficients of variation (CV) were calculated, then PCA and HCA were applied to analyzing Farfarae Flos from different origins. Results: The method for content determination was in agreement with methodological requirements. The results showed that CV values of different components differed greatly, and the hyperoside showed the largest CV value at 0.91. PCA and HCA analysis revealed the wild and cultivated Farfarae Flos were different, and the wild samples contained more chlorogenic acid, caffeic acid, rutin, hyperoside, 3,4-dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid, 4,5-dicaffeoylquinic, and tussilagone. Conclusion: There exist the significant differences among Farfarae Flos from various origins, and their CV values are also different. In addition, the wild samples also differ from those of the cultivated ones. To guarantee the safety and effectiveness in clinical, the impact of chemical differences of different Farfarae Flos on the clinical efficacy should be further studied.

Determination of organic acids and glycosides in different species of Lonicerae Flos / 中草药

Objective: To determine and compare the contents of seven kinds of organic acids (neochlorogenic acid, chlorogenic acid, 4-dicaffeoylquinicacid, caffeic acid, 3, 4-dicaffeoylquinic acid, 3, 5-dicaffeoylquinic acid, and 4, 5-dicaffeoylquinic acid) and secoxyloganin in Lonicerae Flos. Methods: The contents were determined by HPLC. The analysis was carried out with gradient elution on a Phenomenex C18 colunm eluted with methanol and 0.1% phosphoric acid. The detection wavelength was set at 225 nm. Results: Neochlorogenic acid, chlorogenic acid, 4-dicaffeoylquinicacid, caffeic acid, 3, 4-dicaffeoylquinic acid, 3, 5-dicaffeoylquinic acid, and 4, 5-dicaffeoylquinic acid in Lonicerae Flos and secoxyloganin in their linear ranges showed good linear relationship, the average recoveries (n = 9) were 99.41%, 99.05%, 99.01%, 98.71%, 98.79%, 98.82%, 98.89%, and 99.16%, and RSD values were 0.56%, 0.37%, 0.93%, 1.11%, 0.90%, 0.87%, 0.71%, and 1.04%. Conclusion: The method is rapid, simple, accurate, and can be used for quality evaluation of the seven kinds of organic acids in Lonicerae Flos from different germplasm and secoxyloganin.