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OBJECTIVE：To establish a method for simultaneous determination of calycosin glucoside ，ononin，calycosin， formononetin，astragaloside Ⅳ，isoastragaloside Ⅱ，cycloastragenol and isoastragaloside Ⅰ in Astragalus membranaceus before and after bidirectional solid fermentation with Cordyceps kyushuensis ，and to investigate the effects of fermentation on the contents of above 8 components in A. membranaceus . METHODS ：HPLC-DAD-ELSD was adopted. The determination was performed on Agilent 5 TC-C18 column with mobile phase consisted of 0.1％ formic acid aqueous solution-acetonitrile （gradient elution ）at the flow rate of 1 mL/min. The column temperature was set at 30 ℃. DAD detection wavelength was set at 260 nm，ELSD evaporation tube temperature was 100 ℃，atomizer temperature was 80 ℃，carrier gas flow rate was 1.6 L/min；injection volume was 15 μL. RESULTS：The eight components had a good linear relationship within their respective ranges of concentration （all R2＞0.999 0）； RSDs of precision ，stability and repeatability tests were all less than 3％（n＝3 or n＝6）；the recoveries was 97.88％-101.32％， and RSDs were 1.22％-2.39％（n＝6）. Setting the content of components in unfermented A. membranaceus as 100％，after bidirectional solid fermentation with C. kyushuensis ，the change rates of 8 components were －98.51％，－96.41％，－94.74％， －96.40％，289.20％，20.25％，－75.05％，562.46％，respectively. CONCLUSIONS ：After fermentation with C. kyushuensis ，the contents of active components as astragaloside Ⅳ，isoastragaloside Ⅰ and isoastragaloside Ⅱ can be increased significantly in A. membranaceus .
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Objective: To optimize the infiltration process of Astragalus (Astragalus membranaceus var. mongholicus) medicinal materials by Box-Behnken response surface method. Methods: Based on the HPLC-DAD-ELSD and response surface design method, the qualified rate of decoction pieces, the content of index components and bending inspection were used as comprehensive inspection indicators, and the three factors of infiltration were selected for response surface experimental design to optimize the infiltration process of Astragalus medicinal materials parameter. Results: The best infiltration process was as following: infiltration temperature was 20 ℃, with water addition of 1:0.988 for 6 h. Under this process, the qualified rate of Astragalus pieces was 95.81%, the content of calycosin-7-glucoside was 0.072%, and the content of astragaloside IV was 0.276 %. Combining fingerprint analysis and heat map analysis, the material basis of A. membranaceus var. mongholicus changed during the infiltration process. The infiltration parameters should be strictly controlled during the infiltration process to ensure uniform quality of the pieces. Conclusion: The optimized Astragalus medicinal material infiltration process is stable and feasible with good reproducibility, which can provide a reference for the mass production process development of Astragalus medicinal slices.
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OBJECTIVE: To analyze and identify the compositions using HPLC-DAD/ELSD-ESI-TOF/MS for establishing the multi index quantatitive HPLC fingerprint of the Jinming Tablet, in order to providing scientific basis for its quality control. METHODS: The Agilent SB C18 column (4.6 mm×250 mm, 5 μm) was used with a mobile phase of acetonitrile-0.2% acetic acid in gradient elution, the flow rate was 0.8 mL•min-1, the sample voume was 30 μL, the column temperature was maintaine at 25 ℃, the detection wavelength was set at 280 nm. ESI-TOF/MS positive and negative ion mode scanning was used to qualitatively analyze the components in methanol water extract from Jinming Tablet. RESULTS: Twenty compounds in Jinming Tablet extract could be primarily identified by HPLC-DAD on-line detection, in which five compounds were determined; based on the analysis of multiple batches of samples, the multi index chromatographic fingerprint was established, meanwhile combined with similarity analysis to achieve the quality evaluation of Jinming Tablet. CONCLUSION: The HPLC-DAD/ELSD fingerprint film analysis method for Jinming Tablet is established, which could provide reference for the quality control of the material basis of other compound.
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Objective In this paper, HPLC-DAD-ELSD technique was used to establish the chromatography fingerprint of Ilex kudingcha. Methods Shimadzu C18 column (250 mm × 4.6 mm, 5 μm) was used. Acetonitrile-water as the mobile phase; The flow speed was 1.0 mL/min. The detection wavelength was 210 nm and the column temperature was 35 ℃. Results The chromatography fingerprint of Ilex kudingcha from 10 different origins was established. In the chromatography fingerprint with HPLC-DAD of Ilex kudingcha, 18 common peaks were damarcated and the similarities of Ilex kudingcha were between 0.911-0.970, and the chromatography fingerprint with HPLC-ELSD of Ilex kudingcha, 11 common peaks were damarcated and the similarities of Ilex kudingcha were between 0.914-0.962. Chlorogenic acid, kaempferol-3-O-β-D-rhamnoside, isorhamnetin-3-O-β- D-rhamnoside, kudinoside C, kudinoside A, kudinoside E, kudinoside D were confired by LC-MS and binding control. Conclusion The method of precision, reproducibility, stability is accurate, which can be used as the evaluation of the quality of Ilex kudingcha.
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To study the influence of three different drying methods (including 50 ℃-drying, 80 ℃-drying and －70 ℃-freeze-drying methods) on steroidal saponins and homoisoflavonoids in Ophiopogon japonicus,a HPLC-DAD-ELSD-MSn method was investigated to screen and identify the differential components. Through comparing the HPLC chromatograms with that of fresh O. japonicus, 50 ℃-drying medicine was similar with fresh medicine whereas the other two drying methods had great influence on the components of O. japonicus. In this study, 36 differential components were screened, among which 24 constituents(13 homoisoflavonoids and 11 steroidal saponins) were identified via HPLC-LTQ-Orbitrap MS.As a result, it was revealed that different drying methods had significant influences on the components of steroidal saponins and homoisoflavonoids. Among them, 50 ℃-drying method was the most suitable drying approach when the stability of components, cost and practicability were considered.
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Objective: To establish the chromatography fingerprint of Polygoni Cuspidati Rhizoma et Radix with hyphenated technique of HPLC-DAD-ELSD and to evaluate Polygoni Cuspidati Rhizoma et Radix from 10 different origins. Methods: Luna C18 (2) column (250 mm × 4.6 mm, 5 μm) was used. Mobile phase was acetonitrile-H2O; Flow speed was 1.0 mL/min; Temperature of column was set at 35℃; Detective wavelength was at 254 nm; Injection volume was 10 μL. The temperature of drift tube was 109℃ and the flow speed was 3.0 L/min. Results: The chromatography fingerprint of Polygoni Cuspidati Rhizoma et Radix from 10 different origins was established. In the chromatography fingerprint with HPLC-DAD of Polygoni Cuspidati Rhizoma et Radix, 19 common peaks were demarcated and the similarities of Polygoni Cuspidati Rhizoma et Radix were between 0.938-0.993. In the chromatography fingerprint with HPLC-ELSD of Polygoni Cuspidati Rhizoma et Radix, 14 common peaks were demarcated and the similarities of Polygoni Cuspidati Rhizoma et Radix were between 0.905-0.999. Polydatin, resveratrol, emodin-8-O-β-D-glucoside, physcion-8-O-β-D-glucoside, emodin, and physcion were identified. Conclusion: The method is accurate and stable, which can be used as the evidence for the quality evaluation of Polygoni Cuspidati Rhizoma et Radix.
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Objective:To establish an HPLC-DAD-ELSD method for the simultaneous determination of neomycin sulfate and hy-drochloric dyclonine in compound Twaln ointments. Methods:The assay was performed on an Agilent ZOR BAXSB-C18 column(250 mm × 4. 6 mm, 5μm) with acetonitrile-water as the mobile phase with gradient elution at a flow rate of 1. 0 ml·min-1 . The detection wavelength of DAD was 282 nm. The evaporator temperature of ELSD was set at 50℃ and the nebulizer temperature was set at 60℃with the gas flow rate of 1. 6 L·min-1 . The column temperature was kept at 35℃. Results:The linear range of neomycin sulfate was 141. 54-323. 52 μg·mL-1(r=0. 999 6) with the average recovery of 98. 87%(RSD=0. 95%, n=9). The linear range of hydro-chloric dyclonine was 28. 00-64. 00 μg·mL-1(r=0. 999 6) with the average recovery of 99. 57%(RSD=1. 10%, n=9). Conclu-sion:The method is accurate, sensitive and reproducible, and under the same chromatographic conditions, the determination of all the active ingredients in compound Twaln ointments is achieved, which provides basis for the quality control.
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AIM@#To develop and validate a high performance liquid chromatography (HPLC) coupled with diode array and evaporative light scattering detectors (DAD-ELSD) method for the quantitative determination and fingerprint analysis of ten active constituents in three chemical classes (namely, xanthone glycosides, steroidal saponins, and alkaloids) in Zhimu-Huangbai herb pair (ZB).@*METHOD@#Chromatographic separation was performed on a Diamonsil C18 column (4.6 mm × 250 mm, 5 μm, Dikma) by gradient elution using acetic acid in acetonitrile solution at a flow rate of 1.0 mL·min(-1) at 260 nm. The drift tube temperature of ELSD was set to 60 °C and nebulizer gas pressure was 4.0 Bar. Method validation was performed to assure its linearity, limits of detection and quantification, precision, repeatability, stability, and accuracy.@*RESULTS@#The HPLC-DAD-ELSD method allowed the quantification of ten compounds (phellodendrine, jatrorrhizine, palmatine, berberine, neomangiferin, mangiferin, timosaponin E-I, timosaponin B-II, timosaponin B, and timosaponin A-III), and was successfully applied to fingerprint analysis for ten batches of ZB samples.@*CONCLUSION@#This was the first time to apply the combination of DAD and ELSD for the simultaneous determination of ten active ingredients in ZB. The results showed that the combination of quantitative analysis for marker ingredients and chemical fingerprint for the TCM herb pair provides a potentially powerful, widely introduced, and internationally accepted strategy for assessment of complex TCM formulas.
Subject(s)
Alkaloids , Chromatography, High Pressure Liquid , Methods , Drugs, Chinese Herbal , Glycosides , XanthonesABSTRACT
Objective:To develop an HPLC-DAD-ELSD method for the determination of nitidine chloride, 5-ethoxychelerythrine, bergeninum and ardisiacrispin A in Shangtong tinctures ( STD) . Methods: A Hypersil C18 column was used as the chromatographic column, the flow rate was 0. 8 ml·min-1 . For nitidine chloride and 5-ethoxychelerythrine, the mobile phase A consisted of acetoni-trile,the mobile phase B consisted of 0. 1% formic acid-triethylamine (pH 4. 5),and the DAD detection wavelength was at 273 nm. For bergeninum and ardisiacrispin A, the mobile phase consisted of methanol-water(25∶75), the temperature of drift tube was set at 95℃, and the gas flow (N2) was set at 2. 5 SLPM·min-1. Results:There was a good linear relationship between the concentration and peak area for nitidine chloride and 5-ethoxychelerythrine within the range of 0. 021-0. 426 μg (r=0. 999 5) and 0. 075-1. 494 μg (r=0. 999 8), respectively. The average recovery was 99. 22%(RSD=0. 64%) and 98. 61%(RSD=0. 46%), respectively. There was a good linear relationship between the concentration and peak area for bergeninum and ardisiacrispin A within the range of 0. 215-4. 304 μg(r=0. 999 3) and 0. 286-5. 728 μg(r=0. 999 7), respectively. The average recovery was 99. 15%(RSD=0. 77%) and 99. 25%(RSD=0. 56%) accordingly. Conclusion:The method is accurate, sensitive and reproducible, and can be used in the de-termination of nitidine chloride, 5-ethoxychelerythrine, bergeninum and ardisiacrispin A in STD.
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Objective To establish the HPLC-DAD-ELSD fingerprint of Radix astragali,provide new methods for science quality control of the medicinal materials.Methods Application of HPLC-DAD-ELSD techniques were connected in series.The mobile phase A: 10% acetonitrile,B: 90% acetonitrile,detecting wavelength: 265 nm,flow rate: 1 mL/min,column temperature: 35 ℃,sample size: 20 ?L,gain: 20,tube: 55 ℃,neb: 65%,air pressure: 2.068 5?105 Pa.The mutual mode was established depending on ten Astragalus samples from different growing areas in Gansu.The software "Similarity Evaluation System for Chromatographic Fingerfrint of Chinese Materia Medica" was applied to analyzing.ResultsThe established method is good for the separation of saponins,flavonoids from Radix Astragali,and simultaneous determination of the two different components in one sample injection.The similarity of different batches of medicinal materials is fit for the requirement.Conclusion The method is workable to simultaneously determine saponins and flavonoids fingerprint from Radix Astragali,and to control its quality.