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OBJECTIVE To establish methods to identify the chemical components of Gantaishu capsule， and determine the contents of 6 index components including glycyrrhizic acid. METHODS The chemical components of Gantaishu capsule were determined by HPLC-TOF/MS； the contents of 6 index components including glycyrrhizic acid were determined by UPLC-MS/MS. RESULTS A total of 41 chemical components were identified in Gantaishu capsules. The linear ranges of glycyrrhizic acid， mangiferin， luteolin， costunolide， oleanolic acid and berberine were 200-10 000 ng/mL（r were all greater than 0.999）. The limits of quantification were 200， 20， 10， 1， 10， 0.5 ng/mL， and the limits of detection were 100， 10， 5， 0.5， 5， 0.25 ng/mL， respectively； RSDs of precision， stability （24 h） and reproducibility tests were all less than 5.0% （n＝6 or n＝3）； the recoveries were 99.05%-101.08% （RSD were all less than 2.0%， n＝6）. The contents of them were 2.42-2.66， 0.85-1.16， 0.35-0.46， 6.18- 6.46， 0.99-1.29， 5.22-5.56 mg/g. CONCLUSIONS The established methods for identification and content determination are rapid and simple， and can be used for the identification of chemical components and the content determination of index components in Gantaishu capsule.
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OBJECTIVE：To establish a method for online detection of antioxidant active components in Glycyrrhiza uroalensis decoction pieces ，and to identify it. METHODS ：The free radical scavenging rate of 1，1-diphenyl-2-trinitrobenzene hydrazine （DPPH）was determined to evaluate the antioxidant activity of G. uralensis decoction pieces. HPLC-UV-DPPH method was used to screen the anti oxidant active components of G. uralensis decoction pieces. HPLC-TOF/MS was used to obtain mass spectrum data and Qualitive Analyst B 06.00 Build 6.0.633.0 software was used to analyze data. Through contrast analysis of UV absorption spectrum，online chromatogram ，mass spectrum information of G. uralensis and the retention time of each compound ，accurate molecular weight ，antioxidant active components were identified by referring to relevant literature. Validation test was also conducted. RESULTS ：DPPH free radical scavenging rate in 8 batches of G. uralensis decoction pieces ranged 55.71％-60.17％. Seven antioxidative active compounds ，including avolomotor ，8-isopentenyl naringin ，yellow lupulin weitone ，isoflavone B ，3′， 4′-dimethoxy3-hydroxy-6-methyl flavone ，glycyrrhizin E and glycyrrhizin H ，could be screened from G. uralensis decoction pieces. After validation ，the peak area of inverted peak generated by online reaction was positively correlated with DPPH free radical scavenging rate. CONCLUSIONS ：Established method is simple and accurate ，and can be used to quickly screen and identify the main antioxidant components of G. uralensis decoction pieces ；the peak area of inverted peak can be used to evaluate the antioxidant active components of G. uralensis decoction pieces.
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Related substances in pharmaceutical formulations are associated with their safety, efficacy and stability. However, there is no overall study already published on the assessment of related substances in the Compound Ketoconazole and Clobetasol Propionate Cream. In this work, a reliable HPLC-TOF-MS qua-litative method was developed for the analysis of related substances in this preparation with a quick and easy extraction procedure. Besides the active pharmaceutical ingredients, two compounds named ke-toconazole impurity B′ optical isomer and ketoconazole impurity E were identified. Furthermore, a new HPLC method for qualitative and quantitative assessment on related substances and degradation pro-ducts, which were found in the stability test, was established and validated. The single standard to determine multi-components method was applied in the quantitative analysis, which was an effective way for reducing cost and improving accuracy. This study can provide a creative idea for routine analysis of quality control of the Compound Ketoconazole and Clobetasol Propionate Cream.
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Abstract Lasia spinosa (L.) Thwaites is a widely used ethnomedicinal plant in Bangladesh. In this study, we investigated phenolic contents, volatile compounds and fatty acids, and essential oil components of extracts prepared from aerial parts of the plant. The main volatile compounds were methyl ester of oleic acid, palmitic acid and stearic acid as determined by GC/MS. Phenolic contents of the extracts were determined qualitatively and quantitatively by HPLC/TOF-MS. Six phenolic compounds (syringic acid, morin, gentistic acid, 4-hydroxybenzoic acid, cinnamic acid, and apigenin) were found in the extracts. GC/MS analysis of steam distilled essential oil showed camphor, α-pinene and δ-3-carene as the main constituents. In DPPH radical scavenging assay, the highest free radical scavenging activity was observed for the methanol extract with an IC50 value of 0.48 ± 0.04 mg/mL, whereas, in metal chelating activity on ferrous ions (Fe2+) assay, the highest chelating activity was observed for hexane extract (IC50 = 0.55 ± 0.08 mg/mL). The extracts and essential oil were tested against five severe human pathogenic bacteria using disc diffusion assay and subsequent MIC values were also determined. All the extracts (except methanol extract) and the essential oil were found to possess potential antimicrobial activity with corresponding inhibition zone and minimum inhibitory concentration (MIC) ranging from 9-23 mm and 62.5-500 µg/mL. This study has been explored the plant Lasia spinosa can be seen as a potential source of biologically active compounds.
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Chelating Agents/analysis , Free Radical Scavengers , Phenolic Compounds/analysis , Volatile Organic Compounds/analysis , Fatty Acids/analysisABSTRACT
Objective To compare chemical composition in the different parts (leaf, branch, and fruit) of Rhodomyrtus tomentosa, and to study the chemical constituents from fruits of R. tomentosa. Methods High performance liquid chromatography/time-of-flight mass spectrometry (HPLC-TOF-MS) method was executed to analyze the samples. Principle component analysis (PCA) and partial minimum variance discriminant analysis (OPLS-DA) in MassLynx XS software were used to analyze the obtained data. The chemical constituents of fruits were isolated and purified by column chromatography, including silica gel, Sephedex LH-20 and re-HPLC, and the structures were elucidated based on their NMR and MS data. Results The PCA results indicated that the constituents existed in leaf were significantly different from those in branch and fruit, while constituents in branch and fruit were similar. Furthermore, based on OPLS-DA, combined with chromatographic retention regulation, accurate molecular mass, isotopic matching and literature searching, four marker compounds from leaves had been found and identified as myricitrin (1), myricitrin-3-O-L-furanoarabinoside (2), iridin (3), and 3,3’-didemethyl-9-oxo-pinoresinol (4). Besides, five compounds were isolated from fruits and identified as maslinic acid (5), ethyl gallate (6), gallic acid (7), resveratrol (8), and piceatannol (9). Conclusion This research provides an effective strategy for analyzing chemical difference from different parts of R. tomentosa, which can be applied to study the chemical difference from different parts of other species.
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Objective To establish qualitative and quantitative analysis methods for identification and determination of multi-constituent of Chanfukang granules (CFKG) using high performance liquid chromatography-time-of-flight mass spectrometry (HPLC-TOF-MS) and high performance liquid chromatography-triple quadrupole mass spectrometry (HPLC-MS/MS),and a preliminary pharmacokinetic study was carried out.Methods An Agilent ZORBAX Eclipse Plus C1s column (100 mm × 3.0 mm,1.8 μm) and a Waters XBridge(R) BEH C18 column (150 mm × 4.6 mm,2.5 μm) were used with 0.1％ formic acid aqueous solution-acetonitrile as mobile phases by gradient elution.The compounds were detected by electrospray ion source in both positive and negative mode with multiple reaction monitoring (MRM) mode.SD rats were ig with CFKG (0.5,5 g/kg).Blood samples of 0.2 mL were taken from jugular vein at different time points and plasma components and contents were determined by HPLC-MS/MS.Results Twenty-four constituents were identified by HPLC-TOF-MS qualitative analysis and thirteen constituents were quantitatively detected by HPLC-MS/MS.The established HPLC-MS/MS method had a good separation,and all the legal verification met the requirements.The content of stachydrine,Leonurine,and astragaloside Ⅳ were high,which may be the main active ingredient The pharmacokinetic results showed that,only when the dose was 5 mg/kg,stachydrine and Leonurine can detected.Elimination of stachydrine was slow and elimination of Leonurine was fast.Conclusion A rapid and efficient method for studying the chemical constituents was established,which could provide reference for the quality control of Chanfukang granules.
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According to the Chinese Pharmacopoeia 2015, only processed Aconitum tubers can be clinically applied, and the effect of processing is unclear. This research aimed to explore the effect of processing on cardiac efficacy of alkaloids in Aconitum tubers. First, the chemical ingredients in unprocessed and processed Aconitum tubers were identified and compared by using high performance liquid chromatography time-of-flight mass spectrometry (HPLC-TOF/MS) and multivariate pattern recognition methods. Then the representative alkaloids in Aconitum tubers, aconitine, benzoylaconine, and aconine, which belong to diester-diterpenoid alkaloids, monoester-diterpenoid alkaloids, and amine-diterpenoid alkaloids, respectively, were selected for further validation of attenuated mechanism. Subsequent pharmacological experiments with aconitine, benzoylaconine, and aconine in SD rats were used to validate the effect of processing on cardiac functions. After processing the Aconitum tubers, it was found that the contents of diester-diterpenoid alkaloids were reduced, and those of monoester-diterpenoid alkaloids and amine-diterpenoid alkaloids were increased, suggesting that diesterditerpenoid alkaloids were transformed into monoester-diterpenoid alkaloids and amine-diterpenoid alkaloids. Through further decocting the aconitine in boiling water, it was confirmed that the three alkaloids could be progressively transformed. Pharmacological experiments with aconitine, benzoylaconine, and aconine in SD rats showed that aconitine at a dose of 0.01 mg/kg and aconine at a dose of 10 mg/kg enhanced the cardiac function, while benzoylaconine at a dose of 2 mg/kg weakened the cardiac function. The effect of processing is attributed to the transformation of the most toxic diester-diterpenoid alkaloids into less toxic monoesterditerpenoid alkaloids and amine-diterpenoid alkaloids.
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@#The study aimed to separate and identify the metabolites of hypericin in the bile and necrotic tissues in rats. After intravenous injection of 10 mg/kg hypericin, 0-12 h bile of normal rats and 24 h necrotic liver of rats with reperfused hepatic infarction were collected, and metabolites of rats were analyzed by high performance liquid chromatography coupled with electrospray tandemtime of flight mass spectrometry(HPLC-TOF/MS). The prototype(M0)and three glycosylation metabolites(M1, M2, M3)of hypericin in rat bile and the parent compound in rat necrotic liver were detected and identified. Results indicated that prototype and glycosylation of hypericin were the major metabolic form in rat bile and the parent compound was found only in necrotic tissues.
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Objective To analyze chemical constituents of Xionggui Decotion by rapid‐resolution (high performance) liquid chromatography‐time of flight mass spectrometry (HPLC‐TOF‐MS) .Methods A Shelloseido column(100 mm × 3 .0 mm ,3 μm)was used to separate .The mobile phase consisted of water containing 0 .1% methane acid and acetonitrile was used as gradient elute .The flow rate was 0 .4 ml/min .TOF‐MS was applied for qualitative analysis under positive ion mode . Results Under LC/MS condition ,47 major constituents in Xionggui Decotion were identified by time of flight mass spectrome‐try and structure‐relevant fragment ions .Conclusion A simple and reliable method using HPLC‐TOF‐MS was established to i‐dentify the chemical constituents of Xionggui Decotion .
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Objective To introduce an accelerated solvent extraction (ASE) -HPLC-TOF/MS method for simultaneous determination of Polyphyllin I, Polyphyllin II, Polyphyllin VI, Polyphyllin VII, Dioscin and Gracillin in Paris Polyphylla. Methods The extraction was performed by ASE under the following conditions: extracting solvent 70% ethanol; extracting temperature 120°C; pressure 1. 17 MPa (1 700 psi); and static time 6 min. The separation was carried out on a MGC18 column (3.0 mm× 100 mm, 3. 0 μm). Elution was done with a linear gradient mobile phase system consisting of 0. 1% formic acid (V/V) water and acetonitrile. The flow rate was set at 0. 8 ml/min, the sample injection volume was 3 μl, the column temperature was kept constant at 25°C, nebulizer gas pressure was 275. 8 kPa (40 psi), drying gas flow rate was 10 L/min, and gas temperature was 350°C. Results The calibration curves for Polyphyllin I, Polyphyllin II, Polyphyllin VI, Polyphyllin VII, Dioscin and Gracillin were linear within the range of 0. 500 0-50. 00 μg/ml (r = 0. 999 8), 0. 422 3-42. 23 μg/ml (r = 0. 999 7), 0. 612 0-61. 20 μg/ml (r = 0. 999 9), 0. 714 0-71. 40 μg/ml (r = 0. 999 5), 0. 448 0-44. 80 μg/ml (r = 0. 999 9) and 0. 436 0-43.60 μg/ml (r = 0. 9997), respectively; and the average recoveries (n=6) were 98. 9%, 98. 0%, 102. 5%, 101. 9%, 103. 1% and 97. 9%, respectively. Conclusion The introduced method is solvent-saving and time-saving; it can also be used for high-throughput analysis and determination of steroidal saponins in Paris Polyphylla.
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Objective: To identify the chemical components in traditional Chinese herbal medicine Ginseng by high performance liquid chromatography diode array detector (HPLC-DAD) and High performance liquid chromatography-time of flight mass spectrometry (HPLC-TOF/MS). Methods: An Agilent Zorbax XDB-C18 column (250 mm X 4.6 mm, 5 μm) was used for isolation and identification of chemical components in Ginseng with a mobile phase of acetonitrile and water in gradient elution. A gradient program was used as follows: 1-35 min, 19% A; 35-55 min, 19%-29% A; 55-70 min, 29% A; 70-110 min, 29%-40% A; 110-150 min, 95% A. The flow rate was set at 1.0 ml/min, the injection volume was 20 μl, the column temperature was set at 20°C. The time-of-flight mass spectrometer was equipped with an EIS ion source. Scanning mass range was between m/z 100-1 350. Results: Thirty-nine chemical compounds in Ginseng were identified unequivocally. Conclusion: Chromatographic demonstration of 39 chemical compounds of Ginseng in one run is achieved by HPLC-TOF/MS, which provides a foundation for further study on the metabolism and action mechanism of traditional Chinese herbal medicine.
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Objective: To rapidly identify the chemical components in a traditional Chinese herb Xiaochaihu decoction by high performance liquid chromatography-time of flight mass spectrometry (HPLC-TOF/MS). Methods: An Agilent ZORBAX SB-C 18 column (100 mmX3.0 mm,3.5 μm) was used for rapid separation and identification of chemical components in Xiaochaihu decoction, with a mobile phase of methanol and 0.1% formic acid aqueous solvent in gradient elution. The flow rate was 0.6 ml/min and the injection volume was 4 μl. The detector was time-of-flight mass spectrometer with an ESI ion source. Scanning mass range was m/z 200-1 500. Results: Thirty-eight chemical compounds were identified from Xiaochaihu decoction in one run. Conclusion: Thirty-eight chemical compounds have been identified in Xiaochaihu decoction by HPLC-TOF/MS in one run, which paves a way for further understanding the metabolism and mechanism of the chemical compounds in Xiaochaihu decoction.
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AIM: To establish methods for identifying the antetype of Compound Qingkailing(Calculus bovis,Concha margaritifera,Radix isatidis,Cornu bubali) in rat serum by HPLC-MS~n and HPLC-TOF-MS. METHODS: Addition of methanol to serum after Qingkailing given intravenously to rats was used to precipitate protein.The HPLC-TOF-MS provided the exact molecular weight and the HPLC-ESI-MS~n provided the m/z of multilevel fragment.The medicine antetypes were identified by combining the two methods. RESULTS: Four main medicine antetypes in rat serum,such as geniposide,baicalein,wogonoside and cholic acid,were identified. CONCLUSION: It is a rapid and exact method that can be used to identify other complicated traditional Chinese medicine in vivo.