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Objective: The main objective of the research work is to develop and validate a rapid UHPLC method for the estimation of assay and its related substances of Trichostatin A (TSA) in pharmaceutical samples. Methods: The UHPLC method developed for chromatographic separation between TSA and its related compounds on Poroshell 120 SB C18(50×4.6) mm; 2.7 µm RRLC column using Agilent RRLC (UHPLC) system with linear gradient elution. Results: The developed UHPLC method has shown excellent chromatographic separation between TSA and its related compounds within 12 min run time, during validation experiments, specificity study revealed that the peak threshold was more than the peak purity and no purity flag was observed. Repeatability, intra, and inter-day precision results were well within the tolerable limits. Limits of detection concentrations were found between 0.075 to 0.077 ppm and the limit of quantitation is between 0.252 to 0.258 ppm for related compounds and TSA. The related substances method recoveries were found between 80 and 120 % and assay method recovery was found between 98.0 to 102.0%. Conclusion: The developed method capability was proven for the assay of TSA and its related compounds in pharmaceutical samples and the method shows eco-friendlier than routine, conventional HPLC methods in terms of analysis time, cost and HPLC effluent waste.
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This present study was to investigate the metabolism and excretion of characteristic polyphenols such as flavonoids and coumarins in urine and feces of rats after intragastric administration of ethanol extracts of Citri Reticulatae Pericarpium Viride and Citri Reticulatae Pericarpium. The urine and feces of rats were collected after intragastric administration of 70% ethanol extracts of Citri Reticulatae Pericarpium Viride and Citri Reticulatae Pericarpium. Rapid resolution liquid chromatography coupled with quadrupole tandem mass spectrometry (RRLC-QqQ-MSn) was applied to compare the contents of polyphenols in ethanol extract, urine and feces. By comparing with reference substance, 30 polyphenols were identified from the ethanol extracts of Citri Reticulatae Pericarpium Viride and Citri Reticulatae Pericarpium, including flavone glycosides, flavones, flavonone glycosides, flavonones, flavonol glycosides, polymethoxyflavones, coumarins, and limonoids and so on. The detection of various types of compounds showed differences in contents between the intestinal metabolism and excretion in the feces after systemic circulatory metabolism and renal excretion. The results showed that the polymethoxyflavones and flavonones were primarily excreted through urine, and the flavonone glycosides and limonoids were primarily excreted through feces. However, coumarins were hardly detected in feces and urine, indicating that coumarins may be metabolized in the body.
Subject(s)
Animals , Rats , Citrus , Drugs, Chinese Herbal , Feces , Flavonoids , Tandem Mass SpectrometryABSTRACT
The qualitative analysis method of RRLC-Q-TOF-MS/MS was established for determine the chemical constituents in Qige Keli. Kramosil C₁₈ column (4.6 mm×150 mm, 3.5 μm) was used at the temperature of 30 °C. The mobile phase was 0.2% formic acid and acetonitrile by gradient elution, with a flow rate at 1.0 mL·min⁻¹, and the injection volume was 10 μL. The high-resolution quadrupole time-flight mass spectrometry was used as detector with electrospray ion source in both positive and negative models. On the basis of medicinal materials, reference materials, literature reports, and mass spectrometry data, the chemical composition in the Qige Keli was identified. A total of 44 compounds were identified, including 3 flavonoids, 21 flavonoid glycosides, 8 organic acids, 6 lactones, and 3 saponins. The results laid the foundation for the quality control of Qige Keli and the further research on pharmacodynamic materials.
Subject(s)
Acids , Drugs, Chinese Herbal , Chemistry , Flavonoids , Glycosides , Lactones , Phytochemicals , Saponins , Spectrometry, Mass, Electrospray Ionization , Tandem Mass SpectrometryABSTRACT
To develop a rapid resolution liquid chromatography (RRLC) method for the simultaneous determination of epimedoside A, epimedin A1, epimedin A, epimedin B, epimedin C, icariin, baohuosideⅡ, icarisideⅠ, sagittatoside B, 2"--rhamnosyl icarisideⅡ, and baohuosideⅠin epimedium total flavone capsule. At the same time, the effects of the above 11 compounds on osteogenic differentiation of MC3T3-E1 cells were investigated by detecting the content of alkaline phosphatase (AKP). The results showed that baohuoside Ⅱ had the highest activities, and both the activities of baohuoside Ⅱ and icariside Ⅰ were stronger than those of icariin.In this study, the content determination method of flavonoid glycosides was established, and the anti-osteoporosis effect of monomers was compared, providing technical support for the study of the pharmacodynamic and mechanism of Epimedium total flavone capsule.
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Aims To analyze the intervention effect of Captopril on serum endogenous metabolites alternations in spontaneous hypertension rats ( SHR) and to investi-gate possible therapeutic mechanism .Methods The rapid resolution liquid chromatography/quadrupole-time of flight tandem mass spectrometry ( RRLC-Q-TOF-MS) and technology coupled with partial least squares discriminant analysis ( PLS-DA ) processed by SIMCA-P software were used to distinguish significantly different variables and identify potential biomarkers . Results Compared to the normal group , metabolic profiling changed significantly in model group and Cap-toril group .Totally 4 metabolins and their metabolic pathways were detected , which were closely related to the endothelial function .Conclusion Metabolomics reveals possible therapeutic mechanism of Captopril for protecting endothelial function from overall metabolism of the body .It also shows unique potential in terms of interpretation of the complex mechanisms of drugs .
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Objective To establish a rapid and effective supercritical fluid extraction (SFE) and rapid resolution liquid chromatography method coupled with diode-array detector (RRLC-DAD) to quantify the chromones in a species. Methods The effects of four parameters including ethanol concentration (50%–90%), pressure (25–45 MPa), temperature (40–60 oC), and time (30–90 min) on the chromones yields, namely prim-O-glucosylcimifugin, cimifugin, 5-O-methylvisammioside, and sec-O-gluco-sylhamaudol, were investigated using SFE system with orthogonal array design (OAD). Furthermore, the extracts were analyzed using rapid resolution liquid chromatography coupled with diode-array detector (RRLC-DAD) system to confirm the results. Results Under the optimized conditions, i. e., 35 MPa of pressure, 60 °C of temperature, 70% ethanol, and 60 min of time, the yields of prim-O-glucosylcimifugin, cimifugin, 5-O-methylvisammioside, sec-O-glucosylhamaudol, and total chromones were 3.514, 0.132, 6.242, 0.342, and 10.231 mg/g, respectively. In comparison with ultrasonic assisted extraction (UAE), SFE was able to yield a 20.7% increase in the total chromones from Saposhnikoviae Radix. Conclusion SFE is an alternative and promising method to extract chromones from this species, and the established RRLC-DAD method could serve as a rapid and effective method for the identification of chromones from Saposhnikoviae Radix.
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To study pharmacokinetic characteristics of epimedin A, B, C and icariin after intermuscular administration of Chuankezhi injection to rat. The established RRLC-MS/MS method was applied for simultaneous determination of four analytes in rat plasma and calculating their pharmacokinetic parameters. As a result, each analyte showed a good linear relationship in the concentration range of 1-1 000 μg•L⁻¹.The intra-day precise was 96.9%-107.5% with RSD<5.99%, inter-day precise was 92.3%-105.0% with RSD<10.16%. The relative recovery of four analytes was 88.1%-101.1% with RSD<7.9% and their absolute recovery was 72.0%-86.6% with RSD<6.3%. After intermuscular administration of Chuankezhi injection, the plasma concentration of four flavonoid glycosides rapidly arose to peaks at about 10 min, and then quickly declined in rat. Tmax of epimedin A, B, C and icariin was 0.21, 0.19, 0.16 and 0.49 h, respectively, and their mean elimination half-life(t1/2z) was 0.60, 0.62, 0.47 and 0.49 h. The established method was validated to be sensitive, rapid and specific for determination of the four analytes. Serum concentration of 4 species of epimedium flavonoids in Chuankezhi injection was low, and their absorption and elimination seem quickly, displaying similar pharmacokinetic characteristics in this study.
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Objective: To analyse the quantification of protopine from Corydalis Decumbentis Rhizoma (CDR) extract in brain tissues of rats. Methods: A rapid, sensitive, and accurate analytical method based on rapid resolution liquid chromatography (RRLC) coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (Q-TOF-MS) was developed for the quantification of protopine in brain of rats. A simple liquid-liquid extraction process was employed for the sample preparation. Chromatographic separation was achieved using 1.8 μm porous particle columns. Results: The calibration curve of protopine was linear in the range of 12-897 ng/mL. The relative standard deviations of intra- and inter-day precision values were less than 10%. The extraction recoveries were 96.4%, 99.6%, and 98.5%, for protopine at the concentration of 598.0, 119.6, and 12.0 ng/mL, respectively, and internal standard (1.27 μg/mL) was (98.60 ± 0.02)%. Conclusion: The validated method is successfully applied for the determination of protopine in brain tissue of rats after ig administration of CDR extract. © 2014 Tianjin Press of Chinese Herbal Medicines.
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Objective]To establish a rapid resolution liquid chromatography-mass spectrometry(RRLC-MS) method for sibutramine hydrochloride, ephedrine hydrochloride and furosemide, which is the illegally added substances in diet pills. [Method]The chromatographic conditions: Agilent SB-C18 column(2.1mm×50mm, 5μm). The mobile phase consists of acetonitrile(containing 0.1% formic acid) and 0.1% formic acid solution, gradient elution with flow rate of 0.2mL·min-1; The MS conditions: electrospray ionization(ESI) source, with positive and negative ions, multiple reaction monitoring(MRM) mode to detect contents of prohibited substances in diet pills. [Result]Under the above conditions, all seven diet pills were detected with the sibutramine hydrochloride and ephedrine hydrochloride. The concentration range of the two substances was from 0.39 to 4.29μg·g-1 and from 1.63 to 8984.18μg·g-1 respectively. One drug contained furosemide, while the content was 8.71μg·g-1. [Conclusion]The method established was fast, convenient, had high sensitivity and high accuracy, which can detect prohibited substances quantitatively and qualitatively in diet pills.
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OBJECTIVE: To establish an analysis method for identification of gelatine ingredients in Chinese patent medicines. METHODS: Trypsin was used for hydrolysis of gelatins. The characteristic getalins analyses were identified by rapid resolution liquid chromatography (RRLC) coupled to triple quadruple mass spectrometry (QQQ-MS). Trypsin was used for hydrolysis of gelatins. RESULTS: The established present method was specific, precise and reliable. Gelatine ingredients as prescribed wereas detected for several samples. However, instead of gelatin ingredients as prescribed, unprescribed gelatine ingredients were as detected for some samples. CONCLUSION: The methodology validation is performed and the results are satisfied. The method can be used for identification of donkey-hide gelatin, oxhide gelatin, deer-horn glue and tortoise-shell glue in Chinese patent medicines, and provides a scientific reference for the study of quality control method of gelatin ingredients in Chinese Patent Medicines.
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Objective To identify the chemical components in Artemisiae argyi folium by rapid resolution liquid chromatog-raphy-time of flight mass spectrometry (RRLC-TOFMS).Methods The separation was performed on an Agilent Eclipse C18 column (2.1 mm ×100 mm,1.8 μm ).The mobile phase consisted of acetonitrile (A) and 0.1% formic acid (B) were in gradient elu-tion.The flowing rate was 0.35 ml/min, the injection volume was 1μl and the temperature of column was 40℃.Time of flight mass spectrometer ( TOFMS) with electro spray ion source ( ESI) was applied to qualitative analysis under the positive ion mode, and mass scan range was m/z 100-1 500 .Results 31 chemical compounds in Artemisiae argyi folium were identified unequivocally .Conclu-sion A rapid and efficient RRLC-TOFMS approach for identifying the chemical constituents of Artemisiae argyi folium had been suc-cessfully established,which paved a way for quality control and further in vivo studies of Artemisiae argyi folium.
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Objective: Under the guidance of eighteen antagonisms of Chinese materia medica, to analyze the effect of Aconiti Radix Cocta combined with Ampelopsis Radix in different ratios on the characteristic constituents and to explore the in vitro compatibility mechanism of Aconiti Radix Cocta and Ampelopsis Radix. Methods: A certain weight of Aconiti Radix Cocta was combined with various amounts of Ampelopsis Radix, and a rapid resolution liquid chromatography coupled with quadruple-time-of-flight mass spectrometry (RRLC-Q-TOF-MS) method based on chemical profiling approach was used to determine the content of alkaloids in decoctions and sediments. The aconitine alkaloids were established and were subjected to PLS-DA and ANOVA to provide intergroup difference and distribution characteristics. In addition, the method of UV spectrophotometer was adopted to determine the content of starch and tannins in Ampelopsis Radix. Results: There were certain differences among each group, especially between the single Aconiti Radix Cocta and Aconiti Radix Cocta combined with Ampelopsis Radix at the ratio of 1:3. With the increasing of the ratio of Ampelopsis Radix in the combination, the content of alkaloids appeared to increase in the decoction while decrease in the sediment. After determination, the mass fractions of starch and tannic acid in Ampelopsis Radix were 25.44% and 0.31%, respectively. Conclusion: The aconitine alkaloids could be deposited with the starch in Ampelopsis Radix, meanwhile decreased in the decoction, which establishes the foundation for the further research on the chemical mechanism of Aconiti Radix Cocta combined with Ampelopsis Radix.
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Objective: To analyze the chemical constituent changes of Aconiti Cocta Radix (ACR) before and after combination with Trichosanthes Fructus (TF) in different ratios and to provide the material bases for the incombination. Methods: The rapid resolution liquid chromatography with quadrupole and time-of-flight mass spectrometry (RRLC-QTOF/MS) was used to analyze the extract solution from ACR and TF combination. The chromatographic separation was achieved on an Agilent Zorbax Eclipse Plus C18 column (100 mm × 2.1 mm, 1.8 μm) with 0.1% formic acid water solution-0.1% formic acid in acetonitrile solution as mobile phase for gradient elution. Mass Hunter Workstation and partial least squares-discriminant analysis (PLS-DA) were utilized for the identification of components and chemical biomarkers. Peak-area ratio of the analyte/internal standard was used to represent the contents of the variance analysis. The amounts of chemical markers were indicated by the ratio of compound and internal standard peak area. Results: By an overall analysis of RRLC-QTOF/MS, a total of 65 compounds were detected in the extracts from ACR in combination with TF, including 57 compounds with confirmed structures and 15 compounds with significant differences before and after combination, and the relative dissolution rates were affected by the changes of the combination ratios. Conclusion: The sensitive and accurate RRLC-QTOF/MS method could fully reflect the chemical constituent changes of ACR before and after combination with TF. On the basis of in vitro chemical constituent and contents, it is still unclear whether the combination of ACR and TF should be prohibited, which requires further in vivo investigation.
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OBJECTIVE: To observe the plasma metabolic changes of rats after acute hepatic injury induced by CCl4 and the regulating action of Erzhi pills (EZW, traditonal Chinese medicines) on abnormal metabolism, and to find the potential biomarkers. And to explore the mechanisms of Erzhi pills in treatment of acute hepatic injury induced by CCl4. METHODS: Endogenous metabolites in plasma were determined with RRLC-Q-TOF/MS. RESULTS: Nine potential metabolic biomarkers were identified qualitatively. Two metabolic pathways were identified using ingenuity pathway analysis (IPA). CONCLUSION: Changed metabolites can be recovered by Erzhi pills in different degrees, and the treatment effect of Erzhi pills may be related with the regulation of two metabolic pathways. Unique pathway analysis may be effective in drug-target identification, which helps to further understand mechanism of drug action, and improve research efficiency.
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OBJECTIVE: To develope a method for determination of four aflatoxins (B1, B2, G1, G2) in fruit traditional Chinese medicines by rapid-resolution liquid chromatography coupled with electrospray ionization triple quadruple tandem mass spectrometry. METHODS: After being extracted with 70% methanol and purified by immunoaffinity columns, the four aflatoxins were separated on a Shim-pack XR-ODS column using gradient elution with the mobile phase of CH3CN-CH3OH-H2O mixtures monitored under MRM mode in ESI positive scan. RESULTS: There was a good linear relationship over the range of 0.102 1-10.21 ng · mL-1 for AFB1, 0.061 2-7.65 ng · mL-1 for AFB2, 0.193-9.65 ng · mL-1 for AFG1, 0.121-7.55 ng · mL-1 for AFG2 (r > 0.999 8), respectively. The average recoveries ranged from 77.0%-102.4%. CONCLUSION: This method provides a simple and feasible way to determine aflatoxins.
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Objective: A novel method for the analysis of alkaloids in Nelumbo nucifera leaves was established by SPE-RRLC-Q-TOF mass spectrometry. Methods: The crude sample was extracted by 1% HCl with ultrasound-assisted extraction, then purified by SPE column, and eluted with ammonia-methanol (5:95). After concentration, the residue was dissolved by methanol solution. The real sample was analyzed by RRLC-Q-TOF. A Welch Materials C18 column was applied in the RRLC separation using acetonitrile and water as mobile phase. The elutes were detected by Q-TOF to obtain the MS spectra with extract molecular weights under positive ion mode. Results: Nine alkaloids were identified. Conclusion: This method can be used to rapidly determine the alkaloids of N. nucifera leaves.