| Objective:HPLC fingerprint was established and the concentration of ginsenosides in Black ginseng was determined by QAMS.LC-Q-TOF-MS method was used for qualitative analysis of the components in serum,urine and feces of rats after intragastric administration.The concentrations of seven ginsenosides in the blood of black Ginseng were determined by LC-MS/MS,and the pharmacokinetic regular was expounded.It is hoped to clarify the basis,mechanism and scientific connotation of the pharmacodynamic substance of Black ginseng.Methods:1.Establishment of HPLC fingerprint of Black Ginseng and determination of ginsenosides in black Ginseng by QAMSWaters System C18 column（250 mm×4.6 mm,5μm）was used.The mobile phase was acetonitrile（A）-pure water（B）.Gradient elution;The detection wavelength was 210 nm.The flow rate was 1.0 m L·min-1.Column temperature30℃.Injection volume 10μL;HPLC fingerprint of Black ginseng was established and the concentration of 9 ginsenosides in 11 batches of black ginseng samples were determined by QAMS.2.The chemical constituents of Black Ginseng in serum,urine and feces of rats were analyzed by UPLC-Q-TOF-MSA Waters Acquity UPLC BEH C18（2.1 mm×100 mm,1.7μm）column was used;The mobile phase was acetonitrile（A）-0.1%formic acid aqueous solution（B）.Gradient elution;Flow rate:0.3 m L·min-1;Column temperature:30℃;Injection volume:10μL.Electrospray ionization source（ESI）;The detection mode was positive（ESI+）/negative（ESI-）ion detection mode,and the components in serum,urine and feces of the rats after given the extract of black Ginseng were analyzed qualitatively.3.The concentration and pharmacokinetics of seven ginsenosides in plasma of rats were determined by UPLC-MS/MS simultaneouslyShimazu Nexera X2 LC-30AD ULTRA high performance liquid chromatography system was used for separation.The mobile phase-A:H2O;B:Methanol.The column temperature was 40℃,the flow rate was 0.3 m L·min-1,and the injection volume was 5μL.Gradient elution.Analysis was performed by 5500QTRAP mass spectrometer（AB SCIEX）in positive ion mode（ESI+）.MRM mode was used to determine the blood concentration of seven ginsenosides in rats after given the extract of black Ginseng.DAS 2.0 software was used to fit the pharmacokinetic parameters and draw the drug time curve.Results:1.Establishment of HPLC fingerprint of Black Ginseng and determination of ginsenosides in black ginseng by QAMSTwelve common peaks were determined by fingerprint,and the similarity was above 0.900.Cluster analysis and principal component analysis could roughly divide11 batches of black ginseng samples into three categories.The results showed that there was no significant difference between the concentration measured by QAMS and the concentration measured by external standard method.2.The chemical constituents of Black Ginseng in serum,urine and feces of rats were analyzed by UPLC-Q-TOF-MSSeventeen ginsenosides were identified in the extract of Black ginseng.after given the extract of black Ginseng,12,8 and 11 Prototype ginsenoside were detected in serum,urine and feces of rats,and 3 metabolites were detected in feces.3.The concentration and pharmacokinetics of seven ginsenosides in plasma of rats were determined by UPLC-MS/MS simultaneously（1）.Compared with the other five ginsenosides,the ginsenosides Rc and Rf in black ginseng were difficult to be absorbed and eliminated slowly in rats.（2）.The absorption of ginsenosides Rh4 and Rk3 in rats showed multiple absorption peaks.（3）.Compared with the reported studies,it was found that most of the ginsenosides in black ginseng were absorbed and eliminated slowly in vivo compared with the white and red ginseng.Conclusion:In this study,the HPLC fingerprint and the liquid method to determine the the concentration of ginsenoside black ginseng were established.The experiment proved that the method had good economic applicability and accuracy,and could provide new methods and ideas for the quality control of black ginseng in the future.At the same time,the absorption and metabolism of ginsenosides in the rats,as well as the pharmacokinetic rules of several conventional ginsenosides and the unique and rare ginsenosides of black ginseng were described for the first time,which can provide reliable scientific evaluation for the basic research on the pharmacodynamic substances of black ginseng and provide a basis for the further development and utilization of black ginseng. |
