| For the purpose of investigating and clarifying the compatability principle for traditional Chinese medicine of tonifying Qi combined with activiting blood circulation, the standard extract of panax ginseng (EPG) for tonifying qi and different standard extract of Salvia miltiorrhiza (ESM) and Carthamus tinctorius (ECT) for activiting blood circulation were chosen for research, and the pharmacokinetics studies on the absorption, distribution and excretion of EPG, ESM and their combined compatibility in rat plasma, tissues, urine and feces were proceeded by RRLC-ESI-MS, and the synergistic reaction for combined compatibility of tonifying qi combined with activiting blood circulation for absorption, distribution and excretion in rat were investigated. The results were as follows:1. A rapid RRLC-ESI-MS method for simultaneous quantification of nine phenolic acids, namely danshensu, protocatechuic acid, protocatechuic aldehyde, vanillic acid, caffeic acid, ferulic acid, rosmarinic acid, lithospermic acid and salvianolic acid B in Danshen and ESM was established. All calibration curves showed good linear regression (r≥0.9973) within test ranges. The overall intra-day and inter-day variations (R.S.D.) ranged from 2.4 to 5.7%. The overall recoveries ranged from 93.0% to 106.8% with R.S.D. less than 9.6%. This developed RRLC-ESI-MS method provided a suitable quality control method for the determination of major bioactive constituents in Danshen and its preparations.2. A sensitive HPLC-CAD method was developd for simultaneous determination of seven major ginsenosides, namely ginsenosides Rg1, Re, Rb1, Rc, Rb2, Rb3 and Rd in Panax ginseng C. A. Meyer and EPG. This CAD method was evaluated in sensitivity, linearity and reproducibility compared to ELSD and UV. It was found the developed HPLC-CAD method had improved sensitivity, linearity and reproducibility compared to ELSD. This method was successfully applied to analyze seven ginsenosides in ten samples of Panax ginseng and three batches of EPG. The validation results indicated the improved method could be utilized as another approach for quality control of Panax ginseng and EPG.3. A RRLC-ESI-MS method was established for simultaneous quantification of five phenolic acids (PAC, PAL, RA, LA, Sal B) in rat plasma. The validated results of methodology were in accordance with the relevant regulations, and all data and results showed that this RRLC-ESI-MS method could be applied to determine the concentration of the five phenolic acids in rat plasma precisely.This method was successfully applied to the determination of the concentration of PAC, PAL, RA, LA and Sal B in rat plama after i.v. administration of ESM (50 mg·kg-1) and ESM-EPG (50 mg·kg-1:50 mg·kg-1), and PAL, PAC and Sal B in rat plasma after i.g. administration of ESM (800 mg·kg-1) and ESM-EPG (800 mg·kg-1:800 mg·kg-1), respectively, and the pharmacokinetic parameters of the analytes after i.g. administration and i.v. administration were also calculated.After i..v. administration, PAC, PAL, RA, LA and Sal B were detected; compared ESM-EPG compatibility to ESM, for PAL, LA and Sal B, Cmax and AUC increased differently, whereas for RA, except for slight reduction of AUC, t1/2, Cmax and Vz increased differently. It might be concluded that, after i.v. administration, compared ESM-EPG compatibility to ESM, there were some enhancement effects on the absoption of PAL, RA, LA and Sal B, and Cmax of the metabolite PAC also increased.After i.g. administration, PAC, PAL and Sal B were detected, and the bioavailability for PAL and Sal B were 5.23% and 4.12%; after combined with EPG, the pharmacokineteics parameters of PAL and Sal B changed differently, the AUC and Cmax increased in different extent, and the absolute bioavailability of PAL and Sal B were increased to 8.06% and 5.61%, respectively, furthermore for the metabolite PAC, Cmax also increased. 4. A HPLC-UV method was established for the determination of PAC, VAC and Sal B in rat tissues. The validated results of methodology were in accordance with the relevant regulations.This method was successfully applied to study the distribution of PAC, VAC and Sal B in rat tissues (heart, liver, spleen, lung and kidney) after i.g. administration of ESM (800 mg·kg-1) and ESM-EPG (800 mg·kg-1:800 mg·kg-1). The results indicated that after i.g. administration, PAC, VAC and Sal B could be detected in rat heart, liver, lung and kidney; while PAC and VAC in rat spleen.The distribution results indicated that, after i.g. administration, compared ESM-EPG compatibility to ESM, the concentrations of the three phenolic acids in rat tissues (heart, liver, spleen, lung and kidney) were decreased in different extent, which suggested that there were some depressant effects on the distribution of the two metabolites (PAC and VAC) and Sal B prototype.5. A RRLC-ESI-MS method was developed for the determination of PAL in rat urine. The validated results of methodology were in accordance with the relevant regulations, and all data and results showed that this RRLC-ESI-MS method could be applied to determine the concentration of PAL in rat urine precisely.This method was successfully applied to the determination of the concentration of PAL in rat urine after i.g. administration of ESM (800 mg·kg-1) and ESM-EPG (800 mg·kg-1:800 mg·kg-1). It was found that, the accumulative excretion dose of PAL was very low, whereas PAL was not detected in rat feces, which indicated that urine might be the main excretion path of PAL, and other amounts of PAL were proposed to be excreted as the metabolites of PAL.After i.g. administration, the accumulative excretion rate of PAL in rat urine was 1.99%, after combined with EPG, the accumulative excretion rate of PAL was decreased to 1.47%. Considering the former pharmacokinetic results in rat plasma, the bioavailability of PAL was improved, and the accumulative excretion rate was decreased, which indicated that EPG might promote the absorption of PAL. 6. A RRLC-ESI-MS method was developed for the determination of the concentration of Sal B in rat feces. The validated results of methodology were in accordance with the relevant regulations, and all data and results showed that this RRLC-ESI-MS method could be applied to determine the concentration of Sal B in rat feces.This method was applied to the determination of the concentration of Sal B in rat feces after i.g. administration of ESM (800 mg·kg-1) and ESM-EPG (800 mg·kg-1:800 mg·kg-1). It was found that, the accumulative excretion dose of Sal B in rat feces was relatively high, whereas Sal B was not detected in rat urine, which indicated that feces might be the main excretion path of Sal B prototype, and other amounts of Sal B were proposed to be excreted as the metabolites of Sal B. The results also indicated that the oral absorption of Sal B was bad.After i.g. administration, the accumulative excretion rate of Sal B was 60.36%, and after combined with EPG, the accumulative excretion rate of Sal B was decreased to 39.39%. Considering the former pharmacokinetic results in rat plasma, the bioavailability of Sal B was improved, the accumulative rate was decreased, which indicated that EPG might promote the absorption of Sal B.7. A RRLC-ESI-MS method was established for simultaneous determination of six ginsenosides (Rg1, Re, Rb1, Rc, Rb2 and Rd) in rat plasma. The validated results of methodology were in accordance with the relevant regulations, and all data and results showed that this RRLC-ESI-MS method could be applied to determine the concentration of six ginsenosides in rat plasma precisely.This method was successfully applied to the determination of ginsenosides Rg1, Re, Rb1, Rc, Rb2 and Rd in rat plama after i.v. administration of EPG (50 mg·kg-1), EPG-ESM (50 mg·kg-1:50 mg·kg-1) and EPG-ECT (50 mg·kg-1:50 mg·kg-1), and ginsenosides Rb1, Rc and Rb2 in rat plasma after i.g. administration of EPG (800 mg·kg-1), EPG-ESM (800 mg·kg-1: 800 mg·kg-1) and EPG-ECT (800 mg·kg-1:800 mg·kg-1), respectively, and the pharmacokinetic parameters of the analytes after i.g. and i.v. administration were also calculated. After i.v. administration, ginsenosides Rg1, Re, Rb1, Rc, Rb2 and Rd were detected in rat plasma. Compared EPG-ESM or EPG-ECT compatibility to EPG, the concentration of the six ginsenosides in rat plasma were decreased in differen extend; and the pharmacokinetics parameters results showed the AUC and Cmax of the six analytes were also decreased in different extend.After i.g. administration, ginsenosides Rb1, Rc, Rb2 were detected in rat plasma, the bioavalibility of them were 0.47%,0.67% and 0.56%. After combined with ESM, the absolute bioavailability of ginsenosides Rb1, Rc, Rb2 were increased to 0.67%,0.93% and 1.11%, respectively; whereas combined with ECT, the absolute bioavailability of ginsenosides Rb1 were decreased to 0.39%, and the absolute bioavailability of ginsenosides Rc, Rb2 were increased to 1.33% and 0.92%.8. A RRLC-ESI-MS method was established for simultaneous determination of six ginsenosides (Rg1, Re, Rb1, Rc, Rb2, Rd) in rat urine. The validated results of methodology were in accordance with the relevant regulations, and all data and results showed that this RRLC-ESI-MS method could be applied to determine the concentration of six ginsenosides in rat urine precisely.This method was successfully applied to the determination of six ginsenosides in rat urine after i.g. administration of EPG (800 mg·kg-1), EPG-ESM (800 mg·kg-1:800 mg·kg-1) and EPG-ECT (800 mg·kg-1:800 mg·kg-1), respectively. It was found that the acumulative excretion dose of the six ginsenosides in rat urine were relatively low.After i.g. administration, compared EPG-ESM or EPG-ECT compatibility to EPG, the excretion tendency of the six ginsenosides were changed, and the acumulative excretion doses of the six ginsenosides were decreased remarkably.9. A RRLC-ESI-MS method was established for simultaneous determination of six ginsenosides in rat feces. The validated results of methodology were in accordance with the relevant regulations, and all data and results showed that the RRLC-ESI-MS method could be applied to determine the concentration of six ginsenosides in rat feces precisely.This method was successfully applied to the determination of six ginsenosides in rat feces after i.g. administration of EPG (800 mg·kg-1), EPG-ESM (800 mg·kg-1:800 mg·kg-1) and EPG-ECT (800 mg·kg-1:800 mg·kg-1), respectively. It was found that the acumulative excretion doses of the six ginsenosides in rat feces were realtively high, which indicated that feces were the main excretion path of the six ginsenosides. The results also indicated that the oral absorption of the six ginsenosides was bad.After i.g. administration, compared EPG-ESM or EPG-ECT compatibility to EPG, the excretion tendency of the six ginsenosides were changed, the acumulative excretion doses of six ginsenosides of EPG in rat feces were decreased in different extent, which indicated that there were some inhibition effects on the excretion of the six ginsenosides in rat feces. Considering the pharmacokinetic results in rat plasma and the excretion results in rat urine, the bioavailabilities of the three ginsenosides were improved, and the excretion doses were decreased, which indicated that ESM or ECT might promote the absorption of ginsenosides. |
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