Studies On Proteomics Of Shengmai Injection And In Vivo Processes Of Its Active Components

Traditional Chinese Medicine (TCM) injection, a novel dosage form of TCM in China, is widely used for treatment of critical illness such as infectious, cardiovascular and cerebrovascular diseases because of its high bioavailability and significant effectiveness. However, the active ingredients, mechanism of actions and in vivo processes of TCM injection are not clearly understood, which results in the unreasonable clinical utilization of many TCM injections. Therefore, to improve its safety and effectiveness of clinical application, it is necessary to carry out more fundamental research of TCM injection regarding to its material basis of efficacy, molecular targets, cellular mechanisms and in vivo processes.Shengmai injection (SMI), an effective TCM injection composed of Panax ginseng, Ophiopogon japonicas and Schisandra chinensis, is widely used in clinic for the treatment of myocardial infarction, cardiac shock and infectious shock. However, its material basis of efficacy, mechanisms and in vivo process still remain to be elucidated. Therefore, the proteomics of SMI and pharmacokinetics (PK), tissue distribution, excretion, pharmacodynamics (PD), the combined pharmacokinetics-pharmacodynamics (PK-PD) model of its main active components were studied in this thesis. Focusing on dynamic interplay between SMI and biological entity, this study provided the scientific basis for rational clinical application and re-development of SMI. The main contents and contributions of this thesis were summarized as follows:1. Proteomics study of SMI was performed with two-dimensional gel electrophoresis (2-DE) and matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS/MS) techniques for separating and indentifying differentially expressed proteins in rats with IR injury and SMI treatment. The differentially expressed proteins were then validated by Western blot. Results from function analysis indicated that these differentially expressed proteins were mostly associated with energy metabolism. According to change of expression level of each differential protein in all experimental groups, the relationship between multi-metabolic pathways was explored as the regulatory action of SMI on myocardial energy metabolism. It revealed that SMI protected heart against IR injury through optimizing myocardial energy metabolism, improving cardiac efficiency and increasing myocardial energy supply. This study elucidated protective mechanism of SMI against IR injury on a molecular basis, and provided biological interpretation for the integrated regulatory effect of SMI.2. The in vivo processes of the main active components, ginsenoside Rgl, Re, Rf, Rg2, Rb1, Rd, Rc, ophiopogonin D, schisandrin, schisandrol B and schizandrin B of SMI, were studied in rats. A sensitive and reliable analytical method of LC-MS/MS was developed to determine all compounds in serum, tissue, urine, feces and bile of rats after single intravenous administration of SMI to rats. Then, the pharmacokinetic models of all compounds were established and their concentrations-time processes in vivo were explained. Moreover, the pattern of tissue distribution and excretion of the main active components was elucidated. This study revealed in vivo processes of main active components in SMI, and it provided scientific data for clinical application of SMI.3. The pharmacodynamic models of ginsenoside Rgl, ginsenoside Rbl and schisandrin were established to evaluate their effects of inducing real-time nitric oxide (NO) release in myocardial ischemia rats using NO in serum as PD marker. The results demonstrated that ginsenside Rgl and ginsenside Rbl could induce real-time NO release in rats, however schisandrin showed no significant influence on NO levels in serum under the same condition. Based on this result, the combined PK-PD model was studied using ginsenoside Rgl and ginsenoside Rb1 as PK marker and NO in serum as PD marker after single intravenous administration of SMI to rats. A simple and accurate method to determine nitrite and nitrate, the metabolites of NO, in serum based on HPLC with fluorescence detection was established and evaluated in this study. A combined PK-PD model of ginsenoside Rgl and Rbl after intravenous administration of SMI to rats was developed. This study provided scientific data for clinical application of SMI and offered new strategy and method for the combined PK-PD model study of TCM.