The Pharmacokinetic Study Of Principal Components In Yupingfeng Decoction In Rats

Traditional Chinese medicine (TCM) is a kind of effective agent that was summed up based on long-term clinical therapy experience according to TCM theory. Common form of administration for TCM is compound administration. It is a typical complex system.TCM is a great treasure of medicine in China. According to the current records, the type of TCM reaches 12 807 kinds. But so far, TCM has not really entered the mainstream of the international pharmaceutical market. Most we know about TCM are experiential, and active ingredients or real part of TCM is also not totally clear. So, it is hard to work out a truly internal quality control standard for TCM. Pharmacodynamic, safety evaluation and clinical research for TCM are also lagging behind. Faced the rapid development of modern medicine, if we want to promote TCM to international market, we must first clarify the relationship between the chemical composition and the pharmacodynamic mechanism. It is necessary to deeply investigate the TCM in the body, pharmacokinetics study of TCM. So we will understand the absorption, distribution, metabolism, excretion, pharmacology, toxicology of the active components of TCM. YPF came from Danxi's Mastery of Medicine in Yuan dynasty, composed of Astragalus, Atractylodes and Divaricate Saposhnikovia Root. In this compound recipe, Astragalus, Atractylodes and Divaricate Saposhnikovia Root are sovereign, minster and assistant respectively. The proportion of them are 3:1:1.It is a typical agent in TCM that can strengthen and consolidate body resistance and mainly used in clinical recurrent respiratory infections, bronchial asthma and other respiratory diseases. In this study, YPF was choosed as a tool drug. According to published pharmacodynamic study results, under the guidance of TCM theory, Astragaloside IV, Atractylenolide-Ⅰand Prim-O-glucosylcimifugin were selected as object of study to discuss new method for quality control and do pharmacokinetics study of them. They are principle active components in Astragalus, Atractylodes and Divaricate Saposhnikovia Root respectively. At last, we wish to obtain the pharmacokinetics parameters of Astragaloside in different group and discuss the compatibility mechanism in the view of pharmacokinetics aspect. The specific content of this study consists of two parts:Part 1:The determination of Astragaloside IV,AtractyIenolide-Ⅰand Prim-O-glucosylcimifugin in YPF decoctionIn this study, a new method was established to quantify Astragaloside IV, Atractyl enolide-Ⅰand Prim-O-glucosylcimifugin in YPF decoction simultaneously. In the proc ess of establishing this method, some different mobile phase systems were compared, s uch as different proportion of methanol-water and acetonitrile-water to choose chrom atographic conditions. Optimize instrument through instrument tuning, and then scan re ference standards solution under the model of positive and negative, compared with pub lished study report, selected mass spectrometer (MS) parameters. The final chromatogra phic and MS conditions are:Hedera ODS-2 (2.0mm×150mm,5μm) column, mobile ph ase methanol:water(68:32,v:v), flow rate:0.2mL/min, electrospary ionization (ESI+), sel ective ion monitoring(SIM) ion mass spectrum:807.50,287.05,469.05, internal standar d:Telmisartan. Methodological study shows that RSD of Precision results are less than 5% and recovery of these 3 componets are 95.47%,101.94% and 103.70 respecitively. This method is convenient, fast, stable, can simultaneously determinate three principal components in three herbs.The result is accurate and reliable. TCM is based on the prin cipal of sovereign, minster and assistnt. So the efficacy of TCM is the combined effect o f components. This study quantified principal components of three herbs simultaneouss ly, so it can relatively full control the quality of YPF. This study quantified the three acti ve components in YPF decoction, this provided the theoretical basis for pharmacokine tics study of these components in rats.Part 2:Simutaneously determinate AstragalosideⅣand AtrActylenoli de-Ⅰin rats by HPLC-MS and Pharmacokinetics studyIn this study, a new method that can simultaneously quantify AstragalosideⅣand Atractylenolide-Ⅰin rats serum was established. In the process of establishing this me thod, we refer to the established method that can determine AstragalosideⅣand Atrac tylenolide-Ⅰin YPF decoction, optimized the MS conditions; Evaluated different prop ortion of methanol-water, acetonitrile-water mobile phase system, selected chromatogra phic conditions. We evaluated different proportion of diethylether, ethylacetate on liqu id-liquid extraction,different eluted solvents on Solid phase extraction, methanol, ace tonetrile on Protein precipitation. And evaluated diazepam and Telmisartan as internal st andard.At last, internal standard and serum sample preparation method were confirmed. The confirmed chromatographic and MS conditions:are Hedera ODS-2 (2.0mm×150m m,5μm) column for separation. Mobile phase:0.01 glacial acetic acid and methanol, gr adient elution, flow rate:0.2mL/min, electrospray ionization(ESI+), selective ion monito ring(SIM) ion pairs:807.35,287.05, internal standard:diazepam.Methodological study shows that inter-day,intra-day Precision are less than 15% and recovery of these two co mponets are consistant and more than 70%.This method is convenient, fast, stable, can simultaneously determine AS and AT in rat serum. The result is accurate and reliable.Sovereign, minster, assistant and courier are the principle of formulating prescription of TCM, Sovereign drug is recognized as principal components that can treat disease in TCM theory. Therefore, we focus on the pharmacokinetics character of principal components in sovereign drug when we do pharmacokinetics study. In this study, we designed three groups. group 1:Sovereign drug (AS), group 2:Sovereign drug+minster drug (AS+AT), group 3:Sovereign drug+minster drug+assistant drug (AS+AT+POG).According to the published study result, we choose 20 mg/kg as the dose of AT. And then, according to the proportion of AS,AT and POG in YPF decoction, we calculated the dose of AT(2.16495 mg/kg) and POG (569.9 mg/kg). However,569.9 mg/kg for POG is too big, so we choose LD50 (116.9 mg/kg) of POG as final dose. Prepare CMC-Na suspension and intragastric administration suspention, do pharmacokinetics study. Finally, we obtained the pharmacokinetics parameter of AS in different groups.AS group(Tmax:2.1±0.55,Cmax:34±7.58,AUC0-24:263.14±100.59,MRT:6.38±1.54,t1/2:4.42±1.83);AS+AT group(Tmax:2.4±1.78,Cmax:53.6±12.70,AUC0-24:549.81±190.67,MRT:8.24±1.84,t1/2:5.274±1.78);AS+AT+POG group(Tmax:3.6±0.55,Cmax:59.6±20.74,AUC0-24:475.00±70.54,MRT:7.24±0.79,t1/2:5.48±0.62).