Study On The Technology, Fingerprint And Pharmacokinetics Of Qinggan Huayu Granule

Primary liver cancer is one of the most malignant tumors in digestive system, produced by the liver cells or intrahepatic bile duct epithelial cells, the mortality rate is only after esophageal cancer and gastric cancer.Primary liver cancer has the incidence of occult,which progress quickly,most patients are diagnosed of terminal, to lose the chance of operation, and purely palliative therapy. By the therapy of traditional Chinese medicine.it is not only can effective control progress,reduce the recurrence and metastasis, but also can improve the life quality of patients, prolong survival time, therefore the research of treatment primary liver cancer by traditional Chinese medicine has more and more attention.This study is supported by the national science and technology major projects, major new drug candidate drug research topic:Qinggan Huayu Capsule in the treatment of primary liver cancer(2012ZX09103201-021).Qing gan hua yu recipe is an effective recipe in treatment of primary hepatic carcinoma,it is formed by more than thirty years clinical experience of Professor Yao Shukun,it is composed of Radix Scutellariae, flavescent sophora root, largehead atractylodes rhizome, zedoary, trigonous, Scutellaria barbata, Hedyotis diffusa, licorice and other drugs,it has the effect of heat-clearing and detoxifying, invigorating spleen for eliminating dampness,promoting blood circulation to remove blood stasis, soften hard.Preliminary study shows that Qinggan can be used in most cases of primary liver cancer patients, curative effect is exact, mechanism studies showed that it has action characteristics with multiple links, multiple target molecule network,it can improve immune function of patients, prolong survival time, improve the quality of life.and has the extremely value to further research, the subject optimize the extraction process of Qinggan Huayu Recipe,made it into granule, it is convenient for patients to take,it take quality control by TCL qualitative identification and detennination of the content of effective components.Combining with the analytical method of fingerprint,it establish the standard fingerprint of Qinggan Huayu granules,To categorize the main chromatographic peaks,and makes a preliminary pharmacokinetic study,which is to know main effective components of Qinggan Huayu granule of distribution and absorption in vivo.Qinggan Huayu granules preparation process of extractionAccording to the characteristics of traditional Chinese medicine and the physicochemical propertiesof traditional Chinese Medicine, main effective components,the optimum extraction process:Radix Scutellariae, matrine as the principal drug, which are alonely extraction process of water extract and alcohol extract,curcuma zedoary and Rhizoma Atractylodis Macrocephalae have more volatile oil,they are extracted by steam distillation of volatile oil, it is packed with beta-cyclodextrin,Burreed tuber, it first mix Scutellaria barbata, Hedyotis diffusa, licorice with dregs of curcuma zedoary and Rhizoma Atractylodis Macrocephalae extracted volatile oil,then water-extraction, water extraction liquid for alcohol refining process the extract is concentrated, dried, dextrin as accessories,90% ethanol as wetting agent to prepare granules. And it is to optimizates the water extraction, alcohol extraction process, the extraction process of volatile oil, inclusion process, water extraction and alcohol precipitation process, granule molding process.Study on quality control of Qinggan Huayu granulesfour herbs in this topic of scutellaria root, Sophora flavescens, Burreed tuber, Scutellaria barbata were qualitative identification by TCL, the results showed that the TCL identification method established in this study is simple, the spots were clear, high reproducibility, four herbs of Qinggan Huayu granules can effectively identify.HPLC method was developed for determination the content of baicalin,chromatographic conditions were as follows:with eighteen alkyl silane bonded silica as filler, Kromasil C18 column (250mm* 4.6mm,5 m); mobile phase:methanol-water-phosphoric acid (43:57: 0.2); the column temperature:28 C; the detection wave length:280nm; flow rate:1mL/min; sample size:10uL. Baicalin in the range 1.6～9.6μg sample injection volume and peak area showed a good linear relationship, the linear correlation coefficient r=0.9995,6 samples with the average recovery rate was 96.88%RSD=0.71%.HPLC method for simultaneous method was established for determination of oxymatrine and matrine content, chromatographic conditions were as follows:octadecylsilane bonded silica as a filler; Kromasil C18 column (250mm × 4.6mm,5μm), mobile phase:organic phase methanol-acetonitrile (1:1); the aqueous phase,0.1% aqueous ammonia solution; gradient elution program:the organic phase,0～40min (10%～60%),40～41min (60%～10%),41～ 50min (10%); the flow rate was 1.0mL/min; column temperature was 30 ℃; detection wavelength of 210nm; injection volume was 10μL. Oxymatrine and matrine respectively into within 0.44～1.54μg and 0.11～0.66μg range of sample injection volume and peak area showed a good linear relationship, the linear correlation coefficient r=0.9995, Oxymatrine six samples the average recovery was 100.00%, RSD= 2.40%, matrine six samples the average recovery was 99.84%, RSD= 1.45%.Study on fingerprints of Qinggan Huayu granulesThis study was designed to compare different mobile phase, gradient elution, detection wavelength conditions, finally determined the high-performance liquid phase fingerprint Qinggan Huayu granule, chromatographic conditions were as follows:octadecylsilane bonded silica as a filler; Kromasil C18 column (250mm × 4.6mm,5μm), mobile phase: acetonitrile-0.2% phosphoric acid, gradient elution; detection wavelength:300nm; flow rate: 0.8mL/min; injection volume:lOuL; column temperature:28 ℃. Under these conditions a total of 25 were selected as fingerprint peaks characteristic peak with prescription single taste Fingerprint than right, right 25 common peaks were assigned to clarify the source of 25 common peaks, to further improve the quality of Qinggan particle control standards.Preliminary pharmacokinetic studies of Qinggan particle dynamicsThis experiment studied one of the effective components of Qinggan Huayu granule oxymatrine in pharmacokinetics, the chromatographic analysis method:using eighteen alkyl silane bonded silica as filler; Kromasil C18 column (250mm* 4.6mm,5 m), mobile phase: organic phase methanol-acetonitrile (1:1); the aqueous phase,0.1% aqueous ammonia solution; gradient elution program:the organic phase,0～40min (10%～60%),40～41min (60%～10%),41～50min (10%); the flow rate was 1.0mL/min; column temperature was 30 ℃; detection wavelength of 210nm; injection volume was 20μL. Plasma samples were extracted with acetonitrile protein precipitation line.The methodology studies suggest that oxymatrine in 0.55-55.0 g/mL concentration range of linear relationship is good, the low, medium and high average recovery rate of 3 concentrations were 100.56%,100.07%,102.13%, RSD were 2.36%,2.60%,1.05%, intra-day precision RSD days were 2.60%,2.66%,1.06%, plasma substances on the determination of oxymatrine without interference.Phase detection method applied to the high performance liquid, SD rats by intragastric administration, relevant pharmacokinetic parameters:Cmax was 31.6733±1.3512 g/mL, T1/2 was 2.589786±0.2604h,Tmax was 1.5h, Ke was 0.2690±0.02591/h,AUCO-10 was 138.6983±5.1950g/mL*h, AUCO-oo was 146.1711±5.6706 infinity g/mL*h.