Study On Fingerprint Chromatograms And Flavone Pharmacokinetic Of Arisaematis Rhizoma

Arisaematis Rhizoma, belonging to the Arisaema Mart. in Araceae, is recorded to be tuber of Arisaema erubescens (wall.) Schott., Arisaema heterophyllum B1. and Arisaema amurense Maxim, in Chinese pharmacopoeia. Its nature is warm, toxic, bitter and spicy tasted. It links lung, liver and spleen, with the effects of eliminating phlegm and detumescence. External has the effects of riding boils swollen and snake bite. In recent years, it was better effects of applying the Arisaematis Rhizoma compatible treatment of cancer, coronary heart disease.Arisaematis Rhizoma, having much more basic source, has been used for a long time. But its useful is very confusion. The research has revised the quality standard for Arisaematis Rhizoma in Chinese Pharmacopoeia of the2010version. But there is not report about fingerprint chromatograms of Arisaematis Rhizoma so far. The schaftoside and ischaftoside are separated from Arisaematis Rhizoma, the group has preliminary study the pharmacokinetic characteristics of the schaftoside and ischaftoside from Arisaema amurense Maxim in rats, however the study pharmacokinetic characteristics of Arisaematis Rhizoma fiavonoids parts has not been reported.In this paper, the fingerprint chromatogram of Arisaematis Rhizoma is study by high performance liquid chromatography and the pharmacokinetic characteristics of Arisaematis Rhizoma flavonoid is study by ultra-high performance liquid electrospray ionization tandem triple quadrupole mass spectrometry (UPLC-MS/MS). The aim is to better control the quality of Arisaematis Rhizoma and to provide research basic. The main contents including the following six parts, as follows:1. Literature reviewThe paper made a systematic summary for fingerprint chromatograms and pharmacokinetic of traditional chinese medicine on research progress, research status, research method and biological sample processing with test methods and so on.2. HPLC Fingerprint Chromatograms of Arisaematis RhizomaThe analysis was performed on a LichrospherC18(200mm×4.6mm,5μm)with the gradient elution of acetonitrile-water (containing0.1%acetic acid) at a flow rate of1.0ml/min. The detection wavelength was set at270nm. With adenosine as the reference, the similarity of11batches. medicinal herbs is above0.9in the aid of chromatographic fingerprint similarity evaluation system2004A version. According to the retention time and UV spectrum, to identify8common peaks include adenine, hypoxanthine, xanthine, uridine, guanosine, adenosine, schaftoside and ischaftoside. And by contrast the chromatograph with Pinellia pedatisecta Shcott. and Pinellia ternate, the results show there are exist significant differences.3. Simultaneous determination of the schaftoside and ischaftoside in rat plasmaTwo set methods were developed to determination of schaftoside and ischaftoside in the rat plasma. Using UPLC-MS/MS and selecting the multi-reaction monitoring mode (MRM), the schaftoside and ischaftoside of the parent ion at m/z:563[M-H]" and fragment ions at m/z:383[M-H-180]",353[M-H-210]-are treated as a quantitative analysis of ion. Plasma after protein precipitation with methanol, then separated on a Sepax BR-C18(2.1mm×50mm,1.8μm) column and the mobile phase was set at a flow rate of0.3mL/min. Column temperature is set40℃. The mobile phase consisted of methanol-0.05%acetic acid (25:75, V/V) is treated as mobile phase in method one. The linear relationship was good in the range of0.2-100ng/mL for schaftoside and ishaftoside. Methanol-0.05%acetic acid (18:82, V/V) is treated as mobile phase in method two. The linear relationship was good in the range of0.2～100ng/mL for schaftoside and ishaftoside. Endogenous substances did not interfere with sample determination. The extraction recoveries were over than80%and the RSD%of within-day and between-day precision were less than6%. The RSD%of stability are both less than3%. The method is sensitive, rapid and consists with the requirements of the analysis of biological samples.4. Pharmacokinetic characteristics of the schaftoside and ischaftoside in ratsSprague-Dawley rat were given schafoside, ischaftoside and mixtures orally at a dose of4mg/kg, respectively. Blood samples were withdrawn from the rat vein within12hours and separated plasma. Plasma-drug concentration was detected by the UPLC-MS/MS after protein precipitation with methanol. The main pharmacokinetic parameters were obtained by applying DAS2.0software to process the data.Tmax, t1/2and cmax of schaftoside were1.167±0.376h and2.375±0.997h,2.3±0.756h and2.222±0.971h,33.259±3.912μg/L and57.711±26.78μg/L, respectively. Tmax, t1/2and cmax of ischaftoside were1.625±1.137h and2.375±0.997h,2.592±1.175h and2.345±0.853h,36.535±19.53μg/L and42.176±16.102μg/L, respectively. The results showed that the peak concentrations of schaftoside and ischaftoside after mixture administrations were1.7times and1.1times of monomer, respectively and which suggested that the two components can affect their absorption in the body each other.5. Pharmacokinetic characteristics of the Arisaema heterophyllum B1. flavonoid in ratsThe flavonoid parts of Arisaema heterophyllum Bl.was obtained by medicine extracted with60%ethanol, AB-8resin adsorption and collecting the part of30%ethanol elution. The content was determined by HPLC. The rats were given schaftoside, T-II, T-IV and ischaftoside orally at a dose of4mg/kg,1.3mg/kg,0.9mg/kg and2.2mg/kg, respectively. Blood samples were withdrawn from the rat vein and separated plasma. Plasma-drug concentration was detected by the UPLC-MS/MS after protein precipitation with methanol. The main pharmacokinetic parameters of each component were obtained by applying DAS2.0software to process the data. Tmax, t1/2and cmax of schaftoside, T-II, T-IV and ischaftoside were2.625±0.75h,3.455±1.70h and76.005±31.28μg/L;1.875±0.75h、2.37±0.45h and40.684±23.50μg/L;1.875±0.75h,2.601±1.27h and28.21±10.10μg/L;2±0.71h,2.854±1.56h and51.801±19.74μg/L, respectively. Comparison the parameters of the schaftoside and ischaftoside after monomer, mixture and parts administration, the results showed that the components can affect their absorption in the body each other.6. Tissue distribution preliminary study of the schaftoside and ischaftoside in ratsSprague-Dawley rats were given mixtures of schaftoside and ischaftoside orally at a dose of4mg/kg. Tissue samples including heart, liver, spleen, lung and kidney were collected immediately after cervical dislocation at1.5,2and8h after administration. Samples of tissue were homogenized and analyzed the drug concentrations by UPLC-Q/TOF MS after protein precipitation with methanol. The results showed that the drug in tissues collected was almost distributed. The distribution of heart is less, liver and spleen tissues are higher, lung and kidney tissues are followed at1.5h after administration. The distribution of liver and spleen tissues are higher at2h. The concentration of drug in the kidney achieves the highest, liver was followed and spleen was lower significantly at8h.