Study On Preparation And Bioavailability Of Puerarin Microemulsion

Puerarin is a major effective ingredient extracted from the traditional Chinese medicine Ge-gen.It has been reported that puerarin exhibits many pharmacological effect of cardiact blood vascular diseases.It has poor solubility and totally only 3.799% of puerarin can be absorbed by oral administration.However its clinical applications have been hindered by its low solubility.Puerarin is currently solubilized in 50% 1,2-propanediol as clinincal formulation.But this method bring many adverse effects. So our purpose is to develop the formulation of microemulsion drug delivery system for Puerarin.The uptake and transport of Puerarin-Microemulsion in Caco-2 cells were investigated.Finally,the studies on pharmacokinetics of Pue-ME were determined.The HPLC methods for in vitro analysis of puerarin were set up among methanol-water,0.1MHCL,PBS and ethanol-wter.The method was accuracy,sensitive and suitable for the determination of puerarin.The solubility of Puerarin in various vehicles was measured such as surfactant,cosurfactant,oilphase,aqueous media. Studies on stability of Puerarin were carried out in 0.1M HCL and PBS for 12h.The optimum formulation of PUE-ME was screened by solubility experiments, compatibility tests and pseudo-ternary phase diagrams,with the time of formulating microemulsion,the consequence of visual examination and particle size as parameters. The optimum formulation was composed of IPM(10%),Brij-96V(32.5%),ethanol (32.5%) and water(25%).The particle diameter was(16.46±2.95)nm.The content of puerarin in the optimum formulation was 77.11mg/ml.Forming time less 1 min.The release of puerarin from microemulsion was significantly faster than that of the conventional tablet in simulated gastric and intestinal fluid.Results indicated that release of puerarin was more than 90%at 12h.Release in PBS was much faster than that in 0.1M HCL.Release profile of puerarin from tablet and microemulsion followed first-order kinetics and Ritger-peppas model,respectively.Caco-2 cell monolayer model was established according to Caco-2 cell cultivated in millicell plate for 21d.The morphological and growth characters of the monolayer were examined.The polarization and compactification of the monolayer was determined by the alkaline phosphatase activity,transepithelial electrical resistance and apical-to-basolateral of sodium fluorescein across cell monolayer.The Caco-2 cell model was used to study the uptake,transport of Pue-ME.The effect of ME on the transport of drug was investigated.The results indicted that ME could significantly improve the uptake,transport and efflux kinetics of Puerarin(P＜0.05),especially when ME was diluted.In the presence of CsA,potent inhibitors of P-glycoprotein,P_appAP-BL /P_appBL-AP ratio was decreased in ME but have no effectent on transport.A specific and accurate HPLC method was developed for the determination of puerarin in SD rats plasma.Puerarin was extracted by protein precipitation and then the supematant was injected into a UltimateXB-C18 column(250×4.6mm,5μm).The detection wavelength was set at 250nm and the mobile phase consisted of methanol: water(22:78) at a flow rate of 1.0ml/min.The blank plasma had no interference on the determination of puerarin.A good linear relationship was obtained between the peak area of puerarin and the concentration of puerarin over the range of 40 to 2000 ng/ml. The LLOQ was 40ng/ml and the total chromatographic analysis time was within 10 min. The method is precise and fast for the determination of puerarin in plasma after oral administration of Pue-ME to healthy SD rats.The pharmacokinetic parameters of Puerarin in suspension and dilusion-ME were investigated after oral administration to rats at a single dose of 30mg/kg.The date was disposed using the program 3P87.The C_max of purerarin from D-ME is greater than that of the puerarin suspension.The AUC_0.24h of puerarin from D-ME is about 7-fold higher than that of puerarin from suspension.Compared with the puerarin suspension,the relative bioavailability of D-ME was 783%.The D-Me possessed later Tmax.This study suggested that the oral bioavaiability of Pue ME was greater than Pue suspension.