| Objective:(1)To study the protective effect of Kinsenoside on intrahepatic cholestasis and the underlying mechanism.(2)To explore the pharmacokinetic properties and determine the oral bioavailability of Kinsenoside in beagle dogs.Methods:(1)Sprague-Dawley rats were administrated ANIT with or without subsequent treatment with kinsenoside.Blood biochemical indices,bile flow rate,bile acids concentration and liver histopathology were measured to evaluate the protective effect of KD.Then,the expression of Fxr and the related bile acids transporters and enzymes were determined by q-PCR and western blot to discover the underlying mechanisms.(2)A robust LC-MS/MS method was constructed for Kinsenoside determination in beagle dog plasma.The stability of Kinsenoside in beagle dog whole blood and plasma was fully evaluated.The currently developed method was systematically validated,and the validated method was applied to a pharmacokinetic and bioavailability study of Kinsenoside in beagle dogs.Results:(1)Kinsenoside dose-dependently reverse ANIT-induced changes in serum markers,histopathology and bile acids concentration.Furthermore,the gene and protein expression demonstrated that Kinsenoside can active Fxr and increase bile acids secretion via Bsep and other bile acids efflux transporters.(2)Compared with decreasing the pH or adding esterase inhibitors,lowering temperature played a more crucial role in stabilizing KD.Therefore,chemical hydrolysis is the major reason for instability of Kinsenoside in dog whole blood and plasma.At the same time,the absolute oral bioavailability for KD was determined to be around 30%at three different concentrations by using the validated method.Conclusions:(1)Kinsenoside attenuates the ANIT-induced intrahepatic cholestasis through regulation of Fxr in rats.(2)Kinsenoside has a better bioavailability and is suitable for developing an oral dosage form. |
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