Pharmacokinetic, bioavailability, metabolism and plasma protein binding evaluation of NADPH-oxidase inhibitor apocynin using LC–MS/MS

•The paper described the first LC–MS/MS for estimation of apocynin in rat and human plasma.•The method is highly sensitive (LLOQ = 1.0 ng/mL) and selective for estimation of apocynin in the plasma.•The validated method has been successfully applied to evaluate in vitro and in vivo ADME studies.

Apocynin is a major active constituent of Picrorhiza kurroa that exhibits potent anti-inflammatory activity by inhibiting superoxide-generating NADPH oxidase enzyme. To elucidate detailed pharmacokinetic profile of apocynin, high-performance liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) method was developed in rat and human plasma. To the best of our knowledge, this is the first method for complete validation of apocynin in biological matrix using LC–MS/MS. Apocynin was rapidly absorbed after oral administration at 50 mg/kg in rats and peak plasma level achieved within 5 min. Moreover, plasma levels were observed up to 48 h. The bioavailibity of apocynin was found to be 8.3%. In vitro plasma protein binding was found to be 83.41–86.07% and 71.39–73.34% in rat and human plasma, respectively. Apocynin was found stable in gastric (pH 1.2), intestinal (pH 6.8) and physiological (pH 7.4) fluids including microsomal (rat and human) stability studies. Further, apocynin did not convert to its dimeric form diapocynin in any of these studies. The data presented here provide crucial information about apocynin to support its pharmacological efficacy and further development as a potential anti-inflammatory drug candidate.