Liquid chromatography–hydride generation–atomic fluorescence spectrometry determination of arsenic species in dog plasma and its application to a pharmacokinetic study after oral administration of Realgar and Niu Huang Jie Du Pian

Abstract

A high performance liquid chromatography-hydride generation-atomic fluorescence spectrometry (HPLC-HG-AFS) method was developed for the simultaneous determination of four arsenic species (As(III), dimethylarsinic acid (DMA), monomethylarsonic acid (MMA) and arsenate As(V)) in dog plasma. Good separation of the four arsenic species was achieved within 15min on an anion-exchange column with isocratic elution using 15mmol/L KH(2)PO(4) (pH 5.9) as eluent at a flow rate of 1.0mL/min. The assay was linear over the range of 1.25-200, 1.56-200, 1.34-172, and 2.50-200ng/mL with the detection limits of 0.80, 1.00, 0.86 and 2.00ng/mL for As(III), DMA, MMA and As(V), respectively. The method was validated for selectivity, precision, accuracy and recovery and then applied to a comparative pharmacokinetic study of the arsenic species in beagle dogs after a single oral administration of Realgar (24.32mg/kg, equivalent to 11.31mgAs/kg) alone or Niu Huang Jie Du Pian (a patent traditional Chinese medicine (TCM), 380mg/kg, equivalent to 28.45mgAs/kg), respectively. DMA was found to be the predominant species in the dog plasma after dosing, with As(V) appeared as the quickly eliminating one. No traces of MMA and As(III) were detected at any sampling time points. The main pharmacokinetic parameters found for DMA p.o. administration of Realgar and Niu Huang Jie Du Pian were as follows: C(max) (14.7±4.2) and (57.0±32.0)ng/mL, T(max) (2.4±0.5) and (2.5±0.5)h, AUC(0-36) (151.1±12.9) and (635.9±418.2)ngh/mL, AUC(0-∞) (206.0±44.5) and (687.2±425.1)ngh/mL, t(1/2) (16.2±7.9) and (9.4±2.2)h, respectively. The influence of compounding in Niu Huang Jie Du Pian on the pharmacokinetics of arsenics was shown with increased transformation of DMA and its faster elimination rate.