Prime Pharmacokinetic Study On NZL In Broiler Chickens

2-(3-methyl-4-(2’-nitrophenoxy)phenyl)-1,2,4-triazine-3,5(2H,4H)-dione(NZL)is a new chemical entity synthesized by our team,possessing remarkable anticoccidial activity.An Ultra Performance Liquid Chromatography(UPLC)method for the determination of NZL in serum was established and then applied to the protein binding rate in vitro and the pharmacokinetic study.The protein binding rate in vitro was determined by equilibrium dialysis,demonstrating a very high degree of concentration-independent binding.Meanwhile,the pharmacokinetic parameters were well related to the first-order kinetic model and the metabolites were identified as 2-(3-methyl-4-(2’-acetaminophenoxy)phenyl)-1,2,4-triazine-3,5(2H,4H)-dione(for m/z 351)and the oxidate of NZL(for m/z 355)respectively,indicating that the metabolic pathways of NZL in vitro were amination-acetylation and oxidation.Furthermore,its nonclassical bioisosterism whereas any of its analogues,triclabendazole sulfoxide(TCB-SO),was used as the internal standard(IS)for the determination of NZL in plasma.By investigating the effect of mobile phase’s pH on the retention of the four substances(i.e.NZL,DIC,TOL and TCB-SO),simultaneous separation of these analytes could be achieved.Acid-ammonium acetate buffer was used as aqueous phase that successfully overcame the peak-overlapping and peak-tailing of analytes.Favorable extracting efficiency could be achieved by the mixture of ethyl acetate and acetonitrile(3:1,v/v):89.8%,84.4%,75.0%and 74.6%for NZL,Diclazuril,IS,and Toltrazuril respectively.In addition,good linearity of 0.08～50 μg/mL could be obtained.