| Objective: A high-performance liquid chromatographic method (HPLC) for the determination of mebendazole (MBZ) and its metabolites hydroxy-mebendazole (MBZ-OH) and amino-mebendazole (MBZ-NH2) in European eels (Anguilla anguilla) plasma and tissues was developed. Pharmacokinetics and residues of mebendazole and its metabolites were investigated to provide reasonable dosage regimen and scientific proof for the clinical application of mebendazole.Methods: The European eels (Anguilla anguilla), were exposured with MBZ at a dose of 1mg·L-1 for 36h and kept at a water temperature of 25±1℃. Plasma and tissues samples were collected at intervals during and after treatment and analyzed for MBZ and its metabolites by HPLC. The samples of the plasma and tissues were extracted with ethylacetate and analyzed on a C18 chromatographic column with acetnonitrile : acetate acid-sodium acetate buffer, pH 4.5, as mobile phase. The concentration-time data of MBZ and its metabolites in plasma and tissues were analyzed with DAS pharmacokinetic software.Results: The lowest detective limit was 3 ng·mL-1 with this method. A linear range of applicability of 0.01-1.00μg·mL-1 was found for mebendazole and its metabolites. The average extraction recovery of MBZ and its metabolites MBZ-OH and MBZ-NH2 in plasma were 98.93%, 97.92%, 102.91%, respectively. The average extraction recovery of MBZ and its metabolites MBZ-OH and MBZ-NH2 in muscle were 90.45%, 98.88%, 95.11%, respectively. The average extraction recovery of MBZ and its metabolites MBZ-OH and MBZ-NH2 in skin were 71.75%, 84.51%, 79.59%, respectively. The coefficients of variation of intra-day and inter-day were less than 10%.The results were estimated using non-compartmental analysis based on statistical moment theory. The main pharmacokinetic parameters of MBZ for plasma were as follows: elimination half-life (t1/2) was 6.051 h. Clearance (CL) was 0.060 L·h-1·kg-1. Apparent volume of distribution (Vd) was 0.528 L·kg-1. The area under the curve (AUC), the mean residence time (MRT), the time to peak concentration and the peak concentration were 16.499 mg·L-1·h-1, 27.749 h, 36 h, 0.675μg·mL-1, respectively. The main pharmacokinetic parameters of MBZ-OH for plasma were as follows: t1/2 was 2.093 h. CL was 0.174 L·h-1·kg-1. Vd was 0.526 L·kg-1. AUC, MRT, the time to peak concentration and the peak concentration were 5.739 mg·L-1·h-1, 18.227 h, 7 h, 0.204μg·mL-1, respectively. The main pharmacokinetic parameters of MBZ-NH2 for plasma were as follows: t1/2 was 67.242 h. CL was 0.022 L·h-1·kg-1. Vd was 2.144 L·kg-1. AUC, MRT, the time to peak concentration and the peak concentration were 45.227 mg·L-1·h-1, 100.392 h, 36 h, 0.586μg·mL-1, respectively. The eliminated equations in tissues were as follows: MBZ: Cmuscle=3.330e-0.059t, Cskin=0.706e-0.007t and correlation exponential were 0.995, 0.974 respectively. MBZ-OH: Cmuscle=2.438e-0.110t, Cskin=0.178e-0.019t and correlation exponential were 0.962, 0.978 respectively. MBZ-NH2: Cmuscle=0.380e-0.003t, Cskin=1.223e-0.004t and correlation exponential were 0.965, 0.951 respectively.The results showed that MBZ given to European eels via the water was absorbed and distributed rapidly and metabolized into MBZ-OH and MBZ-NH2. The drug concentration was high and distributed extensively in tissues. The penetration into tissues was strong and this drug could be effective in the treatment of whole body and deep tissues infections in European eels. The residual quantity of MBZ and its metabolites and elimination rate were different significantly in every tissue. The concentration of mebendazole and its metabolites in skin was apparently higher than muscle tissue and MBZ-NH2 in skin showed the slowest depletion, and the present study showed that skin can easily be adopted in tissue residues surveillance for MBZ in European eels. It was proposed that withdrawal time should not be less than 48days after bath-treated of MBZ in European eels at 25±1℃, according to the maximum residue limit of 0.02mg·kg-1.
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