MMSPE-RP-HPLC method for the simultaneous determination of methimazole and selected metabolites in fish homogenates

A simple and rapid high-performance liquid chromatography method with ultraviolet detection was developed for the determination of the thyreostatic compound methimazole (1-methyl-2-mercaptoimidazole, MET) and metabolites 2-mercaptoimidazol (SHMET), N-methylthiourea (MTU) and N-methylhydantoin (MEH) from zebrafish (D. rerio) whole body homogenates using mixed mode solid phase extraction technique for sample pre-treatment. The highly polar compounds were separated on a difunctionally bonded silica based reverse phase column using gradient elution. Retention factors ranged between 1.53 and 5.66. The method was linear between 0.1 and 30 microg/ml, the detection limits were 0.4 ng for MET and SHMET, 0.6 ng for MTU and 2.6 ng for MEH. Extraction method was exhibited average recovery rates of 85.2-97.6%.