Determination of clenbuterol in urine using headspace solid phase microextraction or liquid–liquid–liquid microextraction

Abstract

Headspace solid phase microextraction (HS-SPME) and liquid–liquid–liquid microextraction with automated movement of the acceptor phase (LLLME/AMAP) techniques are described for the extraction of clenbuterol (CB) in urine. HS-SPME technique involves the extraction of the drug by SPME fibre in a headspace mode with a cooling device at the upper part of the vial to cool the headspace of the vial where the fiber is suspended. This cooling effect will enhance the absorption of analytes by the fiber. As the cooling system is surrounding the vial, the headspace will also be cooled in addition to the cooling of the fiber. After extraction the derivatization of the extracted drug with hexamethyldisilazane (HMDS) was performed by suspending the fiber in the headspace of another vial saturated with the vapor of HMDS. This derivatized compound was analyzed by gas chromatography with mass spectrometric detection (GC/MS). LLLME/AMAP technique involves the extraction of CB, a basic drug, from an alkaline solution into the organic solvent residing in the pores of the hollow fiber and then back extracted into the acidic acceptor solution inside the lumen of the hollow fiber. The acceptor solution was repeatedly moved in and out of the hollow fibre assisted by a syringe pump. This repeated movement provides fresh acceptor phase to come in-contact with the organic phase and thus improving the efficiency of extraction. Quantification was performed using high performance liquid chromatography with ultraviolet (HPLC/UV) detection. In both the techniques, experimental parameters have been studied to achieve greater sensitivity. Linearity was observed over the range of 1–1000 ng ml −1 ( R 2 = 0.9990) and 50–3000 ng ml −1 ( R 2 = 0.9956) with detection limits of 0.23 ng ml −1 and 3.9 ng ml −1, respectively, for HS-SPME-GC/MS and LLLME/AMAP-HPLC/UV method. R.S.D. values of 3.9% (HS-SPME-GC/MS) and 5.8% (LLLME/AMAP-HPLC/UV) indicated good precision of the techniques. Absolute recovery was found to be 0.007% and 18%, respectively, for HS-SPME-GC/MS and LLLME-HPLC/UV methods. Finally, the techniques have been applied for the analysis of CB in urine samples. No effort has been made to compare the method with official method.