Automated In-Tube Solid-Phase Microextraction Coupled with Liquid Chromatography/Electrospray Ionization Mass Spectrometry for the Determination of β-Blockers and Metabolites in Urine and Serum Samples

Abstract

The technique of automated in-tube solid-phase microextraction (SPME) coupled with liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) was evaluated for the determination of β-blockers in urine and serum samples. In-tube SPME is an extraction technique for organic compounds in aqueous samples, in which analytes are extracted from the sample directly into an open tubular capillary by repeated draw/eject cycles of sample solution. LC/MS analyses of β-blockers were initially performed by liquid injection onto a LC column. Nine β-blockers tested in this study gave very simple ESI mass spectra, and strong signals corresponding to [M + H]⁺ were observed for all β-blockers. The β-blockers were separated with a Hypersil BDS C₁₈ column using acetonitrile/methanol/water/acetic acid (15:15:70:1) as a mobile phase. To optimize the extraction of β-blockers, several in-tube SPME parameters were examined. The optimum extraction conditions were 15 draw/eject cycles of 30 μL of sample in 100 mM Tris-HCl (pH 8.5) at a flow rate of 100 μL/min using an Omegawax 250 capillary (Supelco, Bellefonte, PA). The β-blockers extracted by the capillary were easily desorbed by mobile-phase flow, and carryover of β-blockers was not observed. Using in-tube SPME/LC/ESI-MS with selected ion monitoring, the calibration curves of β-blockers were linear in the range from 2 to 100 ng/mL with correlation coefficients above 0.9982 (n = 18) and detection limits (S/N = 3) of 0.1−1.2 ng/mL. This method was successfully applied to the analysis of biological samples without interference peaks. The recoveries of β-blockers spiked into human urine and serum samples were above 84 and 71%, respectively. A serum sample from a patient administrated propranolol was analyzed using this method and both propranolol and its metabolites were detected.