Simultaneous determination of 33 β2-agonists in swine urine and muscle using dual-template magnetic molecularly imprinted polymer clean-up coupled with UPLC-MS/MS

β 2 -agonists are a class of synthetic phenethanolamine compounds that pose multiple risks to public health. In this study, a dual-template magnetic molecularly imprinted polymer (Dual-T Fe₃O₄@MIP) approach was developed to enable the simultaneous detection of 33 β₂-agonists in swine urine and muscle tissues, with preliminary validation of its applicability in complex biological matrices. Briefly, samples were sheared and homogenized, followed by enzymatic hydrolysis using β-glucoronidase/arysulfatase in ammonium acetate buffer at 55 °C for 4 h. Thereafter, β 2 -agonists in the hydrolysate were extracted with the Dual-T Fe 3 O 4 @MIPs and eluted with an acidic methanol solution. The target compounds were quantified by UPLC-MS/MS equipped with an HSS T3 column (100 mm × 2.1 mm, 1.8 μm). The method was further validated in urine and muscle samples, demonstrating excellent linearity (R 2  ≥ 0.9944), high sensitivity (limit of detection ranging from 0.010 to 0.034 μg/kg, limit of quantitation ranging from 0.024 to 0.100 μg/kg), satisfactory recovery (ranging from 82.1% to 107%), and good precision (≤9.1%). Compared with the conventional solid-phase extraction method, which requires 3 h, the proposed method achieved purification in less than 30 min, representing a substantial improvement in analytical efficiency. The proposed approach is reliable, rapid, and user-friendly, making it suitable for routine analytical applications.