•Magnetic boron nitride nanosheets (Fe3O4@BNNSs) composite was synthesized.•Fe3O4@BNNSs as the MSPE adsorbent was applied for the first time.•MSPE coupled with HPLC-MS/MS was used to analysis of PGRs.•The method was effective to determine PGRs in tomatoes. A novel magnetic boron nitride nanosheets (Fe3O4@BNNSs) composite-based magnetic solid-phase extraction (MSPE) method was employed to analyse six plant growth regulators (PGRs) in tomatoes combined with high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The novel Fe3O4@BNNSs composite was prepared via an in situ chemical coprecipitation process and characterized by scanning electron microscopy (SEM), Fourier transform infrared spectrometry (FT-IR), X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD), and vibrating sample magnetometry (VSM). Several factors that may affect the extraction efficiencies were optimized. Under the optimal factors, low limits of detection (0.002–0.010 ng g−1), good linear ranges (0.05–10 ng g−1) and satisfactory precisions (intra-day: 1.2%-3.9%; inter-day: 2.1%-6.9%) were achieved. The established approach was successfully employed to extract and determine PGRs in tomatoes, and the spiked recoveries were between 85.2 and 109.0%.