Aim: To develop a novel method for the bioanalytical extraction of trigonelline (TRG) from human plasma samples using a magnetic nanocomposite and to evaluate its pharmacokinetic profile. Materials & methods: Magnetic bentonite/β-cyclodextrine (β-CD) nanoparticles, coupled with a validated ion-pairing reversed-phase high-performance liquid chromatography method, were used to determine TRG concentration from plasma samples following a single oral administration. Results: The developed reversed-phase high-performance liquid chromatography method was accurate, precise, specific, selective and reproducible. TRG showed rapid absorption, middle rate of elimination and mean residence time of ∼24 h. The data were best fitted on a two-compartment model in which tmax was 1.0 h, Cmax 0.115 μg/ml, area under the curve (AUC)0-24 1.72 μg/ml.h, Cl 0.0293 l/h/kg, t1/2α 0.79 h, t1/2β 13.68 h and ka 1.63 h-1. Conclusion: The findings of this study could provide useful information to promote the future study of TRG and aid optimal dose finding. © 2021