The incorporation of a chip-based high field asymmetric waveform ion mobility spectrometry (FAIMS) separation in the ultra (high)-performance liquid chromatography–high resolution mass spectrometry (UHPLC–HRMS) determination of the (R/S) ibuprofen 1-β-O-acyl glucuronide metabolite in urine is reported. UHPLC–FAIMS–HRMS reduced matrix chemical noise, improved the limit of quantitation approximately two-fold and increased the linear dynamic range compared to the determination of the metabolite without FAIMS separation. A quantitative evaluation of the prototype UHPLC–FAIMS–HRMS system showed better reproducibility for the drug metabolite (%RSD 2.7%) at biologically relevant concentrations in urine. In-source collision induced dissociation of the FAIMS-selected deprotonated metabolite was used to fragment the ion prior to mass analysis, enhancing selectivity by removing co-eluting species and aiding the qualitative identification of the metabolite by increasing the signal-to-noise ratio of the fragment ions.