Analysis of isomeric opioids in urine using LC-TIMS-TOF MS

In the present work, a fast separation, identification and quantification workflow based on liquid chromatography coupled to trapped ion mobility in tandem with mass spectrometry (LC-TIMS-MS) is described for the analysis of common isomeric drugs of abuse and their metabolites in human urine. In particular, the analytical performance of LC-TIMS-MS is shown for identification based on retention time, collision cross section and accurate mass for three sets of common isomeric opioids and their deuterated analogs in urine. The LC-TIMS-MS analysis provided limits of detection of 1.4-35.2 ng/mL with demonstrated linearity up to 500 ng/mL, enabling discovery and targeted monitoring (DTM) of opioids in urine, with high precision in retention times (RT) (< 0.3%), collision cross sections (CCS) (< 0.6%) and mass accuracy (< 1 ppm) across multiple measurements using external calibration. A good agreement was observed between theoretical and experimental CCS from candidate structures optimized at the DFT/B3LYP level. The need for complementary liquid and mobility separations prior to mass analysis is shown for the analysis of complex mixtures, with mobility resolving power of 80-130. The reproducibility and high speed of LC-TIMS-MS analysis provides a powerful platform for drug and metabolite screening in biological matrices with higher precision and confidence than traditional LC-multiple reaction monitoring (MRM) approaches.