Rapid Identification of 44 Steroids in Human Urine Samples using HPLC-ESI-QTOF-MS

Objective: Detailed analysis of un-processed and un-derivatized free and conjugated urinary steroids is useful to avoid miscalculations and to diagnose sports doping and adrenal problems including abnormal steroidogenesis, congenital deficiency of related enzymes, cancer, and other disease conditions. Hence, the present study was conducted to develop a soft ionization method to identify the maximum number of urinary steroids using ultra-performance liquid chromatography coupled with quadrupole time of flight mass spectrometer (HPLC–Q-TOF-MS). Material and Methods: HPLC–Q-TOF-MS was carried out for the qualitative detection of steroids and their conjugates in urine samples. The method provides high sensitivity and fast analysis of steroids and their glucuronides without hydrolysis or sample preparation or extraction of steroids. Results: Using the method, 44 steroids belonging to C-18, C-19, and C-21 classes and their conjugates were resolved and identified using positive and negative modes of ionizations by their characteristic ionization and collision energy induced dissociation behaviors. Conclusion: The method is time-saving and good to compare samples from different peoples with control or healthy ones as it doesn’t require any kind of pre-treatment or sample processing. It provides a complete picture of steroids metabolism and catabolism. It can be good for doping control or to explore the effects of other drugs. However, in qualitative analysis one may miss the significant information unless direct methods of steroids analysis to be employed.