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Tramadol (TRM) and pregabalin (PGB) are frequently used in combination for neuropathic pain management. Accordingly, a selective and sensitive high-performance liquid chromatography–electrospray ionization–mass/mass spectrometric (ESI–LC–MS/MS) method is presented for determination of TRM and PGB, whether in pure forms or human biological fluids (plasma/urine), using gabapentin (GBP) (IS) as the internal standard. Chromatographic separation was effected in total run time of 2.5 min, on Phenomenex Luna® Omega 1.6 um polar C18 (LC 150 × 2.1 mm) column with a mobile phase of methanol/water (70:30, v/v), 0.1% (v/v) formic acid at a flow rate of 0.3 mL/min. Ionization of the analytes was obtained using electrospray in the positive ion mode (ESI+). The MS/MS detection was performed by monitoring the fragments for TRM, PGB and GBP on a triple quadrupole mass spectrometer. Assay calibration was over the range of 10–1000 ng mL−1 for TRM and PGB with the correlation coefficients over 0.999 in pure form, human plasma and urine spiked with the studied compounds. Validation data showed the inter-run relative standard deviations (RSDs) were less than 4.3% for TRM and 3.8% for PGB, whereas the intra-run RSDs were less than 3.7% for TRM and 3.6% for PGB. The mean extraction recoveries for TRM and PGB were in the ranges of 86.51–93.38% and 86.20–92.42%. This method was successfully performed on real plasma and urine samples taken from neuropathic patients and proved to be an applicable method for routine therapeutic drug monitoring of the proposed drug combination.

ESI–LC–MS/MS for Therapeutic Drug Monitoring of Binary Mixture of Pregabalin and Tramadol: Human Plasma and Urine Applications