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AbstractThe study aimed to develop a sensitive and high-throughput liquid chromatography coupled with tandem mass spectrometry method to quantify concentrations of tramadol and paracetamol simultaneously in human plasma. Sample preparation involved single-step protein precipitation using methanol and two deuterated internal standards, tramadol D6 and paracetamol D4. Agilent Poroshell 120 EC-C18 (100 × 2.1 mm, 2.1 µm) analytical column was employed to achieve chromatographic separation. Detection was in positive ion multiple reaction monitoring mode. A tailing factor (Tf) of 1.5 from the column dead time and signal-to-noise (S/N) ratio >10, were obtained for analytes and internal standards. The standard curve was linear over the concentration range of 2.5–500.00 ng/mL for tramadol and 0.025–20.00 μg/mL for paracetamol. A small injection volume of 1 µL, low flow rate of 440 µL/min and short analysis time of 3.5 min reduced the solvent consumption, analysis cost and system contamination. The results of method validation parameters fulfilled the acceptance criteria of bioanalytical guidelines. The method was successfully applied to a bioequivalence study of fixed-dose combination products of tramadol and paracetamol in Malaysian healthy subjects.

Etoricoxib is a non-steroidal anti-inflammatory drug (NSAID) used to treat pain and inflammation. The objective of the current study was to develop a sensitive, fast and high-throughput HPLC-ESI-MS/MS method to measure etoricoxib levels in human plasma using a one-step methanol protein precipitation technique. A tandem mass spectrometer equipped with an electrospray ionization (ESI) source operated in a positive mode and multiple reaction monitoring (MRM) were used for data collection. The quantitative MRM transition ions were m/z 359.15 > 279.10 and m/z 363.10 > 282.10 for etoricoxib and IS. The linear range was from 10.00 to 4000.39 ng/mL and the validation parameters were within the acceptance limits of the European Medicine Agency (EMA) and Food and Drug Analysis (FDA) guidelines. The present method was sensitive (10.00 ng/mL with S/N > 40), simple, selective (K prime > 2), and fast (short run time of 2 min), with negligible matrix effect and consistent recovery, suitable for high throughput analysis. The method was used to quantitate etoricoxib plasma concentrations in a bioequivalence study of two 120 mg etoricoxib formulations. Incurred sample reanalysis results further supported that the method was robust and reproducible.