A simple analytical method using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in atmospheric chemical ionization mode (APCI) for the simultaneous estimation of acetylsalicylic acid (ASA, CAS 50-78-2) and its active metabolite salicylic acid (SA, CAS 69-72-7) in human plasma has been developed and validated. ASA and SA were analyzed simultaneously despite differences in plasma concentration ranges of ASA and SA after oral administration of ASA. In spite of having different chemical, ionization and chromatographic properties, ASA and SA were extracted simultaneously from the plasma sample using acetonitrile protein precipitation followed by liquid-liquid extraction. The analytes were separated on a reversed phase column with rapid gradient program using mobile phase consisting of ammonium acetate buffer and methanol. The structural analogue diclofenac was used as an internal standard. The multiple reaction monitoring (MRM) transitions m/z 179 --> 137 for ASA, m/z 137 --> 65 for SA and m/z 294 --> 250 for IS were used. The assay exhibited a linear dynamic range of 0.02-10 microg/mL for ASA and 0.1-50 microg/mL for SA. The between-batch precision (%CV) ranged from 2.1 to 7.9% for ASA and from 0.2 to 5.2% for SA. The between-batch accuracy ranged from 95.4 to 96.7% for ASA and from 94.6 to 111.3% for SA. The validated method was successfully applied for the evaluation of pharmacokinetics of ASA after single oral administration of 650 mg test formulation versus two 325 mg reference formulations of ASA in human subjects.

A simple LC-MS/MS method was developed and validated for the estimation of sarpogrelate in 50 µL of rat plasma. The analyte and internal standard (IS) were extracted from rat plasma by acetonitrile precipitation and they were separated on a reversed-phase C₈ column with gradient program. The MS acquisition was performed with multiple reaction monitoring mode using m/z 430.2 to m/z 135.0 for analyte and m/z 448.2 to m/z 285.3 for IS. The calibration curves were linear over the range of 1-1000 ng/mL with the correlation coefficient greater than 0.999. With dilution integrity up to 20-fold, the upper limit of quantification was extendable up to 15,000 ng/mL. The method was successfully applied to the analysis of rat plasma samples after single dose oral administration of sarpogrelate at 5 mg/kg to rats for the determination of its pharmacokinetics. Following oral administration the maximum mean concentration in plasma (C(max), 11514 ng/mL) was achieved at 0.25 h (T(max)) and the area under curve (AUC₀-₂₄) was 11051 ± 3315 ng h/mL. The half-life (t(¹/₂)) and clearance (Cl) were 2.9 ± 1.1  h and 490 ± 171 mL/h/kg, respectively. We believe that development of a method in rodent plasma would facilitate the ease of adaptability of sarpogrelate in human plasma.

A specific, precise, rapid and reliable stability indicating UPLC method has been developed and validated for simultaneous separation and estimation of Epalrestat and Pregabalin in bulk drugs and marketed formulations. Chromatographic separation was achieved on a agilent Zorbax SB C18 (2.1 x 100mm, 1.8µm) column using isocratic mode of elution with 0.1% Formic acid and Acetonitrile in the ratio 60:40 v/v at a flow rate of 1.0 mL/min and analytes were monitored at 226nm. The retention times of Epalrestat and Pregabalin were about 0.97 and 1.27 minutes respectively. The developed method was validated for specificity, linearity, precision, accuracy, ruggedness and stress degradation studies were established. The results of specificity and stability studies indicate that there was no interference of diluent, excipients, and degradation products at respective retention times of analytes. The detector response was linear in the range of 10-150% level with respect to test concentration of Epalrestat and Pregabalin. Correlation coefficient (R2) was found not less than 0.999 for both analytes. The percentage assay of Epalrestat and Pregabalin were about 100.99% and 100.77% respectively. Hence, the developed method was rapid, specific, accurate and economical, and it can be used for routine analysis of these drugs in bulk and marketed formulations.

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