Detection of ASA metabolites in human plasma. (A), Metabolism of ASA. (B), Liquid chromatography–mass spectrometry (LC–MS) detection of ASA metabolite standards. (C), Experimental design. Human subjects were each given a single 325 mg dose of ASA, and blood was drawn both before (0 h) and either 4 (n = 5), 8 (n = 3), or 24 (n = 3) h later. Plasma was isolated from blood, and 0.5 mL was extracted for LC–MS analysis. (D), Plasma salicylate levels. Error bars indicate SEM. * p = 0.02, ** p = 0.001. (E), Plasma salicylurate levels. Error bars indicate SEM. * p = 0.07, ** p < 0.001. (F), Plasma gentisic acid levels. Error bars indicate SEM. * p = 0.03, ** p < 0.001. Note: ASA and salicyl acyl glucuronide were not detected in any of the samples.