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A sensitive and robust ultra-high-performance liquid chromatography-tandem mass spectrometry method has been developed and validated for the quantification of acetaminophen, dexchlorpheniramine, caffeine, cotinine and salicylic acid in postmortem blood samples from children younger than 4 years. The sample was prepared by a protein precipitation with ice-cold methanol/acetonitrile mixture (85:15, v/v). The organic phase was evaporated to dryness and the residue was dissolved in the mobile phase. Separation, with gradient elution and an acidic mobile phase, was achieved on an Acquity UPLC® HSS T3 column. The compounds were quantified using a multiple reaction-monitoring mode. Two transitions were monitored for each compound and one for the deuterated internal standards. The mass spectrometric detection in the positive ion mode was performed for all the compounds except salicylic acid which was detected in the negative ionization mode. The limits of quantification were as follows: acetaminophen 0.30 mg/L, dexchlorpheniramine 0.0050 mg/L, caffeine 0.099 mg/L, cotinine 0.00035 mg/L and salicylic acid 1.3 mg/L. Between-assay and within-assay precisions were ≤15% (biases: -10% to 26%) and ≤10%, respectively. Extraction recoveries varied from 93% to 137%. The matrix effects in blood, corrected with deuterated internal standards, were 100% ± 10% for all compounds except dexchlorpheniramine (111%) and caffeine (138%).

Determination of Acetaminophen, Dexchlorpheniramine, Caffeine, Cotinine and Salicylic acid in 100 μL of Whole Blood by UHPLC–MS/MS