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Determination of cotinine by LC-MS-MS with automated solid-phase extraction.

Cotinine is the primary metabolite of nicotine and the preferred biomarker for assessing cigarette smoke exposure. Several liquid chromatography-tandem mass spectrometry (LC-MS-MS) methods have been described for measuring cotinine in biological fluids. Sample preparation typically involves manual solvent evaporation and reconstitution steps. This study describes a novel LC-MS-MS method for the quantification of cotinine by using electrospray ionization with multiple reaction monitoring and cotinine-d3 as internal standard, coupled with an automated solid-phase extraction (SPE) procedure. The assay was linear over the analytical range of 0.5-1,000 ng/mL. The limits of detection and quantification were 0.13 and 0.20 ng/mL, respectively. Intra-assay and inter-assay imprecision of cotinine in all samples was <5 and <10% (coefficient of variation), respectively. The analytical recovery of cotinine spiked into plasma was >95-100%. Matrix effects in serum and plasma were <10%. A rapid, sensitive and specific LC-MS-MS method was developed and validated for the determination of cotinine in human plasma, using a straightforward automated SPE protocol. The application of this method to an epidemiological study has demonstrated its utility for batch analyses of a large sample set (>500 samples).