Measurement of cotinine in urine by liquid chromatography tandem mass spectrometry

Ann Clin Biochem 2007;44:455-462
© 2007 Association for Clinical Biochemistry


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Original Articles

Carrie Ann Chadwick and
Brian Keevil

Department of Clinical Chemistry, Royal Liverpool University Hospital, Prescot Street, Liverpool L7 8XP, UK;
Department of Clinical Biochemistry, Wythenshawe Hospital, Southmoor Road, Manchester M23 9LT, UK

Background: Cotinine is the major metabolite of nicotine. Itis also a specific biomarker for nicotine exposure in cigarettesmokers. The measurement of urine cotinine concentration willenable: (1) the assessment of the smoking status of lung transplantpatients and (2) tobacco abstinence to be studied in patientsduring treatment under smoking cessation programmes.

Methods: We have developed and validated a method for the measurement of urinary cotinine using reversed phase high-performance liquid chromatography (HPLC) coupled to tandem mass spectrometry (LC-MS/MS). This technique utilizes online ion exchange coupled with an analytical column to eliminate ion suppression effects. The chromatography was performed using a WatersTM 2795 AllianceHT LC system.

Results: Cotinine and d3-cotinine had a retention time of 2.5 min and the cycle time from injection to injection was 4 min. The transition identified for cotinine was m/z 177.1>79.6 and for d3-cotinine m/z 180.2>79.6. This method was linear up to 1000 µg/L. Mean recovery of the assay was 112% with a range of 107-117% (n=9). The limit of quantitation for this assay was 2.5 µg/L and the limit of detection was 0.156 µg/L. The intra- and inter-assay imprecision was <12% and <10% respectively over a concentration range of 22-660 µg/L.

Conclusions: We have developed a robust and rapid assay formeasuring and analysing urine cotinine by LC-MS/MS, by utilizinga technique, which has reduced ion suppression effects. Ultimately,the method will facilitate the assessment of lung transplantpatients’ smoking status.

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