Received on June 9, 2004
Accepted on September 20, 2004

Drug Monitoring and Toxicology

Simultaneous and Sensitive Measurement of Anabasine, Nicotine, and Nicotine Metabolites in Human Urine by Liquid Chromatography/Tandem Mass Spectrometry

¹ Environmental Biomarker Shared Resource, The Cancer Institute of New Jersey, New Brunswick, NJ, and Environmental and Occupational Health Science Institute University of Medicine and Dentistry of New Jersey (UMDNJ)
² Toxicology, University of Medicine and Dentistry of New Jersey (UMDNJ), Piscataway, NJ

^* To whom correspondence should be addressed. E-mail: weisel{at}eohsi.rutgers.edu.

Background: Determination of nicotine metabolism/pharmacokinetics provides a useful tool for estimating uptake of nicotine and tobacco-related toxicants, for understanding the pharmacologic effects of nicotine and nicotine addiction, and for optimizing nicotine dependency treatment.

Methods: We developed a sensitive method for analysis of nicotine and five major nicotine metabolites, including cotinine, trans-3'-hydroxycotinine, nicotine-N'-oxide, cotinine-N-oxide, and nornicotine, in human urine by liquid chromatography coupled with a TSQ Quantum triple quadrupole tandem mass spectrometer (LC/MS/MS). Urine samples to which deuterium-labeled internal standards had been added were extracted with a simple solid-phase extraction procedure. Anabasine, a minor tobacco alkaloid, was also included.

Results: The quantification limits of the method were 0.1-0.2 µg/L, except for nicotine (1 µg/L). Cotinine-N-oxide, trans-3'-hydroxycotinine, nicotine, and anabasine in urine were almost completely recovered by the solid-phase extraction, whereas the mean extraction recoveries of nicotine-N'-oxide, cotinine, and nornicotine were 51.4%, 78.6%, and 78.8%, respectively. This procedure provided a linearity of three to four orders of magnitude for the target analytes: 0.2-400 µg/L for nicotine-N'-oxide, cotinine-N-oxide, and anabasine; 0.2-4000 µg/L for cotinine, nornicotine, and trans-3'-hydroxycotinine; and 1.0-4000 µg/L for nicotine. The overall interday method imprecision and recovery were 2.5-18% and 92-109%, respectively.

Conclusions: This sensitive LC/MS/MS procedure can be used to determine nicotine metabolism profiles of smokers, people during nicotine replacement therapy, and passively exposed nonsmokers. This method avoids the need for a time-consuming and labor-intensive sample enrichment step and thus allows for high-throughput sample preparation and automation.