Simultaneous Determination of Five Tobacco-Specific Nitrosamines in Mainstream Cigarette Smoke by Isotope Dilution Liquid Chromatography/Electrospray Ionization Tandem Mass Spectrometry

Abstract

Tobacco-specific nitrosamines (TSNAs) have been previously implicated as a source of carcinogenicity in tobacco and cigarette smoke. Accurate quantification of these chemicals is needed to help assess public health risk. We have developed and validated a specific and sensitive method to simultaneously measure five TSNAs in the particulate phase of mainstream tobacco smoke. Cigarette smoke particulate, produced using standardized machine smoking protocols, was collected on a Cambridge filter pad. The particulate matter was extracted using methylene chloride, back extracted into aqueous solution, further purified by solid-phase extraction, and analyzed by liquid chromatography/electrospray ionization tandem mass spectrometry using isotopically labeled analogues as internal standards. Limits of detection for this method ranged from 0.05 to 1.23 ng/mL using an injection volume of 20 microL. A linear calibration range spanning 2.5-2500 ng/mL was adequate to measure TSNA levels in cigarette smoke. The method achieved excellent reproducibility and accuracy. The identity of each TSNA was established by chromatographic retention time, analyte-specific fragmentation patterns, and relative peak area ratios of two product/precursor ion pairs. This new method provides higher sensitivity, specificity, and throughput than earlier methods for TSNA determination.

Rapid and sensitive gas chromatography–ion-trap tandem mass spectrometry method for the determination of tobacco-specific N-nitrosamines in secondhand smoke

“…In all cases, MDLs and MQLs were significantly below the levels determined in the SHS samples (see below). Furthermore, compared to other methods which involve a preconcentration step and clean-up procedure, MDLs for this method are much lower than for the GC/TEA method, are similar to those found using LC/MS/MS [15][16][17] and are better than those reported by Zhou et al [19] using GC-IT-MS/MS. Lower MDLs may be attained by using an accelerated solvent extraction (ASE) with lower solvent volumes, or employing a preconcentration step using rotary evaporator or solid phase cartridges (SPE).…”

“…For example, Caldwell and Conner [26] suggested that artifacts form via nicotine nitrosation during smoke collection. In contrast, other groups [15,27] did not observe artifact formation during collection of TSNAs on untreated Cambridge Filters. It has been reported that the formation of artifactual nitrosamines can be reduced with the addition of ascorbic acid to the sample collection system [26].…”

“…However, the GC-TEA method has two main disadvantages: (1) it cannot differentiate the co-eluted nitroso-compounds although it is nitroso-specific, and (2) extensive sample preparation, including pre-concentration and clean-up using liquidliquid (L-L) extraction and/or solid phase extraction (SPE), is necessary due to the limited sensitivity of the method. Recently, LC-MS/MS methods have been developed for TSNA analysis, which provide greater sensitivity and selectivity than the GC-TEA method [15][16][17][18]. However, sample matrix effects, can lead to poor analyte recoveries and decreased accuracy and precision.…”

“…However, sample matrix effects, can lead to poor analyte recoveries and decreased accuracy and precision. Sample clean up using L-L or SPE [15,18] can reduce July 13, 2009 5 these effects, but this increases analysis time and limits sample throughput. Another method uses stable isotopically-labeled standards [15][16][17].…”

“…Sample clean up using L-L or SPE [15,18] can reduce July 13, 2009 5 these effects, but this increases analysis time and limits sample throughput. Another method uses stable isotopically-labeled standards [15][16][17]. Although this approach is effective, it significantly increases the cost of the analysis, making it impractical for routine use.…”