Establishment of a two-dimensional liquid chromatography-tandem mass spectrometry system for detection of four tobacco-specific N-nitrosamines

In this work, we developed a system for the qualitative and quantitative analysis of four tobacco-specific N-nitrosamines (TSNAs). This system was based on strong cation exchange/reversed phase liquid chromatography-tandem mass spectrometry (SCX/RPLC-MS/MS). Benefiting from an efficient two-dimensional separation, the system could well circumvent tremendous matrix effects and provide excellent detection of ultra-low concentrations of TSNAs (ng mL⁻¹ level, after pretreatments) in Chinese Virginia cigarette mainstream smoke samples. Based on the features of TSNAs, one heart-cutting was used to transfer fractions from the first dimension to the second. A side flow was added between the two dimensions to gain retention in RPLC of TSNAs from SCX fractions. The system was evaluated using standard TSNA samples and mainstream cigarette smoke samples. The total separation time was within 20 minutes. The limits of quantitation (LOQs) for N′-nitrosonornicotine (NNN), 4-(methylnitrosoamino)-1-(3-pyridyl)-1-butanone (NNK), (R,S)-N-nitrosoanatabine (NAT) and (R,S)-N-nitrosoanabasine (NAB) were 39, 47, 58 and 9 pg mL⁻¹, respectively, which could well meet the requirements of all kinds of commercial cigarette samples. With Kentucky reference cigarettes as a standard sample, the relative deviations (r) of the measured values of TSNAs from the standard values were between −1.6% and 4.6%. And the relative standard deviations (RSD) of intra- and inter-day analysis were below 4.9% and 5.9%, respectively. These results revealed that the SCX/RPLC-MS/MS system we built could efficiently achieve minimal sample matrix effects and make highly sensitive analysis of pg mL⁻¹ level TSNAs in extremely complicated samples easy, fast and accurate. Finally, we successfully applied the system to the evaluation of TSNA yields in Chinese Virginia cigarettes.