TSRC, Tob. Sci. Res. Conf., 2014, 68, abstr. 76

Determination of heterocyclic aromatic amines in cigarette smoke by UPLC-MS/MS

Altria Client Services, Richmond, Virginia, USA

As stated in the Federal Register (Vol. 77, No. 64) Docket No. FDA-2012-N-0143, Heterocyclic Aromatic Amines (HAAs) are included in the “Established List of the Chemicals and Chemical Compounds Identified by FDA as Harmful and Potentially Harmful Constituents [HPHCs] in Tobacco Products and Tobacco Smoke.” Currently no standardized method exists for determination of the listed HAAs (AC, MeAC , PhIP, IQ, Trp-P-1, Trp-P-2, Glu-P-1 and Glu-P-2) in tobacco smoke. Therefore, the objective of this work was to develop a highly sensitive, selective and validated method for the determination of these HAAs in tobacco smoke using ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). For this new analytical method, five cigarettes were smoked under both ISO and Health Canada Intense conditions and the mainstream smoke was collected using a 44 mm Cambridge filter pad. The filter pad was extracted using an aqueous acidic solution (0.1 N hydrochloric acid) for 30 minutes and the smoke extract was subjected to solid phase extraction (SPE) clean up using an Oasis MCX cartridge to remove potential interferences prior to UPLC-MS/MS analysis. The chromatographic separation and the MS/MS parameters were optimized for the eight analytes and four isotopically labeled internal standards to achieve accurate quantitation of low levels of HAAs in cigarette smoke. The method was validated for linearity, accuracy, precision, limits of detection (LOD), limits of quantitation (LOQ), robustness and sample extract stability. The method exhibits good linearity (coefficient of determination of R2>0.995) in the concentration range of 4-200 ng/cigarette for all analytes, with a LOQ of 4 ng/cigarette. This validated method was used to determine the levels of HAAs in 3R4F, 1R5F, and CORESTA monitor (CM7) reference tobacco products.