Analysis of coumarin and angelica lactones in smokeless tobacco products

Differences in health risks between different styles of smokeless tobacco products (STPs) have prompted interest in their relative levels of toxic chemicals. We report here the development of methods for the analysis of STPs for coumarin and for -angelica lactone (-AL), both of which have been included in various published lists of tobacco toxicants. We have also determined the concentrations of these lactones in commercial STPs from the US and Sweden, representing 80-90% of the 2010 market share for all the major STP categories in these two countries: 65 products (plus two reference products) for coumarin and 66 commercial products for -AL. For coumarin, methanol extracts of the STPs were analysed by HPLC/MS/MS. The lower limit of quantification (LOQ) and limit of detection (LOD) were, respectively, 100 and 30ng coumarin/g of STP on a wet weight basis (WWB). Alpha-AL was determined via direct headspace GC/MS. The LOQ and LOD were 65 and 30ng/g WWB respectively. Coumarin was detected In 3/33 Swedish snus, 5/13 US chewing tobaccos, 16/16 moist snuffs and 5/6 dry snuffs. Concentrations in those samples with quantifiable coumarin contents ranged from 186 to 1656ng/g WWB. Concentrations of coumarin measured in this study were consistent with levels naturally found in tobacco. None of the STPs analysed would significantly contribute to coumarin exposure in consumers compared with dietary sources, and estimated exposure levels were 1000x lower than the European Food Safety Authority Tolerable Daily Intake. Hence the relevance of coumarin to the toxicity of STPs and its inclusion in the FDA's list of harmful and potentially harmful compounds list is questionable. Measurements of -AL in these STPs found that the majority did not have quantifiable contents, however, for three STPs concentrations of -AL were above the LOQ (116-140ng/g WWB) and for four other STPs concentrations of -AL could be estimated between the LOD and LOQ. Beta-angelica lactone was tentatively identified in three of the STPs but the levels could not be reliably quantified. The levels of -AL in tobacco products are reported here for the first time, but the relevance of -AL to the toxicity of STPs is also highly questionable given that it has GRAS status as a permitted food additive.