Metabolomics-based analysis of miniature flask contents identifies tobacco mixture use among the ancient Maya

A particular type of miniature ceramic vessel locally known as "veneneras" is occasionally found during archaeological excavations in the Maya Area. To date, only one study of a collection of such containers successfully identified organic residues through coupled chromatography-mass spectrometry methods. That study identified traces of nicotine likely associated with tobacco. Here we present a more complete picture by analyzing a suite of possible complementary ingredients in tobacco mixtures across a collection of 14 miniature vessels. The collection includes four different vessel forms and allows for the comparison of specimens which had previously formed part of museum exhibitions with recently excavated, untreated containers. Archaeological samples were compared with fresh as well as cured reference materials from two different species of tobacco (Nicotiana tabacum and N. rustica). In addition, we sampled six more plants which are linked to mind-altering practices through Mesoamerican ethnohistoric or ethnographic records. Analyses were conducted using UPLC-MS metabolomics-based analytical techniques, which significantly expand the possible detection of chemical compounds compared to previous biomarker-focused studies. Results include the detection of more than 9000 residual chemical features. We trace, for the first time, the presence of Mexican marigold (Tagetes lucida) in presumptive polydrug mixtures.