Carbon, nitrogen, and oxygen stable isotopes in modern tooth enamel: A case study from Gorongosa National Park, central Mozambique

The analyses of the stable isotope ratios of carbon (δ¹³C), nitrogen (δ¹⁵N), and oxygen (δ¹⁸O) in animal tissues are powerful tools for reconstructing the feeding behavior of individual animals and characterizing trophic interactions in food webs. Of these biomaterials, tooth enamel is the hardest, most mineralized vertebrate tissue and therefore least likely to be affected by chemical alteration (i.e., its isotopic composition can be preserved over millions of years), making it an important and widely available archive for biologists and paleontologists. Here, we present the first combined measurements of δ¹³C, δ¹⁵N, and δ¹⁸O in enamel from the teeth of modern fauna (herbivores, carnivores, and omnivores) from the well-studied ecosystem of Gorongosa National Park (GNP) in central Mozambique. We use two novel methods to produce high-precision stable isotope enamel data: (i) the “oxidation-denitrification method,” which permits the measurement of mineral-bound organic nitrogen in tooth enamel (δ¹⁵N_enamel), which until now, has not been possible due to enamel’s low organic content, and (ii) the “cold trap method,” which greatly reduces the sample size required for traditional measurements of inorganic δ¹³C_enamel and δ¹⁸O_enamel (from ≥0.5 to ≤0.1 mg), permitting analysis of small or valuable teeth and high-resolution serial sampling of enamel. The stable isotope results for GNP fauna reveal important ecological information about the trophic level, dietary niche, and resource consumption. δ¹⁵N_enamel values clearly differentiate trophic level (i.e., carnivore δ¹⁵N_enamel values are 4.0‰ higher, on average, than herbivores), δ¹³C_enamel values distinguish C₃ and/or C₄ biomass consumption, and δ¹⁸O_enamel values reflect local meteoric water (δ¹⁸O_water) in the park. Analysis of combined carbon, nitrogen, and oxygen stable isotope data permits geochemical separation of grazers, browsers, omnivores, and carnivores according to their isotopic niche, while mixed-feeding herbivores cannot be clearly distinguished from other dietary groups. These results confirm that combined C, N, and O isotope analyses of a single aliquot of tooth enamel can be used to reconstruct diet and trophic niches. Given its resistance to chemical alteration, the analysis of these three isotopes in tooth enamel has a high potential to open new avenues of research in (paleo)ecology and paleontology.