Nitrogen isotopes in tooth enamel record diet and trophic level enrichment: Results from a controlled feeding experiment

Abstract

Nitrogen isotope ratios (δ15N) are a well-established tool for investigating the dietary and trophic behavior of animals in terrestrial and marine food webs. To date, δ15N values in fossils have primarily been measured in collagen extracted from bone or dentin, which is susceptible to degradation and rarely preserved in deep time (>100,000 years). In contrast, tooth enamel organic matter is protected from diagenetic alteration by the mineral structure of hydroxyapatite and thus is often preserved over geological time. However, due to the low nitrogen content (<0.01 %) of enamel, the measurement of its nitrogen isotopic composition has been prevented by the analytical limitations of traditional methods. Here, we present a novel application of the oxidation-denitrification method that allows measurement of δ15N values in tooth enamel (δ15Nenamel). This method involves the oxidation of nitrogen in enamel-bound organic matter to nitrate followed by bacterial conversion of nitrate to N2O, and requires ≥100 times less nitrogen than traditional approaches. To demonstrate that δ15Nenamel values record diet and trophic behavior, we conducted a controlled feeding experiment with rats and guinea pigs (n = 37). We determined that nitrogen concentration in tooth enamel (x̄ = 5.0 ± 1.0 nmol/mg) is sufficient for δ15Nenamel analyses with ≥5 mg untreated enamel powder. The nitrogen isotope composition of enamel reflects diet with an enrichment (Δ15Nenamel-diet) of ca. 2–4‰. δ15Nenamel values differ significantly between dietary groups and clearly record a shift from pre-experimental to experimental diet. The small sample size required (≤5 mg) by this method permits analyses of sample size-limited, diagenetically robust tooth enamel, and, as such it represents a promising new dietary proxy for reconstructing food webs and investigating the trophic ecology of extant and extinct taxa.