Ca isotope and dental microwear texture proxies were calibrated on teeth of extant vertebrates with known plant- and animal-based natural as well as custom-made diets. For this purpose animals (rats, guinea pigs, quails and iguanas) were raised in controlled feeding experiments also designed to simulate diet and trophic level switches and also wild animals with well-constrained feeding ecology from modern ecosystems were analyzed. The diet-to-bioapatite Ca isotope fractionation of dental tissues and bone was determined for different mammal, bird and reptile taxa fed the same distinct plant-, meat-, and insect-based pelleted diets. It seems to be similar across different vertebrate taxa, which permits an application to fossil and extinct taxa. Calcium and stable strontium isotopes enable us to distinguish both meat- and insect-feeders from plant-feeders. Diet-related Ca isotopes and enamel surface textures both have a high preservation potential in fossil teeth, which was confirmed by chemical and mechanical in vitro alteration experiments. Moreover, these methods facilitate minimal-invasive micro-sampling of enamel for Ca isotope and non-destructive 3DST analysis of teeth which is even applicable to a single tooth. For the first time, Ca isotope and 3DST analysis were combined to reconstruct the diet of extinct vertebrate taxa and their trophic level in fossil food webs. In Permo-Carboniferous food webs comprising the earliest presumed herbivores such as edaphosaurids, diadectomorphs and casaeids higher Ca isotope signatures in their bones and teeth were measured than for sympatric carnivorous pelycosaurs such as Dimetrodon or sphenacodontids, which is in line with herbivory of the former. This confirms that the first transition from animal- to plant-feeding among terrestrial tetrapods occurred around 300 million years ago or even earlier.
To broaden the versatility of the dietary toolbox, additional new isotopic diet and trophic level proxies such as Zn isotopes and N isotopes in enamel-bound organic matter were explored and validated on teeth from controlled feeding experiments and then for successfully applied to fossil teeth. For instance, Zn isotopes were used as trophic level proxy to assess the feeding ecology of the oldest anatomical modern human from SE Asia and its sympatric fauna, to trace a meat-rich diet of Neanderthals as well as to shed new light on potential food competition of the great white shark with the extinct Megalodon shark. Enamel-bound nitrogen isotopes display a similar trophic level effect of ca. 3-4 permille both in controlled rodent feeding experiments as well as for large mammals from modern African ecosystems. Enamel-bound N isotopes record the same dietary information as collagen-bound N of the same individuals and these diet-related N isotope compositions can be preserved over geological time scales in fossil teeth of Cenozoic mammals and megalodon sharks and even Mesozoic dinosaurs.
Food processing wears down teeth, thus affecting tooth functionality and evolutionary success. Therefore, another dietary proxy system microwear texture analysis of teeth was further explored which characterizes tooth surface wear at the microscopic level, enabeling us to distinguish soft- and hard-object feeders but also more subtle dietary differences and seasonality. In controlled feeding experiments fundamental factors influencing dental wear such as different natural diets (fruits, insects, seeds, vertebrates), amount, size and type of external mineral abrasives, amount of phytoliths, plant matter hydration state as well as the time to form and overwrite a diet-related surface texture were systematically assessed. Furthermore, mechanical and chemical alteration experiments were pursued to test the resistance of diet-related surface textures against taphonomic processes such as fluvial transport, aeolian sediment impact or acidic attack. A first catalogue of badly preserved enamel surface textures was compiled and can help to identify alteration of diet-related dental wear by postmortem surface modifications due to taphonomic or anthropogenic (i.e. excavation, preparation or conservation) effects.
