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Thesis Format



Master of Science




Dr. Fred J. Longstaffe


The oxygen (O) isotope composition of mammalian collagen has the potential to provide information about the drinking water and hence geographic location and climate during the life of modern and ancient animals. An accurate and reproducible O-isotope analysis of collagen, however, has been elusive. Here, the O-isotope compositions of collagen from cortical bone of four megaherbivores were compared using two extraction procedures: the traditional Longin (HCl) method and the less commonly employed EDTA method. Preservation of O-isotope compositions was evaluated by measuring collagen peptide-chain masses, specifically the glutamine deamidation level (GDL). The EDTA method yielded collagen with a GDL of 1 – characteristic of unaltered collagen – and reproducible O-isotope results (±0.8‰). Much lower GDL values (0.2-0.95) and poorer reproducibility (±1.6‰) was obtained for the Longin method. This method uses strong acids to demineralize bioapatite releasing collagen, catalyzing reactions such as glutamine deamidation leading to O-isotope exchange with reagents, and should be avoided.

Summary for Lay Audience

Bones can provide crucial information about the life of a mammal – the food they ate, the water they drank, and the region where they lived. These details can be determined from the proportion of heavy to light isotopes of carbon, nitrogen and oxygen captured and preserved in bones during life. Such measurements are obtained from bones using the harder mineral portion called bioapatite or the softer tissue called collagen. This study focused on measurement of oxygen isotopes in collagen, which should reflect the meteoric water where the animal lived. Typically, collagen is extracted from bone using strong acids such as HCl, which does not affect the original carbon and nitrogen isotope measurements that provide information about diet. Reactions during collagen extraction using HCl and heat, however, change the original oxygen isotope compositions, which otherwise would have carried information about drinking water origin. This study focused on limiting these changes by using a collagen extraction method that employed a weaker acid (EDTA) and avoided heating. The success of the new method was demonstrated by measuring the chain masses of peptides (smaller versions of proteins) that make up the collagen and by the acquisition of reproducible oxygen isotope data for the collagen. The results from this study can be applied to a variety of applications including paleoclimatic and forensic research, both of which can benefit from reproducible oxygen isotope compositions in bone collagen.

Creative Commons License

Creative Commons Attribution 4.0 License
This work is licensed under a Creative Commons Attribution 4.0 License.

Available for download on Sunday, April 07, 2024