Characterization of Proteomes Extracted through Collagen-based Stable Isotope and Radiocarbon Dating Methods

Isotope analyses on "collagen" extracted from ancient bone have been routinely used for dietary and chronological inferences worldwide for decades. These methods involve the decalcification of biomineralized tissues with acid, often followed by processes to remove exogenous contaminants, and then gelatinization of what is often described as the "collagen" fraction. However, little is known about the relative content of collagen to the many other noncollagenous proteins (NCPs) potentially present. Some of these NCPs have great longevity in ancient bone, and some, for example, fetuin-A, are useful for obtaining better taxonomic information than collagen. This study uses Orbitrap Elite LC-MS/MS to characterize the proteomes of the acid-soluble and base-soluble fractions, which are usually discarded, and the gelatinized "collagen" fraction obtained from both stable isotope and radiocarbon methods applied to several ancient bovine bones. The results showed that all fractions tested contain numerous NCPs, but the base-soluble fraction for both methods contains the greatest number of NCPs with the highest relative abundances. This study confirms that not only do the waste fractions obtained from the "collagen" extraction procedure of stable isotope or radiocarbon dating methods yield a plentiful resource of NCPs that is currently being overlooked but that they also provide proteomes as complex as those obtained from standard proteomics methods.