IN VITRO DECOMPOSITION OF BONE COLLAGEN BY SOIL BACTERIA: THE IMPLICATIONS FOR STABLE ISOTOPE ANALYSIS IN ARCHAEOMETRY*

Abstract

To understand biogenic collagen type I decomposition and to establish how diagenesis may bias archaeometric data, modern mammalian bone was inoculated with a selection of ubiquitous soil bacteria. The presence of exogenous microbial biomass in the inoculated specimens was then checked microscopically prior to collagen extraction. The experimentally degraded bone collagen showed altered amino acid compositions, attributable to the selective breakdown of certain amino acids by the bacteria. While both the bulk collagen extract and the single amino acids exhibited shifts to more negative δ13 C‐values, enrichment was recorded for general δ13 N, and a depletion trend relative to unaltered collagen was observed for individual amino acid δ13 N. One explanation for the enrichment of the global δ15N‐values is cleavage of peptide bonds, which leaves 15 N within the substrate, while the change of 13C is mostly due to the altered amino acid composition. On the other hand, possible repolymerization of cleavage products under experimental conditions may also be responsible for the depletion trend of individual amino acid δ13C‐ and δ15 N‐values. This paper discusses the results as a basis for the development of a method for the reconstruction of the isotopic abundance of the original collagen, using the amino acid composition of the degraded product, the contribution of individual amino acids to its global δ‐values and of isotope discriminations implied in the microbial decomposition.

Experimental chemical degradation compared to natural diagenetic alteration of collagen: implications for collagen quality indicators for stable isotope analysis

“…Such alterations of δ-values would be significant in the case of dietary or habitat reconstruction, since a decrease of δ 15 N by 2‰ closely corresponds to one trophic level (Ambrose 1993), and an increase of δ 13 C by 3‰ would indicate a certain proportion of C 4 -plants or marine resources in the food of an individual coming from an otherwise C 3 -plant dominated habitat. Strikingly, the direction of these shifts of the isotopic ratios in the highly degraded samples 16 and 32 is opposite to the one reported after experimental microbial degradation of modern marten bone collagen, where a decrease of the δ 13 C-values and a fundamental increase of the δ 15 N-values was evidenced (Balzer et al 1997). Contrary to microbial decomposition, which obviously selectively metabolizes amino acids which play a role in the microbial metabolism, the kinetics of peptide bond cleavage are a likely explanation for the phenomenon, although also chemical hydrolysis finally leads to a non-random loss of certain amino acids.…”

“…In this process, the amino acid profile of collagen can be considerably altered, since microorganisms preferably tend to metabolize amino acids with a higher number of carbon atoms as a source of energy (Grupe and Turban-Just 1998;Grupe et al 2000). It has been shown that the collagen of modern bone which had experimentally been inoculated with soil bacteria exhibits modified stable isotopic ratios for carbon and nitrogen after incubation (Balzer et al 1997). The δ…”

“…The δ 15 N-values, however, exhibited a shift to more positive ratios in the magnitude of a whole trophic level effect, apparently a consequence of the cleavage of peptide bonds in the course of which the heavier 15 N preferably remains in the substrate (Bada et al 1989;Balzer et al 1997;Grupe et al 2000).…”

Reconstruction of breastfeeding and weaning practices using stable isotope and trace element analyses: A review

“…Empirical (Grupe, 1995) and experimental (Balzer et al, 1997) studies have shown that certain microbes preferentially degrade specific amino acid residues with large side chains in postmortem bone collagen such as phenylalanine, aspartic acid, methionine, glutamic acid, and histidine. It is possible that sulfur atoms are affected by diagenesis without evident changes in the concentration and isotope ratio of carbon and nitrogen because sulfur in bone collagen exists only as the methionine residue with a relatively much lower concentration (Eastoe, 1955).…”

Bone Diagenesis and its Implication for Disease Diagnosis: The Relevance of Bone Microstructure Analysis for the Study of Past Human Remains

“…crystallinity changes, porosity, and protein content), the histological study of bone integrity, simply or combined with other parameters, has been frequently used, not only to assess diagenetic changes (e.g., Hackett, 1981;Garland, 1987;Maat, 1993;NielsenMarsh & Hedges, 1999;Pfeiffer & Varney, 2000;Jackes et al, 2001;Turner-Walker & Syversen, 2002;Jans et al, 2004;Guarino et al, 2006;Turner-Walker & Jans, 2008;Abdel-Maksound, 2010;Hollund et al, 2012;Turner-Walker, 2012;Hollund et al, 2013), but also as a pre-screening tool to infer the degree of biomolecular (e.g., Hagelberg et al, 1991;Cipollaro et al, 1998;Zink et al, 2005), isotopic (e.g., Schoeninger et al, 1989;Balzer et al, 1997;Maurer et al, 2014) or paleopathological preservation (e.g., Stout, 1978;.…”