Actualistic Testing of the Influence of Groundwater Chemistry on Degradation of Collagen I in Bone

Abstract

Recent experiments have heightened our understanding of reactions which can stabilize biomolecules during early diagenesis, yet little remains known about how groundwater chemistry can aid or hinder molecular preservation within a bone through geologic time. To elucidate this issue, we conducted actualistic experiments of bone decay employing varied fluid compositions to simulate a suite of groundwaters. Modern domestic chicken (Gallus gallus) femora were placed in a matrix of compositionally- and texturally-mature, fluvially-deposited sand. To simulate groundwater flow, deionized water or solutions enriched in calcium carbonate, phosphate, or iron were percolated through separate trials for a period of 90 days. After completion of the experiment, degradation of the bones was examined via histologic thin sectioning and two immunoassays against collagen I, the primary bone structural protein: immunofluorescence and enzyme-linked immunosorbent assay. Collagen loss was found to be greatest in the iron trial and least in the calcium carbonate trial, the latter of which experienced partial permineralization with calcite over the course of the experiment. Specifically, the iron trial was found to retain only ~35 ng of collagen I per 100 ng of protein extract, whereas the calcium carbonate trial retained ~90 ng of collagen I. Further, in the iron and calcium carbonate trials, cementation of sediment onto bone surfaces preferentially occurred over more porous regions of the epiphyses, perhaps stimulated by greater release of decay compounds from these regions of the bones. Of the two trials exhibiting intermediate results, the phosphate trial induced slightly greater decay of collagen than the deionized water control, which retained ~60 ng and ~80 ng of collagen I per 100 ng of protein extract, respectively. These results demonstrate that highly acidic conditions during early diagenesis can overwhelm any preservative effects of free radical-mediated stabilization reactions, whereas early-diagenetic permineralization can drastically slow biomolecular decay (ostensibly by hampering microbial access to the interior of a bone), thereby increasing the likelihood of a bone to retain biomolecules and/or their decay products through protracted diagenesis. Future variations of this actualistic experiment employing varied durations, solute concentrations, bacterial communities, pH values, and/or host sediments could provide further important insights into the ways in which early-diagenetic environments control the initial decay of biomolecules within bone and other tissues.