RAMAN SPECTROSCOPY REVEALS PHYLOGENETIC AND PHYSIOLOGICAL SIGNALS FOR CARBONATE CONTENT IN BONE MINERAL

Bone consists of an organic component, collagen, and a mineral calcium phosphate, commonly equated with hydroxylapatite, but that often includes other cations (e.g., Fe, Mg, Sr, Zn) and carbonate (CO₃) as an anion. However, several fundamental questions regarding bone mineral composition and regular composition variables have not been addressed before. We hypothesized that carbonate content was lower in slower growing and metabolically less active species (such as fishes and amphibians) and higher in fast growing and metabolically active species (mammals and birds). Acknowledging the fundamentally different ossification parameters we further asked if dermal bone and chondral bone have the same carbonate content. Using Raman spectroscopy, we compared taxonomically highly diverse samples of vertebrates covering all major groups and the two dominant ossification modes – dermal and endochondral ossification.

We found carbonate content in vertebrates to be unexpectedly high and variable, with values ranging from 5% to 11%. Fishes and amphibians showed lower carbonate content (5%-7%) than mammals, reptiles, and birds (8%-11%). While dermal and chondral bone tend to be similar in value we found that in large-mouth bass, cat, common brushtail possum, grebe, and sharp-shinned hawk carbonate content in dermal bone was higher than in chondral bone. This phenomenon was not restricted to endothermic species, and could be connected to nutrient prioritization during ontogeny. Further, we found that cold-blooded crocodiles, descended from endothermic terrestrial archosaurs, had similarly high carbonate values as highly endothermic birds, their closest living relatives. This suggests that metabolic rate may not correlate with carbonate content.

The results of our study indicate substantial proportions of phosphate can be substituted by carbonate in bone mineral. It further hints at a simple correlation of bone deposition rate and carbonate content, with species depositing more bone per time unit having a higher carbonate content.