A comparison of pretreatment methods for the analysis of phosphate oxygen isotope ratios in bioapatite

The integrity of the biological phosphate oxygen isotope (δ(18)O(p) ) signal is thought to be contingent upon the complete removal of competing sources of oxygen such as associated organic matter. A range of pretreatment methods to purify phosphate material from competing sources of oxygen has been reported, with contradictory evidence on the usefulness and efficiency of one or another. Yet, a systematic comparison of these techniques for bioapatite phosphate has not been conducted.

Chemical and thermal pretreatment techniques were tested for their effectiveness at removing organic matter and the likelihood that they modify original δ(18)O values. The test was performed in inorganic (synthetic apatite and a phosphorite rock) and organic (bone and tooth tissues) phosphate materials for which we had an expectation of the actual original δ(18)O(p) value. Analysis of nitrogen content (wt.%), scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectroscopy were employed.

We detected variable efficiency at removing organic matter between pretreatment methods with no correlation to any specific structural change. The δ(18)O(p) results showed considerable variation between samples pretreated with the different methods and the untreated samples, with a compositional range of up to 4.5 ‰ in the bone samples. Variations of the δ(18)O(p) values within error were found for tooth enamel, phosphorite rock and inorganic apatite.

We recommend a cautious approach when interpreting and comparing δ(18)O(p) data from bone samples treated with different pretreatment protocols. In general, the untreated samples seem to show δ(18)O(p) values closer to the expected ones. According to our results, pretreatment is completely unnecessary in highly mineralized tissues.