Isotopic analysis of phosphate oxygen from bones and teeth (O-18(p)/O-16(p), delta O-18(p)) is a common tool used to investigate modern and ancient ecosystems and their climate. However, existing methods have expanded to use pretreatments for organic removal, require large sample sizes, or require extended precipitation timing. All together, these factors could affect accuracy and precision of delta O-18(p) measurement by promoting the formation of oxygen-bearing or nitrogen-rich contaminants. However, the nature and occurrence of contamination are not fully explored. Here we sought to develop a method of silver phosphate precipitation that tests the effect of different sample treatments and reduced sample sizes while preserving sample isotopic composition. Our protocol (SPORA) precipitates Ag3PO4 crystals from similar to 1.5 mg of starting material while purifying phosphate from contaminants, like nitrogen or carbonate. Isolation and purification of phosphate are achieved with an anion exchange resin, followed by precipitation of silver phosphate using an updated silver ammine solution that targets small amounts of phosphate in solution. We used a variety of phosphate oxygen reference materials and biogenic apatite materials, such as modern and fossil specimens with varying collagen content, to test the SPORA protocol and its effects on the resultant phosphate oxygen isotopic composition. Results were then compared to those from another published silver phosphate precipitation method (i.e., Rapid University of Chicago Dilute, Rapid UC). Overall, delta O-18(p) values of standards and biogenic apatites were similar between protocols (R-2 = 0.99, p << 0.05). In addition to isotope composition comparisons, UV-Vis spectroscopy and Fourier Transform Infrared (FTIR) analyses discerned phosphate recovery and material composition of crystals precipitated via different protocols, respectively. We found that the resin i) may retain similar to 10% of phosphate with no isotopic effects and ii) the SPORA protocol produces Ag3PO4 with more accurate delta O-18(p) measurements by preventing the formation of contaminant oxygen phases, silver oxide (Ag2O) and silver carbonate (Ag2CO3)(,) that confound the phosphate oxygen isotope composition. The SPORA Ag3PO4 precipitation procedure overcomes analytical limitations such as sample size and collagen contamination, conditions that other procedures for delta O-18(p) analysis cannot address simultaneously. The SPORA protocol can be used on a large array of bioapatite materials for paleoecological, paleoclimatic, and archeological applications, while reducing the required sample size and ensuring pure Ag3PO4 for isotopic analysis.
