THE EFFECTS OF ISOTOPICALLY VARIABLE WATER USED FOR TOOTH ENAMEL PRETREATMENT PRIOR TO STABLE ISOTOPIC ANALYSIS

Pretreatment of tooth enamel and other carbonate-bearing minerals prior to isotopic analysis is common practice, intended to remove organic matter and secondary carbonate from primary bioapatite without altering the isotopic composition. The purpose of this experiment was to determine if the water in aqueous solutions used to pretreat tooth enamel has an effect on the isotopic composition of the carbonate fraction of bioapatite. We hypothesized that pretreating enamel with labeled aqueous solutions would have no effect on the isotopic composition of bioapatite, beyond the removal of secondary carbonate. Our standard pretreatment method is: 1) soak enamel powder in 3% H₂O₂ for 30 min to remove organic matter, 2) rinse 5x with DI water and dry, 3) soak enamel powder in 1.0 M acetic acid Ca-acetate buffer (pH≈4.5) for 1 hr to remove secondary carbonate and 4) rinse 5x with DI water and dry. We also prepared a 0.1 M unbuffered acetic acid solution to use in step (3). To determine the effect of the labeled aqueous solutions, each step in the process was prepared using two naturally labeled waters: one ¹⁸O-enriched (δ¹⁸O = +16.4‰ V-SMOW) and the other ¹⁸O-depleted (δ¹⁸O = –10.0‰ V-SMOW). Enamel samples were drilled from 10 fossil elephant teeth collected from the middle Pliocene Hadar Fm. from Dikika, Ethiopia. The homogenized enamel powder was pretreated with the following methods using labeled solutions: untreated, H₂O₂ only, H₂O₂ + 1.0 M acetic acid Ca-acetate buffer, H₂O₂ + 0.1 M acetic acid. Results from these analyses indicate that pretreatment solutions prepared with isotopically distinct waters do significantly affect the δ¹⁸O­­­­ values of bioapatite after pretreatment. Enamel pretreated with ¹⁸O-enriched acidic solutions resulted in δ¹⁸O values ~0.5‰ more enriched than enamel pretreated with ¹⁸O-depleted acidic solutions. δ¹³C results indicate that both acidic pretreatment procedures are successful in removing organic carbon and secondary carbonate; we observed differences at each step of the analysis, but not between the different acid treatments. Although our results relate directly to tooth enamel pretreatment, they may have implications for any pretreatment of carbonate-bearing minerals in which oxygen isotopic composition is sought for terrestrial paleoclimate and paleoecological reconstructions.