Improving the accuracy of ¹⁸O and ¹⁷O values of O₂ measured by continuous-flow isotope-ratio mass spectrometry with a multipoint isotope-ratio calibration

Rationale: Stable isotope analysis of O-2 is a valuable tool to identify O-2-consuming processes in the environment; however, reference materials for O-2 isotope analysis are lacking. Consequently, a one-point calibration with O-2 from ambient air is often applied, which can lead to substantial measurement uncertainties. Our goals were to develop a simple multipoint isotope-ratio calibration approach and to determine measurement errors of delta O-18 and delta O-17 values of O-2 associated with a one-point calibration.Methods: We produced O-2 photosynthetically with extracted spinach thylakoids from source waters with delta O-18 values of -56 parts per thousand to +95 parts per thousand and delta O-17 values of -30 parts per thousand to +46 parts per thousand. Photosynthesis was chosen because this process does not cause isotopic fractionation, so that the O isotopic composition of the produced O-2 will be identical to that of the source water. The delta O-18 and delta O-17 values of the produced O-2 were measured by gas chromatography coupled with isotope-ratio mass spectrometry (GC/IRMS), applying a common one-point calibration.Results: Linear regressions between delta O-18 or delta O-17 values of the produced O-2 and those of the corresponding source waters resulted in slopes of 0.99 +/- 0.01 and 0.92 +/- 0.10, respectively. In the tested delta range, a one-point calibration thus introduced maximum errors of 0.8 parts per thousand and 3.3 parts per thousand for delta O-18 and delta O-17, respectively. Triple oxygen isotopic measurements of O-2 during consumption by Fe2+ resulted in a delta O-18-delta O-17 relationship (lambda) of 0.49 +/- 0.01 without delta scale correction, slightly lower than expected for mass-dependent O isotopic fractionation.Conclusions: No significant bias is introduced on the delta O-18 scale when applying a one-point calibration with O-2 from ambient air during O-2 isotope analysis. Both O-2 formation and consumption experiments, however, indicate a delta O-17 scale compression. Consequently, delta O-17 values cannot be measured accurately by GC/IRMS with a one-point calibration without determining the delta O-17 scale correction factor, e.g. with the O-2 formation experiments described here.