A comparison of extraction systems for plant water stable isotope analysis

Rationale The stable isotope ratios of water (δ 2 H and δ 18 O values) have been widely used to trace water in plants in a variety of physiological, ecohydrological, biogeochemical and hydrological studies. In such work, the analyte must first be extracted from samples, prior to isotopic analysis. While cryogenic vacuum distillation is currently the most widely used method reported in the literature, a variety of extraction‐collection‐analysis methods exist. A formal inter‐method comparison on plant tissues has yet to be carried out. Methods We performed an inter‐method comparison of six plant water extraction techniques: direct vapour equilibration, microwave extraction, two unique versions of cryogenic vacuum distillation, centrifugation, and high‐pressure mechanical squeezing. These methods were applied to four isotopically unique plant portions (head, stem, leaf, and root crown) of spring wheat ( Triticum aestivum L.). Extracted plant water was analyzed via spectrometric (OA‐ICOS) and mass‐based (IRMS) analysis systems when possible. Spring wheat was grown under controlled conditions with irrigation inputs of a known isotopic composition. Results The tested methods of extraction yielded markedly different isotopic signatures. Centrifugation, microwave extraction, direct vapour equilibration, and high‐pressure mechanical squeezing produced water more enriched in 2 H and 18 O content. Both cryogenic vacuum distillation systems and the high‐pressure mechanical squeezing method produced water more depleted in 2 H and 18 O content, depending upon the plant portion extracted. The various methods also produced differing concentrations of co‐extracted organic compounds, depending on the mode of extraction. Overall, the direct vapor equilibration method outperformed all other methods. Conclusions Despite its popularity, cryogenic vacuum distillation was outperformed by the direct vapor equilibration method in terms of limited co‐extraction of volatile organic compounds, rapid sample throughput, and near instantaneous returned stable isotope results. More research is now needed with other plant species, especially woody plants, to see how far the findings from this study could be extended.