Preservation methods for the isotopic composition of dissolved carbon species in non-ideal conditions

Rationale: The stable carbon isotope compositions of dissolved inorganic carbon (δ¹³C_DIC) and dissolved organic carbon (δ¹³C_DOC) are readily affected by post-sampling microbial activity if not adequately preserved. Existing preservation methods require rapid chilling, analysis, and/or toxic chemicals, all challenging to use safely in the field and during remote field seasons. Therefore, a preservation method that is safe but also effective over a range of storage times is needed when sampling waters at very remote sites. Methods: Two samples, with different dissolved inorganic carbon (DIC) and dissolved organic carbon (DOC) concentrations, were filtered with a 0.2-μm filter and preserved with six different methods, mercuric chloride, copper sulfate, phosphoric acid, benzalkonium chloride, zinc chloride, hydrochloric acid, and a filter-only control. These samples were held at 4°C, 22°C, or 35°C. Regular measurement of the DIC and DOC δ¹³C values were made over the following 60 days for δ¹³C_DIC and 66 days for δ¹³C_DOC. Results: Over the course of the experiment, mercuric chloride, copper sulfate, zinc chloride, and benzalkonium chloride resulted in δ¹³C_DIC fractionation at both 4°C and 22°C. Only filtering to 0.2 μm at the time of collection, with or without acidification with phosphoric acid, resulted in minimal isotopic fractionation at both 4°C and 22°C and over the entirety of the experiment. For δ¹³C_DOC values, only filtering to 0.2 μm minimized fractionation for both bulk and vial storage over 66 days at 22°C. Conclusions: Filtering to 0.2 μm at the time of collection is more effective than the use of toxic chemicals for measuring δ¹³C_DIC and δ¹³C_DOC values. Phosphoric acid is as effective as only filtering for δ¹³C_DIC and may be ideal depending on sampling considerations. These results demonstrate not only that water samples can be preserved for δ¹³C_DIC and δ¹³C_DOC analysis for long periods, but that preservation is best accomplished with non-toxic or low-toxicity methods.