A method to determine the carbon isotope ratio of soil organic phosphorus

Despite the importance of soil organic phosphorus (OP) for plant nutrition, its dynamics in ecosystems remain elusive due to the lack of multiple stable P isotopes. Here, we developed a method that isolates the soil OP pool to measure its carbon isotope signature. To develop the method, we, first, tested three extractants (0.5 M H₂SO₄, HCl, and NaOH), which are commonly used for soil OP extraction, to evaluate their capacity to preferentially extract OP. Next, we isolated OP from the extract by coprecipitation and adsorption, and for this purpose, we evaluated different pH treatments (ranging from pH 1.5 to 10), and iron- or aluminum hydroxide additions. Our results show that H₂SO₄ extracted the largest amount of soil OP, and these extracts had the lowest organic carbon-to-organic phosphorus (OC:OP) ratio compared to HCl and NaOH extracts. The pH adjustments of the extracts to pH 4 – 7.5 removed ≥ 95% of the extracted OP from the solution. The molar OC:OP ratio of the precipitates was the lowest (11 – 16) at pH 7.5, showing a strong preferential OP removal due to the pH alteration of the extracts. Metal hydroxide addition (combined with pH treatment) did not further improve the preferential OP precipitation. Finally, we determined the carbon isotope ratio (δ¹³C) of the isolated OP pool. Overall, the method developed here provides a simple and effective approach to determine the carbon isotope ratio of the soil OP pool, which opens new avenues to study soil OP dynamics.