Conversion of unstable compounds can contribute to the auxin pool during sample preparation

Auxin is a powerful regulator of plant growth and development (Heisler and Byrne, 2020). Accurate quantification of the main form of auxin, indole-3-acetic acid (IAA), is therefore essential for the study of auxin biology. Synthetic forms of the IAA-related compounds indole-3-pyruvic acid (IPyA) and indole-3-acetaldehyde (IAAld) have been shown to convert nonenzymatically to IAA, particularly in aqueous environments (Ernstsen et al., 1986; Tam and Normanly, 1998; Quittenden et al., 2009; Dai et al., 2013). These compounds are present in plants, and IPyA is now considered the main intermediate between tryptophan and IAA (Zhao, 2018). If substantial amounts of IPyA and/or IAAld convert to IAA during sample preparation, IAA quantification may be severely compromised. Here, a cysteamine-based derivatization method for converting IPyA and IAAld to stable forms (Novák et al., 2012) was used to demonstrate that endogenous IAA-related compounds can convert nonenzymatically to IAA during pea (Pisum sativum) and Arabidopsis (Arabidopsis thaliana) sample preparation. In derivatized samples, quantified IAA content was up to 40% lower than in underivatized controls. As a result, we recommend that harvested samples be derivatized soon after the completion of a short extraction period.