Improvement of Electrospray Ionization Response Linearity and Quantification in Dissolved Organic Matter using Synthetic Deuterated Internal Standards

Aquatic dissolved organic matter (DOM) is an ultra-complex mixture of compounds covering a wide range of masses and with an unknown extent of isomeric complexity, making its structural elucidation and quantification highly challenging. Electrospray ionization high-resolution mass spectrometry (ESI-HRMS) has advanced DOM analysis, but accurate concentration determination remains limited by the lack of response factor correction. Here, we address this limitation by introducing novel deuterated compounds as internal standards that mimic DOM structures. Using a d5-labeled compound free of isobaric interferences in DOM, we assessed ionization suppression in various aquatic sample extracts, and improved concentration-based linearity in a coastal DOM reference material. Our results show that deuterated carboxylic acid-rich standards enable ‘pseudo-quantification’ by correcting for ionization suppression and instrument drift. Applying this approach, we estimate that ionizable acids constitute 15-30% of DOM in river, coastal, and deep-ocean samples using an Orbitrap system and 11-24% using a 15T FT-ICR-MS, highlighting platform differences. Additionally, we establish a ~1 ng L⁻¹ feature detection limit for DOM compounds in seawater via a standard LC-MS gradient method. These findings demonstrate that deuterated standards provide a simple, practical way to improve DOM semi-quantification, enhancing our understanding of its chemical composition in environmental studies.