Extreme isomeric complexity of dissolved organic matter found across aquatic environments

Abstract

The natural aquatic environment contains an enormous pool of dissolved reduced carbon, present as ultracomplex mixtures that are constituted by an unknown number of compounds at vanishingly small concentrations. We attempted to separate individual structural isomers from several samples using online reversedphase chromatography with selected ion monitoring/tandem mass spectrometry, but found that isomeric complexity still presented a boundary to investigation even after chromatographic simplification of the samples. However, it was possible to determine that the structural complexity differed among samples. Our results also suggest that extreme structural complexity was a ubiquitous feature of dissolved organic matter (DOM) in all aquatic systems, meaning that this diversity may play similar roles for recalcitrance and degradation of DOM in all tested environments.Dissolved organic matter (DOM) is the dominant form of organic carbon in most aquatic environments. Upon mineralization, it is an important precursor of outgassing of CO 2 from inland waters (Tranvik et al. 2009), it carries substantial amounts of nutrients and energy from land to sea (Medeiros et al. 2016), and it persists in the deep ocean for millennia (Dittmar and Stubbins 2014). It is an ultra-complex mixture of compounds that provides the ultimate test of the capabilities of analytical chemistry (Rodgers et al. 2005;Dittmar and Stubbins 2014), and investigations into its nature have been confounded by its extreme molecular complexity. High-resolution mass spectrometry (HRMS) is able to resolve many thousands of molecular masses from complex natural mixtures of organic compounds (Marshall et al. 1998;Riedel and Dittmar 2014;Hendrickson et al. 2015), but is unable to differentiate between structural isomers of a molecular formula.Most recent research has utilized advanced visualization or multivariate statistical approaches to interpret HRMS data (Wu et al. 2004;Sleighter et al. 2010;Kellerman et al. 2015), but when the HRMS data are not combined with more This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. Scientific Significance StatementMolecular complexity is an inherent feature of dissolved organic matter (DOM), confounding investigations into its nature. Recent research has suggested that this complexity may explain the persistence of DOM due to the implied low abundance of individual compounds. Here, we use chromatographic separation and collision induced dissociation of deprotonated molecules to demonstrate the extreme isomeric complexity of individual molecular formulas in DOM and show that isomeric complexity occurs across diverse aquatic environments. 21Limnology and Oceanography Letters 3, 2018, 21-30

Mathematical chromatography deciphers the molecular fingerprints of dissolved organic matter

“…In the following, we use the term chromatographic feature to mean a nonresolved peak, and reserve the word peak for true analyte peaks (which are rare in DOM) or resolved mass spectrometry peaks, which usually contain information from numerous isomers with the same formula. 9,13 The use of the term elution profile or chromatographic feature of components or formulas does not imply single analyte peaks, but always refers to complex mixtures unless specifically stated otherwise. All data processing and modelling was carried out using PLS_Toolbox (v8.61, Eigenvector Research Inc.) in MATLAB (v9.7, MathWorks Inc.).…”

“…From the molecular perspective, the dominant molecular formulas were usually present in several components (Fig. S12 †), highlighting the isomeric diversity hidden behind each formula, 13,32 and revealing for the first time that isomers can be grouped by behaviour across environments.…”

“…After fractionation, the complexity of each obtained mass spectrum decreases and the interpretation may be simplified, although the smearing of isomeric mixtures throughout the elution demonstrates that the mass spectral peaks are not isomerically pure. [10][11][12][13] While manual fraction collection is labour-intensive, the online coupling of HPLC and HRMS fully automates the measurement procedures. 14,15 Compared to direct infusion HRMS, the formulaic complexity of HPLC-HRMS datasets increases by orders of magnitude.…”

“…Since chromatograms of DOM represent unresolved humps, the abundance of single analytes cannot easily be deduced. 13,18 Instead, complex mass spectra in broadly eluting fractions must be analysed. However, the visual analysis of raw data is overburdened by the large number of spectra, and statistical analyses capable of extracting information from direct infusion datasets (such as principal component analysis) require unfolding of the multidimensional data.…”

Molecular Level Analysis Reveals Changes in Chemical Composition of Dissolved Organic Matter From South Texas Rivers After High Flow Events

“…Although no direct measurement was conducted, a recent study suggested that riverine DOM after storm events could potentially have high structural diversity (Wagner et al, 2019), based on modeling results (Zark et al, 2017;Hawkes et al, 2018;Zark and Dittmar, 2018). Based on the central limit theorem, these modeling studies assumed a random combination of atoms in DOM molecules and suggested that each formula assigned in ultrahigh-resolution mass spectrometry potentially represents 10-100 different isomers (Zark et al, 2017;Hawkes et al, 2018). Our results, however, suggested an opposite pattern.…”