The One-Sample PARAFAC Approach Reveals Molecular Size Distributions of Fluorescent Components in Dissolved Organic Matter

Molecular size plays an important role in dissolved organic matter (DOM) biogeochemistry, but its relationship with the fluorescent fraction of DOM (FDOM) remains poorly resolved. Here high-performance size exclusion chromatography (HPSEC) was coupled to fluorescence emission-excitation (EEM) spectroscopy in full spectral (60 emission and 34 excitation wavelengths) and chromatographic resolution (<1 Hz), to enable the mathematical decomposition of fluorescence on an individual sample basis by parallel factor analysis (PARAFAC). The approach allowed cross-system comparisons of molecular size distributions for individual fluorescence components obtained from independent data sets. Spectra extracted from allochthonous DOM were highly similar. Allochthonous and autochthonous DOM shared some spectra, but included unique components. In agreement with the supramolecular assembly hypothesis, molecular size distributions of the fluorescence fractions were broad and chromatographically unresolved, possibly representing reoccurring fluorophores forming noncovalently bound assemblies of varying molecular size. Samples shared underlying fluorescence components that differed in their size distributions but not their spectral properties. Thus, in contrast to absorption measurements, bulk fluorescence is unlikely to reliably indicate the average molecular size of DOM. The one-sample approach enables robust and independent cross-site comparisons without large-scale sampling efforts and introduces new analytical opportunities for elucidating the origins and biogeochemical properties of FDOM.