Fluorescence Excitation−Emission Matrix Regional Integration to Quantify Spectra for Dissolved Organic Matter

TLDR

Excitation–emission matrix fluorescence spectroscopy is widely used to characterize dissolved organic matter in water and soil, but interpreting the >10,000 wavelength‑dependent data points remains challenging. The authors developed fluorescence regional integration, which partitions EEMs into five excitation–emission regions, integrates the volume beneath each region, normalizes it to the region’s area and dissolved organic carbon concentration, and applies this method to standard fulvic acid and fifteen DOM fractions from wastewater and rivers, complemented by 13C NMR, FTIR, and UV–Vis analyses. They found that DOM fractions fluoresced in one or more regions, with hydrophobic neutral fractions showing the highest cumulative EEM volume, and that aromatic carbon signatures in wastewater biomass DOM, revealed by NMR and FTIR, enabled EEMs to distinguish wastewater effluent from drinking water DOM.

Abstract

Excitation−emission matrix (EEM) fluorescence spectroscopy has been widely used to characterize dissolved organic matter (DOM) in water and soil. However, interpreting the >10,000 wavelength-dependent fluorescence intensity data points represented in EEMs has posed a significant challenge. Fluorescence regional integration, a quantitative technique that integrates the volume beneath an EEM, was developed to analyze EEMs. EEMs were delineated into five excitation−emission regions based on fluorescence of model compounds, DOM fractions, and marine waters or freshwaters. Volumetric integration under the EEM within each region, normalized to the projected excitation−emission area within that region and dissolved organic carbon concentration, resulted in a normalized region-specific EEM volume (Φi,n). Solid-state carbon nuclear magnetic resonance (13C NMR), Fourier transform infrared (FTIR) analysis, ultraviolet−visible absorption spectra, and EEMs were obtained for standard Suwannee River fulvic acid and 15 hydrophobic or hydrophilic acid, neutral, and base DOM fractions plus nonfractionated DOM from wastewater effluents and rivers in the southwestern United States. DOM fractions fluoresced in one or more EEM regions. The highest cumulative EEM volume (ΦT,n = ΣΦi,n) was observed for hydrophobic neutral DOM fractions, followed by lower ΦT,n values for hydrophobic acid, base, and hydrophilic acid DOM fractions, respectively. An extracted wastewater biomass DOM sample contained aromatic protein- and humic-like material and was characteristic of bacterial-soluble microbial products. Aromatic carbon and the presence of specific aromatic compounds (as indicated by solid-state 13C NMR and FTIR data) resulted in EEMs that aided in differentiating wastewater effluent DOM from drinking water DOM.