Fluorescence of dissolved organic matter in natural waters

Fluorescence spectroscopy has recently found increasing use in investigations of dissolved organic matter (DOM) in terrestrial and marine aquatic environments since Coble et al. (1990; Nature 348, 432-435) applied the technique of excitation-emission-matrix (EEM) contouring to natural waters. Fluorescence of natural waters mainly originates from two groups of organic compounds: humic substance (humic type fluorescence, HTF), and unknown compounds which exhibit spectra similar to those of the aromatic amino acids tryptophan and tyrosine (protein type fluorescence, PTF). The aim of this study was to use fluorescence spectroscopy, including EEM spectroscopy, to investigate the distribution and spectral characteristics of fluorescent DOM in natural waters. Concentrations of dissolved organic carbon (DOC) were measured using an ultra-violet (UV)/persulphate oxidation system, and DOM was also investigated using UV/visible absorption spectroscopy. A number of field surveys were undertaken, in order to investigate DOM in rivers, estuaries, and the coastal zone. The Beaulieu, Test, and Itchen estuaries, in the Solent region of southern England, and the Tweed estuary, in Northumberland, were studied. These estuaries provided contrasting surrounding terrestrial environments and levels of anthropogenic influence. The Test and Itchen estuaries are subject to considerable inputs of sewage and industrial effluent or wastewater, but the Beaulieu and Tweed estuaries are relatively undisturbed. The rivers along the south coast of England were also studied in four seasonal surveys. Ultrafiltration, using an Amicon 3000 nominal molecular weight cutoff membrane, provided "truly dissolved" and "colloidal" fractions, which provided insight into the molecular weight distribution of HTF, DOC, and absorbance. The distribution and spectral characteristics of fluorescence in the Northeast Atlantic Ocean was also investigated.HTF exhibited two main peaks on EEM spectra, at excitation wavelengths of ca. 230 nm and 330-345 nm, each with peak emission wavelengths of ca. 420-445nm. These double peaks may represent the first and second absorption bands of the component HTF (mainly fulvic acid) fluorophors. The position of the peak emission wavelength with varying excitation wavelength showed similar characteristics over a wide range of sample types, which may have been a result of the overall similarity of the underlying humic substance fluorophores. Marine samples exhibited peak excitation and emission generally at shorter wavelengths than freshwater samples, possibly because of the smaller, less aromatic, fluorophors in marine humic substance. PTF in surface Atlantic Ocean, Adriatic Sea and Tweed estuary waters exhibited EEM peaks at ca. 230/320-340 nm and 275-280/320-340nm, at wavelengths similar to that of tryptophan bound within proteins. PTF in sewage effluent and water overlying the sediment in the Atlantic Ocean exhibited peaks at similar wavelengths to that of free tyrosine, at ca. 230, 275/300-310nm. In most of the rivers and estuaries in this study, free tryptophan and tyrosine do not represent a significant source of PTF, although they may be important contributors in regions affected by sewage discharge. Tryptophan bound to proteins or humic substance is probably the most widespread PTF fluorophor in natural waters. Indirect evidence for tryptophan providing much of the PTF in the Beaulieu estuary was provided by results from HPLC analyses.