Applications of fluorescence spectroscopy to environmental chemistry

The work presented in this thesis consists of three parts. The first is a photophysicalstudy of the mechanism of fluorescence quenching by stable nitroxyl radicals, whichare becoming an important analytical tool for the study of reactive transients insurface waters (1, 2). In part two, quenching of dissolved organic matter (DOM)fluorescence by nitroxides is employed to investigate the electrostatic propertie~ ofDOM in aqueous solution, with the goal of elucidating the apparent ionic strength andpH dependence of metal-fulvic acid binding constants. In part three, the intrinsicoptical properties (absorbance, fluorescence, and fluorescence efficiency) of DOM areexamined in a coastal region to understand how these properties vary with source. ageand sunlight exposure time.Nitroxide-fluorophore adducts were employed to investigate the mechanism by whichnitroxyl radicals quench fluorescence (3). Fluorescence quantum yields and lifetimeswere measured for a series of adducts, and quenching rates were shown to be quite high(kq≈108-1010 s-1), even at distances of ≈12 Å. Forster or Dexter energy transfermechanisms are unable to account for the observed rates and lack of solventdependence in quenching. An excellent correlation is observed between kq and thenon-radiative relaxation rate. These results confirm that nitroxyl radicals are verynon-selective in their quenching abilities, and suggest that the best analytical probeadducts will include a fluorophore with an appreciable non-radiative relaxation rate.Diffusional quenching by charged and neutral nitroxides was employed to explore theelectrostatic properties of fulvic (FA) and humic (HA) acids. Cationic nitroxides werefound to be up to 16 times more effective than neutral analogues in quenching thefluorescence of humic materials. This result is attributed to the enhanced coulombicattraction of cations to the anionic FA or HA surface, and is interpreted as an estimateof surface electrostatic potential. Reduction of molecular charge at low pH andshielding of charge at high ionic strength (I) produced diminished enhancements.consistent with this interpretation. The potential was found to be particularly sensitive to ionic strength. suggesting that this electrostatic effect should be ofparticular importance in transition zones. such as estuaries, where I increases from<5 mM to 0.7 M as river water and seawater mix. High molecular weight fractions ofHA have a higher apparent surface potential than lower molecular weight fractions.indicating that larger humic molecules may have an enhanced ability to bind metalions.Optical properties of colored DOM may vary with source and age of the matertal.Absorption spectra can be characterized by their log-linearized slopes (S) as well as bytheir absolute intensities. The slope, S, is found to be much greater (steeper decrease inabsorbance with increasing wavelength) for blue-water samples than for rtverine andcoastal samples, indicating that the visible-light absorbing fraction of DOM may bepreferentially removed from surface waters. Fluorescence quantum yields were verysimilar for a wide variety of samples, but do show some minor differences; changes inquantum yield with excitation wavelength within a single sample are an indication ofthe heterogeneity of the chromophore mixture present in these materials. To bettercompare fluorescence data, complete excitation/ emission matrix spectra werecollected. When normalized to their respective absorbance spectra, these provide a full'map' of fluorescence quantum efficiency over the entire uv-visible range. Thistechnique is showing promise as a way to identify important spectral regions in thesecomplex chromophore mixtures. DOM isolated on C-18 columns had somewhatdifferent optical charateristics than whole water samples, suggesting selectiveisolation of absorbing material.

Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at the Massachusetts Institute of Technology and the Woods Hole Oceanographic Institution February 1992