Humic substances (HS) and chromophoric dissolved organic matter (CDOM) are ubiquitous, widely impacting environmental processes, yet despite decades of study the link between structure and the unique optical properties evident in HS/CDOM remains elusive. Model compounds derived from a solely microbial source, as well as terrestrial sources from both aquatic environments and soil systems, exhibit many of the same optical properties despite their disparate methods of generation and sources. All show a pH dependent absorbance, exhibit increasing absorbance as wavelength decreases and a loss of absorbance upon borohydride reduction.

The link between colored humic substances is their ability to form electronic interactions that extend long wavelength absorbance. The underlying processes by which charge transfer bands or electronic interactions in HS/CDOM are generated are investigated by optical and potentiometic titrations of untreated and borohydride reduced material. Borohydride reduction targets carbonyl functional groups such as aromatic ketones and quinones. The reduction of these groups affects the optical properties by reducing long wavelength absorbance and causing a blue shift in the fluorescence emission spectra.

A direct comparison of divergent sources of fulvic and humic acids including an aquatic fulvic acid, Suwannee River Fulvic Acid (SRFA) and a microbial source of fulvic acid, Pony Lake Fulvic Acid (PLFA), the soil derived humic acids, Elliott (EHA) and Leonardite Humic Acids (LHA), an aquatic humic acid Suwannee River Humic Acid (SRHA) as well as Lignin Alkali Carboxylate (LAC) highlights differences between sources of humic material as exemplified by borohydride induced optical changes such as absorbance intensity in the UV and visible range, difference (DA), fractional difference, spectral slope (S), fluorescence excitation-emission matrix spectra (EEMS), and differential emission spectra (DF) as well as quantum yield.

Traditional Raman spectroscopy, although capable of providing relevant chemical functional group information, cannot be applied to untreated CDOM because of high fluorescence background. Surface enhanced Raman scattering (SERS) provides the capability of overcoming the florescence background, thus providing useful Raman spectral data. SERS spectra of model compounds and CDOM were collected using roughened silver electrodes. Functional groups were identified from selected borohydride reduced CDOM SERS spectra.