The Neuse River, North Carolina, U.S., exemplifies a typical de facto potable reuse scenario, where drinking water sources are located downstream of treated wastewater effluent discharges. The study results imply that planned potable water reuse, whether in an indirect or direct potable reuse scenario, might provide better control over water quality than the status quo conditions. Using fluorescence excitation-emission matrix (EEM) measurements, anthropogenic influence of a wastewater treatment plant (WWTP) discharge was observed in samples near the location of drinking water treatment plant (WTP) intakes, eight or more miles downstream of the WWTP, implying that anthropogenic compounds were not fully removed or degraded by natural processes in this reach of the river. PARAllel FACtor (PARAFAC) analysis supported a two-component model of humic-like and nonhumic-like dissolved organic matter (DOM). A nonmodeled anthropogenic feature was also indicated. Significantly, the quenched fluorescence of humic-like DOM (static and/or dynamic quenching) by nonhumic-like DOM-previously demonstrated for probe molecules but first reported here in a natural/anthropogenic-influenced system-offers exciting insight into studies of humic/nonhumic interactions with important implications for pollutant fate and transport, sensing applications, and water treatment. A molecular spectroscopic explanation for dual fluorescing peaks in amino acids and humic substances is postulated.