The formation of reactive oxygen species (ROS) from effluent organic matter (EfOM) was investigated under simulated solar irradiation. In this study, EfOM was isolated into three different fractions based on hydrophobicity. The productivity of ROS in EfOM was measured and compared with that of natural organic matter (NOM) isolates, including Suwannee River humic acid/fulvic acid (SRHA/FA) and Pony Lake fulvic acid (PLFA). The hydrophilic (HPI) component had a greater quantum yield of 1O2 than those of the hydrophobic (HPO) and transphilic (TPI) fractions because the HPI contained peptides and proteins. Regarding O2•-, the phenolic moieties acted as electron donating species after photochemical excitation and therefore electron transfer to oxygen. A positive correlation was found between the phenolic concentrations and the steady state O2•-concentrations. H2O2 accumulated during the irradiation process from superoxide as precursor. Potentially, due to the presence of proteins or other organic species in the HPI fraction, the decay rates of H2O2 in the dark for both the effluent wastewater and the HPI fraction were significantly faster than the rates observed in the standard NOM isolates, the HPO and TPI fractions. Autochthonous NOM showed a higher •OH productivity than terrestrial NOM. The [•OH]ss was lowest in the HPI fraction due to the lack of humic fraction and existence of soluble microbial products (SMPs), which easily reacted with •OH. Overall, the HPO and TPI fractions were the major sources of superoxide, H2O2 and •OH under simulated solar irradiation. The HPI fraction dominated the production of 1O2 and acted as a sink for H2O2 and •OH.