Long-term effects of zinc oxide nanoparticles on soluble microbial product composition and transformation in submerged membrane bioreactors

The increasing release of engineered nanoparticles into wastewater systems has raised concerns regarding their long-term effects on biological treatment processes and soluble microbial product (SMP) formation. However, the effects of zinc oxide nanoparticles (ZnO NPs) on the composition, diversity, and persistence of low-molecular-weight (low-MW) SMPs in membrane bioreactors (MBRs) remain poorly understood. In this study, the long-term effects of ZnO NP exposure on SMP characteristics were investigated in submerged anoxic-aerobic MBRs operated with and without ZnO NP addition. SMP production, extracellular polymeric substances (EPS), molecular weight distribution, fluorescence properties, and low-MW SMP identification were analyzed using HPLC-SEC, FEEM fluorescence spectroscopy, and GC-MS. ZnO NP exposure increased polysaccharide production in both loosely bound and tightly bound EPS fractions, indicating stress-induced enhancement of the extracellular protective matrix. High-MW SMP fractions and protein- and fulvic acid-like fluorescence intensities also increased under ZnO NP stress. GC-MS analysis revealed that ZnO NP exposure increased the number and diversity of detectable low-MW SMPs, with a shift toward aromatics, esters, and alcohols, while the relative abundance of alkanes and alkenes decreased. Several compounds were detected exclusively in the ZnO NP-exposed reactor, suggesting stress-induced metabolic transformation and altered biodegradation pathways. Although many SMPs were partially or fully removed during treatment, some recalcitrant compounds persisted in the effluent, including long-chain hydrocarbons and aromatic derivatives. The results demonstrate that ZnO NP exposure substantially alters SMP composition and promotes the formation of more diverse and persistent low-MW organic compounds in MBR systems. These findings provide new insights into nanoparticle-induced stress responses and their implications for SMP dynamics, effluent quality, and membrane fouling in biological wastewater treatment systems.