Is longer sonication worth the energy? Trade-offs between trimethoprim removal, toxicity, and biodegradability with initial assessment of impact on resistance genes.

Pharmaceutical contamination by antibiotics poses a major environmental challenge, particularly as regulators set limits for treated wastewater effluents. This study presents a hitherto unreported application of pulsed high-frequency sonication for degrading trimethoprim (TMP) and associated antimicrobial resistance genes (ARGs) in synthetic and reject wastewater. Optimization with synthetic binary samples revealed that sonication achieved >80% TMP removal at calorimetric-based energy per order (EEO) of 116.54 kW/m3, but mineralization remained low (∼10%). Prolonged treatment (5-6 h) fully eliminated TMP, yielding heavily hydroxylated and probably substituted products with short-chain alcohol groups, which exhibited lower biodegradability than demethylated/demethoxylated metabolites (e.g. m/z 245) dominant at 1-4 h. Notably, 30 min sonication produced hydroxylated TMP (m/z 307) with slight cytotoxicity to HeLa cells. The diaminopyrimidine (DAP) moiety of TMP, which drives its antimicrobial activity, remained largely intact in most metabolites identified in all the treatment times. This indicates that extending the treatment time would not provide additional benefits. Rather, sonication-based degradation should be tuned to promote site-specific attacks on antibiotic molecules, thereby inhibiting antimicrobial activity and minimizing the risk of ARGs development in treated water. Combining high-frequency sonication with low dose ozonation (via Venturi injection) effectively removed TMP (>98%) from low-solids reject wastewater, alongside substantial reductions in solids and COD. However, at higher solids levels, the process may induce cell lysis and release of TMP and ARGs from particulates. These findings position high-frequency ultrasound assisted ozonation as an effective treatment for wastewater streams with low solids, though careful control is needed to mitigate ARG mobilization in turbid matrices.