This study presents the synthesis and characterization of undoped Fe3O4 and 3% Cu-doped Fe3O4 nanoparticles via a cost-effective and environmentally friendly co-precipitation method. A comprehensive suite of nanomaterial characterization techniques, including energy dispersive spectroscopy (EDS), scanning electron microscopy (SEM), X-ray diffraction (XRD), high-resolution transmission electron microscopy (HR-TEM), and X-ray photoelectron spectroscopy (XPS), was employed to confirm the elemental composition, structural parameters, and chemical states of the prepared materials. Additionally, fragmentation analysis using LC–MS illustrated the structural stability and breakdown of the nanoparticles under specific conditions. Both the materials were characterized by vibrating sample magnetometer (VSM), Brunauer-Emmett-Teller (BET) for investigating the magnetic, and surface characteristics respectively. Furthermore, the synthesized nanoparticles were evaluated for their photocatalytic degradation efficiency towards methylene blue and eriochrome black T dyes. A systematic investigation of various parameters, including the pH of the dye solution, dye concentration over time, catalyst dose, dye concentration versus degradation, catalyst reusability, and kinetic study, was conducted to understand the factors influencing the photocatalytic activity of both undoped and Cu-doped Fe3O4 nanoparticles. The LC-MS analysis predicted the probable degradation pathways of MB and EBT dyes. This work provides insights into the sustainable synthesis, comprehensive characterization, and practical application of Fe3O4-based nanoparticles for water treatment applications, highlighting their potential as efficient and environmentally benign Photocatalyst for wastewater remediation. An investigation was conducted to investigate the antimicrobial properties of both undoped and doped nanoparticles. The minimum inhibitory concentration (MIC) of the synthesized nanoparticles were determined against the bacterial strains E. coli, P. aeruginosa, S. aureus, and S. pyogenes, as well as the fungal strains C. albicans, A. niger, and A. clavatus.