This study evaluates the effectiveness of iron-modified biochar (Fe-BC) in fixed-bed heterogeneous Fenton processes for levofloxacin (LFX) removal, a widely-used fluoroquinolone antibiotic. The objective is to optimize parameters such as pH, oxidants (H2O2 and S2O82−), and biochar forms (functionalized and raw) using factorial analysis of mixed data (FAMD) and response surface methodology (RSM). These optimizations identified the ideal conditions for maximal LFX removal. The most effective removal with Fe-BC occurred at 2.5 mM H2O2 and pH 7.5, while the optimal S2O82− conditions were 1.6 mM at pH 2.8. Both Fe-BC and raw biochar (RBC) showed the highest adsorption at pH 5.8. In adsorption-only, RBC and Fe-BC reduced LFX to 530 μg/L and 335 μg/L, respectively, in 60 min. The oxidation process further decreased LFX levels to between 8.9 μg/L and 0.1 μg/L using S2O82− and H2O2, respectively. The research expanded upon a kinetic model, incorporating the calculation of kinetic constants for both adsorption and oxidation processes, to deepen our understanding of the intricate degradation dynamics at play. Identifying by-products was crucial in elucidating degradation pathways. These findings are vital for environmental remediation, demonstrating the efficiency of Fe-BC in removing harmful antibiotics from water. This research highlights the potential of modified biochar in environmental clean-up, especially for water contaminated with antibiotics. The results emphasize the importance of optimizing treatment conditions for effective antibiotic removal, contributing valuable insights to the field of environmental remediation.