The electrochemical two-electron oxygen reduction reaction (2e− ORR) offers a sustainable route for the on-site production of hydrogen peroxide (H2O2), yet developing cost-effective and high-performance catalysts remains highly challenging. In this study, porous biochar (BC) was first derived from corncob powder via KOH activation, followed by loading of uniformly dispersed cobalt nanoparticles through impregnation and carbothermal reduction methods to yield catalysts denoted as Co/BC-X (with X representing the Co : C molar ratio). The characterization of the catalysts by various analytical methods confirmed the formation of uniformly dispersed cobalt nanoparticles anchored on porous biochar with abundant oxygen-containing functional groups (OFGs), significantly enhancing the electrocatalytic production of H2O2. Density functional theory (DFT) calculations suggested the modulation of the electronic structure of the carbon atoms through synergy between Co nanoparticles and OFGs, optimizing the adsorption free energy of the OOH* intermediate closer to the theoretical optimal value and steering the ORR predominantly along the 2e− pathway. Among catalysts, the optimized Co/BC-3.3 catalyst achieved outstanding electrocatalytic performance toward H2O2 production with high H2O2 selectivity (∼96%), an electron transfer number of ∼2.06 at 0.45 V vs. RHE, a low Tafel slope of 72.42 mV dec−1 and excellent stability. In an H-type cell, the H2O2 production rate reached 1602 mmol gcat−1 h−1 using Co/BC-3.3, highlighting the effectiveness of the proposed strategy for sustainable and cost-effective conversion of biomass waste into high-performance electrocatalysts, along with providing fundamental insights into tuning the electronic structure of carbon-based materials for efficient on-site H2O2 electrosynthesis.