This study presents the synthesis, characterization, and utilization of marine macroalgae-derived bio-carbon catalysts (BC and KOH-AC) for the efficient conversion of waste cooking oil (WCO) into biodiesel. The biochar (BC) was produced through slow pyrolysis of macroalgal biomass, which was subsequently activated with potassium hydroxide (KOH) to produce a KOH-modified activated carbon (KOH-AC) catalyst. Advanced characterization techniques, including SEM, EDX, XRD, FTIR, and TGA, were used to examine the physicochemical characteristics of the catalysts. The synthesized catalysts were utilized to produce biodiesel from WCO, and the results revealed that the highest biodiesel yields, 98.96%, and 47.54%, were obtained using KOH-AC and BC catalysts, respectively, under optimal reaction conditions of 66 °C temperature, 12.3 M/O molar ratio, 130 min time, and 3.08 wt.% catalyst loading via RSM optimization. The kinetic and thermodynamic parameters, such as k, Ea, ΔH, ΔS, and ΔG, were determined to be 0.0346 min−1, 43.31 kJ mol−1, 38.98 kJ mol−1, − 158.38 J K−1 mol−1, and 92.58 kJ mol−1, respectively. The KOH-AC catalyst was recycled up to five times, with a significant biodiesel yield of 80.37%. The fuel properties of the biodiesel met ASTM (D6751) specifications, ensuring that it has excellent fuel characteristics and can be used as an alternative fuel.