In this research study, orange peel-based biocatalysts developed from different acid protonation were used as a metal-free catalyst for hydrogen production from sodium borohydride (NaBH4). In order to prepare the orange peel-based biocatalyst with higher catalytic activity, experiments were conducted with pure orange peel, different acid molar concentrations, and calcination temperatures. The physical morphology, surface texture, and chemical interaction were thoroughly analyzed by XRD, FTIR Raman, FESEM, BET, and TGA. As a result of the experiment, it was determined that the highly acid-treated biocatalyst (40% H3PO4, 40% H2SO4, 40% HCl) and calcinated at 450 °C for 1 h had higher catalytic activity. As a result, bio-hydrogen production at 35 °C and 70 °C methanolysis with 3% NaBH4 catalyzed by a mixture of acid-treated catalysts were found as 46,213 and 63,842 ml min−1g.cat−1, respectively. However, with the increase of molar concentration of biocatalyst with 40% individual acid prolonged samples, the HGR rates will not have a satisfactory value in comparison with the 40% mixture of the acid-treated catalyst due to less number of active sites. © 2023 Hydrogen Energy Publications LLC