This study aims to evaluate the characteristics and feasibility of the autoclave assisted-alkali hydrogen peroxide delignification and enzymatic hydrolysis pretreated sugarcane bagasse for Mucor circinelloides cultivation to produce biofuels. The experimental setup consists of unpretreated sugarcane bagasse (SCB), delignified SCB, hydrolyzed SCB, and delignified-hydrolyzed SCB. The characterization was done using FTIR, XRD, and HHV calorimeter. The pretreatment of sugarcane bagasse using autoclave-assisted alkali hydrogen peroxide delignification was able to remove the lignin and hemicellulose. At the same time, the following enzymatic hydrolysis was able to increase the digestibility of sugarcane bagasse’s cellulose, making it suitable for Mucor circinelloides cultivation. The cultivation of Mucor circinelloides was done in the 100 ml sugarcane bagasse hydrolysate medium for 72 hours and a 250 rpm stirring rate produced approximately 9.1 grams of lipid.