ABSTRACTAlgae Spirogyra have the potential to be developed as a raw material for bioethanol production. This study aims to determine the effect of the hydrolysis process of algae Spirogyra on total sugar production. The hydrolysis process of Spirogyra hyalina was carried out using a variation of heating duration and type of enzyme. This study also aimed to determine the effect of fermentation duration and the addition of nutritional fermentation on the reducing sugar, pH changes, microbial biomass and ethanol production from algae Spirogyra hyalina by using Zymomonas mobilis. The fermentation process was carried out in anaerobic conditions using a modified fermentor for 96 hours. Results showed the differences the hydrolysis process of algae Spirogyra hyalina affected on sugar levels. The highest sugar level was achieved by a combination of α-amylase and β-amylase enzymes. Fermentation duration and nutritional fermentation affected reducing sugar, pH substrate, microbial biomass and ethanol levels produced from the fermentation of algae Spirogyra by using Zymomonas mobilis.