Abstract. In conventional practice, starch is extracted and purified from a native source prior to glucose production using two steps: liquefaction and saccharification. Sequential protocols, liquefaction, and starch extraction and purification necessitate high-energy process conditions, material handling equipment, and other substantial cost inputs. In this work, native starch in ground fresh cassava root pulp and cassava flour was subjected to low-energy, simple single-step enzymatic hydrolysis at 37°C and 60°C for glucose production. Commercial purified cassava starch was similarly treated for comparison. Cassava root pulp reached a maximum of 30% starch conversion to glucose within 4 h of hydrolysis at 60°C and a maximum of 55% conversion within 96 h at 37°C. Cassava flour yielded mixed results, with slightly less conversion than root pulp at 60°C, reaching only 27%, but outperformed root pulp at 37°C, reaching 72% conversion. Purified cassava starch yielded 96% and 100% conversions at 60°C and 37°C, respectively. At 37°C, the reaction rate constants were 0.023 h-1, 0.021 h-1, and 0.025 h-1, respectively, for cassava root pulp, flour, and purified cassava starch, while at 60°C, the reaction rate constants were 0.816 h-1, 1.230 h-1, and 1.102 h-1, respectively. Activation energy for the substrates was similar at 134.6 kJ mol-1 for root pulp, 157.4 kJ mol-1 for flour, and 143.1 kJ mol-1 for purified starch. Keywords: Cassava root, Cassava starch, Enzymes, Glucose, Reaction kinetics, Synergistic enzymatic hydrolysis.