Progression toward terminal oxidation of starch via hybrid organic–enzymatic bioanode for renewable carbohydrate conversion

Abstract

The performance of starch-based biofuel cells is fundamentally limited by inefficient utilization of the fuel's inherent energy density. Sequential oxidation of carbohydrate fuels, such as glucose, the monomeric unit of starch, using minimal catalyst systems has therefore emerged as a promising strategy to improve biofuel cell efficiency. Here, we extend a previously reported three-catalyst, organic–enzymatic glucose oxidation cascade to polymeric carbohydrate fuels by integrating enzymatic starch depolymerization with downstream electrochemical oxidation. To our knowledge, this represents the first demonstration of a defined hybrid organic–enzymatic cascade designed to drive progression of starch-derived glucose equivalents toward terminal oxidation using a minimal catalyst system. Colorimetric assays confirm that α-AMY and AMG efficiently hydrolyze soluble starch, achieving 92.3 ± 5.9% conversion of glucose equivalents within one hour at 60 °C. Subsequent electrochemical oxidation through a modified version of our established cascade employing 4-amino-TEMPO, 2-keto-3-deoxygluconate aldolase from Picrophilus torridus (ptKDGA), and oxalate decarboxylase from Bacillus subtilis (OxDC) produced current densities up to 4.21 ± 0.06 A m⁻², corresponding to 74% of the current density observed for glucose under identical conditions, with the increase in current density after starch hydrolysis found to be statistically significant (p < 0.001). Coulombic efficiency measurements yielded 36.1 ± 8.1% (n = 3), indicating progression beyond conventional partial oxidation pathways.