Substrate concentration: A more serious consideration than the amount of 5-hydroxymethylfurfural in acid-catalyzed hydrolysis during bioethanol production from starch biomass

5-hydroxymethylfurfural (5-HMF) yield during bioethanol production from starch was determined using spectrophotometry and chromatography. Increasing acid concentration and time favored 5-HMF production with HCl while yield decreased after 45-minute hydrolysis time for HNO3 and H2SO4 hydrolyzed samples. Impacts of glucose (substrate) concentration and produced 5-HMF on bioethanol yield were studied with different sulphuric acid concentrations and different α-amylase and amyloglucosidase activities. A central composite rotational design was utilized to determine the conditions of hydrolysis for optimum glucose production. The results showed that maximum glucose yield occurred at 0.5 M acid concentration and 45-minute hydrolysis time, while maximum yield was achieved at 120 and 280 units of α-amylase and amyloglucosidase activities respectively. It was shown that 5-HMF did not exhibit much inhibition on ethanol yield at low acid concentrations but became pronounced at higher acid concentrations, while high glucose concentrations had a pronounced negative effect on ethanol yield and fermentation efficiency.