In glycolysis, two molecules of ATP are consumed during two phosphorylation reactions to form fructose 1, 6−biphosphate. In return four molecules of ATP are produced by substrate level phosphorylation (conversion of 1, 3−biphosphoglycerate to 3−phosphoglycerate and phosphoenol pyruvate to pyruvate). Two molecules of
NADH2 are formed at the time of oxidation of glyceraldehyde 3−phosphate to 1, 3−biphosphoglycerate. The net reaction of glycolysis is as follows :
Glucose +2NAD++2ADP+2H3PO4→ 2 Pyruvate +2NADH+2H++2ATP
Each NADH is equivalent to 3ATP, so the net gain in glycolysis is 8ATP.