Strain, medium, and chemicals
All experiments were performed with an industrial yeast strain S. cerevisiae 4126. Yeast extract peptone dextrose (YPD) medium composed of 5 g/L yeast extract powder, 3 g/L peptone (Sangon Biotech, Shanghai) and different levels of glucose (30, 120, 280 g/L) was used. YPD medium containing 30 g/L and 120 g/L glucose for pre-cultures in flasks and fermenters was sterilized by the usual method (121°C, 20 min), while the YPD medium composed of 280 g/L glucose for continuous ethanol fermentation was sterilized at 110°C for 15 min to minimize the Maillard reaction between amino acids and reducing sugars. YPD medium containing of 120 g/L and 280 g/L glucose was perceived as low gravity medium and very high gravity (VHG) medium, respectively. The following metabolites: GLC, G6P, F6P, FBP, 6PG, R5P, T6P, TRE, GAP, GOLP, 2+3PG, PEP, PYR, OAA, CIT, AKG, SUC, FUM, MAL, ATP, ADP, AMP, NAD+, NADH, NADP+, NADPH, AcCoA (refer to the list of abbreviations) used as standards for LC-MS/MS analysis were purchased from Sigma-Aldrich.