It is well described that the enterobacterium Escherichia coli produces acetate when growing on glucose. When glucose is exhausted, E. coli reorganizes its metabolism to consume this acetate. The resulting glucose/acetate transition (and hence, the switch from glycolysis to gluconeogenesis) is a natural model to study dynamic adaptation. Exponential growths on glucose and on acetate as the only substrate have been well described in the literature. However, experimental data and knowledge are scarce about the acetate metabolism during and after the growth on glucose. Here, 13C-metabolic flux analysis (13C-MFA) and quantitative metabolomics approaches combined with quantitative transcriptomics were used to unravel the physiological and molecular properties of the acetate metabolism during the different stages of the glucose/acetate transition. We first demonstrated that during the glucose consumption phase, the acetate accumulation resulted from both a constant flux of acetate production and an adjustable flux of acetate consumption. Second, the transition was proved to be a fast process, with the metabolite and transcriptional patterns being set up in minutes after glucose exhaustion. Third, acetate consumption is rapidly observed after glucose exhaustion but this metabolism is catabolic without biomass formation