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The ability to accurately determine intracellular concentrations of metabolites is of key importance in studying signaling and metabolic reaction networks and their regulation in vivo. In the last few years, much time has been invested in studying the intracellular metabolites concentration profiles of E. coli in the stationary phase. However, there are few studies related to the intracellular metabolites concentration profiles of E. coli in the exponential phase. In this study, metabolic regulation in E. coli was studied in terms of the change in intracellular metabolites (e.g., glucose-1-phosphate, fructose-1,6-bisphosphate, phosphoenolpyruvate, pyruvate, acetyl-coenzyme A, 6-phosphogluconate, ribulose-5-phosphate, xylulose-5-phosphate, erythrose-4-phosphate) concentration at two different growth phases, in batch
culture. Samples were taken every 2 h after batch cultivation started. Culture sample vials (5 mL) were plunged rapidly into an equal volume of 60% aqueous methanol (-40°C). The quenched biomass was centrifuged for 10 min at 3000×g and -9°C. An extraction procedure using 100% methanol (-48°C) was performed to extract intracellular metabolites from E. coli K12 strain W3110. All measurement were carried out on an HPLC system coupled with an ion trap mass pectrometer equipped with a turbo-ion spray source.