Gram-negative bacteria such as Escherichia coli respond to decreased levels of carbon by entering a stationary phase associated with increased protein degradation and decreased synthesis. Although mass spectrometry approaches have identified changes in a number of metabolites during starvation, this process requires constant extraction of metabolites and MS analysis, which can be technically demanding. Link et al. developed a method that allows the real-time monitoring of 300 compounds in live cells ranging from bacteria to mammalian cells at 15- to 30-second intervals. A cultivation vessel was attached to a time-of-flight (TOF) mass spectrometer, which ensures the constant cycling and sampling of cells through direct injection into the spectrometer. This approach produces findings comparable to those of an established manual metabolomics approach but has much higher temporal resolution. The authors observed metabolic dynamics in E. coli at different time points during the two hours of starvation and 30 minutes after adding glucose. They focused on two clusters of metabolites whose levels changed during the analysis: a group of metabolically costly molecules such as valine and tryptophan that accumulated during starvation and decreased after glucose addition and a second group of energetically cheaper compounds such as succinate that were scarce during starvation and more abundant under fed conditions. The authors proposed that the different dynamics in these two groups were due to changes in amino acid degradation. Mathematical modeling using the profiling data coupled with protein synthesis inhibitor experiments revealed that these changes were due to feedback inhibition of the biosynthesis of the costly amino acids resulting in the utilization of the cheaper amino acids for protein synthesis. Overall, the utility of this method to provide real-time analysis of the metabolic status of a cell may reveal new potential regulatory mechanisms under different environmental conditions.