Raman 18 O‐labeling of bacteria in visible and deep UV‐ranges

Raman stable isotope labeling with 2 H, 13 C or 15 N has been reported as an elegant approach to investigate cellular metabolic activity, which is of great importance to reveal the functions of microorganisms in native environments. A new strategy termed Raman 18 O‐labeling was developed to probe the metabolic activity of bacteria. Raman 18 O‐labeling refers to the combination of Raman microspectroscopy with 18 O‐labeling using H 2 18 O. At an excitation wavelength of 532 nm, the incorporation of 18 O into the amide I group of proteins and DNA/RNA bases was observed in Escherichia coli cells, while for an excitation wavelength electronically resonant with DNA or aromatic amino acid absorption at 244 nm 18 O assimilation was detected using chemometric tools rather than visual inspection. Raman 18 O‐labeling at 532 nm combined with 2D correlation analysis confirmed the assimilation of 18 O in proteins and nucleic acids and revealed the growth strategy of E. coli cells; they underwent protein synthesis followed by nucleic acid synthesis. Independent cultural replicates at different incubation times corroborated the reproducibility of these results. The variations in spectral features of 18 O‐labeled cells revealed changes in physiological information of cells. Hence, Raman 18 O‐labeling could provide a powerful tool to identify metabolically active bacterial cells.