FIGURE 4.

Reactivity of 2-MBT to NO in the absence of cells. (A) [NO] in the bioreactor in the absence of cells was measured. At t = 0, DPTA was added and after 10 min different concentrations of 2-MBT or the same volume of DMSO was added to the bioreactor. The solid lines represent the averages from at least 3 biological replicates, and the shaded regions represent the standard errors of the means. (B) 34 μM of PAPA or 34.7 μL of 10 mM NaOH were added to a bioreactor containing MOPS and 50 μM of 2-MBT or DMSO in the absence of bacteria. After 2 h, which correspond to the amount of time it took for NO to deplete under cell-free settings (Supplementary Figure S1), samples from the bioreactor were withdrawn for LC-MS analysis. Representative extracted-ion chromatograms (EIC) at m/z of 167.9 of 3 replicate experiments are plotted. Representative total-ion current chromatogram (TIC) is provided in Supplementary Figure S5 and no significant signal intensities for the 2-MBT dimer or other oxidized species were observed in the mass spectra. (C) Bacterial growth in the bioreactor containing MOPS and 2-MBT or DMSO that had or had not been exposed to NO was measured hourly. The solid dots represent the average OD₆₀₀ from 3 biological replicates, and the error bars the error of the means.