What do acid-base titrations of live bacteria tell us? A preliminary assessment

To gain insight into the non-equilibrium processes that affect the titration curves of bacteria, we performed pH stat experiments with suspensions of live cells of the Gram-negative bacterium Shewanella putrefaciens. The experiments lasted for 5 hours, during which acid or base addition was monitored. Periodically, the electrophoretic mobility of the cells, as well as the buffer capacity and the concentrations of cations and dissolved organic carbon (DOC) of the solution, were measured. At the end of the experiments, the viability of the cells was determined. In a limited number of cases, final solutions were screened for the presence of cell-wall constituents using gel electrophoresis. The results showed a very different behavior of the cell suspensions under acid and alkaline conditions. At pH 4, acid addition ceased after 20 minutes. The cells remained intact but were no longer viable at the end of the experiment, while little change in the buffer capacity of the solution was observed. The data at pH 4 were consistent with protonation of cell wall functional groups. At pH 8 and 10, base addition continued during the entire duration of the experiments. The cells remained viable, and the buffer capacity and DOC concentration of the solutions increased with time. Gel electrophoresis indicated that proteins and lipopolysaccharides had been released to solution at pH 10. In contrast to pH 4, the buffering capacity of the bacterial cells under alkaline conditions did not appear to be limited by the initial availability of ionisable functional groups in the cell wall. This preliminary study shows that a complex set of processes, including active metabolic responses, control the acid-base activity of live cell suspensions.