Separation and Quantification of Murein and Precursors from Enterobacter cloacae after Treatment with Trimethoprim and Sulphadiazine

Abstract

Abstract The intracellular concentrations of the soluble murein precursors UDP-Mur-NAc-pentapeptide in the cytoplasm, the membrane-bound lipid precursor disaccharide pentapeptide and the muropeptides of Enterobacter cloacae cultures treated with trimethoprim (12·5μg mL−1) and sulphadiazine (250 μg mL−1) were determined by using capillary zone electrophoresis analysis. In the presence of trimethoprim, UDP-Mur-NAc-pentapeptide as well as disaccharide pentapeptide accumulated. In the case of sulphadiazine-treated cells, the concentration of UDP-Mur-NAc-pentapeptide roughly paralleled the control cells but sulphadiazine caused a slow incremental accumulation of disaccharide pentapeptide. The muropeptide composition of the murein indicated that the differences between the peptidoglycans produced by the control cells and the cells grown in the presence of either trimethoprim or sulphadiazine alone or in combination were quite marked. The results suggest that the enhanced activity of trimethoprim plus sulphadiazine against E. cloacae is caused by an additional effect on the inhibition of the bacterial peptidoglycan biosynthesis and that this additional effect is a fundamental part of the antibacterial action of the antimetabolites. This effect leads to changes of cell morphology and resultant changes in bacterial cell permeability.