Understanding antimicrobial activity in live cells

The critical rise of antimicrobial resistance (AMR) requires the discovery of new antibiotics, particularly against Gram-negative bacteria. The permeability barrier of bacteria is a vital property to consider within the drug development process. In this work, we describe a new NMR method to assess membrane permeability and membrane interaction affinity of a new family of antimicrobial compounds exploiting the enhanced paramagnetic relaxation enhancement (PRE) effect induced by solvent paramagnetic agents in non-permeable drugs mixed with lipid vesicles. The results exhibited differences in the permeability rate and membrane interaction affinity of drugs among membranes with different lipid composition, confirming that our method is applicable to any type of cell/bacterial membrane. In addition, we have expanded this assay to live cells making use of a NMR-coupled bioreactor. With this setup, we have been able to measure NMR metabolomics to simultaneously monitor membrane permeation, membrane affinity and changes in the bacterial metabolism as a result of the addition of antimicrobial agents, providing evidence on their mechanism of action at the metabolite level. In general, our study introduces a new method for assessing drug permeability in lipid vesicles or live cells that will allow the design of more efficacious drugs and antimicrobial agents in a standard chemistry lab environment.