Abstract: Antibacterial agents are the key drugs for the treatment of bacterial infections and one of the most important foundation drugs in modern medicine.However,the current problems of increasingly serious antimicrobial resistance and insufficient channels for new antimicrobial agent discovery have attracted attention and need to be solved urgently.Antibacterial activity is acted when antibacterial agents permeate across the bacterial outer membrane and accumulate to effective concentration at the target.Therefore,measuring the cellular accumulation of antibacterial agents in bacteria would be helpful to establish the relationships between properties of the intracellular accumulation of compounds and their structures.The guidance for the design of new antibacterial agents could be provided through these relationships.At present,a variety of detection methods have been developed for the determination of antimicrobial agent concentration in bacteria.This paper briefly introduced the structural differences of cell membranes between Gram-positive bacteria and Gram-negative bacteria.Subsequently,radiometric assay,fluorometric assay and new methods based on mass spectrometry,which included LC-MS/MS,SPE-MS,nanofluidics-MS,TOF-SIMS and biological probes were reviewed.Each detection method and their current applications were presented.The advantages and disadvantages of the above analysis methods were summarized,and prospects were made for future development,with the aim of providing suggestions for the application of the assays of antimicrobial agent accumulation in bacteria,and reference for the design and development of new antibacterial agents,especially new drugs against Gram-negative bacteria.

Key words: antimicrobial agents; antimicrobial uptake; detection methods; intracellular accumulation; subcellular localization