Biochemical mechanism of combined effect of ethambutol and sparfloxacin against Mycobacterium smegmatis.

M. smegmatis cells grown in the presence of combination of ethambutol (EMB) and sparfloxacin (SPX) had decreased level of total cellular lipids as compared to control as well as cells grown in the presence of sub-inhibitory concentration (MIC50) of individual drugs. Amongst various phospholipids analyzed, maximum decrease was observed in the content of phosphatidylinositolmannosides (PIMs) of the cells grown in combination of EMB and SPX. In contrast, the subcellular distribution of phospholipids revealed a significant increase in PIMs content of both cell wall and cell membrane of the cells grown in the presence of combination of drugs as compared to control as well as individual drugs. Mycolic acids of M. smegmatis cells were found to be main targets as combination of drugs resulted in significant decrease in total cellular as well as cell wall mycolic acids as compared to control and individual drugs. Changed lipid composition of M. smegmatis cells grown in the presence of MIC50 of EMB, SPX and combination resulted in significant surface changes as was evident from decreased limiting fluorescence (Fmax) intensity of 1-anilinonaphthalene-8-sulfonate (ANS). Thus, the results of this study suggested that ethambutol and sparfloxacin in combination exerted their antimycobacterial effect principally due to their action on phosphatidylinositolmannosides (PIMs) and mycolic acids, which form the permeability barrier of mycobacteria.