In the present study we synthesized, characterized and checked the antimicrobial activity of fatty acid analogs of cholesterol. The synthesized compounds were characterized using IR, (1)H NMR, (13)C NMR and mass spectral data and tested for their antimicrobial activity by disk diffusion assay with slight modifications against Gram-positive, Gram-negative strains of bacteria as well as fungal strains. Minimum inhibitory concentration (MIC) of all the synthesized compounds was also determined. Compounds 7-14 showed inhibitory action against both the groups of bacteria and four strains of fungus. In vitro antimicrobial activity of the test compounds show that the compounds 10 and 13 are excellent antibacterial agents, where as compounds 13 and 14 are the excellent antifungal agents among the eight synthesized compounds.