Morbidity and mortality induced by bacterial infection are significant health problems. To take over this challenge, new antibacterial agent discovery is a must. In this work, 11H-indeno[1,2-b]quinoxaline-11-one was chemically synthesized, and its biological activities were evaluated against Salmonella typhimurium (S. typhi), Klebsiella pneumoniae (K. pneumoniae), and Bacillus subtilis (B. subtilis). The biological activity of the compound was studied by a well diffusion method besides minimum inhibitory concentration (MIC), and resistant development evaluation. Optimization of the geometric structure was performed using DFT calculation and a comparison between calculated and experimental geometric structures and spectral data using the function BLY3P/6-311++G(d,p). MIC values against S. typhi, K. pneumoniae, and B. subtilis were recorded as values of 10, 20, and 20 mM respectively. Within seven days, none of these bacteria have no resistance against the tested compound at 1, 2, 3, and 4x MIC concentrations. The comparison between optimized structures geometry and spectral data (1HNMR and 13CNMR) with experimental data displayed acceptable agreement. Eventually, 11H-indeno[1,2-b]quinoxaline-11-one could be recommended as an effective antibacterial agent.