© 2022 Elsevier B.V.Tetracyclic molecules show important properties such as biological activities and fluorescence sensors. Hence, in this study, we have reported unknown tetracyclic skeleton. N-propargylated C-2 and C-4 disubstituted imidazole derivatives were reacted by 2-aminomethyl piperidine without using any transition metal. The reaction unveiled 16 different imidazo[1,2-a]pyrido[1′,2′:3,4]imidazo[2,1-c]pyrazine skeletons. Cyclization reaction tolerated phenyl, naphthyl, bi-phenyl, 2-thienyl, and many substituents on the benzene ring such as OMe, CF3, and halogens. We have uncovered the exact structure of the tetracyclic skeleton using single-crystal X-ray analysis. Significant fluorescence emission of tetracyclic molecules was investigated, and derivatives bearing electron-withdrawing groups, CF3, and the 2-thienyl group possessed a bathochromic effect (shift to longer wavelength) which was confirmed by not only steady-state experiments but HOMOs and LUMOs calculations. The antimicrobial effects of the synthesized molecules on six different organisms were tested. Considerable activities of molecules 5p and 5n on many organisms were determined. Docking modeling and Lipinski, Ghose, and Veber compatibility studies of two different molecules were performed.