Synthesis, characterization, and supramolecular architectures of two distinct classes of probes for the visualization of endogenously generated hypochlorite ions in response to cellular activity

Abstract

Two distinct classes of compounds, (E)-2-(((3-amino-4-nitrophenyl) imino) methyl)-5-(diethylamino) phenol (SB) and 5-(diethylamino)-2-(5-nitro-1H-benzo[d]imidazol-2-yl) phenol (IM) were synthesized. SB, a bright red colored compound was crystallized in acetonitrile as a triclinic crystal system while IM, yellow colored compound crystallized as a monoclinic crystal system in dimethylformamide by vapor diffusion of diethylether. These compounds were characterized using spectroscopic techniques (IR, UV–visible, 1H, and 13C NMR), and X-ray crystallography. SB and IM displayed classical and non-classical H-bonding involving C-H…O and π…π interactions. These compounds detected hypochlorite ions in aqueous DMSO (1: 9, v/v, HEPES buffer, pH 7.4), and detection was visible via color changes by naked eye. We also performed UV–visible and fluorescence titrations, showing detection limits of 8.82 × 10−7 M for SB and 2.44 × 10−7 M for IM. The fluorometric responses from SB and IM were also studied against different ROS and anions. DFT calculations were performed to strengthen the proposed sensing mechanisms of both SB and IM. Hypochlorite, which is endogenously generated by myeloperoxidase in endosomes, was specifically visualized using SB and IM in lipopolysaccharide-treated RAW264.7 cells. These probes were also used to image the generation of hypochlorite by RAW264.7 cells during phagocytosis of non-fluorescent polystyrene beads.