A Near-Infrared Fluorescent Probe for Detection of Nitroxyl in Living Cells

Nitroxyl (HNO), the one-electron reduced and protonated congener of nitric oxide (NO), has been demonstrated with excellent bio-pharmacological effects in cardiovascular disorder treatment, which is distinctive from that of NO. Despite its high reactivity, the accurate detection of HNO is a challenging issue. To resolve this problem, in this work, a near-infrared (NIR) metal-free fluorescent probe, ER-JN, was designed and synthesized for the detection of intracellular HNO concentration in simulated physiological conditions and living cells. The probe was consisted of two moieties, the BF2-chelated tetraarylazadipyrromethane fluorophore (aza-BODIPY) and the HNO recognition unit, diphenylphosphinobenzoyl group. The probe was purified by silica column chromatography eluting with CH2Cl2 to obtain the green solid product with a yield of 28%. The as-prepared probe exhibited high sensitivity, good selectivity and low cytotoxicity, and was applied to fluorescent bio-imaging of HNO in simulated physiological conditions. When used in detection of HNO, quantum yield of the probe increased from 0.01 to 0.35. The linear range was 0-50 mu M, with the detection limit of 0.03 mu M (S/N = 3). With confocal laser scanning microscope imaging analysis, the probe could be used to detect HNO concentration in living cells. Furthermore, the results of flow cytometry confirmed that the probe could be employed for the qualitative and quantitative detection of intracellular HNO level. In this work, we found that probe ER-JN could not only detect HNO in aqueous solution and in living cells, but also target endoplasmic reticulum. We anticipate that ER-JN will provide experimental bases in studying physiological and pathological functions of HNO in cells, in vitro and in vivo.