Analytical Sciences

A Simple Long-wavelength Fluorescent Probe for Simultaneous Discrimination of Cysteine/Homocysteine and Glutathione/Hydrogen Sulfide with Two Separated Fluorescence Emission Channels by Single Wavelength Excitation

Hanchuang ZHU,* Caiyun LIU,* Hanming ZHANG,* Pan JIA,* Zilu LI,* Xue ZHANG,* Yamin YU,* Wenlong SHENG,** and Baocun ZHU*

*School of Water Conservancy and Environment, University of Jinan, Shandong Provincial Engineering Technology Research Center for Ecological Carbon Sink and Capture Utilization, Jinan 250022, China
**Biology Institute, Qilu University of Technology (Shandong Academy of Sciences), Jinan 250103, China

Small molecular biothiols, such as cysteine (Cys), homocysteine (Hcy), reduced glutathione (GSH), and hydrogen sulfide (H₂S), play crucial parts in regulating the redox balance of life activities, regulating normal physiological activities and preventing various diseases. Quantitative analysis of these important small molecular substances is very important for revealing their diverse physiological and pathological effects. Although many fluorescent probes have been reported to detect biothiols in cells, it is still not sufficiently advanced to detect biothiols with separated fluorescence emission peak by same wavelength excitation. In our work, we designed a simple conjugate of Nile red and NBD (7-nitro-1,2,3-benzoxadiazole) as long-wavelength fluorescent probe NR-NBD for the simultaneous discrimination of these biothiols at single wavelength excitation. Probe NR-NBD could efficiently discriminate Cys/Hcy, GSH and H₂S by two separated fluorescence emission channels and absorption spectra. Importantly, probe NR-NBD has excellent specificity and sensitivity towards the monitoring of endogenous/exogenous Cys/Hcy and GSH/H₂S in living cells and zebrafish.

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