Chemical probe compounds that become fluorescent upon reduction, and methods for their use

Abstract

Chemical stain compounds containing a fluorophore and a reducible quenching unit are disclosed. The reducible quenching unit quenches the fluorophore while in its oxidized state. Upon reduction, the quenching properties of the quenching unit are diminished or eliminated. The chemical compounds can be used in a variety of applications, including the detection of bacterial cells, monitoring the electron transport chain function of bacterial cells, monitoring the oxidation state of non-biological systems, and assaying the effectiveness of antibacterial or antimicrobial agents.

1 . A chemical compound having the structure FU-RQU or FU-L-RQU; where FU is a fluorophore unit, RQU is a reducible quenching unit, and L is a linker unit. 
     
     
         2 . The compound of  claim 1 , having the structure FU-RQU. 
     
     
         3 . The compound of  claim 1 , having the structure FU-L-RQU. 
     
     
         4 . The compound of  claim 1 , wherein:
 the reducible quenching unit has an oxidized state and a reduced state;   the reducible quenching unit in its oxidized state partially or fully quenches the fluorescence of the fluorophore unit.   
     
     
         5 . The compound of  claim 1 , wherein the fluorophore unit is fluorescein, BODIPY (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; boron dipyromethene difluoride), a rhodamine, a cyanine, or a coumarin. 
     
     
         6 . The compound of  claim 1 , wherein the reducible quenching unit is quinone, naphthaquinone, anthraquinone, quinonemethine, copper (Cu(II)), sulfate (SO 4   2− ), or nitrate (NO 3   − ). 
     
     
         7 . The compound of  claim 1 , wherein the reducible quenching unit is quinone, naphthaquinone, and anthraquinone. 
     
     
         8 . The compound of  claim 1 , wherein the linker unit is a linear linker, a branched linker, a cyclic linker, an aromatic linker, a polycyclic aromatic linker, or an unsaturated linker. 
     
     
         9 . The compound of  claim 1 , wherein the linker unit comprises carbon atoms, hydrogen, oxygen, sulfur, chlorine, fluorine, iodine, phosphorous, silicon, nitrogen, other atoms, or combinations thereof. 
     
     
         10 . The compound of  claim 1 , having the structure FU-polyethylene glycol-RQU, FU-O—P(═O)—O—RQU, FU-S(═O)—RQU, or FU-S(═O) 2 —RQU. 
     
     
         11 . (canceled) 
     
     
         12 . A kit comprising a chemical compound having the structure FU-RQU or FU-L-RQU;
 where FU is a fluorophore unit, RQU is a reducible quenching unit, and L is a linker unit;   and a solvent.   
     
     
         13 . The kit of  claim 12 , wherein the solvent comprises water, DMSO, ethanol, methanol, dimethylacetamide, dimethylformamide (DMF), N-methyl pyrrolidinone (NMP), or mixtures thereof. 
     
     
         14 - 18 . (canceled) 
     
     
         19 . A method of monitoring change in the oxidation state of a non-biological system, the method comprising:
 providing a non-biological system;   contacting the non-biological system with at least one chemical compound to form a test system, wherein the chemical compound has the structure FU-RQU or FU-L-RQU; where FU is a fluorophore unit, RQU is a reducible quenching unit, and L is a linker unit;   determining the fluorescence of the test system at a first time point to provide a first fluorescence;   determining the fluorescence of the test system at a second time point to provide a second fluorescence;   determining the difference between the first fluorescence and the second fluorescence to monitor change in the oxidation state of the non-biological system.