Abstract
Changes in adenosine triphosphate (ATP) and peroxynitrite (ONOO–) concentrations have been correlated in a number of diseases including ischemia-reperfusion injury and drug-induced liver injury. Herein, we report the development of a fluorescent probe ATP-LW, which enables the simultaneous detection of ONOO– and ATP. ONOO– selectively oxidizes the boronate pinacol ester of ATP-LW to afford the fluorescent 4-hydroxy-1,8-naphthalimide product NA-OH (λex = 450 nm, λem = 562 nm or λex = 488 nm, λem = 568 nm). In contrast, the binding of ATP to ATP-LW induces the spirolactam ring opening of rhodamine to afford a highly emissive product (λex = 520 nm, λem = 587 nm). Due to the differences in emission between the ONOO– and ATP products, ATP-LW allows ONOO– levels to be monitored in the green channel (λex = 488 nm, λem = 500–575 nm) and ATP concentrations in the red channel (λex = 514 nm, λem = 575–650 nm). The use of ATP-LW as a combined ONOO– and ATP probe was demonstrated using hepatocytes (HL-7702 cells) in cellular imaging experiments. Treatment of HL-7702 cells with oligomycin A (an inhibitor of ATP synthase) resulted in a reduction of signal intensity in the red channel and an increase in that of the green channel as expected for a reduction in ATP concentrations. Similar fluorescence changes were seen in the presence of SIN-1 (an exogenous ONOO– donor).
Highlights
Adenosine-5′-triphosphate (ATP) has been referred to as the “molecular currency”.1,2 ATP concentrations range between 1 and 10 mM, with a 1000:1 ratio between ATP and adenosine diphosphate (ADP) typically prevailing.[3]
We reported AND-logic gate-like dual-analyte fluorescence scaffolds that permit the detection of ONOO− and a second analyte.[34−38] we believe that systems able to detect more than one analyte independently using different emission channels will prove advantageous;[20] this is because, in principle, dual-channel emission should enable the concurrent evaluation of each individual species, whereas AND-logic systems only provide information on the synergy of the two species
Article consistent with the known intramolecular charge transfer (ICT) process seen in 4-hydroxy-1,8-naphthalimide products (Figure S1).[40]
Summary
Adenosine-5′-triphosphate (ATP) has been referred to as the “molecular currency”.1,2 ATP concentrations range between 1 and 10 mM, with a 1000:1 ratio between ATP and adenosine diphosphate (ADP) typically prevailing.[3]. Viously, several single-analyte fluorescent probes have been reported for the selective imaging of ONOO−.15−17 fluorescent probes for the selective detection and visualization of ATP have been developed.[18] to our knowledge no fluorescent probes capable of the simultaneous and independent imaging of ONOO− and ATP have been reported If available, such systems would allow the presumed close relationship between these two critical species to be monitored in real time. As a preliminary proof-of-concept study for cellular applications, changes in ATP and ONOO− associated with acetaminophen (APAP) were evaluated This model was chosen due to the importance of preclinical tools for drug development in the screening of drug-induced liver injury (DILI) associated with drugs like APAP.[19−21] this model was selected because APAP can result in a depletion of ATP and an increase in the levels of ONOO− (Scheme S1).[22,23] We believe ATP-LW could prove popular as a fluorescent tool in fundamental and clinical research. The chemical structure of ATP-LW was fully characterized using 1H NMR and 13C NMR spectroscopy, as well as high-resolution mass spectrometry
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