Molecular Probe to Visualize Effect of Glycolytic Inhibitor on Reducing NADH levels in the Cellular System

1,4-Dihydronicotinamide adenine dinucleotide hydride (NADH) is an essential coenzyme existing in all living organisms. Due to its involvement in different biological process, fluorescence imaging of intracellular NADH levels at different pathological conditions has emerged as an interesting area of research. We report here the exploration of a fluorescent probe MQ-CN-BTZ as dual channel NADH imaging agent (green and red channel) for cellular system. Interestingly, depending on the ratio between probe and NADH concentration in solution phase, the probe showed emission at ~530 nm and ~660 nm when excited at 475 nm. It is to be noted that the probe showed very large Stokes shift of ~180 nm with respect to the longer wavelength emission with good fluorescence response towards NADH. In general, such large Stokes shift is highly beneficial for imaging applications largely due to better separation between emission and excitation spectra, and reduced spectral overlap. Finally, the probe was utilized to image the event of glycolysis pathway by employing 3-bromopyruvic acid (3-BrPA) as a glycolytic inhibitor that inhibits the activity of glyceraldehyde phosphate dehydrogenase (GAPDH) enzyme involved in a crucial step of the glycolysis. As the depletion of the NADH levels corresponds to the inactivity of GADPH upon treatment with inhibitor, we imaged the modulation of NADH concentration in cellular system in the presence of 3-BrPA inhibitor indicating the importance of glycolysis step in elevating NADH levels. Overall, the present study attempts to demonstrate the importance of fluorescent probe for imaging intracellular NADH in the presence of glycolytic inhibitor.