Two donor-acceptor fluorophores were prepared and tested for quantitative determination of HSA in aqueous samples. Fluorophores were non-emissive in polar solvents due to energy loss via nonradiative decays. Complexation of the fluorophores with HSA resulted multi-fold enhancement of emission in the red-near infrared (NIR) region. The emission intensity was linearly correlated to the amount of protein in the solution, which enabled us to develop calibration graphs for quantitative estimation of HSA in synthetic urine samples. Between the two fluorophores, the methoxy substituted fluorophore 1 selectively recognized HSA. It exhibited remarkable fluorescence enhancement with HSA over bovine serum albumin (BSA) and other globular proteins. The selective sensing aptitude of 1 was attributed to its restricted motions in the protein's microenvironment due to multiple non-covalent interactions, preventing energy loss by radiationless decay. The different recognition properties of the fluorophores were estimated by the steady-state fluorescence and molecular docking studies. These findings indicate that this class of fluorophores can be useful for quantitative estimation of HSA in biological urine and blood samples in clinical practice.
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