A highly sensitive and selective probe for Vancomycin (Van) in aqueous and serum samples is developed in this study. The probe is based on a triad consisting of a near-infrared squaraine dye (Seta-640) conjugated to two anthraquinone molecules via Lys-d-Ala-d-Ala peptides. In the absence of Van, the close proximity and efficient electron-transfer from the excited Seta-640 dye to anthraquinone result in significant fluorescence quenching of the dye ("off"-state). When Van is added, the antibiotic molecules bind with high affinity to the -d-Ala-d-Ala ligands in a 2:1 stoichiometry (Van:triad), resulting in fluorescence recovery that is as high as 30 times ("on"-state). Even though bound Van enhances the fluorescence by reducing the rate of (intrinsic) polarity-induced nonradiative decay process, this effect plays only a minor role. Instead, the main reason behind the observed fluorescence recovery after drug binding is the effective inhibition of electron-transfer; plausibly arising from a steric-induced lengthening of the spatial separation between electron donor and acceptor. The probe has detection limits of 7.0 and 96.9 nM in buffer and human serum, respectively, operates in the clinically relevant range, is insensitive to Van crystalline degradation product (CDP-1), and is easy to operate by using a commonly available fluorescence spectrometer.
Keywords: anthraquinone; electron transfer; molecular triad; squaraine dye; vancomycin.