Cyclodextrin supramolecular inclusion-enhanced pyrene excimer switching for time-resolved fluorescence detection of biothiols in serum

Abstract

We report here an efficient pyrene excimer signaling-based time-resolved fluorescent sensor for the measurement of biothiols (cysteine (Cys), homocysteine (Hcy), glutathione (GSH)) in human serum based on thymine–Hg2+–thymine (T–Hg2+–T) coordination chemistry and the inclusion interaction of cyclodextrin. The sensing mechanism of the approach is based on the competitive ligation of Hg2+ ions by Hcy/Cys/GSH and T–T mismatches in a bis-pyrene-labeled DNA strand with the self-complementary 5′ and 3′ ends. The introduction of γ-cyclodextrin can provide cooperation for the molecular level space proximity of the two labeled pyrene molecules, moreover the hydrophobic cavity of γ-cyclodextrin can also offer protection for the pyrene dimer's emission from the quenching effect of environmental conditions and enhance the fluorescence intensity of the pyrene excimer. When the biothiols are not presented, the sensing ensemble is in the “off” state due to the long distance between the two labeled pyrene molecules resulted from the formation of a more stable T–Hg2+–T structure. While in the presence of biothiols, Hg2+ interacts very strongly with thiol groups and the T–Hg2+–T structure is dehybridized, and then the pyrene excimer will be formed due to the self-complementary 5′ and 3′ ends of the DNA probe and the cooperation interaction of γ-cyclodextrin to pyrene dimer, thus resulting in switching the sensing ensemble to the “on” state. In the optimum conditions described, the linear concentration range of 1.0–100μM with the limit of detection (LOD) of 0.36μM for GSH was obtained. Moreover, due to the much longer lifetime of the pyrene excimer fluorescence than those of the ubiquitous endogenous fluorescent components, the time-resolved fluorescence technique has been successfully used for application in complicated biological samples.