TAPP-based fluorescent probes for selective cysteine detection: insights into structure–reactivity relationships

Abstract

Herein, a series of TAPP-based π-conjugated fluorescent probes functionalized with aldehyde, malononitrile, and cyanoacrylate groups were designed and synthesized to investigate structure–reactivity relationships in cysteine (Cys) sensing. The photophysical properties of the probes were systematically examined in THF, DCM, and DMSO. The results revealed that the absorption and emission features are primarily governed by π–π* transitions localized on the TAPP framework, while the terminal acceptor groups mainly modulate the electronic distribution without significantly altering the ground-state transitions. Among the investigated derivatives, only the aldehyde-functionalized probe exhibited a pronounced “turn-on” fluorescence response toward Cys. This behavior is attributed to a reaction-based sensing mechanism involving the formation of a thiazolidine ring between the aldehyde group and Cys. The proposed mechanism is supported by complementary spectroscopic analyses, including ¹H NMR and HRMS, which confirm the consumption of the aldehyde group and the formation of a new product species. The probe displayed rapid response kinetics, reaching a stable fluorescence signal within approximately 50 s, and exhibited high selectivity for glutathione (GSH) over N-acetylcysteine (NAC), other amino acids, inorganic sulfur species, and metal ions. Fluorescence titration experiments showed a linear response over 0–15 µM, with a low limit of detection (LOD) of 0.37 µM. Furthermore, pH-dependent studies demonstrated that the probe operates effectively under near-physiological conditions. Spiking experiments yielded satisfactory recovery values, confirming its practical applicability. Overall, this study not only presents a sensitive and selective fluorescent probe for cysteine detection but also provides mechanistic insight into how terminal group functionality governs reaction-based sensing behavior in TAPP-based systems.