Visible-near-infrared and fluorescent copper sensors based on julolidine conjugates: selective detection and fluorescence imaging in living cells

We present novel Schiff base ligands julolidine–carbonohydrazone 1 and julolidine–thiocarbonohydrazone 2 for selective detection of Cu<SUP>2+</SUP> in aqueous medium. The planar julolidine-based ligands can sense Cu<SUP>2+</SUP> colorimetrically with characteristic absorbance in the near-infrared (NIR, 700–1000 nm) region. Employing molecular probes 1 and 2 for detection of Cu<SUP>2+</SUP> not only allowed detection by the naked eye, but also detection of varying micromolar concentrations of Cu<SUP>2+</SUP> due to the appearance of distinct coloration. Moreover, Cu<SUP>2+</SUP> selectively quenches the fluorescence of julolidine–thiocarbonohydrazone 2 among all other metal ions, which increases the sensitivity of the probe. Furthermore, quenched fluorescence of the ligand 2 in the presence of Cu<SUP>2+</SUP> was restored by adjusting the complexation ability of the ligand. Hence, by treatment with ethylenediaminetetraacetic acid (EDTA), thus enabling reversibility and dual-check signaling, julolidine–thiocarbonohydrazone (2) can be used as a fluorescent molecular probe for the sensitive detection of Cu<SUP>2+</SUP> in biological systems. The ligands 1 and 2 can be utilized to monitor Cu<SUP>2+</SUP> in aqueous solution over a wide pH range. We have investigated the structural, electronic, and optical properties of the ligands using ab initio density functional theory (DFT) combined with time-dependent density functional theory (TDDFT) calculations. The observed absorption band in the NIR region is attributed to the formation of a charge-transfer complex between Cu<SUP>2+</SUP> and the ligand. The fluorescence-quenching behavior can be accounted for primarily due to the excited-state ligand 2 to metal (Cu<SUP>2+</SUP>) charge-transfer (LMCT) processes. Thus, experimentally observed characteristic NIR and fluorescence optical responses of the ligands upon binding to Cu<SUP>2+</SUP> are well supported by the theoretical calculations. Subsequently, we have employed julolidine–thiocarbonohydrazone 2 for reversible fluorescence sensing of intracellular Cu<SUP>2+</SUP> in cultured HEK293T cells