3.1.2. Ligands for Sensing Analysis and for Biological Applications

In order to achieve the goal of zinc(II) detection in complex biological systems, or to design zinc-based architectures with biological activity, the C=N moiety included in flexible ligands can be an easy synthetic solution. A half-*salen*-type *N*,*O*,*N* ligand [107] was prepared in situ by Shyamal Kumar Chattopadhyay in 2019 (Figure 7) and employed to produce a mononuclear zinc(II) complex whose structure was determined by single crystal X-ray diffraction. In an aqueous methanol solution at the physiological pH, the complex exhibits an intense greenish-blue fluorescence whose maximum does not differ substantially with respect to the free ligand while the intensity is about 17-fold stronger. The ability to give fluorescence in aqueous solutions makes the probe promising for DNA binding activity and fluorescence bio-imaging. By DFT calculations, the nature of the electronic transitions was assigned to the π,π\* transitions of the imine and heterocyclic moiety and to a n,π\* transition for the free ligand, and to π,π\* LCT transition in the complex.

**Figure 7.** *N*,*N*,*O* pincers for sensing analysis and biological applications.

Designed for cytotoxic and antibacterial activity by M.R. Prathapachandra Kurup and coworkers in 2020 [108], a potentially *N*,*N*,*O* tridentate ligand worked with copper and zinc salts with unexpected different results. The two complexes (Figure 7) show a very different structural and spectroscopic pattern. In solution, the fluorescence emission intensity of the ligand decreased on complexation with Cu(II) ion, which can be attributed to the decrease in electron density on the ligand due to d orbital being involved. Contrarily, the ligand acts as a bidentate pincer toward zinc(II), coordinating to the metal cation through phenoxo oxygen and imine nitrogen. The bis-chelate metal complex produces an enhancement in the fluorescent intensity in DMF, due to the prevention of the photoinduced electron transfer process preserving ILCT bands.

π Very recently [109], two *N*,*O*,*N* tridentate ligands were used by Fabiao Yu, Guang Chen, and coworkers as fluorescent sensors to monitor intracellular zinc(II) in living cells by fluorescent bioimaging. The elaborate fused-rings aromatic part guarantees a rich π-conjugated system able to give a photoluminescent response. The oxygen atom group of the C=O fragment is the third neutral jaw of the tridentate ligand. Due to the restriction of the isomerization and rotation of C=N upon coordination, the probes show fluorescence enhancement (from 4 to 7-fold at 523 and at 543 nm, respectively) and large Stokes shifts of the emission spectra. A real-time two-photon excitation wavelength apt to biological experiments and deep penetration in tissues was detected.
