Click Chemistry Derived Hexa-ferrocenylated 1,3,5-Triphenylbenzene for the Detection of Divalent Transition Metal Cations

The 1,3-dipolar cycloaddition reaction (click chemistry approach) was employed to create a hexa-ferrocenylated 1,3,5-triphenylbenzene derivative. Leveraging the presence of metal-chelating sites associated with 1,2,3-triazole moieties and 1,4-dinitrogen systems (ethylenediamine-like), as well as tridentate chelating sites (1,4,7-trinitrogen, diethylene triamine-like) systems, the application of this molecule as a chemosensor for divalent transition metal cations was investigated. The interactions were probed voltammetrically and spectrofluorimetrically against seven selected cations: iron(II) (Fe2+), cobalt(II) (Co2+), nickel(II) (Ni2+), copper(II) (Cu2+), zinc(II) (Zn2+), cadmium(II) (Cd2+), and manganese(II) (Mn2+). Electrochemical assays revealed good detection properties, with very low limits of detection (LOD), for Co2+, Cu2+, and Cd2+ in aqueous solution (0.03–0.09 μM). Emission spectroscopy experiments demonstrated that the title compound exhibited versatile detection properties in solution, specifically turn-off fluorescence behavior upon the addition of each tested transition metal cation. The systems were characterized by satisfactory Stern–Volmer constant values (105–106 M–1) and low LOD, especially for Zn2+ and Co2+ (at the nanomolar concentration level).

NMR experiments were carried out using a Varian VNMRS 500 MHz spectrometer ( 1 H at 500 MHz, 13 C{ 1 H} NMR at 126 MHz) equipped with a multinuclear z-gradient inverse probe head.The spectra were recorded at 25 °C and standard 5 mm NMR tubes were used. 1 H and 13 C chemical shifts (δ) were reported in parts per million (ppm) relative to the solvent signal, i.e., Chloroform-d: δH (residual chloroform) 7.26 ppm, δC (residual chloroform) 77.16 ppm; DMSO-d6: δH (residual DMSO) 2.50 ppm. 1 H DOSY (Diffusion Ordered SpectroscopY) NMR experiments were performed using a stimulated echo sequence incorporating bipolar gradient pulses 3 and with convection compensation. 4The gradient strength was logarithmically incremented in 15 steps from 25% up to 95% of the maximum gradient strength.NMR spectra were analyzed with the MestReNova v12.0 software (Mestrelab Research S.L).The hydrodynamic radius from 1 H DOSY NMR experiment was estimated using the unmodified Stokes-Einstein equation 5,6 : where D is the measured diffusion coefficient for compound 3 (9.0810−11 m 2 s −1 ), kB is the Boltzmann constant (1.380648510 −23 kgs −2 K −1 ), T is the temperature for the 1 H DOSY NMR spectrum acquisition (298 K), rH,solv is the calculated hydrodynamic radius of compound 3 (ca.1.2 nm), η is the viscosity of the solvent (DMSO) at temperature T (0.001991 kgm −1 s −1 ).APCI-HRMS (q-TOF) measurements were recorded using Synapt G2-S HDMS mass spectrometer (Waters) equipped with the atmospheric-pressure chemical ionization (APCI) ion source and quadrupole-Time-of-Flight (q-TOF) mass analyzer.Methanol (Honeywell, HPLC-MS Chromasolv, purity ≥ 99.9%) was used as a solvent and mobile phase with the flow rate of 100 µl/min.Sample was dissolved and injected directly into the APCI source.Injection volume was 1 µl.The measurements were recorded in the positive and negative ion modes with the resolving power of TOF analyzer 20 000 FWHM.The lock-spray spectrum of Leucine-enkephalin was generated by the lockspray source and the correction was performed for recorded mass spectra in the mass range of m/z = 50 -1200.The exact mass measurements were performed within 3 mDa mass error.Nitrogen was used as the desolvation and cone gas, and their flow values were set to 600 L/h and 100 L/h, respectively.Nebulizer gas pressure was set to 5.0 bar.Source and probe temperatures were set to 120˚C and 550˚C, respectively.Corona current was set to 13.0 µA, sampling cone voltage and source offset were set to 40 V.The instrument was controlled and data were processed using the MassLynx V4.1 software package (Waters).
UV-vis measurements were performed with a WVR UV-1600PC spectrometer, with the spectral resolution of 2 cm −1 .For the UV-Vis measurements, the wavelengths for the absorption maxima λmax were reported in nm.
Emission spectra were recorded with a HITACHI F-7100 FL spectrometer, parameters, scan speed: 1200 nm/min, delay: 0.0 s, EX slit: 5.0 nm, EM slit: 5.0 nm, PMT voltage: 400 V.The wavelengths for the emission maxima (λem) were reported in nm.
Cation binding experiments between compound 3 (chemosensor) and cations (analytes; Mn 2+ , Cd 2+ , Ni 2+ , Zn 2+ , Co 2+ , Fe 2+ , Cu 2+ ) were performed employing the emission spectra measurements.Cations were introduced in the form of their corresponding salts: The experiments were performed in the DMSO/H2O = 1:1 v/v solvent system as follows.Stock solution of 3 (2•10 −4 M) in DMSO was diluted with adequate volume of DMSO (to reach volume of 0.5 mL).This was followed by addition of proper amount of stock solution (1•10 −5 M) of given cation, and finally H2O was added to reach volume of the sample of 1 mL.Final concentration of 3 in each sample equaled 2•10 −7 M. Excitation wavelength (ex) was 270 nm.][9][10] Data for the calculations were collected for emission wavelength (em) of 356 nm.KSV values were taken as a slope value of 1/C(cation) versus 1/ΔI linear regression plots.LOD values were estimated from the intercept and slope of the linear regression plots of (I-Imin)/(Imax-Imin) versus log(Ccation) as follows.At first, the x(y=1) value was calculated.Next, LOD was taken as 10 x(y=1) .Spectrofluorimetric studies on systems' stoichiometries (continuous variation method, Job's plot method 11 ) were performed in DMSO/H2O = 1:1 v/v solvent system with the samples comprising total number of moles of the receptor (compound 3) and analyte (cations) of 4•10 −8 mol.Samples varied in the molar fraction (xcation) of the analyte, ranging from 0.00 to 0.94.

S7. DFT calculations methodology
Theoretical calculations were performed using Orca 5.0 program. 12Molecules were optimized using BP86 method 13 with def2-TZVP basis set 14 augmented with def2/J auxiliary basis set 15 with RI approximation 16 .The calculations were performed in the presence of the solvent field with the polarizable continuum model (PCM) using the CPCM polarizable conductor calculation model. 17The parameters of water were used.The starting geometries were constructed using Avogadro program. 18Following geometry optimization, the vibrational frequencies were calculated, and the results showed that optimized structures are stable geometric structures (no imaginary frequencies).Gibbs free energies at 25 °C were obtained from the frequency calculations.
The calculations were performed for the following molecules: a) a single water molecule, b) Cd 2+ ion hydrated by six water molecules, c) three chosen representative fragments of the receptor, d) complexes of Cd 2+ with the chosen fragments of the receptor and water molecules.The initial geometries of the complexes were constructed in such a way that Cd 2+ would adopt octahedral geometry with either receptor atoms and water molecules as ligands.The binding energies of the complexes were calculated from the equation: Where: ΔGbindfree energy of the binding, ΔGcomplex -free energy of the complex, n -number of water molecules included in the complex, ΔGwater -free energy of a single water molecule, ΔGligand -free energy of the receptor fragment, ΔGion -free energy of Cd 2+ hydrated by six water molecules.

Figure S10 .
Figure S10.Stern-Volmer plots for the emission quenching of 3 by the addition of metal cations.Parameters of the linear regression, as well as calculated Stern-Volmer constant (KSV) values are also provided.
Figure S12.Job's plots for interactions between 3 and metal cations.