Synthesis, Characterization, Antibacterial Activity of Binuclear Metal Complexes with Macrocyclic Schiff Base Ligand and DFT Studies.

1. Introduction:

   Schiff bases constitute a broad class of organic compounds characterized by the presence of an imine (-C=N-) functional group [1], [2], [3]. They are typically formed through the condensation reaction between aliphatic or aromatic amines with aldehydes or ketones containing a reactive carbonyl group [4]. The formation of Schiff bases generally proceeds via nucleophilic attack of the amine on the carbonyl carbon, followed by elimination of water. Depending on the electronic properties of the substrate, this process can be accelerated by either an acid or base catalyst, or by the application of elevated temperature [5], [6]. The broad applicability of Schiff bases is primarily attributed to the strong donor ability of the azomethine nitrogen atom, which acts as an effective electron donor and enables Schiff bases to coordinate readily with metal ions, thereby attracting considerable scientific interest in both ligand design and metal-complex synthesis [7]. Schiff bases are substantial due to their stability, chelating properties, and biological applications [8]. 

The rise of antibiotic-resistant organisms has increased the importance of identifying new antibacterial medicines with improved selectivity and reduced toxicity [9]. Schiff bases and their metal complex have also been shown to have antibacterial, antifungal, and anticancer properties [10]. They are effective antibacterial agents, Metal complexation can significantly improve biological activity and alter microbial susceptibility [11], [12]. Schiff-base macrocyclic ligands and their metal complexes represent an essential class of compounds that have been extensively investigated. Macrocyclic tetradentate Schiff base ligands are significant in coordination chemistry due to their four donor sites, which provide robust and versatile binding [13], as shown in Figure 1.

The synthesis of tetraazamacrocyclic complexes has been significantly aided by condensation reactions between diketones and primary diamines in the presence of metal ions.The metal facilitates the process by directing the steric course toward cyclic molecules rather than polymeric structures. Macrocyclic complexes exhibit greater thermodynamic stability and enhanced selectivity as ion binders compared to their open-chain analogs [14], [15]. Their cyclic framework generates stable metal complexes with various structural features, making them significant in catalysis, bioinorganic chemistry, and medical applications [16]. Macrocyclic Schiff ligands, particularly when coordinated with transition metals, have attracted significant attention because of their structural diversity and their ability to influence the physicochemical and biological properties of the resulting metal complexes [17]. In the present study, a novel Schiff base ligand and its symmetrical binuclear azomethine complexes of Mn(II), Fe(II), Co(II), Ni(II), Cu(II), and Zn(II) were successfully synthesized and characterized, These complexes were prepared through the condensation of Terephthalaldehyde and 1, 6- hexamethyle-nediamine (Figure 2).

The tetradentate Schiff base ligand and all the six complexes were identified and characterized using NMR, IR, UV-Visible spectroscopy, atomic  absorption, molar conductivity, and magnetic moment measurement. They were examined for their antibacterial efficacy against Escherichia coli, Staphylococcus aureus, and Klebsiella Pneumonia to assess the influence of metal-ion identity and macrocyclic coordination on biological effectiveness. Furthermore, Density Functional Theory (DFT) calculations were carried out using the B3LYP functional. The 6-311++G(d,p) basis set was used for non-metal atoms (C, H, and N), while the LANL2DZ basis set was employed for the transition metal ions. Gaussian 16 software was used for geometry optimization and the calculation of structural parameters, including bond lengths and bond angles. The frontier molecular orbitals (FMOs),(HOMOs and LUMOs) were also investigated. The global reactivity descriptors were derived from FMO energy-level analysis. Providing optimized geometries, HOMO–LUMO energy gaps, and electrostatic potential assessments of the molecules. The thermodynamic parameters were calculated in both the gas and solvent phases to evaluate the effect of solvation on the stability of the studied compounds. 

2. Materials and Methods:

All solvents and reagents used in this work were of analytical quality. Most chemicals were supplied by the University of Duhok, including MnCl₂·4H₂O, FeCl₂· 4H₂O, CoCl₂· 6H₂O, NiCl₂· 6H₂O, CuCl₂· 2H₂O, ZnCl₂, hexamethylenediamine, ethanol, and diethyl ether. Additionally, Terephthalaldehyde was purchased from Bide Pharmatech Ltd, glacial acetic acid from Scharlau, and DMF from Thomas Baker. The ¹H and ¹³C-NMR spectra of the synthesized ligand were recorded on a Bruker Ultra Shield 500 MHz NMR spectrometer in CDCl3 at 298 K. FTIR spectra were obtained using a Shimadzu FTIR8400S spectrophotometer in the 4000–400 cm⁻¹ range with KBr discs. Magnetic susceptibility measurements were performed in the solid state at 298 K using a Sherwood Scientific magnetic  balance. Electronic absorption spectra were recorded at 25 ^oC in DMF using a Jenway 6800 double beam UV–Vis spectrophotometer 10⁻³ M with 1 cm quartz cells. The melting and decomposition temperatures of the ligand and its complexes were determined using an electrothermal melting point apparatus. Molar conductivity measurements were carried out using a 4520PH conductivity meter for 10⁻³ M DMF solutions at 25^oC. Metal ion contents of the complexes were finally  quantified using an AA-670G Atomic absorption spectrophotometer. The disc diffusion method was used to measure antibacterial activity.  The geometries and energies of these compounds were assessed using Gaussian 16W. 

2.1 Synthesis of Schiff Base Ligand [C₂₈H₃₆N₄]:

The synthetic route of the macrocyclic Schiff base ligand by mixing (0.02mol, 2.6822g) Benzene-1, 4-dicarboxaldehyde in 70ml of ethanol with (0.02 mol, 2.323g) hexamethylenediamine in 50ml of ethanol, under continuous magnetic stirring. Subsequently, three droplets of glacial acetic acid were added as a catalyst. The reaction mixture was refluxed for 8 hours at 80 ^oC. The precipitate formed was then filtered, thoroughly washed with ethanol and diethyl ether, and dried in vacuo over anhydrous CaCl2. A pale-yellow solid was obtained, yielding 4.0751 g (95.09%) and a melting point of 176 ^oC. H-NMR, 500 MHz; CDCl3) δ 8.27 (s,1H), 7.747 (s, 2H), 3.62 (t, 2H), 1.77 – 1.35 (m, 4H). 13C-NMR (125 MHz, CDCl3) δ 160.28, 138.07, 128.23, 77.03, 76.77, 61.83, 30.81, 27.17. 

Figure 1: The macrocyclic formation reaction, where A and B represent aliphatic or aromatic(dialdehyde and diamine),n is the number of moles of M(Ⅱ) transition metals.

2.2 Synthesis of Binuclear Metal (II) Complexes:

2.2.1 Synthesis of [Ni2(C28H36N4) Cl4] Complex:

The complex was prepared by reacting (0.001 mol,0.428g) of the ligand in 15 mL of Chloroform, which was then placed in an ultrasonic bath for approximately 20 minutes. An ethanolic solution of NiCl2· 6H2O (0.002mol, 0.4753 g) was added dropwise to the ligand solution. The mixture was refluxed for approximately 9 hours at 80oC. The olive- green resultant solid complex was filtered, thoroughly washed with ethanol and diethyl ether successively, and subsequently dried at room temperature. Yield (0.6076 g), (88.35%), m.p (224) Dec.

Figure 2:Synthesis of Schiff Base Ligand.

2.2.2 Synthesis of the additional metal ions complexes [M₂(C₂₈H₃₆N₄) Cl₄]: 

A series of transition metal complexes with the general formula [M2LCl4], M = Mn(II), Fe(II), Co(II), Ni(II), Cu(II), Zn(II),were synthesized following the same procedure described for the nickel complex. The corresponding metal salts were used in 0.002 mol quantities: M=MnCl₂.4H₂O (0.3985g), FeCl₂.4H₂O (0.3976g), CoCl₂.6H₂O (0.475g), CuCl₂.2H₂O (0.340g), ZnCl₂ (0.2725g). Each metal salt solution was added dropwise to a chloroform solution of the ligand (0.001mol) that had been previously subjected to ultrasonication, followed by reflux. The solids were filtered, washed with Ethanol and diethyl ether, dried at room temperature. All complexes were subsequently subjected to physicochemical and some spectroscopic characterization. The synthetic route adopted for the preparation of the binuclear Schiff base complexes is illustrated in Figure 3.” 

2.3 Computational Aspects:

In chemistry and physics, density functional theory (DFT) is a valuable technique for determining the electronic structure of atoms, molecules and solids [18]. Electronic structure calculations play a crucial role in elucidating chemical structure and reactivity [19]. By advancing the development of computational chemistry, density functional theory (DFT) has been widely employed owing to its accuracy and low computational cost in calculating a broad spectrum of molecular properties, yielding reliable results that align with experimental data [20]. The geometries of the free ligand and its transition-metal complexes were optimized using the Gaussian 16 program. Geometry optimization calculations were carried out within the framework of density functional theory (DFT) employing the B3LYP functional. The 6-311++G (d,p) basis set was applied to the ligand atoms (C, H, and N), whereas the LANL2DZ basis set was used for the metal centers (Mn, Fe, Co, Ni, Cu, and Zn). This computational level was selected because it has been widely employed in studies of organic, coordination, and organometallic systems, offering a practical balance between computational cost and accuracy in describing ground-state geometries and electronic structures [21], [22]. We employed the computational protocol to determine the optimal bond lengths, bond angles, thermodynamic parameters and frontier molecular orbital energies (FMOs). Examining the energy of the (HOMO and LUMO)is especially crucial as variation in their energy separation illustrates how metal coordination influences the reactivity and stability of the complex [23], [24]. This is particularly relevant when  considering the HOMO–LUMO energy gap associated with  these orbitals. To identify the electrophilic and nucleophilic regions, the molecular electrostatic potential is employed [25]. 

2.4 Antibacterial Efficacy:

Antibacterial activity was assessed using clinical isolates collected from Sheryan Hospital. The antibacterial activity of  the free ligand and its complex was evaluated against Staphylococcus aureus (a Gram-positive bacterium), Klebsiella pneumoniae, and Escherichia coli (two Gram-negative bacteria). Antibacterial activity was evaluated using the disc diffusion method according to the Clinical and Laboratory Standards Institute (CLSI) guidelines [26]. Each bacterial strain was cultured into 5 mL of Brain  Heart Infusion Broth using a single well-isolated colony. The cultures were subsequently incubated at 37 ^oC for 18–24 h. To cover the surface of the Muller-Hinton agar (MHA) plates with bacteria in three different orientations we used a sterile cotton swab that had been submerged in the bacterial suspension. The plate was rotated 60 degrees between streaks, and the edges were cleaned thoroughly to ensure the germs were spread evenly. The ligand and its complexes were dissolved in DMF (10⁻³ M), and 5 mm filter paper discs were dipped into the solutions, which were then allowed to dry in a sterile environment. Discs were aseptically arranged on inoculated agar plates at distances of 30–36 mm to prevent overlapping inhibition zones. Every single dish maintained a precise temperature of 37 ^oC for more than 24 hours in an incubator. The circumference of the inhibition zone encircling the disc was measured after the incubation period [27].

*=Decomposition, M% = metal percent, Cal =Calculated, M.wt = Molecular weight

Table 2: The experimental and calculated FT-IR spectra of the SB ligand and its compounds.

ν = Stretching , δ = bending.

3.2 NMR Study of the Schiff Base Ligand:

The chemical structures of the synthesized Schiff Base ligand were elucidated using ¹H and ¹³C NMR spectroscopy (500 MHz) in CDCl3. The ¹H-NMR spectrum demonstrates successful condensation and structural characterization of the synthesized Schiff base ligand. In azomethine systems generated from aromatic aldehydes and amines, the absence of an aldehydic proton signal (typically near 9-10.0 ppm in the starting materials) is a primary diagnostic of complete imine production, a pattern that has been reported in several Schiff base studies [41]. The presence of a -CH=N proton signal at δ 8.27 ppm(s,1H) [42]. The signal at δ 7.74 ppm corresponds to the aromatic hydrogens [43]. A triplet signal observed at 3.60-3.63 ppm is attributed to the methylene protons adjacent to the imine nitrogen CH=N-CH2 [34]. The signal from 1.35-1.77ppm in the 1H-NMR spectrum of the Schiff base ligand results from protons of the methylene -CH2- group [44]. The ¹³C NMR spectra of the Schiff base ligands were obtained using CDCl₃ as the solvent. In the spectra, the signal area at 160.28 ppm confirms the formation of azomethine [45].The chemical shifts of the aromatic carbon in this Ligand were determined to be 128.23, 138.07ppm [5]. Strong signals from (76.77,77,03) δ belong to the CDCl3 Solvent [46]. The CH2-N=C contributed to the signal observed at 61.83ppm in the Schiff Base Ligand [47]. The ¹³C NMR spectrum of this molecule demonstrated peaks at δ 27.17 and 30.81 ppm ascribed to the methylene carbons CH2-CH2-[34], as shown in Figures 4 and 5.

3.3 Electronic spectra – magnetic moment:

The electronic spectra of the Schiff base ligand and its metal complexes were recorded in DMF solution, 0.001 M. The results are summarized in Table 3. The corresponding UV–Visible spectra of the ligand and its metal complexes are presented in Figure 6. The UV-Vis spectrum of the free Schiff base ligand exhibited absorption peaks within the range of(47846-35587)cm⁻¹ , (34246-29239 )cm⁻¹ , which can be attributed to π → π^∗ and n → π^∗ transitions associated with the azomethine group and aromatic ring of the ligand [48], [49], [50]. The Mn(II) complex displays a magnetic moment of 5.7 B.M, which correlates closely with the calculated spinonly value for a high-spin d5 configuration featuring five unpaired electrons. The spectrum shows several intense absorption bands in the ultraviolet region (44444-33222) cm⁻¹, which are attributed to π → π^∗ transitions, while the bands observed at higher wavelengths (32258-28901) cm⁻¹ are assigned to n → π^∗ transitions. The absorption band at 28089 cm⁻¹ is attributed to CT. In Mn(II) complex, the d-d transitions are generally very weak and are often not clearly observed because the electronic transitions are still spin-forbidden. Even though in a tetrahedral environment, due to relaxation of the parity selection rule, they remain spin-forbidden and therefore weak. Therefore, the spectra are not detectable in the visible spectrum [51]. The electronic spectrum of the Fe(II) complex exhibits absorption peaks in the ultraviolet region (44849-35971)cm⁻¹, which are assigned to π → π^∗, while those at (34013-32573) cm⁻¹ correspond to n → π^∗ transition. The bands in the (28571-25316) cm⁻¹ may be assigned to ligand to metal charge transfer transition. In the visible region, the complex exhibits a weak broad band around 11947 cm⁻¹, corresponding to the 5E → 5T2 transition characteristic expected of a high-spin Fe(II) center d6 configuration tetrahedral geometry.

Figure 4. ¹H-NMR spectrum of Schiff Base Ligand.

Figure 5. ¹³C-NMR Spectrum of Schiff Base Ligand.

Table 3. Electronic transitions and magnetic moments.

C.T= Charge Transfer, Td= Tetrahedral, Sq=square planer

The magnetic moment value is 4.6 B.M, corresponding to four unpaired electrons [52], [53]. The UV-Vis spectrum of Co(II) complex exhibits intense absorption bands (43478-34246) cm⁻¹ , which are attributed to π → π^∗ transition, and a band at 29325 cm⁻¹ is assignable to the n → π^∗ transition. The absorption observed at 27932 cm⁻¹ is assigned to a charge transfer transition. In the visible region, two weak, broad bands appear at 14836 cm⁻¹ and 16556 cm⁻¹ , arising from overlapping d-d transitions of cobalt in tetrahedral high-spin Co(II). The expected spin-allowed transition is a ⁴A₂(F)→⁴T₁(P) transition. The absence of ⁴A₂(F)→ ²T₂(F) and ⁴A₂(F) → ⁴T₁ (F) transitions is due to their low energy, which is out of the range of the instrument [54]. The magnetic moment value of 3.8 B.M for the Co(II) complex corresponds to three unpaired electrons [55]. In tetrahedral Co(II) complexes, the d-d transitions are typically broad and often overlap, resulting in fewer observable bands compared to octahedral Co(II) complexes, which usually display three well-resolved transitions. The Nickel complex spectrum shows several intense absorption bands in the ultraviolet region (43859-31250) cm⁻¹, which are assigned to π → π^∗ and n → π^∗ transitions. The band around 28490 cm⁻¹ is attributed to a charge-transfer transition. In the same spectrum, two weak peaks at (21186, 13404) cm⁻¹ were ascribed to the (d–d) transition ³T₁(F → ³T₂(F), ³T₁(F)→³A₂ in a tetrahedral geometry. The spin-only value for a d8 system with two unpaired electrons is 2.8 B.M, whereas tetrahedral Ni (II) complexes often exhibit higher magnetic moments due to orbital contribution.  

The magnetic value of 4.3 B.M. excludes a square-planar geometry, which is usually diamagnetic for Ni(II) complexes, and supports a tetrahedral coordination environment, which is higher than spin only due to the orbital contribution [56], [57], [58]. The electronic spectrum of the divalent Cu-complex showed bands at lower wavelength (44247-32786) cm⁻¹ are assigned to π → π^∗ and n → π^∗ transitions. The very weak band appears at (29411-27932) cm⁻¹, which may be attributed to a ligand-to-metal charge-transfer transition. A very weak broad band appears at 23201 cm⁻¹ and 19047 cm⁻¹, which is assigned to the 2B1g → ²A_1g and ²B_1g → ²E_g transitions [56]. The observed magnetic moment value of 1.82 B.M corresponds to one unpaired electron, as expected for a d9 configuration. Together with the electronic spectral pattern, this supports a square-planar geometry around the copper(II) ion. The Zn(II) metal complex has no discernible peak for the d10 configuration, indicating the absence of (d-d) electronic transitions. As expected for the d10 system they were found to be diamagnetic. A tetrahedral geometry has been attributed to the Zn(II) complexes based on these findings [59]. 

3.4 Antibacterial Activity:

The biological features of the complexes are influenced by the ligands chelating characteristics, the nature of the donor atoms, the overall charge of the complexes, the characteristics of the metal ion, the composition of the counter ions that stabilize the complex, and the geometrical configuration of the complex [60]. The C=N group, known as the azomethine group in Schiff bases, is responsible for biological activity due to its involvement in hydrogen-bond formation with the active centers of cellular constituents, thereby hindering normal cellular development. Consequently, numerous Schiff bases have been reported to exhibit antibacterial activity [61]. All synthesized compounds were evaluated for in vitro antibacterial activity against one Gram-positive bacterium (S.aureus) and two Gram-negative bacteria (E. coli and K. pneumoniae). The disc diffusion technique was used to determine the antibacterial efficacy of synthetic ligands and their corresponding coordination complex [62]. The synthesized Schiff base ligand and its metal complex were assessed for their antibacterial efficacy against bacterial strains, using Gentamicin as a positive control. The outcomes presented in Figure 7 were derived from a comparison of the biological characteristics of the ligand and its metal chelates. The common antibiotic Gentamicin was used as the standard antibacterial agent, respectively. The ligand and its complexes were dissolved in dimethylformamide (DMF) to obtain 0.001 M solutions. DMF was also evaluated under the same experimental conditions and showed no observable antibacterial activity. The results show that most complexes were more potent than the free ligand against bacteria. Furthermore, the Cu(II) complex exhibited the most potent antibacterial activity among the tested compounds, specifically against S. aureus with an inhibition zone of 13mm (lower than the Gentamicin standard) and K. pneumoniae and E. coli 8,7mm (higher than the Gentamicin standard) [61].

Other complexes also showed moderate to good activity, indicating that metal coordination is crucial for enhancing biological efficacy [49]. The Mn(II), Co(II), and Zn(II) complexes exhibit more action against S. aureus compared to Gram-negative bacteria, demonstrating inhibition zones of around 9–11 mm for S. aureus. In contrast, the free ligand (L) exhibits minimal activity against S. aureus (∼6 mm), whereas DMF shows no inhibitory effect, thereby confirming that the solvent did not influence the observed antibacterial activity. This pattern substantiates the idea that metal coordination augmented the biological activity of the Schiff base ligand. Comparable Schiff base ligand-versus-complex behavior is well documented for Schiff base systems, wherein the metal complexes frequently surpass the parent ligand in diffusion-based antibacterial evaluations. The chelating ligand increases lipophilicity, thereby facilitating penetration of lipid membranes in Gram-positive bacteria. The findings suggest that chelation enhances antibacterial efficacy [50], [63]. Lipidomic compounds are capable of traversing the fat bilayer that envelops the cell. This could be the determining factor in antibacterial action. The positive charge is more readily distributed among donor groups when ligand orbitals overlap, which reduces the polarity of metal ions in coordination. Additionally, it enhances the liposuction of complexes by increasing electron delocalization within the chelate ring.The enhanced lipophilicity of compounds enables them to penetrate lipid membranes, thereby inhibiting bacterial function [64]. 

has the highest thermodynamic stability because it exhibits the most negative Gibbs free energy. The Co(II) and Fe(II)  complexes have intermediate stability with Gibbs free energy values of (1656.963 and 1613.738) a.u. respectively. These results indicate efficient coordination between the ligand and metal centers, but with less stabilization relative to the Cu(II) complex. The Mn (II) complex is a little less stable (1574.629) a.u. than the other two. This is because it has a half-filled d5 shape, which usually means that the ligand field stabilization energy is lower. The Ni(II) and Zn(II) complexes, on the other hand, have the least negative Gibbs free energy values (1477.454 and 1498.174) a.u. Respectively, which  means they are less thermodynamically stable. This behavior is anticipated for Zn(II) since its d10 electronic structure does not provide ligand field stabilization. ZPVE magnitude and Cv rise post-coordination, demonstrating metal–ligand interactions add vibrational modes and molecular complexity. The complexes have more vibrational and rotational degrees of freedom than the ligand, increasing entropy. It enhances Gibbs free energy and stability [67].

3.5.2 The optimized geometry:

We employed the B3LYP/6-311++G(d,p)/LANL2DZ method and Gaussian 16 software to perform DFT calculations elucidating the influence of metal ions on the electronic characteristics of ligands. This computational approach identifies molecular structure, bond lengths, bond angles, thermodynamic parameters, and electronic characteristics, among other variables. DFT computations were employed to investigate these properties (Figure 8). The outcome reveals that modifying the central atom has a significant influence on the energy of the peripheral molecular orbitals [21]. The lengths and angles of the bonds of the prepared complexes are illustrated in Tables 6, 7, and 8. In these complexes, a Macrocyclic Structure is formed by four nitrogen-donor atoms from the chelate ligand. The measured bond lengths and angles indicate four-coordinate metal complexes with an azomethine, necessitating the participation of four electrondonating sites. These complexes have a metal-ligand proportion of 2:1 characterized by a tetradentate chelate ligand referred to as a Schiff Base Ligand. A similar pattern is observed in the gas phase, where all complexes remain thermodynamically favored relative to the free ligand. As presented in Table 5 a comparable stability pattern is also observed in the gas phase. The Cu(II) complex remains the most stable, followed by the Co(II), Fe(II), Mn(II), Zn(II), and Ni(II) complexes. While the Gibbs free energy values calculated in the gas phase are slightly less negative than those in solution, the relative order of stability is maintained. This finding suggests that solvation enhances the thermodynamic stabilization of the complexes, mainly through dielectric effects and weak solute–solvent interactions, but does not substantially modify their inherent stability sequence. Similar observations have been reported in published DFT investigations of Schiff-base systems, where solvent-phase calculations provide additional stabilization while preserving the preferred energetic arrangement identified in the gas phase [68]. atoms from the azomethine group (N3-N6) forming the corners. The bond length (M1-N3) of (2.00-2.87), (M1-N4) of (2.41-2.97),(M2-N5) of (1.97-2.24),(M2-N6) of (2.01-2.24), (M1-Cl7) of (2.24-2.44), (M1-Cl8) of (2.26-2.35), (M2-Cl9) of (2.29-2.45),(M2-Cl10) of (2.28-2.42), The measured bond lengths correspond with the bond distances documented in analogous four-coordinated systems in the literature [54], [69], [70]. A separate part of the analysis examined the calculated bond angles of the compounds. The bond angles of (N3-M1- N4), (N5-M2-N6), (Cl7-M1-N4 or Cl7-M1-N3) and (Cl10- M2-N5 or Cl10-M2-N6), are contained in the intervals [(84.9- 100.62) ,(82.92-100.89), (81.458-100.62) & (82.9-165.56)] [70], [71], [72], [73]. 

Figure 6. UV-Vis Spectra of the synthesized compounds: (a) is the Schiff Base Ligand, (b) Mn (II), (c) Fe (II), (d) Co (II), (e) Ni(II), (f) Cu(II), (g) Zn(II) Schiff Base Complexes. 

Figure 7. The antibacterial efficacy of a ligand and its metal complex in (mm).

Table 4. The thermodynamic behaviors of the ligand and its complexes in the solvent.

Table 5. The thermodynamic behaviors of the ligand and its complexes in the gas phase.

3.5.3 Electronic properties:

Frontier molecular orbitals (FMOs) are widely recognized as the most important orbitals for describing molecular electronic structure and chemical behavior. It is widely recognized that the frontier molecular orbitals (FMOs) are the principal orbitals. In a chemical process, the two most essential orbitals are the HOMO and the LUMO, or the Highest occupied molecular orbital and the lowest unoccupied molecular orbital [74]. Subsequently, the electrical transport properties of molecules are determined by the frontier orbitals, specifically the HOMO and LUMO. The frontier orbital gap is the energy difference between the HOMO and LUMO energy levels [75], [76]. The calculated HOMO and LUMO energy values for the metal complexes, along with derived quantum chemical parameters, are presented in Table 9, as shown in Figure 9. The HOMOLUMO energy gap of a molecule is commonly used as an indicator of various properties, including its kinetic behavior, chemical stability, optical polarizability, and chemical hardness and softness [77], [78], [79]. The electron distribution of any molecule is less variable and demonstrates minimal polarization when the HOMO–LUMO energy gap is substantial. These compounds are designated as hard molecules. When there is minimal variation in HOMO–LUMO energy, polarization is strong, electron distribution is readily manipulated, and the molecules are classified as soft molecules [80]. Assessing the HOMO–LUMO energy gaps of molecules is essential for providing information. This resulted in the determination that the most stable and hardest complex was the Zn(II) complex. The calculations indicated that the molecule identified as the Cu(II) complex had the  greatest softness and reactivity, with lower kinetic stability than the others. Based on the calculation The complexes can be arranged in order of increasing energy gap: Zn(II) >Fe(II) >Ni(II) >Co(II) >Mn(II) >Cu(II) [81]. This indicates that the Zn(II) complex has the largest energy gap and is therefore expected to be the hardest and least reactive species among the studied complexes. Using HOMOLUMO energies obtained from DFT calculation at the B3LYP level employing 6-311++G(d,p) basis sets for nonmetal atoms and LANL2DZ for the metal centers, were used to calculate a variety of metrics for the complexes, including the energy gap(E) [82], [83]. Chemical hardness(η) [84], electrophilicity index(ω) [85], [86], and Electronegativity (X) [87]. The ionization potential (IP) and Nucleophiles are characterized by parameter(N) [88]. Tetracyanoethylene (TCE) is employed as a reference owing to its inherently low HOMO energy among polar organic molecules [89]. The HOMO and LUMO energies are used to calculate these quantum-chemical characteristics using formulas. Where TCE serves as the reference The energies of the HOMO and LUMO electron orbitals correspond to the ionization potential (IP), respectively. The Mn(II) complex has the lowest ionization potential, making it the most effective electron donor. The chemical reactivity of a molecule depends on its chemical structure. The Cu(II) complex has the highest electronegativity among the complexes, indicating the most excellent electron-withdrawing ability. The Nucleophilicity index [90]. It is essential and all the synthesized compounds (1–6) range from (3.60 to 4.90) a.u. The maximum value of N for Mn(II)complex is 4.90 a.u, the strongest Nucleophile among all [91], [92]. The electrophilicity values of all synthesized complexes are classified as the most powerful, as they all exceed 1.5 eV [93]. 

Figure 8. The optimized structure of the ligand and complexes (different colors indicate Blue -Nitrogen, White -Hydrogen,
Grey- Carbon, and Green -Chlorine).

Table 6. The geometrical bond length of the synthesized compounds.

Electrophilicity Index (ω) ∶ ω = µ²/2η                                          (1)

Ionization energy (IP) = IP = −EHOMO                                      (2)

Energy Gap (∆EGAP) ∶ ∆E = ELUMO −EHOMO                      (3)   

Chemical Hardness (η): 

η = ELUMO −EHOMO/2                                                            (4) 

Electronegativity (X) = 1/2 (ELUMO +EHOMO)                      (5) 

So ftness (S) = 1/2η                                                                     (6) 

N = EHOMO(Nu)EHOMO (TCE)                                              (7)

Table 7: Demonstrates the spatial bond angles of the synthesized compounds as determined using the B3LYP/6-311++G(d,p) method.

Table 8. The geometric bond angles of the synthesized compounds.

Figure 9. The HOMO and LUMO plots of the studied complexes.

Table 9. The electronic features of the prepared compounds (in a.u, one a.u. = 27.211 eV) at the level of DFT/B3LYP theory. 

4. Conclusions:

This research successfully synthesized a novel macrocyclic Schiff Base Ligand via condensation. It was then coordinated with several transition-metal ions, forming well-defined metal complexes. Comprehensive spectroscopic and analytical characterizations were consistent with the proposed structures. Electronic absorption spectra and magnetic susceptibility data confirmed the expected coordination geometries around the metal centers. The data demonstrate that the ligand forms stable binuclear complexes and acts as a tetradentate ligand. Molar conductivity investigations showed that complexes with the formula [M2LCl4], where M=Mn(II), Fe(II), Co(II), Ni(II), Cu(II), and Zn(II) were neutral, non-electrolytes, and all consisting of a tetrahedral geometry except Cu (II) which shows square planargeometry. Frontier molecular orbitals and optimal geometries were predicted by the DFT with the B3LYP method, using 6-311++G (d, p) and LANL2DZ basis sets for the ligand and its metal chelates, respectively. Among all the compounds, the Zn(II) complex exhibited the highest hardness and lowest reactivity, as shown by the HOMO–LUMO gap values. The antimicrobial test showed that most complexes have higher antibacterial activity than the free ligand.

5. Acknowledgements:

The authors would like to thank the Molecular Topology and Drug Design Research Unit, Department of Physical Chemistry, Pharmacy Faculty, University of Valencia, for providing access to a supercomputer with the Gaussian 16 software.

Funding: There is no financial support or sponsorship from
any institute.

Data Availability Statement: The data supporting the findings of this study are available upon reasonable request from
the corresponding author.

Declarations:

Conflict of interest: The authors confirm that there is no conflict of interest.

Ethical approval: This study does not involve human or animal subjects; therefore, ethical approval was not required.

Author Contributions: Marjan Marwan Islam: Conceptualization, methodology, software, validation, formal analysis, investigation, resources, data curation, and writing—original draft preparation. Salwan Idress Mohamed: review, editing, and supervision.