Synthesis, Spectroscopic Characterization, Biocidal evaluation Molecular Docking & DFT Investigation of 16-18 membered Macrocyclic Complexes of Cobalt (II)

Abstract

Macrocyclic ligands (MacL₁-MacL₃) and Co(II) complexes were synthesized via template condensation of o-phenylenediamine with various aromatic dicarboxylic acids. The elemental analysis, FT-IR, mass spectrometry, ¹H NMR, ¹³C NMR, UV-vis, SEM analysis, powder X-ray diffraction, thermogravimetric (TG) analysis, electrochemical studies, and DFT analysis were used to characterize these synthesized ligands and their cobalt (II) complexes. TGA analysis to determine the stability and decomposition kinetic parameters. In element analysis, the percentage of different elements present and also the stoichiometry of compounds were confirmed. The proposed framework for tetraaza macrocyclic cobalt (II) complexes was supported by spectral analysis, which also revealed distorted octahedral geometry surrounding the central metal atom. The molecular structure of cobalt (II) complexes was also optimized theoretically, and their electronic or thermodynamic parameters were obtained from density functional theory (DFT). The synthesized ligands and their cobalt (II) complexes were tested against bacteria: Escherichia coli, Bacillus subtitles. Candida albicans were tested for antifungal properties. It was found that ligands and complexes show good antimicrobial results. Finally, using the Auto Dock VinaPyRx programme, molecular docking studies were used to evaluate the biological significance of the synthesized ligands to identify the probable and efficient binding mechanisms between the various ligands and the active site of the receptor protein.

Graphical abstract

Antimicrobial, DFT, and Docking representation of Cobalt (II) macrocyclic complexes

Supplementary Information

The online version contains supplementary material available at 10.1007/s12039-022-02109-2.

Introduction

The current scenario of macrocyclic ligands with amide group and their transition metal complexes have been developing an active area of research interest from the last few decades, attracting the attention of the researchers due to the broad biological actions of transition metal complexes.^1–4 An important class of tetraamide macrocyclic (MacL_1-3) ligands are derived from the condensation of o-phenylenediamine with various aromatic dicarboxylic acids. Such type of ligands and their transition metal complexes have a wide-ranging field of pharmacological activities such as antitumor;^5–7 antibacterial;^8–10 antifungal;^11,12 antioxidant;^13,14 anti-inflammatory¹⁵ and analgesics¹⁶ activities. The formation of the macrocyclic ligands implicates the metal ion in the cavity of macrocyclic ligands and forms stable complexes, which makes the reactant groups in the desired conformation for the ring closer.¹⁷ The cyclization reaction is supported by the formation of metal-ligand bonds and the arrangement of multidentate ligands around the central metal atom, both favored by the formation's enthalpy, which also overcomes the unfavourable entropy.¹⁸ In general, the formation of stable macrocyclic complexes depends upon the size, ring structure, and rigidity of the cavity of the macrocycle. The strength and magnitude of crystal field effects are especially crucial in the case of transition metal complexes.¹⁹ The transition metal macrocyclic complexes formed by the macrocyclic ligands having the heterocyclic moieties are of prodigious importance due to their biocidal activities and good binding affinity with proteins.²⁰ Hence, the biological activities of the macrocyclic complexes containing heterocyclic moieties in the macrocyclic ring can increase the selectivity of ligands for a given ion and enhance the metal complex biological properties.²¹

The substituted pyridine is a fortunate framework and the most important class of heterocyclic compounds. It shows the many interesting biological activities and is widely tolerated by people owing to their enormous biological activities. People widely tolerate it and show extensive applications in remedial chemistry.²² The macrocyclic complexes and their derivatives derived from the o-phenylenediamine have been studied, and due to their pharmacological and biological properties, the o-phenylenediamine containing macrocyclic complexes gives a broad class in synthetic inorganic chemistry.²³ Following a thorough review of the literature, the DFT was used in the current study to calculate a number of macrocyclic molecules properties. DFT is unquestionably the best option for theoretical calculations. Cobalt(II) macrocyclic complexes possess significant antibacterial and antifungal as compared to other transition metal complexes.²⁴ Here in the presented work, the macrocyclic ligands were synthesized by condensation of o-phenylenediamine with substituted pyridine and phthalic acid, and their coordination behaviour with cobalt(II) compounds was studied (Scheme 1). Numerous spectral analyses, including elemental analysis, FT-IR, ¹H NMR, ¹³C NMR, mass spectrometry, UV-vis, SEM analysis, X-ray diffraction, DFT, and TG analysis, were used to explain the structure of these ligands and their complexes. The MacL_1-3 ligands and their complexes were optimized theoretically. Molecular docking and antibacterial activity have also been investigated in the current work.

Scheme 1.

Synthetic route of the synthesis of tetraamide ligand (MacL₁) and their cobalt(II) complex.

Experimental

Material and methods

The solvents and other metals salt used were of high purity, purchased from Sigma-Aldrich, and utilized just as it arrived. The phthalic acid, pyridine-2,3-dicarboxylic acid, and pyridine-2,6-dicarboxylic acid from (fluka). All the solvents are of AR grade.

Synthesis of ligand (MacL₁-MacL₃)

The 2:2 molar ratios are used to carry out the reaction. Ethanolic solution of o-phenylene diamine (1.08 g, 0.01 M) was placed in a magnetically stirred round-bottomed flask, and ethanolic solution of Phthalic acid (1.66 g, 0.01 M), pyridine-2,3-dicarboxylic acid (1.67g, 0.01 M) and pyridine-2,6-dicarboxylic acid (1.67g, 0.01 M were refluxed at room temperature for about 8-10 h and to make the solution acidic add the few drops of conc. HCl (to facilitate the condensation reaction). The colored ppt are formed with % yield of 76%, 78%, and 75%, respectively. The formed precipitate was separated by filtration and repeatedly rinsed with ethanol and dried in vacuo. The synthetic compounds were once more recrystallized in benzene and dried in vacuo.

MacL₁: Off white solid; Yield 76%; M.p.; 107 °C; Analytical calculated for C₂₈H₂₀N₄O₄; C, 70.58; H, 4.23; N, 11.76; Found; C, 69.80; H, 4.04; N, 11.60; MS(ESI), m/z: 476.15 [M+H]⁺.

MacL₂: Off white solid; Yield 78%; M.p.; 142 °C; Analytical calculated for C₂₆H₁₈N₆O₄; C, 65.27; H, 3.79; N, 17.56; Found; C, 64.92; H, 3.45; N, 17.10; MS(ESI), m/z: 478.14 [M+H]⁺.

MacL₂: Off white solid; Yield 75%; M.p.; 169 °C; Analytical calculated for C₂₆H₁₈N₆O₄; C, 65.27; H, 3.79; N, 17.56; Found; C, 65.12; H, 3.15; N, 17.08; MS(ESI), m/z: 478.14 [M+H]⁺.

Synthesis of complexes [Co(N₄ MacL₁)Cl₂]– [Co(N₆ MacL₃)Cl₂]

The 1:1 molar ratio are used to carry out the reaction. The complexes were synthesized by template method. A weighed amount of CoCl₂.6H₂O (0.023 g, 0.0001 mol) dissolved in ethanol and added to the ethanolic solution of ligand MacL₁ (0.047 g, 0.0001 mol) and mixed slowly with constant stirring. The resulting solution was refluxed for about 10-13 h at 45 °C. The solid Brown colored precipitate was obtained, separated by filtration, washed thoroughly with ethanol, and dried under vacuo (% yield=78%). Recrystallized from benzene. The same procedure has been used for the synthesis of Co(N₆MacL₂)Cl₂ and Co(N₆ MacL₃)Cl₂ by replacing MacL₁ with MacL₂ and MacL₃, respectively, with % yield of 76% and 75%, respectively (Scheme 1).

[Co(N₄ MacL₁)Cl₂]: Reddish pink solid; Yield 78%; M.p. 147 °C; Analytical calculated for C₂₈H₂₀Cl₂CoN₄O₄; C, 55.47; H, 3.32; N, 9.24; Found; C, 54.91; H, 3.08; N, 8.82; MS(ESI), m/z: 606.02 [M+H]⁺.

[Co(N₆ MacL₂)Cl₂]: Brown solid; Yield 76%; M.p. 134 °C; Analytical calculated for C₂₆H₁₈Cl₂CoN₆O₄; C, 51.34; H, 2.98; N, 13.82; Found; C, 51.04; H, 2.51; N, 12.97; MS(ESI), m/z: 608.30 [M+H]⁺.

[Co(N₆ MacL₂)Cl₂]: Brown solid; Yield 76%; M.p. 134 °C; Analytical calculated for C₂₆H₁₈Cl₂CoN₆O₄; C, 51.34; H, 2.98; N, 13.82; Found; C, 51.09; H, 2.62; N, 13.60; MS(ESI), m/z: 608.30 [M+H]⁺.

The analytical data and physical properties

The Rast Camphor method was used to determine the molecular weights. Conductivity Bridge 305 was used to evaluate conductivity in dry DMF (dimethylformamide). The CHNS Analyzer was used to determine the percentage of the various components (C, H, and N) present: ELEMENTAR Vario EL III at STIC Kerala, the FT-IR was recorded on FT-IR; Perkin Elmer Spectrum IR Version 10.6.2 using as KBr disc India. NMR spectra were recorded in DMSO-d₆ using a JEOL400 ECZS NMR Spectrometer, and A SCIEX Triple TOF 5600 mass spectrometer was used to certify the mass spectra. The complexes in DMSO had their electronic spectra (10^-3 M, 200–800 nm) measured at room temperature using a Perkin Elmer -25 spectrophotometer. Co(II) complexes ESR spectra were captured at liquid nitrogen temperature (LNT) using a JEOL JES - FA200 ESR Spectrometer with the X and Q bands. The powder X-ray diffraction was recorded on Bruker D8 Advance Twin- Twin at STIC Kerala, India. The morphological study was examined using a JEOL JSM6510 scanning electron microscope (SEM). TGA study has determined the stability and decomposition kinetic characteristics. Thermogravimetric analyzer (Hitachi STA 7200) TGA/DTA study was carried out with N₂ flow (20 mL min^-1) and heating rate of 10 °C/min.

Results and Discussion

The biological active tetraamide macrocyclic ligands (MacL₁-MacL₃) and their corresponding complexes [Co(II)(N₄ MacL₁)Cl₂], were derived from o-phenylenediamine and aromatic dicarboxylic acid by template condensation reactions (Scheme 1). All the complexes are colored solids. All the synthesized macrocyclic ligands and cobalt (II) complexes were soluble in most of the organic solvents like benzene, methanol, DMF, and DMSO and sparingly soluble in water. They are non-electrolytic in nature since the observed molar conductance values for 10⁻³, mol L^-1 solutions in dry DMF are in the range of 13-32 ohm^-1 cm², also suggesting that the anions are coordinating with the central metal atom in these complexes. The physical properties and analytical data for all the macrocyclic ligands and cobalt (II) complexes are given in Supplementary Information (Table S1, SI). The analytical data confirmed the proposed stereochemistry of the ligands and complexes. Numerous spectroscopic, physicochemical, and biological characterizations were done on the synthesized macrocyclic ligands and the related macrocyclic complexes.

FT-IR

The absence of -NH₂ stretching vibrations of amino acids and -OH groups of dicarboxylic acids in the FT-IR spectra of macrocyclic ligands and their complexes is the first factor that conforms the synthesis of macrocyclic ligand. In the spectra of all the ligands band present in the region of 3244-3279 cm^-1 assigned to ν(NH) of the amide group. The bands in the areas of 1648-1707, 1544-1587, 1252-1275, and 628-684 cm^-1 in the IR spectra of the ligands and complexes can be attributed to the amide I. [ν(C=O)], amide II [ν(C-N) + δ(N-H)], amide III [δ(N-H), and amide IV wagging [Θ (C=O)] vibrations respectively which confirmed the closed cyclic products are present.²⁶ The complexes [Co(N₄MacL₁)Cl₂]-[Co(N₆MacL₃)Cl₂] showed a minor negative value in the (NH) band in the area 3215-3235 cm⁻¹ when compared to their ligands, which is well in keeping with the coordinated N-H stretching vibration.²⁷ This is further supported by the possibility of assigning the (M-Cl) and (M-N) vibrations to all complexes' bands in the range of 330-336 cm^-1 and 480-490 cm^-1, respectively. The complexes absorption bands are attributed to the C-H stretching and bending vibrational modes, which exist in the areas 2815-2975 and 1420-1448 cm^-1, respectively (Table 1, Figure 1).

Table 1.

Characteristic IR Spectral data of ligands (MacL₁-MacL₃) and Co(II) complexes (in cm^-1).

Compound	ν(NH)	Amide				ν(Co-N)	ν(Co-Cl)
		I	II	III	IV
[MacL1]	3244	1648	1558	1252	661	-	-
[MacL2]	3268	1669	1544	1265	665	-	-
[MacL3]	3279	1683	1567	1271	628	-	-
[Co(N4 MacL1)Cl2]	3215	1675	1571	1267	659	490	303
[Co(N6 MacL2)Cl2]	3226	1707	1580	1275	678	486	336
[Co(N6 MacL3)Cl2]	3235	1692	1587	1254	684	480	332

Figure 1.

IR spectrum (a) ligand MacL₁ (b) Complex [Co(N₄MacL₁)Cl₂].

¹H NMR of the tetraamide macrocyclic ligands (MacL_1-3)

In the ¹H NMR spectra of the macrocyclic ligands (MacL₁-MacL₃ were recorded in (400 MHz, DMSO-d₆), the peaks indicated the mode of linkage in the compounds. The ¹H NMR spectra of the precursors showed the signals of the -COOH proton and -NH₂, but were absent in the synthesized ligands, indicating the removal of the -COOH and -NH₂ protons for the formation of a macrocyclic structure. The information well accords with what the infrared data have revealed. The ¹H NMR spectra of the ligands (MacL₁-MacL₃) presented in Table 2 exposed the signals predictable for the given Scheme. From the ¹H NMR spectra of the (MacL₁-MacL₃) it is also concluded that no peak could be assigned for -OH or -NH₂ groups, confirming the formation of proposed macrocyclic ligands after the condensation.²⁸ A signal was detected in the region δ 9.10-9.60 ppm for amide protons in macrocyclic ligands.²⁹ In the spectra of the macrocyclic ligands, the multiplets appearing in the region δ 8.11-8.21 ppm and δ 6.32-6.47 ppm correspond to the 1,4-and 1,2-phenyl ring protons, respectively, while multiplets appeared in the region δ 6.40-7.76 ppm was assigned to protons of a pyridine-2,3-dicarboxylic acid moiety which have different chemical environment (Figure 2).³⁰

Table 2.

¹H NMR data (in ppm) of ligands (MacL₁- MacL₃).

Compound	(CO-NH) (m)	1,4-C6H4 (m)	1,2-C6 H4 (m)	C6H3N–
MacL1	9.32	8.11, 8.21	6.47, 6.32	-
MacL2	9.60	-	-	6.40-7.76
MacL3	9.10	-	-	6.53-7.66

Figure 2.

¹H NMR spectrum of ligand (MacL₁).

¹³C NMR

The results drawn from the FT-IR and ¹H NMR spectra were also supported by the ¹³C NMR spectral data regarding the validity of the suggested Skelton. The carbons linked to the nitrogen atoms showed a chemical shift indicative of their suggested coordination in the ligands (Table 3) (Figure 3).

Table 3.

¹³C and NMR spectral data of macrocyclic ligands (MACL_1-3).

Compound	(CO-NH)	1,4-C6H4	C6H3N-	1,2-C6H4
MacL1	169.08	115.4, 118.1	-	129.5, 133.3, 134.5
MacL2	169	-	150.8, 132.6, 128.2	-
MacL3	164	-	151.3, 131.5, 127.7	-

Figure 3.

¹³C NMR spectrum of ligand MacL₁.

Mass spectrum of the ligands (MacL_1-3) and cobalt(II) complexes

The mass spectra of ligands (MacL₁-MacL₃) and their corresponding cobalt (II) complexes were recorded. The proposed molecular formula of the ligand [(C₂₈H₂₄N₄O₄)] was confirmed by the mass spectrum of the ligand (MacL₁), which displayed a molecular ion peak at m/z 477.21 amu [M⁺] that corresponded to its molecular weight. The line (at m/z=325 amu (C₁₈H₁₄N₂O₂C₂⁺)⁺¹) in Figure 4 is called the base peak, and it is due to the commonest fragment ion. The intensity of all other peaks was measured with respect to this base peak. The other fragmentation peaks were also observed at 163(C₆H₆N₂O₂C₂⁺)⁺¹, 189(C₈H₈N₂O₂C₂⁺)⁺¹, 201(C₁₀H₈N₂O₂C₂⁺)⁺¹, 214(C₁₀H₈N₂O₂C⁺C)⁺¹, 241(C₁₃NO₂HN⁺H₁₀C₂⁺)⁺², 278(C₁₅H₁₃N₂O₂C₂⁺)⁺¹, 289(C₁₆H₁₆N₂O₂C₂⁺)⁺¹, 303(C₁₇H₁₃N₂O₂C₂⁺)⁺¹, 340(C₂₀H₁₄N₂O₂C₂⁺)⁺² and 371(C₁₈H₁₄N₂O₄C₂⁺)⁺¹ amu. The intensity of these peaks confirmed the stability of various fragments. In the mass spectra of all other ligands and their corresponding complexes MacL₂, MacL₃, [Co(N₄MacL₁)Cl₂], [Co(N₆MacL₃)Cl₂], and [Co(N₆MacL₃)Cl₂] show the molecular ion peak at 481, 481, 607,609 and 609, respectively. This is comparable to the molecular mass of compounds.

Figure 4.

Mass spectrum of (a) ligand (MacL₁) (b) Complex [Co(N₄ MacL₁)Cl₂].

Electronic spectral data

The observed magnetic moment of Co(II) complex at room temperature in the range of 4.80–4.87 B.M. corresponds to three unpaired electrons, which is substantially higher than the spin-only value, 3.87 B.M. The spin-orbit coupling may cause this deviation from the spin-only value. The electronic spectra of six coordinated cobalt (II) complexes were recorded using DMSO as solvent. For high spin d⁷ the bands are observed in the at 9663-12,620 (ν₁), 13,242-13,979 (ν₂) 19,498-24,701 cm^-1 (ν₃) and 26,298-32,359 cm^-1 (ν₄). The electronic spectrum of the Co(II) complex exhibits these bands, and the position of the bands is comparable to the characteristic features of distorted octahedral geometry and having the D_4h Symmetry. These bands are corresponding to the following transition: ⁴T_1g(F) → ⁴T_2g(F) (ν₁), ⁴T_1g(F) → ⁴A_2g(F) (ν₂), ⁴T_1g(F) → ⁴T_2g(P) (ν₃), respectively.³¹ The fourth band may be due to charge transfer (Table 4, Figure 5).

Table 5.

(a) Thermal Parameters of tetraamide macrocyclic ligands and their Cobalt (II) complexes by applying Coats and Redfern method at a heating rate 10 °C/ min.

Compound	R (From Graph)	TMAX. (From DTG)	(Ea) (J/mol)	(A) (min.-1)	(∆G) (KJ/mol)	(∆H) (KJ/mol	(∆S) (J/mol)
MacL1	0.70858	438.08	12.99	0.002312	130.780	-3.628	-290.31
MacL2	0.99354	510.00	36.48	0.004827	148.059	-4.203	-282.10
MacL3	0.99618	523.00	54.90	0.006879	148.102	-4.185	-282.19
[Co(N4MacL1)Cl2	0.76722	453.00	7.69	0.001271	137.638	-3.758	-295.54
[Co(N6MacL2)Cl2	0.67599	463.00	5.49	0.000872	142.197	-3.836	-298.55
[Co(N6MacL3)Cl2	0.54652	478.93	10.00	0.001480	145.107	-3.971	-294.68

(b) Thermal Parameters of tetraamide macrocyclic ligands and their Cobalt (II) complexes by applying the Ozawa-Flynn-wall method at a heating rate 10 °C/ min.
Compound	R (From Graph)	TMAX. (From DTG)	(Ea) (J/mol)	(A) (min.-1)	(∆G) (KJ/mol)	(∆H) (KJ/mol	(∆S) (J/mol)
MacL1	0.88182	438.08	21.86	0.0039	128.891	-3.619	-286.00
MacL2	0.99521	510.00	44.46	0.00596	148.943	-4.195	-300.27
MacL3	0.99723	523.00	63.60	0.0080	151.588	-4.284	-281.65
[Co(N4MacL1)Cl2	0.95704	453.00	17.87	0.00310	125.619	-3.748	-269.03
[Co(N6MacL2)Cl2	0.90124	463.00	11.55	0.001839	139.562	-3.837	-293.14
[Co(N6MacL3)Cl2	0.84213	478.93	20.43	0.003047	160.606	-3.961	-327.03

Table 4.

Electronic spectral data and magnetic moments of the Cobalt (II) macrocyclic complexes.

Compound	μeff. (BM)	λmax (cm-1)
[Co(N4 MacL1)Cl2]	4.82	9663, 13242, 19498, 26298
[Co(N6 MacL2)Cl2]	4.83	10948, 13575, 21575, 28705
[Co(N6 MacL2)Cl2]	4.87	12642, 13979, 24701, 32359

Figure 5.

UV-vis spectrum of Co(II) macrocyclic complex (1b).

The EPR spectra of [Co(N₄MacL₁)Cl₂]- [Co(N₆MacL₃)Cl₂] complexes

EPR spectroscopy can be used to investigate the paramagnetic properties of cobalt (II). In cobalt (II), due to the very fast spin-lattice relaxation time, the EPR measurements are possible only at liquid nitrogen temperature to avoid broadening the line at higher temperatures.³² The EPR spectra of the tetraaza macrocyclic cobalt(II) complex were recorded in dry DMF solution. The g|| and g⊥ values are found to be 2.12 and 1.78, respectively Figure 6. The spin-only value for g is 2.0023, and the difference in g values from the spin-only value may be due to the high angular momentum contribution.

Figure 6.

EPR spectrum of Co(II) macrocyclic complex.

Thermogravimetric analysis

The thermogravimetric (TG) analysis of macrocyclic ligands (MacL₁-MacL₃) and their corresponding cobalt (II) complexes was carried out with N₂ flow (20 mL/min) and heating rate of 10 °C/min in the temperature range 45-650 °C. Thermogravimetric curve is given in Figure 7. The thermogravimetric curve for ligand (MacL₃) shows the complete decomposition of 100% mass loss in one step. The [Co(N₆ MacL₃)Cl₂] TG curve is a three-step breakdown with a mass loss of 70%. The fact that there is no weight loss before 180 °C implies that the complex is stable up to 180 °C in the absence of coordinated water molecules. First mass loss of 11.51% occurred between 180 and 220 °C, indicating the breakdown of the Cl₂ group. The 2^nd mass loss of 26.09% was discovered in the temperature range of 250-320 °C, showing the breakdown of the C₁₀H₆N₂ group. In the range of 510-600 °C, C₁₆H₆N₄O₄ loses 52.82 % of its weight throughout the decomposition.³³

Figure 7.

Thermogravimetric analysis of ligand (MacL₃) and Complex [Co(N₆ MacL₃)Cl₂].

The kinetic decomposition parameters in TGA have been calculated using two kinetic approaches: the COATS AND REDFERN method and the Ozawa-Flynn Wall (OFW) method. Furthermore, the thermodynamics parameter such as Arrhenius factor, enthalpy change, Gibbs free energy, and entropy change has been calculated.³⁴

Kinetic Parameters

The kinetics of macrocyclic compounds was studied by data obtained from TGA. The rate constant (k) and the degree of conversion (α), which both relate to thermal degradation, are two functions that describe the rate of reaction:

$$\frac{d\alpha }{\mathit{dt}}=k(T)f(\alpha )$$ 1

In the above equation, absolute temperature (T) in Kelvin, the time duration of a reaction (t) in minutes, the degree of conversion is (α), the differential form of the reaction model is f(α) is, and the reaction rate constant is k(T) is, α can be written as:

$$\alpha =\frac{m_o-m_t}{m_o-m_{\infty }}$$ 2

where m₀ represents the initial mass, m_t represents the mass with respect to time, and $m_{\infty }$ represents the sample's final mass. The Arrhenius equation can be used to express the rate constant k(T):

$$k(T)=A{e}^{\frac{{-E}_a}{\mathit{RT}}}$$ 3

where R= gas constant, E_a = activation energy, and A = pre-exponential factor. The equation for the rate of reaction with linear heating rate can be found by combining equations (1) and (2)

$$\frac{d\alpha }{\mathit{dt}}=\frac{A}{\beta }{e}^{\frac{{-E}_a}{\mathit{RT}}}f(\alpha )$$ 4

Different iso-conversional kinetic approaches are developed based on Equation (4). These kinetic methods used different temperature integral approximations to calculate the kinetic parameters. The following can be stated as the generic equation that applies to all kinetic methods:

$$ln\left(\frac{\beta }{T^B}\right)=constant-C\left(\frac{E_a}{\mathit{RT}}\right)$$ 5

In this study, the most popular kinetic methods, Ozawa-Flynn-Wall (OFW), Coats and Redfern, are utilized to compute the activation energy of the tetra azamacrocyclic ligands and their cobalt(II) complexes.³⁵ The Doyle approximation is the foundation of the OFW technique, which is based on the idea that the variables (A, g(α), and Ea) are independent of the absolute temperature (T), and it is written as follows:

$$ln\beta =ln\left(\frac{{\mathit{AE}}_a}{g\left(\alpha \right)R}\right)-5.331-1.052\left(\frac{E_a}{\mathit{RT}}\right)$$ 6

where g(α) is an ideal reaction model's integral form. The Coats and Redfern approximation is the foundation of the Coats and Redfern method, which is defined by the equation given below:

$$ln\left(\frac{\beta }{T^2}\right)=ln\left(\frac{\mathit{AR}}{E_a}\right)-\frac{E_a}{\mathit{RT}}$$ 7

Analysis

The overall activation energy was found to be around 107.953 kJ/mol. The activation energy obtained through Kissinger method is close to the activation energies obtained from OFW, KAS, and for conversion of 0.1 and 0.2. After the kinetic study, the following equation calculated the thermodynamic parameters such as Arrhenius factor, Enthalpy change, Gibbs free energy, and Entropy change.

$$A=\frac{\beta \times E_a\times e^{\left(\frac{E_a}{{\mathit{RT}}_{\mathit{max}}}\right)}}{R{T}_{\mathit{max}}^2}$$ 8

$$\mathrm{\Delta }G=E_a+R\times T_{\mathit{max}}ln\left(\frac{K_B{T}_{\mathit{max}}}{\mathit{hA}}\right)$$ 9

$$\mathrm{\Delta }H=E_a+R{T}_{\mathit{max}}$$ 10

$$\mathrm{\Delta }S=\frac{\mathrm{\Delta }G-\mathrm{\Delta }H}{T_{\mathit{max}}}$$ 11

where k_B is Boltzmann’s constant (1.38×10^-23 Js^-1), and h is Planck constant (6.626×10^-34Js^-1). The result for Arrhenius the Factor uses different kinetic methods for different heating rates. Further, the result of ∆H, ∆G, and ∆S using the different kinetic methods for a 10 °C/min heating rate. The positive values of ∆H and ∆G show that the thermal decomposition of MacL₂ is an endergonic, non-spontaneous, and unfavourable process (Figures 8, 9).

Figure 8.

The linear fitted curve obtained through the Coats-Redfern method at 10°/min heating rate of (MacL₁-MacL₃).

Figure 9.

The linear fitted curve was obtained through the Ozawa -Flynn-wall method at 10°/min. The heating rate of (MacL₁-MacL₃) (Table 5).

X-ray powder diffraction

The X-ray diffraction analysis of the tetraamide macrocyclic ligands (MacL_1-3) and their complexes [Co(N₄MacL_1-3)(Cl)₂] was obtained in the range of 2θ =10-80° Figure 10. Among all the compounds, the ligands in the diffractogram of ligands show sharp peaks, indicating their crystalline nature. Whereas in the diffractogram of all the cobalt (II) complexes, peaks expansion and less intensity suggest the semi-crystalline or amorphous nature of the complexes. The average crystalline size of the ligands and their corresponding cobalt (II) complexes were calculated by using Scherrer’s formula. The highest intensity peaks were at 2 θ = 22.8167° and 21.7313° for ligand and complex. The presence of metal in the ligand environment is confirmed by the powder X-ray diffraction.³⁵

$$D=k\frac{\lambda }{\beta \mathit{cos}\theta }$$

Where the λ=wave length of the X-rays used, θ = diffraction angle, β= (FWHM) full width at half maximum of the peak position, and K is Scherrer's constant. The ligand and complexes computed average crystalline size were discovered to be 51.34 and 45.68 nm, respectively. The contraction of Cobalt(II) macrocyclic complexes is a sign of macrocyclic ligand coordination with cobalt metal, which results in a contraction of the ligand in complexes. However, many attempts have been tried to form a single crystal but have not been successful in growing the single crystal. The powder X-ray diffraction study shows that the ligands, which have four coordination sites, function as tetradentate chelating agents. Additionally, a hexacoordinated environment for cobalt has been proposed since the Cl^- anions coordinated to the cobalt atom.

Figure 10.

(a) The PXRD diffractogram of ligand (McL₁) (b) The PXRD diffractogram of Complex [Co(N₄MacL₁)Cl₂.

SEM Analysis of Tetraamide Macrocyclic ligands and Cobalt (II) Complexes:

The surface morphology of the tetraamide macrocyclic ligands (MacL_1-3) and their complexes [Co(N₄MacL_1-3)(Cl)₂] have been examined using scanning electron microscope (SEM) analysis, and it is observed that the SEM image of the ligand MacL₁ revealed highly pure and small roads like structure revealing its crystalline nature in Figure 11 (a). While the irregular rough surface clouds indicated the SEM image of complexes the semi-crystalline character of the complexes in the SEM image. From the scanning electron microscope analysis, the change in surface morphology of the compounds also confirms the coordination of metal ions with the donor atom of ligands. The average crystalline size was found to be 50 and 45 nm for the ligand and complex, respectively the contraction in the size of complexes compared to ligand was is in good agreement with the results from the powder X-ray diffraction analysis.³⁶ Although attempts were made to grow single crystals many times but failed.

Figure 11.

SEM image of (a) ligand MacL₁; (b) complex [Co(N₄ MacL₁)Cl₂].

Electrochemical study

The oxidation levels of the metal centers in the cobalt(II) complexes were verified using cyclic voltammetry. In order to take into account the spectral and structural changes that come along with electron transfer, the electrochemical characteristics of metal complexes, particularly those with nitrogen donor atoms were explored. Using an electrochemical analyzer, the redox behaviour of cobalt(II) complexes in DMSO was investigated. Glassy carbon served as the working electrode. Only in relation to the metal core are any of these compounds electroactive. Cobalt(II) is oxidized to cobalt(III) on the cyclic voltametric time scale, and the detection of a single, strong peak that is not displayed by the matching ligand shows that the metal center is symmetrically situated in one complex. When compared to the Ag/AgCl, CI electrode, the cobalt (II) complexes exhibit both metal- and ligand-centered electrochemistry in the potential range of 1.7 V. Peak-to-peak separation of all the complexes is greater than 100 mV, indicating a single, nearly reversible oxidation process. They all display redox potentials between 1.0 and 1.25 V. The cathodic and anodic peak heights (Ipc and Ipa) are identical in all cobalt (II) complexes. The Co(III)/Co(II) pair is responsible for the cobalt (II) complexes' electrochemical behaviour. The positive potential shows that the ligand is tightly linked to a metal in a lower oxidation state (Figure 12).⁴⁵

Figure 12.

Cyclic voltammogram of complex [Co(N₄ MacL₁)Cl₂].

Molecular modeling

The software Avogadro 4.0 was used to perform the geometry optimization. Figure 13 depicts the fully optimized molecular structure of the ligands and Cobalt (II) macrocyclic complexes. The bond lengths of the macrocyclic ligands and the cobalt (II) complexes are compared, with the bond angles form the Cobalt (II) complexes listed in Table 6 and Table 7, respectively. The proposed structure is verified by the bond angle values, which are approximately equal to the octahedral geometry of the complexes. The C=O and C-N bond lengths of the ligand are 1.2051 and 1.39048, respectively. In complexes, these C=O (1.2274 Å) and C-N (1.42685 Å) bond lengths become longer, indicating coordination of the ammine group via the N atom. Selected bond lengths of complexes are Co-N and Co-Cl, confirming the distorted octahedral geometry of the Cobalt(II) complexes.³⁷

Figure 13.

Optimized structure of ligand (MacL₁) and the Co(II) macrocyclic complex. Color code: blue-N; red-O; grey-C; white-H; green-Cl; peach-Co.

Table 6.

The calculated bond lengths (Å) of the ligand and the complexes

Compound	C=O	C-N	N-H	Co-N	Co-Cl
MacL1	1.2051	1.39048	1.01095	-	-
MacL2	1.21695	1.37088	1.02246	-	-
MacL3	1.21309	1.38103	1.01201	-	-
[Co(N4 MacL1)Cl2]	1.22746	1.42685	1.02993	1.98771	2.24491
[Co(N6 MacL2)Cl2]	1.22669	1.43265	1.03269	2.18037	2.23325
[Co(N6 MacL3)Cl2]	1.22627	1.42447	1.03063	2.01901	2.25196

Table 7.

Selected bond angles (°) for the Cobalt(II) complexes.

Atom connectivity	Bond angles (°)	Atom connectivity	Bond angles (°)
N-Co-N	86.7523	N-Co-Cl	89.2919
N-Co-Cl	179.6951	N-Co-Cl	90.6023
N-Co-Cl	89.6435	N-Co-Cl	90.4243
N-Co-N	179.7675	N-Co-Cl	89.5951
N-Co-Cl	90.7574	Cl-Co-Cl	179.8913
N-Co-Cl	89.3485
N-Co-N	86.8228

Density Functional Theory (DFT) Study

The DFT calculations have been carried out by using Avogadro 4.0 with the ORCA program. The frontier molecular orbitals of a molecule are its highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO), and they play a significant role in regulating molecular chemical reactions. It is also confirmed from the previous investigations that the gap in energy (ΔE) between the HOMO-LUMO is explained the stability of the molecule. The energy gap (ΔE) of these frontier molecular orbitals for macrocyclic ligands and complexes are used to predict the stability and strength of these compounds are obtained from DFT calculations. The Koopman’s Theorem specifies that the energy of HOMO-LUMO corresponds to ionization energy (IE) and electron affinity (EA), respectively. According to Koopman's Theorem, calculations of HOMO-LUMO energies values presented in Table 8 can be used to determine quantum chemical parameters^38–41 for example, electrophilicity index (ω), chemical potential (Pi), electronegativity (χ), softness (S) and hardness (η).

Table 8.

Theoretically calculated quantum parameters of the macrocyclic ligands and complexes.

Parameters	(MacL1)	(MacL2)	(MacL3)	1(a)	1(b)	1(c)
EHOMO (eV)	-8.687	-8.438	-8.617	-15.25	-11.44	-17.15
ELUMO (eV)	-1.135	-1.223	-1.017	-9.535	-9.238	-15.28
∆E (eV)	7.552	7.215	7.6	5.715	2.208	1.87
IE (eV)	8.687	8.438	8.617	15.25	11.446	17.15
χ (eV)	4.911	4.830	4.817	12.3925	10.342	16.2195
η (eV)	3.776	3.607	3.8	2.8575	1.104	0.935
S (eV)-1	0.132	0.138	0.131	0.1749	0.4528	0.5347
ω (eV)	3.19	3.23	3.053	26.8721	48.4402	140.68
Pi	-4.911	-4.830	-4.817	-12.392	-10.342	-16.219

The HOMO and LUMO energy gap for ligands is found to be 7.552 eV, 7.215 eV, 7.6 eV. The energy gap is reasonably reduced in complexes, respectively (Figure 14). HOMO and LUMO are distributed throughout the entire π -moiety of the ligand. HOMO is distributed across the entire π-moiety, and LUMO is distributed over cobalt metal in complexes.

Figure 14.

(a) The HOMO-LUMO energy gap of the tetraamide macrocyclic ligands (MacL_1-3). (b) The HOMO-LUMO energy gap of the Cobalt(II) complexes.

Antimicrobial assays

Antibacterial activity

The Inhibition Zone Technique was used to assess the bacterium-fighting ability of the bacteria.²⁵ In this approach, 5 mm diameter Whatman No. 1 paper plates with the supplement agar medium (composed of 5 g peptone, 5 g meat extract, 5 g NaCl, 20 g agar, and 1000 mL distilled water) were used. The Petri dish was filled with pipetted agar media. 5 mL of heated seeded agar was added after it had set. The molten agar was cooled to 40 °C to create the seeded agar, and the bacterial solution was then added. In dimethylformamide, the compounds were dissolved at 80 ppm concentrations. We soaked 5 mm diameter paper discs of Whatman No. 1 filter paper in these solutions at 80 ppm concentrations. The petri plates were dried and positioned at the proper distance on the medium previously seeded with organisms in petri plates. For 24 h, the petri plates were kept in an incubator set at 30±2 °C. The precise measurement in mm of the inhibitory zone that subsequently formed around each disc containing the test chemicals. The synthesized compounds are tested against Bacillus subtitles (+) and Escherichia coli (-) by taking Ampicillin as standard at 80 ppm concentration. The results are shown in Table 10. The results of antibacterial screening show the maximum inhibition by Cobalt (II) complexes compared to ligands because complexes formed as a result of chelation become more lipophilic. When chelating, the polarity of the metal ion disappears, which causes the orbitals of the ligands to overlap and the metal's positive charge to be shared with donor groups. The invasion of complexes into the lipid membranes is efficiently induced by the delocalization of electrons on the chelate ring, which inhibits microbial growth.

Table 10.

The binding affinity (kcal/mol) of ligands (MacL_1-3) with different proteins.

Active Protein (PDB ID)	Binding Affinity (kcal/mol)
	MacL1	MacL2	MacL3
Sars-Cov-2 (2ajf)	-10.4	-9.8	-10.0
C. Albicans (3dra)	-10.7	-10.1	-10.4
X. Campestris (3row)	-10.8	-10.7	-10.4
E. Coli (3t88)	-11.2	-14.0	-9.9
B. subtilis (5h67)	-11.7	-10.5	-10.6

Antifungal activity

The Radial Growth Method²⁵ was used to assess the antifungal activity of the ligands and their complexes with divalent cobalt using Czapeks agar medium, which is (composed of 20 g of starch, 20 g of agar-agar, 20 g of glucose, and 1000 mL of distilled water). The Solutions of the test ligands and Cobalt(II) complexes at 80 ppm concentration in dimethylformamide were prepared. The agar medium was then emptied into the Petridis’s, and by using an inoculum needle, the fungi were placed on the medium. Petri plates wrapped in polythene and placed in an incubator set at 28±2 °C with a few drops of alcohol. By measuring the fungal region's diameter, the fungus's linear growth was determined. The Candida albicans organisms used in these investigations for all synthesized compounds the results are in Table 9 (Figure 15)

Table 9.

Antibacterial and antifungal studies of ligands and their Cobalt(II) complexes.

Antibacterial and antifungal studies of ligands and their Cobalt(II) complexes
Compound	Antibacterial activity (Zone inhibition diameter in mm)		Antifungal activity (Zone inhibition diameter in mm)
	E Coli	Bacillus subtilis	Candida albicans
[MacL1]	08	10	09
[MacL2]	10	12	12
[MacL3]	09	08	08
[Co(N4 MacL1)Cl2]	13	12	15
[Co(N6 MacL2)Cl2]	15	17	20
[Co(N6 MacL3)Cl2]	10	12	11
Ampicillin	20	20
Amphotericin-B	-	-	22

Figure 15.

Zone inhibition diameter(mm) data of macrocyclic ligands and complexes. (a) Antibacterial activity; (b) Antifungal activity.

Antifungal activity results show that the ligand Macl₂ is more active than macL₁ and MacL₃, and all the complexes [Co(N₆ MacL₂)Cl₂] are more active against C. Albicans.

The coordination of metal to the ligands influences the magnetic property, conductance, and solubility. The high solubility of the complexes, which makes it easy to aggregate in bacterial and fungal cells and activate enzymes, is another explanation for their enhanced activity. The antimicrobial potential of Schiff base macrocyclic complexes of Transition metal has been reported by Chandra et. al.¹⁹ However, In the present investigation, N₄ tetraaza macrocyclic complexes of Co(II) show superiority over these complexes.

Molecular docking study

The Molecular Docking study is an effective Computer-aided drug designing method for predicting the binding association and binding site between the drug and active site of bio-molecules (Proteins and CT-DNA). Docking also played a major role in advancing therapeutically significant molecules for over four decades.⁴² In this study, molecular docking was used to examine the particular groove binding characteristics and provide useful details about the way drugs bind in the active site. Molecular docking is an important computational technique in structural biochemistry and computer-aided drug design. Molecular modeling studies were carried in order to validate the results of our studies.⁴³

Discovery studio with Auto Dock VinaPyRx software was used to assess the biological importance of synthesized ligands.⁴⁴ Tetra amide macrocyclic ligands were docked with various receptor protein targets, including E. coli (3t88), C. albicans (3dra), X. campestris (3row), SARS-COV-2 (2ajf), and B. subtilis (5h67). Higher negative binding energy means more affinity of the ligand for receptors, which is the binding affinity utilized to select the best-docked structure from the output. The binding modalities of proteins amino acids are explained using 2D plots. Within the docking compounds, typical hydrogen bonds were discovered, and several hydrophobic non-polar interactions were found between the aromatic carbons of the macrocyclic ligands and the amino acids of the active protein.

The docking site on the protein targets was defined and then run with pyrx AutoDockVina was performed with all ligand structure. The best pose for each run was saved. The interactions of ligands (MacL_1-3) with protein sars-cov-2 (PDB: 2AJF). The interactions between the protein and ligand, including hydrogen bonding and hydrophobic interactions, were analyzed in Figure 16, and out of these ligands, MacL₁ shows a good binding affinity. The MacL₁ ligand was docked with the active site of the protein sars-cov-2 and showed various interactions. A conventional hydrogen bonding interaction (green color) is observed in phe390, asn390, and tyr394 with a bond length of 3.08 Å. Some pi-pi stacked interactions were also observed in phe40 and his401. The best-docked ligand MacL₁ selected has a binding score of -10.4 kcal/mol with binding site of SARS-Cov 2 (2ajf) protein, as shown in Figure 16.

Figure 16.

(a) 2D interaction of ligand MacL₁ with protein 2ajf. (b) 3D image of ligand MacL₁ in the active site of protein 2ajf.

The interactions of ligands with protein C. Albicans (PDB: 3DRA). The interactions between the protein and ligand, including hydrogen bonding and hydrophobic interactions, were analyzed in Figure 17, and out of these ligands, MacL₁ shows a good binding affinity. The MacL₁ ligand was docked with the active site of the protein C. Albicans and showed various interactions. The conventional hydrogen bonding interaction in asn435 with a bond length of 2.93 Å is observed (Light green color in 2D plot). Some pi-pi T-shaped interactions were observed in trp150, pi-sigma with thr154, pi-alkyl ILE434, pi-anion asp153, and van der Waals interactions with arg491 were observed. The best-docked ligand MacL₁ selected has a binding score of -10.4 kcal/mol with a binding site of 3dra protein as shown in Figure 17.

Figure 17.

(a) 2D interaction of ligand MacL₁ with protein 3dra; (b) 3D image of ligand MacL₁ in the active site of protein 3dra.

The interactions between the protein X. Campestris (PDB: 3ROW) and ligand, including hydrogen bonding and hydrophobic interactions, were analyzed, and out of these ligands, MacL₁ shows a good binding affinity, as in Figure 18. The MacL₁ ligand was docked with the active site of the protein X. Campestris and showed various interactions. The van der Waals interactions is observed in asn627, pro630, glu495, and ieu102. The interactions were also observed in pi-alkyl ILE434 and pi-anion asp153. The best binding mode of docked MacL₁ ligands with the binding site of 3ROW protein is shown in Figure 18, with a binding affinity of -10.8 kcal/mol.

Figure 18.

(a) 2D interaction of ligand MacL₂ with protein 3row; (b) 3D image of ligand MacL₂ in the active site of protein 3row.

The interactions between the protein E. Coli (PDB: 3T88) and ligand, including hydrogen bonding and hydrophobic interactions, were analyzed, and out of these ligands, MacL₂ shows a good binding affinity, as in Figure 19. The MacL₂ ligand was docked with the active site of the protein E. Coli and showed various interactions. A conventional hydrogen bonding interaction (green color) is observed in thr1518 and thr394 with a bond length of 2.32 Å. pi-cation and pi-anion interactions were observed in arg341, asp1511, and asp387, arg1465, respectively. Some pi-alkyl interactions were also observed in ala1462, arg390, arg1514, and ala338. The best-docked ligand MacL₂ selected, has a binding score of -14 kcal/mol with binding site of 3t88 protein, as shown in Figure 19.

Figure 19.

(a) 2D interaction of ligand MacL₂ with protein 3t88; (b) 3D image of ligand MacL₂ in the active site of protein 3t88.

The interactions between the protein B. subtilis (PDB: 5H67) and ligand, including hydrogen bonding and hydrophobic interactions, were analyzed, and out of these ligands, MacL₂ shows a good binding affinity, as shown in Figure 20. The MacL₂ ligand was docked with the active site of the protein B. subtilis and showed various interactions. A pi-donor hydrogen bonding interaction is observed in gly144 with a bond length of 3.08 Å. pi-cation, and pi-anion interactions were observed in glu148, lys145, and glu163, respectively. Some pi-alkyl interactions were also observed in lys135 and leu168. The best-docked ligand MacL₂ selected has a binding score of -11.7 kcal/mol with a binding site of 3dra protein, as shown in Figure 20.

Figure 20.

(a) 2D interaction of ligand MacL₁ with protein 5h67; (b) 3D image of ligand MacL₁ in the active site of protein 5h67.

Conclusions

Novel biologically active tetraamide macrocyclic ligands (MacL_1-3) and their Cobalt(II) complexes were synthesized by templet condensation reactions. Analytical and spectroscopic methods characterized the prepared ligands (MacL^1-3) and their complexes. FT-IR and ¹H NMR spectroscopic results clearly indicate that the nitrogen atoms of amide group are coordinated to the Co(II) metal atom. The oxidation state was explained by an electrochemical study. The proposed structure of the synthesized Cobalt (II) macrocyclic complexes were characterise with has various spectroscopic analysis that confirmed octahedral geometry. DFT calculations of tetraaza macrocyclic ligands and their cobalt (II) complexes have been calculated and agreed with experimental results. The experimental results and molecular docking investigations are relevant because ligands can be effective antibacterial agents against known microbes. It has been determined how well (MacL₁- MacL₃) ligands and their Cobalt(II) macrocyclic complexes inhibit the growth of bacterial and fungal strains. According to the study, Cobalt (II) complexes are more active than macrocyclic ligands. All Cobalt(II) macrocyclic complexes have good activity against the B. subtilis.

Supplementary Information

Below is the link to the electronic supplementary material.

Declarations

Conflict of interest

The authors declare no conflict of interest.

Supplementary Information (SI)

Supplementary information associated with this article is available at www.ias.ac.in/chemsci.