Ball milling: a simple and efficient method for quantitative solvent-free synthesis of new potential bioactive Ni(II) and Co(II) complexes

ABSTRCT

A simple and efficient technique for quantitative solvent-free synthesis of Ni (II) and Co (II) complexes by using ball milling as a green strategy is studied. The isolated solid compounds will be characterized using micro-analytical, thermo-gravimetric, magnetic and spectroscopic studies. Theoretical studies were carried out by applying DFT theory to predict the optimized configuration for all isolated solid complexes. Moreover, the in vitro antimicrobial activity assay versus fungi and bacteria via MIC technique has higher potency of Ni (II) than Co (II). The in vitro cytotoxicity of human muscle rhabdomyosarcoma (RD) via MTT technique and the antioxidant by DPPH free radical scavenging has higher inhibitory effect on complexes than free ligand. The cyclic voltammetry of Co(II) in absence/presence of H₂BHNH was investigated to realize the significant effect of complex formation on the electrochemical manners of Co(II).

KEYWORDS:

• Ball milling
• DFT
• cytotoxic activity
• antioxidant activity
• cyclic voltammetry

Introduction

Cancer is one of the most feared diseases in the world today because it is not easy to treat; this is due to cancer that comes as a result of uncontrolled multiplication of modified normal cell of human. So, the potential way of cancer treatment is drug therapy. Whereas, the drugs used to battle cancer join one of two wide types. The first is cytotoxic drugs in which the cell is killed and the second is cytostatic in which the cell stabilizes the drugs. Both types cause a decrease in the tumor size because cancer cell has a high humanity rate, which only preventing them from isolating will cause a decrease in the inhabitants [1–4].

Metals complexes are fundamental cellular components chosen to function in numerous essential biochemical processes for living organisms. Generally, metals have unique characteristics that involve reactivity toward organic substrates, variable coordination modes, and redox activity. For these reasons, the design of special coordination complexes, whichever as drug or pro-drug, is considered the main target to get a powerful tool in cancer diagnosis [5–8]. Based on the interface between molecular bioinorganic chemistry and biological system, the design of coordination complexes for cancer treatment in the current research is extensive in scope, targets multiple cellular and biological properties of many types of tumor. Recently, the improvement of anticancer drugs stimulated from cytotoxicity toward the design of selective agents, which act on definite cellular targets [9–11].

So according to the previous facts, Ni (II) and Co(II) complexes with high biological efficiency will be prepared in an efficient and environmentally friendly way, which is to grind the reactors using ball milling technique as a green approach [12,13]. The current advance in nanotechnology led to the improvement of new metallic nanoparticles that eventually increase the environmental hazards. The main aim is to minimize the hazard of synthetic procedures, which are associated with derivative compounds and chemicals used.

Experimental

Apparatus

The apparatus used to propose the configuration for prepared compounds was displayed in Scheme 1S.

Synthesis of H₂BHNH and its metal complexes

The Schiff base ligand was formed as displayed in Scheme 1. Also, the waste-free and facile solid state ball milling technique was used to prepare Ni (II) and Co (II) complexes as a green strategy of chemistry in pure state with high yield (99%) as shown in Table 1.

Scheme 1. The outline of the synthesis of H₂BHNH and its Ni(II) & Co(II) complexes

Conformational calculations

The quantum chemical parameters were estimated by applying DMOL3 program [14] in Materials Studio package [15]. The configuration of produced ligand and its metal complexes was proposed by applying Density Functional [16].

Antimicrobial activity assay

In microbiology, the MIC technique was generally the initial point for larger pre-clinical evaluations of novel antimicrobial agents [17,18]. So, for drug discovery, the first step is the screening of a records drug candidate for MICs versus tested bacteria or fungi as shown in Scheme 2S.

In vitro cytotoxicity assay

The cytotoxic effect of tested compounds on Rhabdomyosarcoma cell lines (RD) were performed by determining the absorbance of each well at 570 nm via applying ELISA (Enzyme-linked immunosorbent assay) microplate reader (Thermo Electron Corporation Multiskan-EX reader) [19] and mean optical density for each concentration was measured (Scheme 3S). The effect is expressed as % cell viability as shown in this relation:

$\mathrm{\%}CellViability=OD\left(Test\right)/OD\left(Control\right)x100$

Antioxidant Activity

The free radical-scavenging activity of tested compounds was determined by applying Blois method [20] using DPPH as a stable radical scavenging (Scheme 4S). The ability to scavenge the DPPH radical was determined by applying this equation [21]:

$\mathrm{\%}DPPHscavengingactivity=A_{control}-A_{sample}/A_{control}x100$

A_control: The absorbance of control

A_sample: The absorbance of the sample

Cyclic voltammetry

The cyclic voltammetry was estimated for Co(II) in absence/presence of ligand by using three types of electrodes which are reference electrode (Ag/AgCl/KCl), the glassy carbon working electrode (GCWE) and an auxiliary electrode (platinum wire) that immersed in 30 ml 0.l M KCl as supporting electrolyte which connect with potentiostat of the type DY 2100.

Results and discussion

General characteristics

The isolated solid complexes of Ni (II) and Co (II) were physically and analytically characterized to set up their chemical formula and also have non-hygroscopic nature. The non-conducting aspect was proposed for Ni (II) and Co (II) complexes, owing to molar conductivity values for 10⁻³M are 4, 7 Ω⁻¹cm²mol⁻¹, respectively [22].

¹H NMR and IR spectra studies

The ¹H NMR spectrum of H₂BHNH in DMSO is displayed in Figure 1 have two signals at 11.98 and 11.30 ppm assignable to the protons of (OH)_naphthoic & (NH) correspondingly. The intramolecular hydrogen bonding was suggested to be due to the appearance of (OH)_naphthoic signal at high value downfield from TMS. Also, the aromatic and (-N = C-H) protons have multiplet signals displayed in 7.34–8.47 ppm range.

Figure 1. ¹H NMR spectrum of H₂BHNH in DMSO

In the IR spectrum of H₂BHNH, the bands of υ(C = N) & υ(C = O) were displayed at 1622 & 1646 vibrations [23], correspondingly (Table 2). The bands of υ(OH)_naphthoic & υ(NH) were observed at 3449 & 3271 cm^−1, respectively [23]. The intramolecular hydrogen bonding was suggested to be due to the existence of υ(OH) band at 3449 cm⁻¹. Also, there is another confirmation for the existence of intramolecular hydrogen bonding (O-H··O) which is the presence of broad weak bands in the 1900–2050 and 2100–2230 cm⁻¹ regions [23].

Table 1. Analytical and physical data of H₂BHNH and its metal complexes

No.	Compound	(E. F.) (F. Wt.)	Yield	M.P. (°C)	%Found (Calculated)			Λm*
			%		C	H	M
1	H2BHNH	C18H14O2N2 (290.324)	78	260	74.41 (74.46)	4.91 (4.86)	--	-
2	[Ni(HBHNH)2].2H2O	NiC36H30O6N2 (673.374)	99	285	64.11 (64.21)	4.48 (4.49)	8.83 (8.71)	4
3	[Co(HBHNH)2].2H2O	CoC36H30O6N2 (673.594)	99	285	64.15 (64.19)	4.51 (4.49)	8.78 (8.75)	7

* In DMSO (ohm⁻¹cm² mol⁻¹)

Table 2. Most important IR spectral bands of H₂BHNH and its metal complexes

Compound	υ(OH)naphthoic	υ(NH)	υ(C = O)	υ(C = N)	υ(C-O)naphthoic
1	3449	3271	1646	1622	1268
2	-	3207	1630	1608	1282
3	-	3116	1631	1602	1290

In IR spectra of Ni(II) as well as Co(II) complexes the H₂BHNH behaved as a mononegative tridentate ligand coordinating via (C = N)_azo, (C = O) & (OH)_naphthoic with the displacement of the hydrogen atom from it. This manner of complexation was recommended depending on; (i) the absence (OH)_naphthoic group; (ii) the lower shift of υ(C = O) as well as υ(C = N) and (iii) the appearance of new bands at (508, 511) & (416, 430) cm⁻¹ that ascribed to υ(M-O) & υ(M-N) [23], correspondingly.

Electronic spectra, magnetic properties and ligand field parameters

The UV-visible data in DMSO, magnetic moments and ligand field parameters of Ni(II) & Co(II) complexes are collected in Table 3 and represented graphically in Figure S1. The outcomes results suggested octahedral configuration for Ni(II) & Co(II) complexes [24].

Table 3. Magnetic moments, electronic spectra and ligand field parameters of H₂BHNH metal complexes

Compound	μeff. (B.M.)	Band position (cm^−1)	Dq (cm^−1)	B (cm^−1)	β	υ2/υ1	Transition
2	3.14	13341 23812	815	857	0.82	1.62	^3A2 → ^3T1g (F) ^3A2g → ^3T1g (P)
3	5.21	13793 18182	728	856	0.88	2.15	^4T1g → ^4A2g(F) ^4T1g → ^4T1g(P)

XRD

The diffraction patterns of investigated Ni(II) & Co(II) complexes as shown in Figure 2 were determined in of 10° < 2θ <80° region at RT by X-ray diffraction using (Cu, K_α) radiation [25]. The patterns displayed sharp peaks in two investigated complexes reflecting crystalline nature [25]. The crystallite size was determined through FWHM for characteristic peaks by applying the equation of Deby–Scherrer:

$B=0.94\lambda /\left(Scos\theta \right)$

θ: The diffraction angle, Cu/Kα (λ) = 1.5406 A^o

S: The crystallite size

B: The line width at half maximum height

Also, by applying Bragg equation, the values of inner crystal plane d-spacing are calculated as follows:

$n\lambda =2dsin\left(\theta \right)atn=1$

The particle size as well as lattice parameter values are displayed in Table 4 for examined complexes. The particle size appeared within 0.740 & 0.741 nm for Ni(II) & Co(II) complexes, respectively.

Figure 2. XRD of (a) [Co(HBHNH)₂].2H₂O, (b) [Ni(HBHNH)₂].2H₂O complexes

Table 4. XRD data for Co(II) and Ni(II) complexes

Complex	θ^o	D (A^o)	FWHM (β)	S (A^o)	S (nm)	I
2	21.965	2.059	0.2108	7.407	0.740	392
3	21.965	2.059	0.2105	7.410	0.741	154

Scanning electron microscope studies

The SEM of the Co(II) & Ni(II) complexes displayed that the presence of small grains of non-uniform size and the particles were agglomerated with controlled morphological structure as shown in Figure 3. Also, the SEM images of Co(II) & Ni(II) complexes reveal irregular shaped grains [26].

Figure 3. SEM views of (a) [Co(HBHNH)₂].2H₂O, (b) [Ni(HBHNH)₂].2H₂O complexes

Thermal studies of Ni(II) complex

The TG curves of [Ni(HBHNH)₂].2H₂O complex are thermally stable up to 137°C, above that point the dehydration of complex begins [27,28]. This low temperature indicated that the two molecules of water are not coordinated with the nickel ion. In the temperature range 137°C–198°C, the TG curve displayed 5.3% weight loss that related to two water molecules as shown in Figure S2. The degradation of organic moiety started >198°C. In the TG curve, the temperature ranges 198°C–329°C, the weight loss 23.0% was ascribed to the partial decomposition of the chelating agent with the removal of the two loosely bound phenyl moieties (2C₆H₅). The second weight loss stage (329°C–410°C) which could be correlated with the elimination of the two (C₂H₂N₂) portions. Finally, at 618°C the weight loss of 44.4% was mainly ascribed to complete decomposition of the remaining more tightly bound fragment of the organic moiety, besides breaking the chelate bond, leaving nickel oxide (11.2%) of the complex initial mass.

Thermodynamic and kinetic studies

Kinetic parameters

Horowitz–Metzger and Coats–Redfern equations [29,30] were applied to estimate the kinetic as well as thermodynamic parameters of the decomposition steps for Ni(II) complex as shown in Figure 4 and Table 5. The outcomes data from the two applied methods were nearly similar to a certain extent, so the results can be concluded as follows:

Figure 4. Coats–Redfern and Horowitz–Metzger plots for [Ni(HBHNH)₂].2H₂O (a) first step, (b) second step, (c) third step and (d) fourth step

Table 5. Kinetic parameters evaluated by Coats–Redfern and Horowitz–Metzger equations for Ni(II) complex

Compound	step	Method	Ea	A	∆H*	∆S*	∆G*
			kJ\mol	(S^−1)	kJ\mol	kJ\mol.K	kJ\mol
2	1st	HM	131	1.2 × 10^18	127	99.8	92.8
		CR	122	5.9 × 10^16	119	74.9	92.9
	2nd	HM	66	1376592	63	−130.4	119.3
		CR	59	168312	56	−146.1	106.9
	3rd	HM	204	9.6 × 10^13	199	15.9	187.8
		CR	193	1.2 × 10^13	190	4.1	188.3
	4^th	HM	115	7.3 × 10^2	108	−160.1	237.5
		CR	108	27927	105	−161.1	160.9

The higher values of ∆G demonstrated the lower elimination rate of H₂BHNH from one step to another.

The decomposition step was non-spontaneous process due to positive sign of ∆G referred to the higher quantity of residue than the beginning.

The positive values of ΔS referred to the disordered reaction, while the negative values indicated ordered reaction.

DTG curves referred to high activation energies that indicated the high stability of Ni(II) complex.

The decomposition steps are endothermic processes due to positive sign of ∆H.

Computational studies

Optimized molecular structure

The optimized molecular structures for H₂BHNH and its Ni(II) & Co(II) complexes are represented in Figure 5. The bond length as well as bond angle are renowned for the following explanations:

• On complexation the bond angles of H₂BHNH were altered where the major change observed through ‘C(14)-C(8)-O(11), O(13)-C(14)-N(15), N(15)-C(17)-H(31)’ angles.

• The bond angles of the isolated solid complexes were referred to an octahedral geometry with d²sp³ or sp³d² [31].

• The bond angles of the investigated complexes were longer than free ligand due to donor atoms shared in coordination [32].

• On chelation, a large difference in bond lengths observed through C(8)-O(11), O(11)-H(12), C(14)-O(13), N(15)-H(30), N(15)-N(16) and C(17)-H(31) atoms of free ligand than complexes. Anywhere, the bond length of investigated complexes became more longer than the ligand [33].

• In addition, DFT theory was applied to calculate the dipole moment, sum of atomic energies, electrostatic energy, spin polarization energy, binding energy and kinetic energy, total energy, LUMO, HOMO as represented in Figure 6 and Table 6.

Figure 5. Molecular modeling of (a) H₂BHNH, (b) [Ni(HBHNH)₂].2H₂O, (c) [Co(HBHNH)₂].2H₂O

Figure 6. The HOMO and LUMO of (a) H₂BHNH, (b) [Ni(HBHNH)₂].2H₂O, (c) [Co(HBHNH)₂].2H₂O

Table 6. The molecular parameters of the H₂BHNH and its complexes

Compounds	Total Energy (Ha)	Binding Energy (Ha)	Dipole moment (debye)	HOMO (eV)	LUMO (eV)	Spin polarization Energy (Ha)	Exchange-correlation Energy (Ha)	Electrostatic Energy (Ha)	Kinetic Energy (Ha)	Sum of atomic Energies (Ha)
1	−954	−6.874	2.857	−4.975	−2.624	2.021	2.724	−2.255	−9.363	−947
2	−2102	−13.872	2.296	−4.991	−2.925	3.579	6.408	−6.716	−17.144	−2088
3	−2075	−14.027	0.397	−4.201	−3.118	3.596	4.557	−8.557	−13.624	−2061

MEP

Both electrophilic and nucleophilic attack points were resolute by electrostatic potential. The MEP of H₂BHNH and its Ni(II) & Co(II) complexes that were shown in Figure 7 displayed three characteristic colors: blue color that referred to electron-poor area which prefers the nucleophilic attack. On the other hand, red color referred to electron rich which preferred electrophilic attack. While, the green color referred to electrostatic potential area [34].

Figure 7. MEP of (a) H₂BHNH, (b) [Ni(HBHNH)₂].2H₂O, (c) [Co(HBHNH)₂].2 H₂O

Biological studies

Antimicrobial studies

A lot of studies were established an increase in antimicrobial activity following the interaction of active organic compounds with metal ions. The current study used a methodology modified for antimicrobial bioassays against Gram-positive (+)ve as well as Gram (-)ve bacteria. The results obtained were considered appropriate for determining MIC for performing antimicrobial sensitivity testing with good quality reproducibility as well as efficiency.

The antimicrobial activities of H₂BHNH and its Ni(II) & Co(II) complexes (0.2 ml of 10 μg/ml DMSO) versus P. Aeurginosa & B. Subtillis are represented in Table 7. Generally, the studies ligand showed variable antibacterial activity against either B. Subtillis & P. Aeurginosa. Moreover, the activities were dependent upon the type of the organic ligand used. On the other hand, the antimicrobial study of the isolated metal complexes versus the tested bacteria which arranged in the subsequent order: Ni(II) > Co(II) > H₂BHNH. The data also showed the dependence of the antimicrobial activity on the size of metal cation used where the higher the size of the metal ion, the higher the antibacterial activity.

Table 7. Diameter of Inhibited Zones (I. Z. D.) in mm as a criterion of antibacterial activity of the H₂BHNH and its complexes at concentration level of mg/l

Compounds	Bacteria
	B. Subtillis (G + ve)	P. Aeurginosa (G – ve)
	I. Z. D. (mm)	I. Z. D. (mm)
Ampicillin	19	22
1	18	22
2	24	25
3	23	23

Cytotoxicity

The cytotoxicity of H₂BHNH and its Ni(II) & Co(II) complexes assayed at various concentrations (50, 150, 200 and 400 μg/mL) on RD (ATCC) human muscle cell line was shown to have cell inhibition in range of 51–68%, 41–56% and 46–58%, correspondingly (note that: the number of observed viable cells was decreased). The cytotoxic potential of Ni(II) is greater than H₂BHNH as shown in (Figure 8 and Table 8). The cytotoxicity was observed to increase with the increase in concentration.

Figure 8. The cytotoxicity of H₂BHNH and its Ni(II) & Co(II) complexes

Table 8. % Cell viability

Compounds	Dose Concentration (µg/mL)
	50	150	200	400
1	68	61	54	51
2	56	52	47	41
3	58	57	51	46

Antioxidant studies

Various researchers have used scavenging effect of synthesized compounds on DPPH radical as a quick and reliable parameter to measure the in vitro antioxidant activity. DPPH is stable nitrogen radical that bears no similarity to the highly reactive and temporary peroxyl radicals concerned with the peroxidation of lipids. A lot of antioxidants that react quickly to peroxyl radicals may even be inert to DPPH owing to steric seclusion. Also, DPPH was decolorized by reducing agents as well as H transfer that contributes to inaccurate interpretations of antioxidant capacity.

The antioxidant activity of the H₂BHNH and its Ni(II) & Co(II) complexes studied by DPPH method at 20 µL – 140 µL showed scavenging potential (∼19-43%), (∼23-48%) and (∼27-51%), correspondingly (Figure 9 and Table 9) substantiating the fact that complexation leads to enhanced antioxidant activity [35–37]. The discoloration from purple DPPH radical solution to yellow may be ascribed to hydrogen donation from the (O-H) and (N-H) groups in line with earlier reports on different metals [38]. The resultant free radical formed in complexes through the exchange mechanism was more stabilized due to charge delocalization than that in case of the ligand thereby affording increased antioxidant activity of complexes over ligand.

Figure 9. Cyclic voltammogram of different concentration CoCl₂ in absence of H₂BHNH at 298.15 K

Table 9. % DPPH scavenging activity data at T = 60 min

Compounds	20 µL	40 µL	60 µL	80 µL	100 µL	120 µL	140 µL
1	19	24	26	27	33	38	43
2	23	28	31	32	36	43	48
3	27	33	37	43	50	47	51

Cyclic voltammetry of CoCl₂

The cyclic voltammetry technique was applied within the potential window of (0.8 to −0.8 V) using scan rate 0.1 V/sec at 298.15 K to study the electrochemical redox mechanism of Co²⁺ in 0.1 M KCl as a supporting electrolyte and GC as working electrode (Figure 10). In the forward scan, (Co³⁺/Co²⁺) cathodic potential peak (E_pc1) and (Co²⁺/Co¹⁺) cathodic potential peak (E_pc2) come into view at 0.121 V and – 0.325 V, correspondingly. While, in the reverse scan the (Co¹⁺/Co²⁺) anodic potential peak (E_pa2) exists at 0.035 V followed by (Co²⁺/Co³⁺) anodic potential peak (E_pa1) at 0.212 V. Thus, the redox mechanism of the reaction can be represented as follows:

$C{o}^{3+}+e^{-}Red.\to C{o}^{2+}at0.121V$

$C{o}^{2+}+e^{-}Red.\to C{o}^{1+}at-0.325V$

$C{o}^{1+}Oxid.\to C{o}^{2+}+e^{-}at0.036V$

$C{o}^{2+}Oxid.\to C{o}^{3+}+e^{-}at0.214V$

After that, the peak current is given by the Randles-Sevcik equation [39].

$I_p=\left(2.69x{10}^5\right)n^{3/2}AC{\nu }^{1/2}{D}^{1/2}$

I_p: the current in ampere

D: diffusion coefficient in cm²/s

A: surface electrode area (cm²)

C: molar concentration of metal ion (mol/cm³)

Ν: scan rate in volts/sec

Furthermore, the difference potential (ΔE_P) was determined from the following equation [40]

$\mathrm{\Delta }{E}_p=E_{p,a}-E_{p,c}$

The above quantities (E_pa1, E_pa2, E_pc1, E_pc2, I_p) can be used to estimate numerous cyclic voltammetric parameters such as Q, Γ, αn_a & k_s. The surface concentration of the redox ion was determined by applying the following equation [41]

$\mathrm{\Gamma }=i_{p}4RT/n^2{F}^{2}A\nu$

i_P: the peak current

F: faraday constant (96485.33 coulomb)

T: temperature in K

N: the number of e⁻ in redox reactions

R: gas constant (8.314 J.mol⁻¹.K⁻¹)

A: the surface area of working electrode in cm²

Ν: the scan rate in Vsec⁻¹

The charge transfer coefficient of electrons was evaluated by applying the following equation [42]

$\alpha n_a=1.857RT/\left(E_{pc}-E_{pc/2}\right)$

In case of cathodic peak, E_pc/2 is the half-wave potential.

The charge consumed during the redox reaction was determined by applying the following equation [43].

$Q=nFA\mathrm{\Gamma }$

The heterogeneous charge transfer rate constant (k_s) was estimated by applying the following equation:

$k_s=2.18\ast {\left[D_c\alpha n_{a}F\nu /RT\right]}^{1/2}\ast exp\left[{\alpha }^{2}nF\left(E_{p,c}-E_{p,a}\right)/RT\right]$

D_C: the diffusion coefficient of oxidized particles

N: electrons number in redox reactions

R: the gas constant

F: Faraday Constant

T: the temperature in K

Α: charge transfer coefficient

ΔE_P: the difference potential

n_a: scan rate and the number of electrons transfer in the rate determining step

Figure 10. Cyclic voltammogram of CoCl₂ in the presence of different concentration of H₂BHNH at 298.15 K

The previous parameters were estimated for the anodic as well as cathodic peak and tabulated in Table 10. Most of the evaluated parameters for the anodic and cathodic peaks increase as the concentration of CoCl₂ increases. The increase in all peak currents supported the suggested diffusion mechanism [44].

Table 10. Solvation and kinetic parameters D, k_S, Γ and Q of CoCl₂ at 298.15 K for wave (B) in absence of H₂BHNH

M x10^−3	Ep,a (volt)	Ep,c (volt)	∆Ep (volt)	-Ip,a x10^−5 (Amp)	Ip,c x10^−5 (Amp)	Ip,a/Ipc (Amp)	E°	Da x10^−11	Dc x10^−11	αna	Ks x10^−4 (cm/sec)	Γc x10^−8 (mol/cm^2)	+ Qc x10^−5	Γa x10^−8 (mol/cm^2)	- Qa x10^−5
0.66	−0.006	−0.324	0.318	4.37	1.23	3.55	−0.165	61.11	4.84	1.645	8.47	0.417	1.26	1.481	4.49
1.32	−0.008	−0.322	0.314	4.28	1.15	3.73	−0.165	14.90	1.07	1.908	4.13	0.390	1.18	1.453	4.40
1.96	0.008	−0.292	0.300	8.77	1.86	4.72	−0.142	28.12	1.26	2.168	4.17	0.629	1.91	2.975	9.01
2.60	0.035	−0.264	0.299	17.8	4.22	4.22	−0.115	66.02	3.71	2.074	6.92	1.431	4.34	6.040	18.3
3.23	0.037	−0.255	0.292	17.7	4.01	4.41	−0.109	42.36	2.18	2.168	5.06	1.362	4.13	6.007	18.2

Cyclic voltammetry response of Co(II) in presence of H₂BHNH

The electrochemical behavior was studied for complexation between H₂BHNH and CoCl₂ in 0.1 M KCl solution at GCE within (+0.8 to −0.8 V) potential window (Figure 11). The result current redox peaks are decreased by increasing the ligand concentration that favoring the covalency between the cobalt ions and H₂BHNH. Furthermore, the cathodic & anodic potential peaks were shifted. These outcomes results recommend a strong interaction between cobalt and H₂BHNH.

Figure 11. Cyclic voltammogram of CoCl₂ at different scan rates at final adding in absence of H₂BHNH at 298.15 K

In addition to the previous variations in the shape of voltammogram, the calculated values of E_Pc, E_Pa, ∆E_p, i_Pa, i_Pc, (i_Pa/i_Pc), E°, k_s, α_na, Γ and Q for CoCl₂ in the presence of H₂BHNH (Table 11).

Table 11. Solvation and kinetic parameters D, k_s, Γ and Q of CoCl₂ at 298.15 K for wave (B) in presence of H₂BHNH

[M] x10^−3	[L] x10^−3	Ep,a (volt)	Ep,c (volt)	∆Ep (volt)	-Ip,a x10^−5 (Amp)	Ip,c x10^−5 (Amp)	Ip,a/Ipc (Amp)	E°	Da x10^−11	Dc x10^−11	αa	Ks x10^−4 (cm/sec)	Γcx10^−8 (mol/cm^2)	+Qc x10^−5	Γa x10^−8 (mol/cm^2)	- Qa x10^−5
3.20	0.65	0.04	−0.25	0.29	15.2	4.65	3.23	−0.10	31.2	2.96	2.39	6.09	1.579	4.78	5.117	15.5
3.18	1.28	0.03	−0.26	0.29	12.8	4.13	3.01	−0.11	22.4	2.33	2.28	5.33	1.394	4.22	4.322	13.1
3.16	1.91	0.04	−0.30	0.34	11.4	3.87	2.91	−0.13	17.8	2.12	1.33	6.61	1.314	3.98	3.830	11.6
3.14	2.53	0.07	−0.35	0.42	8.89	3.76	2.36	−0.14	11.3	2.01	0.95	12.2	1.274	3.86	3.011	9.12
3.12	3.14	0.08	−0.40	0.48	7.34	3.52	2.08	−0.16	7.76	1.81	0.73	17.3	1.196	3.62	2.488	7.54
3.10	4.61	0.04	−0.41	0.45	2.97	1.67	1.78	−0.18	1.34	0.43	1.26	8.25	0.568	1.72	1.012	3.07
3.06	6.07	0.03	−0.44	0.47	1.52	1.34	1.13	−0.20	0.35	0.29	1.16	7.36	0.449	1.36	0.508	1.54
2.96	8.83	0.04	−0.47	0.47	0.51	1.47	0.33	−0.23	0.05	0.38	0.86	7.74	0.502	1.52	0.170	0.52

Effect of scan rate in (absence/presence) of H₂BHNH

The cyclic voltammograms of Co²⁺ in (absence/presence) for H₂BHNH at different scan rates 10, 20, 50 & 100 mV/s in 0.1 M KCl at 298.15 K are displayed in Figures 12 and 13. The outcomes data showed that both cathodic & anodic peak currents increase in linear manner with the increasing in the scan rate. Also, the cathodic peak was slightly shifted toward the negative potential while, the anodic peak was slightly shifted toward positive direction with the extent of the scan rate. These remarks suggest the diffusion controlled redox mechanism and supported by the linear relation between square root of scan rate and peak currents as displayed in Figures 14 and 15. The ratio of peak current is >1 that continue a correspondence to a quasi reversible system [45]. The solvation and kinetic parameters for CoCl₂ in absence/presence of H₂BHNH at various scan rates were shown in Tables 12 and 13.

Figure 12. Cyclic voltammogram of CoCl₂ at different scan rates at final adding in presence of H₂BHNH at 298.15 K

Figure 13. Relation (ip vs. ν^1/2) for CoCl₂ at final adding in absence of H₂BHNH at 298.15 K and different scan rate

Figure 14. Relation (ip vs. ν^1/2) for CoCl₂ in presence of H₂BHNH by molar ratio (1:1) at 298.15 K and different scan rate

Table 12. Effect of scan rate on CoCl₂ [3.23x10⁻³ M] for wave (B)

ʋ	Ep,a (volt)	Ep,c (volt)	∆Ep (volt)	-Ip,a x10^−5 (Amp)	Ip,c x10^−5 (Amp)	Ip,a/Ipc (Amp)	E°	Da x10^−11	Dc x10^−11	αna	Ks x10^−4 (cm/sec)	Γc x10^−8 (mol/cm^2)	+Qc x10^−5	Γa x10^−8 (mol/cm^2)	- Qa x10^−5
0.1	0.038	−0.25	0.292	17.7	4.01	4.411	−0.109	42.4	2.177	2.169	5.06	1.3618	4.13	6.007	18.2
0.05	0.034	−0.24	0.267	18.1	3.54	5.117	−0.098	88.6	3.383	3.407	4.39	2.4010	7.27	12.29	37.2
0.02	0.024	−0.23	0.213	9.26	1.51	6.151	−0.129	57.9	1.530	3.66	1.15	2.5535	7.74	15.70	47.6
0.01	0.020	−0.22	0.244	7.81	1.16	6.711	−0.101	82.4	1.829	3.976	1.25	3.9480	12.0	26.50	80.3

Table 13.. Effect of scan rate on co-complex for wave (B)

ʋ	Ep,a (volt)	Ep,c (volt)	∆Ep (volt)	-Ip,a x10^−5 (Amp)	Ip,c x10^−5 (Amp)	Ip,a/Ipc (Amp)	E°	Da x10^−11	Dc x10^−11	αna	Ks x10^−4 (cm/sec)	Γc x10^−8 (mol/cm^2)	+Qc x10^−5	Γa x10^−8 (mol/cm^2)	- Qa x10^−5
0.10	0.08	−0.34	0.42	7.34	3.52	2.09	−0.128	7.74	1.787	7.952	31.6	1.194	3.61	2.49	7.56
0.05	0.06	−0.39	0.45	5.58	1.94	2.87	−0.167	8.92	1.075	0.723	6.98	1.313	3.98	3.78	11.6
0.02	0.03	−0.36	0.39	3.20	1.42	2.27	−0.168	7.27	1.403	0.824	3.18	2.366	7.16	5.39	16.5
0.01	0.01	−0.33	0.35	2.85	0.94	3.07	−0.163	11.48	1.216	1.194	1.59	3.122	9.44	9.58	29.2

Conclusion

The bivalent Ni (II) and Co (II) complexes of N’-benzylidene-3-hydroxy-2-naphthohydrazide ligand were synthesized by ball milling as an efficient technique for quantitative solvent-free synthesis. The isolated solid compounds will be characterized using several spectroscopic techniques. The theoretical studies represented in the molecular modeling of isolated solid compounds were investigated using the DFT method which showed the compatibility with the practical outcomes data. Moreover, the in vitro antimicrobial activity assay versus fungi and bacteria via MIC technique has higher potency of Ni (II) than Co (II). The in vitro cytotoxicity on human muscle rhabdomyosarcoma (RD) via MTT technique and the antioxidant by DPPH free radical scavenging have higher inhibitory effect of complexes than free ligand. Voltametric studies explained the effect of Co(II) in absence/presence of H₂BHNH on the electrochemical behavior of cobalt solution through the change in the values of (i_P, E, Γ, α_na, rate k_s, and Q).

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Supplementary material

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