SYNTHESIS, SPECTRAL ELUCIDATION, ANTIBACTERIAL, ANTIOXIDANT AND DNA STUDIES OF ONNO TETRADENTATE SCHIFF BASE METAL(II) COMPLEXES DERIVED FROM 2-AMINOPHENOL DERIVATIVES

SYNTHESIS, SPECTRAL ELUCIDATION, ANTIBACTERIAL, ANTIOXIDANT AND DNA STUDIES OF ONNO TETRADENTATE SCHIFF BASE METAL(II) COMPLEXES DERIVED FROM 2-AMINOPHENOL DERIVATIVES

Kumarasamy Savitha and Subramaniam Vedanayaki ^*

Department of Chemistry, Kandaswami Kandar’s College, Paramathi Velur, Namakkal - 638182, Tamil Nadu, India.

ABSTRACT: The new asymmetrical tetradentate (ONNO) Schiff base (L) 4-chloro- 2- ((E)- (4- ((E)-((2-hydroxy-5-nitrophenyl)imino)methyl)benzylidene) amino)phenol was prepared by the condensation of terephthalaldehyde with 2-amino 4-chlorophenol and 2-amino 4-nitrophenol in 1:1:1 molar ratio in methanol. The homo binuclear metal(II) complexes were synthesized in ligand to metal ratio 2:2 from the template method. The composition, geometry and binding sites of ligand and its complexes were determined by various studies like Elemental, molar conductance, electronic, magnetic moment, FT-IR, ¹H and ¹³C-NMR, ESI-Mass, EPR, Thermal, and PXRD. The above spectral studies show that the Schiff is a dibasic tetradentate (ONNO), and its metal complexes possess a square planar structure. The biological applications of all compounds were tested for antibacterial, antioxidant, and DNA cleavage, and the final reports reveal high inhibition activity for metal(II) complexes compared to the ligand. The DNA binding activities of Cu(II) complex was observed by electronic absorption method. Antitumor activity of Cu(II) complex was screened for their cancer-inhibiting ability against MCF-7 human breast cancer cells by using the MTT experiment.

Schiff base tetra dentate ligand, Square planar, Antimicrobial, Antioxidant, DNA studies, Antitumour activity

INTRODUCTION: The design and preparation of asymmetrical Schiff base ligand derived from the 1:2 ratio condensations of carbonyl group (dialdehyde) with alkyl or aryl primary amines and its derivatives. Tetra dentate Schiff base and its metal complexes played an important role in the progress of coordination chemistry following in a substantial number of publications. The infection treatment was challenged by multidrug resistance in pathogenic organisms and oxidative stress, so the development of newer Schiff base compounds for the management of infections and oxidative stress is warranted ^{1, 2}.

Tetra and polydentate ligands are having nitrogen and oxygen donor atoms in their structure act as chelating agents for the non-transition and transition metal ions, which form stable complexes with transition metal(II) ions ^3-5.

In recent years, the preparation and spectral evaluation of macrocyclic binuclear metal complexes have been more efforts and useful to study the relation between geometry and magnetic exchange interactions. Ligands are used to substrates in the synthesis of many numbers of biologically active and industrial compounds via ring closer, cyclo-addition, and replacement reactions ⁶. Unsymmetrical Schiff base ligand and its complexes were recommended and used as biological models in understanding irregular binding of peptides and catalyst in chemical process ⁷. The macrocyclic chemistry field of metal is developing very fast because it shows the many pharmacological applications and importance in the area of bioinorganic and medicinal chemistry ⁸. Macrocyclic Schiff base complexes give potential applications of many fields in fundamental and applied sciences, stabilization of unusual oxidation states, sensors technology, magnetic resonance imaging, models for biological structures and functions, contract enhancing agents, and metal selective extraction ⁹. Schiff base metal complexes allow systematic studies depends on the modified ancillary ligand, which plays the main role in the catalysis; enzymatic reaction, magnetism, and molecular modeling studies are related to the development of bioinorganic chemistry ¹⁰. Metal complexes are reported to have excellent biological applications such as antimicrobial, antioxidant, anti-insecticidal, antifertility, antitubercular, anti-tumor and anti-inflammatory ^11-18. Schiff base ligand and its binuclear metal complexes can cleave the DNA through three kinds of mechanisms like hydrolytic, oxidation, and photolytic cleavages ¹⁹. Cytotoxicity effect of Cu(II) complex has considerable ability due to their bind and cleave DNA, which leads to cell cycle prevention and apoptosis ^{20, 21}.

Based on the above fact, herein we have reported the synthesis of new asymmetric Schiff base ligand (4- chloro- 2- ((E)- (4- ((E)- ((2-hydroxy-5-nitro-phenyl)imino)methyl) benzylidene) amino) phenol) containing the azomethine (CH=N) and hydroxyl groups (OH) as potent chelating sites and its Co(II), Ni(II) and Cu(II) binuclear metal complexes evaluated by diverse physicochemical technique and their applications were analyzed for antibacterial, antioxidant, DNA studies (cleavage and binding) and antitumor activity (in-vitro cytotoxicity).

EXPERIMENTAL SECTION:

Analytical and Physical Measurements: Terephthalaldehyde, 2-amino 4-chlorophenol, 2-amino 4-nitrophenol, metal salts were purchased from Sigma Aldrich. Ethanol, Methanol, DMSO, DMF, and Acetone were purchased from Loba and Merck chemicals. The purity of all compounds was tested by TLC.

C, H and N elements were carried out on a Thermo Finningan Flash EA 1112 series elemental analyzer. The molar conductance of compounds was measured at room temperature using by ELICO CM 180 Conductivity Bridge. The magnetic moment values of compounds were calculated using Gauy balance calibrated with Hg[Co(SCN)₄] method at room temperature. UV spectral studies were measured using the Perkin-Elmer Lambda-25 spectrometer in the range of 200-800 nm. FT-IR spectra of compounds were recorded on a Shimadzu FT-IR-8300 spectrometer using KBr pellet experiment in range 4000-400 cm^-1. ¹H and ¹³C-NMR spectra were recorded on a BRUKER ADVANCED III 4000 MHz spectrometer using TMS as an internal reference. Electro Spray Ionization of mass spectra was obtained on a Perkin-Elmer R MU-6E instrument in the mode of positive. The ESR spectra were recorded on the JES-FA200 ESR spectrometer with X-band frequency at room temperature. Thermal analyses were measured at 0°-1000 °C using SDT-Q600 V20.9 Build 20 thermal analyzer in an oxygen atmosphere. The Powder-XRD spectra were carried out on a Perkin-Elmer TA/SDT-2960 and Philips 3701 instrument.

Synthesis of Asymmetrical Tetra Dentate Schiff Base Ligand: The mixture of 1mmole of tereph-thalaldehyde with 1mmole of 2-amino 4-chloro-phenol and 1mmole of 2-amino 4-nitrophenol were dissolved in methanol. The solution was kept under stirring for 2 h, the formed light yellow precipitate was separated by filtration, washed and purified by methanol solvent. The Schiff base solid was recrystallized from ethanol. Yellow solid, Molecular weight- 395.80, Melting point-240 °C, Yield- 80%, IR (KBr cm^-1): 3339 (OH), 1623 (C=N), 1289 (C-O); ¹H-NMR (DMSO-d_6, δ, ppm): 8.85 (s, 1H, C=N), 10.1 (s, 1H, OH), 6.5-8.2 (m, Ar—H); ¹³C-NMR (100MHz, CDCl₃): 113, 115, 116, 117, 118, 119, 123, 127, 129, 130,  138, 139, 140, 141, 143, 150, 161; Elemental analysis: C₂₀H₁₄ClN₃O₄ Calculated values: C- 69.14, H- 4.70, N- 7.68: Found values: C- 68.86, H- 4.50, N- 7.98; ESI-Mass: m/z: (M+1)⁺  396.

Synthesis of Binuclear Metal(II) Complexes: The homo binuclear metal(II) complexes were synthesized by using template method ²². The mixture of terephthalaldehyde (2mmole) with 2-amino 4-chlorophenol (2mmole) and 2-amino 4-nitrophenol (2mmole) was dissolved in methanol, which was added to the methanolic solution of metal salts (2mmole) like Co, Ni, and Cu. The mixture was stirred, and a few drops of triethylamine were added to the mixture. It was stirred for 1 h and under reflux for 3 h. The product was partly evaporated, cooled at room temperature, the obtained metal complexes were separated by filtration, washed with methanol and diethyl ether, stored at room temperature. The structure of metal complexes was shown in Scheme 1.

SCHEME 1: METAL COMPLEXES PREPARATION FROM TEMPLATE METHOD

Schiff Base Binuclear Co(II) Complex (Co₂L₂): Brownish black solid, Molecular weight- 994.11, Melting point->300 °C, Yield- 78%, IR (KBr cm^-1): 1602 (C=N), 1340 (C-O), 577 (M-O), 425 (M-N); Elemental analysis: C₄₆H₄₂Cl₂Co₂N₆O₈  Calculated values: C- 55.49, H- 4.25, N- 8.44, M- 11.84: Found values: C- 55.80, H- 4.51, N-4.18, M-11.34; Molar conductance ( ?^-1 cm² mol^-1) 10.5.

Schiff Base Binuclear Ni(II) Complex (Ni₂L₂): Reddish yellow solid, Molecular weight- 992.11, Melting point->300 °C, Yield- 76%, IR (KBr cm^-1): 1598 (C=N), 1338 (C-O), 548 (M-O), 438 (M-N); Elemental analysis: C₄₆H₄₂Cl₂N₆Ni₂O₈  Calculated values: C- 55.52, H- 4.25, N- 8.44, M- 11.80: Found values: C- 55.12, H- 4.13, N- 8.21, M-11.41; Molar conductance ( ?^-1 cm² mol^-1) 11.4.

Schiff Base Binuclear Cu(II) Complex (Cu₂L₂): Black solid, Molecular weight- 1004.86, Melting point->300 °C, Yield- 79%, IR (KBr cm^-1): 1606 (C=N), 1344 (C-O), 529 (M-O), 473 (M-N); Elemental analysis: C₄₆H₄₂Cl₂Cu₂N₆O₈  Calculated values: C- 54.98, H- 4.21, N- 8.36, M- 12.65: Found values: C- 54.35, H- 4.10, N- 8.04 , M- 12.22; Molar conductance ( ?^-1 cm² mol^-1) 13.6.

Pharmacological Studies:

Antibacterial Activity: In-vitro antibacterial activity was performed by the Disc-agar well diffusion experiment. The asymmetric tetradentate Schiff base ligand and its homo binuclear complexes were screened against bacteria of Gram-positive (S. aureus, B. subtillis) and Gram-negative (K. pneumoniae, E. coli). Tetracycline was used as a standard drug for antibacterial activity, and the tested compounds were dissolved in DMSO. The antibacterial activities were maintained in nutrient agar well plates at 4 °C. The 100 µL concentration of culture supernatant was placed on agar well plate, and then it was incubated for 24 h at 37 °C. The antibacterial activity was determined by measuring the diameter of the zone, indicating complete inhibition ²³.

Antioxidant Activity: 4-chloro-2-((E)-(4-((E)-((2-hydroxy-5-nitrophenyl)imino)methyl)benzylidene) amino)phenol and it's metal (Co, Ni, Cu) complexes were investigated by using its free radical scavenging activity on the stable DPPH free radicals described in the literature ²⁴. The scavenging activity investigates the antiradical power of an antioxidant by calculating the decrease in the absorbance wavelength of DPPH at 510 nm. The ligand and its metal complexes exhibit DPPH free radical scavenging activity at different concentrations (like 10, 20, 30, 40, and 50 µL). The percentage of DPPH free radical scavenging ability was measured by using the following formula

I% = Ac – As/ Ac × 100

Where Ac is the absorbance of the control and As is the absorbance of the tested sample. IC₅₀ values were calculated for ligand and its complexes, which showed significant ability, and it is defined as concentration sufficient to generate 50% of maximum scavenging activity. Ascorbic acid was used as a standard.

DNA Binding Studies:

Electronic Absorption Spectroscopy: Electron absorption spectroscopy is one of the most useful techniques to determine the interaction between the metal complexes with DNA from the stock solution of calf thymus (CT) ²⁵. DNA was prepared by 5mM Tris-HCl/20mM NaCl buffer (pH=7.2) at room temperature. The stock solution was stored at 4 °C and used up to 4 days. The buffer solution of CT- DNA gave a ratio of ~1.8-1.9 and the UV absorbance at 260 to 280 nm. The concentration of CT-DNA was determined using the known molar extinction coefficient value of 6600 M^-1 cm^-1 at absorption intensity is 260 nm. The stock solution concentrations of the complex were prepared by dissolving the Cu complex in DMSO solvent. All the experiments were maintained at proper dilution with the corresponding buffer to the required concentration. The binding constant (K_b) was calculated by using the equation:

[DNA]/ (ε_a-ε_f) =[DNA]/ (ε_a-ε_f) +1/K_b(ε_a-ε_f)

Where [DNA] is the molar concentration of DNA is base pairs, ε_a, ε_f are apparent extinction coefficient, the K_b values were obtained from the ratio between the equation of  DNA/ (ε_a-ε_f) versus [DNA] in each case.

Agarose Gel Electrophoresis Assay: The cleavage of supercoiled pBR322 DNA was investigated by gel electrophoresis method ²⁶. The agarose gel electrophoresis studies were carried out incubation of the mixtures containing 20 µL pBR322 DNA, 50 mM of NaCl, 50 mM of metal complexes, and 50 mM H₂O₂ in Tris-HCl buffer (pH=7.4) at 37 °C for 1 h. After incubation, the sample compounds were electrophoresed at 60 °C for 2 h on 1% of agarose gel using TAE (Tris-Acetic acid-EDTA) buffer (pH=8.0). After 0.5 µg/ml of ethidium bromide was used, the gel was stained. Thus all the experiments were performed at room temperature and photographed under UV light at 360 nm.

Antitumor activity: In-vitro cytotoxicity was carried out on an MCF-7 cell line in which cell viability was tested using MTT (3-(4,5-dimethyl thiazol-2-yl)- 2, 5- diphenyltetrazolium bromide) activity ²⁷. In-vitro effect of growth inhibition of Cu(II) complex was measured by spectro-photometric experiment. This experiment was used to determine the MTT conversion into “formazan” by living cells, and it was maintained in a 96-well microplate having DMEM/F12 plain media supplemented with 10% heat-inactivated FBS (Fetal Bovine Serum) having 5% of a mixture of 100 µg/mL of streptomycin and 100 units/ml of penicillin were incubated at 37 °C for 24 h in the presence of 5% CO₂. In this study, various concentrations of 10, 20, 40, 80, 160 and 320 µL of the stock solution were prepared in DMSO, which were added to relevant wells having 100 µL of the DMSO medium. After incubation, 100 µL of MTT stock solution (5mg/10ml of MTT in PBS) was added to each well and further incubated for 4 hrs. After the supernatant was removed, the plates were gently shaken to form solubilize formazan crystals by adding 100 µL of DMSO. The absorbance was measured at wavelength 590 nm by a microplate reader. The activity was performed in triplicate and used to calculate the mean.

RESULTS AND DISCUSSION: The synthesized ligand and their metal complexes were investigated by diverse physicochemical methods. The resultant compounds are soluble in DMF and DMSO, partially soluble in ethanol and methanol.

Elemental and Molar Conductivity Analysis: The composition of C, H, and N are analyzed by CHNS elemental analyzer. The molar conductance of ligand and their complexes were measured at 25 °C in DMF solution (10^-3M) indicates low conductance values (10.5 to 13.6), which shows that the complexes are non-electrolytic nature ^28,29.

Electronic Absorption Spectral Data: The UV-Vis spectral data of the ligand and its complexes were measured within a 200-800 nm wavelength at room temperature in the DMSO solution. The electronic spectra of the mixed ligand exhibit absorption bands at 296 and 384 nm, which is referred to the aromatic benzene rings (?→?*) and imine group (n→?*) respectively ³⁰.

The diffuse reflectance spectrum of the macrocyclic metal complexes of Co(II), Ni(II) and Cu(II) shows bands at 444, 452 and 414 nm, which may be attributed to the ligand to metal co-ordinations (L→M transitions) in square planar geometry of the compounds (given in Table 1). The magnetic moment and absorption spectra is used to determine the geometry of the metal complexes ³¹.

TABLE 1: UV SPECTRUM OF LIGAND AND ITS METAL(II) COMPLEXES

FT-IR Studies: The IR spectrum is used to investigate of the existence of intramolecular hydrogen bonding, the nature of the coordination mode in the metal complexes, and the presence of the tautomeric forms in the solid states. The FT-IR spectral data of the ligand and their complexes were observed in the wavenumber region at 400-4000 cm^-1. The important vibrational frequency of the free ligand exhibits band at 1623 cm^-1, which indicates the formation of the imine group(C=N) ³². However,  on complexation, this band(C=N) is shifted to lower energy range 1606-1598 cm^-1, suggesting the participation of the azomethine nitrogen atom in coordination to the metal ion. The sharp broadband around 3345 cm^-1 in the ligand was assigned to the phenolic –OH group. The absence of ν(OH) band in the metal complexes indicates the co-ordination of the ν(OH) group after deprotonation. The ν(C=O) band at 1289 cm^-1 for ligand were lower frequency than the band at about 1700 cm^-1 for ν(C=O), this frequency change shows the formation of azomethine group (ligand).

The appearance of new bands at 425-473 and 529-577 cm^-1 in the vibrational spectra of the complexes were assigned to ν_(M-N) and ν_(M-O) frequencies respectively ³³. These vibrations were clearly suggesting the co-ordination of the metal ions with the azomethine nitrogen and phenolic oxygens in the complex.

NMR Spectral Studies:

¹H-NMR Spectrum: The ¹H NMR spectrum of the ligand was recorded at 25 °C (RT) in DMSO-d⁶ solvent with TMS as an internal standard. The azomethine (CH=N) proton signal was obtained at δ8.85 ppm; this signal indicates the formation of mixed Schiff base ligand ^{34, 35} shown in Fig. 1a. The new peaks of phenolic –OH protons of the aminophenol and aromatic protons were observed at δ10.0, 10.1 ppm, and δ6.5-8.2 ppm ³⁶.

¹³C-NMR Spectrum: ¹³C- NMR spectrum of the synthesized ligand shifted to δ 161 ppm for an azomethine carbon atom in Fig. 1b. The peaks showed at δ 150 ppm, δ 143 ppm, and δ 113-141 ppm were due to phenolic –OH carbon by aminophenol, substituted Cl atom by aminophenol and aromatic benzene ring carbon atoms ³⁷.

FIG. 1A: ¹H NMR SPECTRUM OF LIGAND

FIG. 1B: ¹³NMR SPECTRUM OF LIGAND

ESI-Mass Spectrum: The electronic impact spectrum of Schiff base ligand and its copper complex showed molecular ion peaks at m/z 396 (M+1)⁺ and m/z 1005 (M+1)⁺, which corresponds to the proposed molecular formula of ligand (C₂₀H₁₄ClN₃O₄) and its Cu complex (C₄₆H₄₂Cl₂Cu₂N₆O₈) respectively ^{38, 39}. The mass spectral data of Schiff base ligand and its copper (II) complex were confirmed by comparing their molecular formula weights with (m/z) mass values, which is in good agreement for these ligand and Cu complex as shown in Fig. 2a, 2b.

FIG. 2: A AND B: ESI-MASS SPECTRUM OF LIGAND AND CU(II) COMPLEX

Electron Resonance Spectrum: The electron spin resonance studies of the binuclear Cu (II) complex gives information about hyperfine and super-hyperfine structures and the nature of the bonding between the copper ion and its ligand ⁴⁰. The epr spectrum of the complex [Cu₂L₂] was displayed at room temperature and which exhibits an axial symmetry at X-band frequencies in the solid-state. The obtained g-values are g_II=2.2391 and g_⟘=2.0563, which are related by G=(g_II-2)/(g_⟘-2)=4.0. If G value is more than 4, i.e. (G>4), which indicates no considerable exchanging interaction between the two copper centers in the complex.

FIG. 3: ESR SPECTRUM OF CU(II) COMPLEX

FIG. 4: THERMOGRAM CURVES OF LIGAND AND ITS METAL(II) COMPLEXES

Whereas if G value is less than 4(G<4), which shows that the exchange interaction occurs in the solid-state complex. From the observed G value of Cu(II) complex is 4.24. It is clear that g_II>g_⟘>2.0023, which indicates d_x2-y2 in the orbital ground state for the square planar structure of the Cu(II) complex shown in Fig. 3.

Thermal Studies: The thermal analysis (TG & DSC) was used to determine the thermal stability of compounds for ligand and its metal complexes in the presence of air atmosphere at the temperature range between 0 to 1000 °C ⁴¹. The thermogram of the ligand and its complexes were shown in Fig. 4. Both TG and DSC analysis of compounds have three steps of decomposition process each. The first, second, and third steps correspond to the elimination of small groups of substituted Cl and methyl atom by 2-aminophenol, the removal of total ligand moiety, and the formation of metal oxide residue.

P-XRD Analysis: The X-ray powder diffraction analysis of the synthesized compounds of ligand and its metal complexes has been carried out to confirm whether the nature of the sample is amorphous (or) crystalline. The structure of Schiff base and its binuclear metal complexes were investigated using P-XRD analysis, which indicates the amorphous orthorhombic crystal nature of compounds within the range 10-90˚C (2θ) and the wavelength is 1.5406 A° ⁴². Thus, all the compounds show crystalline nature from its observed peaks. The Powder-XRD provides the d-value, relative intensity and 2θ value for each peak.

FIG. 5: ANTIBACTERIAL ACTIVITY OF TESTED COMPOUNDS

Biological Applications:

Antibacterial Assay: The antibacterial activity of tetra dentate Schiff base ligand and its Co(II), Ni(II) and Cu(II) complexes were screened against for S. aureus, B. subtillis (gram-positive) and K. pneumoniae, E. coli (gram-negative) bacteria and tetracycline was used as standard drug. The antibacterial results showed that the Schiff base ligand has very low (or) no inhibition activity compared to its metal complexes ^{43, 44}. Thus Cu(II) complex exhibits higher antibacterial activity than the other Co(II) and Ni(II) complexes Fig. 5.

Antioxidant Assay:

DPPH Free Radical Scavenging Activity: The ligand and its Co(II), Ni(II), and Cu(II) complexes were screened for their DPPH free radical scavenging ability using Ascorbic acid as standard ⁴⁵. The ligand has very low activity when compared to all the complexes. The scavenging activity of Cu(II) complex is higher than that of other metal complexes. The IC₅₀ values were determined (shown in Fig. 6) for all compounds, and the IC₅₀ values of [Cu₂ (C₄₆H₄₂N₆O₈Cl₂)] is 5.93 mg/ml showed significant activity compared to other complexes and ligand. The order of scavenging activity of all complexes according to their IC₅₀ values is given below.

Ascorbic acid > [Cu₂ (C₄₆H₄₂N₆O₈Cl₂)] > [Ni₂ (C₄₆H₄₂N₆O₈Cl₂)] > [Co₂ (C₄₆H₄₂N₆O₈Cl₂)] > C₂₀H₁₄ClN₃O₄

FIG. 6: ANTIOXIDANT ASSAY OF LIGAND AND ITS METAL(II) COMPLEXES

DNA- Copper Complex Interaction Studies: The interactions of CT-DNA and metal complexes were determined by the electronic absorption spectroscopy technique. From the studies, it is revealed that the intensity changes of the intraligand ? → ?* transition band occur at 250-280 nm ⁴⁶. The UV absorption experiments of Cu(II) complex in the presence of buffer solution are performed by using a fixed concentration of complex to which increments of the stock solution was added. The interaction of [Cu₂ (C₄₆H₄₂N₆O₈Cl₂)] complex with duplex DNA led to a decrease in the intensities, and a small amount of redshift in the electronic absorption spectra and the concentration of DNA increases with the absorption bands of Cu(II) complex are affected to a considerable extent. This absorption spectra indicate clearly that the addition of CT-DNA to the Cu(II) complex yields hypochromic and redshift (shown in Fig. 7) to the ratio of [DNA]/[Cu] for the [Cu₂ (C₄₆H₄₂N₆O₈Cl₂)] complex and the complex interact with CT-DNA most likely through a binding mode that involves ? → ?^* stacking interaction between the aromatic chromophore and the base pair of DNA.

FIG. 7: ABSORPTION SPECTRA OF Cu(II) COMPLEX WITH DIFFERENT CONCENTRATIONS (100, 120, 1140, 160, 180, 200 µL) H₂O₂ INDUCED DNA DAMAGE/ PRODUCTION ASSAY

Gel electrophoresis activity is a method to determine different binding modes of newly synthesized complexes to supercoiled pBR322 DNA. The natural- derived plasmid pBR322 DNA has three forms of the closed-circle supercoiled form (form-I), nicked form (form-II), and linear form (form-III) ^{47, 48}. The circular plasmid DNA is conducted by electrophoresis; relatively, the fasted migration will be measured for the supercoiled form (form-I). If cleavage occurs on one strand, the supercoils will relax to generate slowed moving open circular form (form-II). If both strands are cleaved a form-III (linear form), it will be produced that moves in between supercoiled form and open circular form. From the results (in Fig. 8) it indicates that the electrophoretic gel separation of plasmid pBR322 DNA, interaction with metal complexes in the presence of H₂O₂. The addition of the complexes with a mixture of form-II and form-III, which form the cleavage of supercoiled DNA. The reports show that the Co(II), Ni(II), and Cu(II) metal complexes induced the cleavage of DNA intensively in the presence of H₂O₂. These observations suggested that all dinuclear complexes effectively cleave the plasmid pBR322 DNA.

FIG. 8: DNA CLEAVAGE STUDIES OF SCHIFF BASE AND ITS METAL(II) COMPLEXES

In-vitro Cytotoxicity Assay: To determine the cytotoxicity effect, the newly synthesized homo binuclear Cu(II) complex was treated with the human breast cancer cell line (MCF-7) by MTT experiments method ⁴⁹. The absorbance value is lower than the control cell, which shows a reduction in the rate of cell proliferation. Contrary, a higher absorbance rate show an increase in cell proliferation cell survival is almost 50% after 24 hrs of incubation with Cu(II) complex. The anticancer results of complex revealed that Cu(II) complex exhibits significant cytotoxic effect. The IC₅₀ value and percentage of inhibition of Cu(II) complex are given in Table 2.

TABLE 2: ANTITUMOUR ACTIVITY OF Cu(II) COMPLEX

CONCLUSION: Macrocyclic homo binuclear Co(II),Ni(II), and Cu(II) complexes derived from terephthalaldehyde with 2-amino 4-chlorophenol and 2-amino 4-nitrophenol. The asymmetrical tetra dentate Schiff base and its binuclear Co(II), Ni(II) and Cu(II) complexes were prepared, evaluated by physicochemical experiments and diverse spectroscopic methods.

The spectral studies indicate that all metal(II) complexes are four coordinated and possess square planar geometry around the metal ion. The biological applications of antibacterial, antioxidant, and DNA cleavage assay of the metal(II) complexes show more inhibition activity than the free ligand. The Cu(II) complex has considerable anticancer and DNA binding abilities.

ACKNOWLEDGEMENT: The authors are thankful to SAIF, IIT Bombay for ESR Analysis, SAIF, IIT Madras for mass analysis, St. Joseph’s College, Trichy for IR analysis, Sastra Deemed University, Thanjavur for Thermal analysis. The authors are very grateful to the Guide and Principal of Kandaswami Kandar’s College, P. Velur, Namakkal (Dt), for providing facilities to perform the work.

CONFLICTS OF INTEREST: The authors declare that there are no conflicts of interest regarding the publication of this research paper.

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    How to cite this article:

    Savitha K and Vedanayaki S: Synthesis, spectral elucidation, antibacterial, antioxidant and DNA studies of onno tetradentate schiff base metal(II) complexes derived from 2-aminophenol derivatives. Int J Pharm Sci & Res 2020; 11(9): 4513-23. doi: 10.13040/IJPSR.0975-8232.11(9).4513-23.

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