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Design and Biological Evaluation of Some Novel Metal Complexes derived from 1-(4-(Imidazole-1-sulfonyl)phenyl)ethanone derivative.

Byline: Sami A. Al-Harbi

Summary: A monobasic bidentate ligand (H2L) was prepared from the reaction of cyanoacetic acid hydrazide with 1-(4-((1H-imidazol-2-yl)sulfonyl)phenyl)ethanone (ISE) in ethanol. A new series of Mn(II), Fe(III), Co(II), Ni(II), Cu(II), Ag(I) and Zn(II) metal complexes of H2L ligand was synthesised. The interaction of the H2L ligand with salicyaldehyde and naphthaldehyde afforded coumarin and benzocoumarin derivatives, respectively. Structural characterisation of the newly synthesised compounds was achieved by elemental analysis, thermogravimetric analysis TGA, electrical molar conductance and spectral techniques, such as IR, UV, 1H NMR, 13C-NMR and ESR, as well as by magnetic moment measurements. The reported ligand (H2L) is monobasic dibasic with an N-O bidentate set, which coordinated with the metal ions to produce the metal complexes. The N atom of CH=N functioned coordinately with the O atom of carbonyl in the ligand.

The prepared novel compounds were assessed for their antimicrobial activities. Fe(III), Co(II), Ni(II) and Ag(I) compounds showed good activity against Gram (-ve) bacteria. The newly synthesised compounds were assessed for their in vitro anticancer activity against both MCF-7 and HCT-116 cell lines; normal fibroblasts of the baby hamster kidney (BHK) cell line were also assessed. Ni(II) and Ag(I) complexes exhibited significant activity towards the MCF-7 and HCT-116 cell lines. Cu(II) and Fe(III) complexes exhibited better activity in the MCF-7 cell line, while Zn(II) and benzocoumarin compounds showed better activity in the HCT-116 cell line compared to doxorubicin (DOX) as a reference drug. Molecular docking was used to illustrate interaction among components within the active sites of dihydrofolate reductase (DHFR).

Keywords: 1-(4-(Imidazole-1-sulfonyl)phenyl)ethanone derivative, Metal complexes, Anti-cancer, antimicrobial activities.

Introduction

Sulfonamides and imidazole derivatives are important classes of drug, the significance of which is drawn from their unique biological properties. Sulfonamides present antiproton [1], anti-thyroid [2], anti-carbonic anhydrase [3] and antibacterial activities [4] as part of their properties. Sulfonamide structures have also been used in the synthesis of novel derivatives that have been reported to have significant antitumor activity both in-vivo and in-vitro [5]. Imidazole derivatives exhibit antiviral, antifungal, antibacterial, anti-inflammatory, anticoagulant and anticancer properties [6-8].

Medical professionals and researchers have long valued sulfonamides and imidazole moieties for their therapeutic properties and have stressed the need to synthesize novel chemotherapeutic agents, both of which have expanded the scope of clinical medicine by presenting new opportunities for novel and unique drug compositions [9]. Schiff base ligands show a vital role as chelating ligands in transition metal coordination chemistry [10, 11]. A large amount of bimetallic Schiff base complexes of different structural types have been prepared and characterized [12]. These complexes vary in their new applications for biological activities [13].

Moreover, since the discovery of cis-platin and continuous research producing new generations of antitumour drugs, multidentate organic compounds containing metal ions, which are analogues in the catalyst of some metal enzymes, [14] have drawn the attention of researchers seeking novel, less toxic, less expensive and more efficient anti-cancer drugs. Of these compounds, Schiff base ligands with sulfonamide and imidazole moieties containing multidentate atoms coordinated to metal ions are among the most promising [14, 15].

In the present study, a novel monobasic bidentate ligand (H2L) containing a combination of two pharmacological sulfonamide and imidazole moieties was synthesized via reactions of 1-(4-((1H-imidazol-2-yl)sulfonyl)phenyl)ethanone (ISE) and cyano-acetic acid hydrazide in absolute ethanol. The ligand was reacted with manganese(II), iron(III), cobalt(II), nickel(II), copper(II), silver(I) and zinc(II) nitrates in the molar ratio 1:1 (metal : ligand), forming stable tetrahedral and octahedral complexes. Additionally, when the ligand reacted with salicyldehyde and naphthaldehyde, it afforded coumarin and benzocoumarin derivatives, respectively. The metal complexes and organic compounds were identified using physicochemical analyses and spectroscopic techniques.

The metal complexes and organic coumarin derivatives of H2L were examined for antimicrobial properties. Eight microorganisms were used for this investigation: Staphylococcus aureus (RCMB 010027), Streptococcus pneumoniae (RCMB 010010) and Bacillis subtilis (RCMB 010067) as Gram (+ve) bacteria, Klebsiella pneumoniae (RCMB 0010093), Escherichia coli (RCMB 010052), Mycobactrium tuberculosis (RCMB010120) and Sallmonella typhimurium (RCMB 010315) as Gram (-ve) bacteria, Aspergillus fumigatus (RCMB 02568) and Candida albicans (RCMB 05036). Iron(III) and silver(I) complexes showed high activity against three Gram-negative bacteria. In addition, the silver(I) complex showed good activity against Escherichia coli, while the other metal complexes showed moderate activity against the specifying organisms.

Investigation of the Schiff base and its complexes, including its coumarin and benzocoumarin derivatives, on human cancer (breast and colon) cell lines showed that nickel(II) and silver(I) complexes and benzocumarin, have significant activity against both breast carcinoma (MCF-7) and colon carcinoma (HCT-116) cell lines. The activity that was observed from the study of molecule docking indicated that coumarin, as well as iron (III) in conjunction with the benzocoumarin, may exert their activity through the inhibition of the DHFR enzyme.

Experimental

Materials

All chemicals used were of analytical grade. 1-(4-((1H-imidazol-2-yl)sulfonyl)phenyl)ethanone (ISE) and cyano acetic acid hydrazide were synthesized based on the method reported in literature [16]. The metal salts, Mn(NO3)2.4H2O, Fe(NO3)3.9H2O, Co(NO3)2.6H2O, Ni(NO3)2.6H2O, Cu(NO3)2.3H2O, AgNO3 and Zn(NO3)2 and organic solvents were purchased from Merck, BDH and Sigma-Aldrich.

Synthesis of the Schiff base ligand

The ligand was synthesized by mixing 1-(4-((1H-imidazol-2-yl)sulfonyl)-phenyl)ethanone (ISE) (2.50 g, 10 mmol) to cyano acetic acid hydrazide (0.99 g, 10 mmol) in ethanol (30 mL). The reaction mixture was heated to reflux for 4 h. The resulting white crystals were filtered off and washed several times with a few amount of ethanol then ether and finally air dried. Fine crystals were obtained by recrystallization from ethanol. The yield was 2.42 g (73.0 %), m.p. 240-242C. The suggested structure of H2L ligand is illustrated in Scheme-1.

Synthesis of metal complexes of the Schiff base ligand (H2L)

An equimolar amount of metal salt in 25 ml ethanol was added drop wise to a 25 ml hot ethanolic solution of the organic ligand (H2L). Only in the case of silver(I) complex the pH of the reaction mixture was adjusted to 2.3. The reaction mixture was stirred for half hour at room temperature. The resultant reaction mixture was further stirred under reflux for three hours during which time colored products precipitated. After that, the volume of the solution of the reaction mixture was reduced, then filtered and the isolated solid was washed with ethanol and ether and finally dried in vacuum over CaO at room temperature for several days.

Synthesis of compounds 8 and 9

A mixture of the reported Schiff base (H2L) (3.31 g, 1.0 mmol) and salicyldehyde and/or naphthaldehyde (1.0 mmol) in ethanol (50 mL) containing (1 mL) of piperidine was stirred under refluxed for 4 h, and during the reflux period a compounds were precipitated. The precipitated compounds were collected by filtration and recrystallized from the proper solvent to give coumarin 8 (ISC) and benzocoumarin 9 (ISB), respectively.

Physical Measurements

FT-IR spectra (KBr) were recorded on (Jasco) spectrophotometer model 4600. Elemental analysis (C, H, N, S) were performed by using Carlo-Erba 1106 elemental analyzer [17-19]. After decomposition of the metal complex in hot concentrated nitric acid and neutralization of the reaction medium the resulting metal ions were determined complexmetrically using EDTA and murexide or Eriochrome Black T (EBT) indicator system. 1H and 13C-NMR spectra were recorded by Varian spectrometer, EM-390, 600 MHz. The electronic absorption spectra (UV-Vis) were obtained in Dimethylformamide (DMF) solution with an Angstrom UV-VIS photometer. Mass spectra measurements were made by use of Hewlett Packard MS 5988 mass spectrometer. The Gouy's methodology played a significant role in the measurement of magnetic susceptibility of the metal complexes at room temperature. The instrument used was (Johnson Matthey, Alfa product Model No. (MKI)).

The magnitude of paramagnetism was assessed by use of the equation ueff = 2.828 (IM.T)1/2 B.M. IM represents molar susceptibility obtained from Pascal's constants for the paramagnetism of the atoms present in the complex. T; represents the absolute temperature [20]. The molar conductance of the complexes was measured using freshly prepared 10-3 M solutions in electrochemically pure DMF at room temperature, using a Conductometer model 1483. Melting points (AdegC, uncorrected) were measured in open capillaries on a Gallen Kemp melting point apparatus (Sanyo Gallen Kemp, Southborough, UK).

The thermogravimetric measurements were performed using a Shimadzu TG 50-Thermogravimetric analyzer in the 25-800 AdegC range and under an N2 atmosphere.

Antimicrobial Investigation

In vitro assessments of antimicrobial activities were conducted by use of a diffusion method known as standardized disc-agar [21]. The antimicrobial activities of newly synthesized compounds activity was examined towards the following series of the sensitive, Gram-positive organisms: S. aureus (RCMB 010027), S. pneumoniae (RCMB 010010), K. pneumoniae (RCMB 0010093), B. subtilis (RCMB 010067), Gram-negative organisms: E. coli (RCMB 010052), Salmonella typhimurium (RCMB 010315), Mycobacterium tuberculosis (RCMB 010120) and fungi: A. fumigatus (RCMB 02568) and C. albicans (RCMB 05036). Ampicillin, gentamicin and amphotericin B were used with Gram-positive and Gram-negative bacteria as well as antifungal [22, 23].

Evaluation of the anticancer activity

The anticancer activity of novel compounds was examined for carcinoma (MCF-7) in conjunction with HCT-116. Performance of cytotoxic activity against the BHK was also assessed [24]. The inhibition of cell growth, (IC50), was determined by using the relation: IC50 = (IC - IT/IC) x 100, where IC represents the average/mean value of optical density of untreated cells (control) and IT represents the value of the absorbance of treated cells. The cytotoxic effect of the cell (CC50) compounds was also calculated: the percentage of the cytotoxic effect of the cell is equal to:

[1- IT / IC] x 100, where IT coupled with IC represents the absorbance of complexes and the control of the cell, respectively [25-27].

Docking and Molecular Modeling

Docking and molecular modeling are calculated according to the methods reported elsewhere [28-30]

Results and Discussion

Characterization of Schiff base ligand (H2L)

Infrared spectrum of the inspected ligand (H2L) confirmed the expected structure. Vibrational assignment was assisted by comparing the frequency of vibration of the H2L ligand to the frequency of vibration of the aldehydic and amine moieties, namely ISE and cynoacetic acid (1-phenylthylidene) hydrazide [31]. Firstly, the vibrational modes of the azomethine group, (-C=N-), were identified by comparing the infrared spectra of (ISE) and cyanoacetic acid hydrazide, in which (C=O) and (NH2) in cyanoacetic acid hydrazide were obscured. The intense band of the (-C=N-) of the H2L ligand was recognized in the azomethine moiety of the Schiff base molecules. Additional bands were consigned in the infrared spectrum of the H2L ligand. The details of the vibrational bands of the H2L ligand were recorded, and related compounds are listed in Table-2.

1H NMR spectrum of H2L ligand exhibits three singlet signals at I' = 2.26, 4.24 and 9.08 ppm for the methyl, methylene and NH groups, respectively. In addition a broad signal for NH of imidazole at I' = 14.22 ppm and multiple signals for aromatic protons and CH=CH of imidazole from I' = 7.60 to 7.76 ppm were detected. 13C-NMR spectrum of H2L showed a signal at I' = 14.38 ppm for the methyl group, I' = 25.25 for the methylene group, eight signals for eleven aromatic and imidazole carbon atoms from I' = 116.71 to 159.90 ppm and one signal at I' = 166.36 ppm corresponding to the carbonyl group. Table-3 lists the 1H NMR spectra chemical shifts (ppm) of ISE, H2L ligand, coumarin 8 and benzocoumarin 9 in dimethyl sulfoxide (DMSO).

In the UV region, electronic transitional bands at 272, 324, 335 and 417 nm were characterized for H2L ligands in DMF solution. These transitions may attributed to the I a I* transition of the aromatic rings [32], the I a I* transition of the azomethine groups, and the n a I* transitions of the oxygen and nitrogen atoms. The H2L ligand mass spectrum showed the parent peak at m/z = 331, which compares well with the formula weight (331.35) of the ligand (Fig. 2).

Table-1: Elemental analysis and some Physical measurements to ligand (H2L) and its complexes.

###Molecular###M.P.###Elemental Analyses; Calc. (Found)%

###Compound No.###M.Wt.###Yield%###Color###AdegC

###formula###C###H###N###S###M

###140-###52.79###4.03###11.19###12.18###-

###ISE###C11H10N2O3S###250.27###71###White

###141###(52.61)###(3.93)###(11.18)###(12.07)###-

###C14H13N5O3S###240-###50.75###3.95###21.14###9.68###-

###H2L###331.35###73###White

###242###(50.59)###( 3.85)###(12.04)###(9.57)###-

###33.54###3.62###16.76###6.40###10.96

1###[MnL(H2O)3(NO3)]###MnC14H18N6O9S###501.33###58.5###White###> 360

###(33.41)###(3.51)###(16.61)###(6.28)###(10.88)

###30.78###2.95###17.95###5.87###10.22

2###[FeL(NO3)2].2H2O###FeC14H16N7O11S###546.23###66.6###Brown###> 360

###(30.66)###(2.84)###(17.81)###(5.75)###(10.11)

###33.28###3.59###16.63###6.35###11.66

3 [CoL(H2O)2(NO3)].H2O###CAdegC14H18N6O9S###505.33###64.1###Blue###> 360

###(33.19)###(3.48)###(16.51)###(6.25)###(11.55)

###33.29###3.59###16.64###6.35###11.62

4###[NiL(H2O)2(NO3)].H2O###NiC14H18N6O9S###505.09###54.8###Green###> 360

###(33.18)###(3.49)###(16.51)###(6.22)###(11.51)

###32.97###3.56###16.48###6.29###12.46

5 [CuL(H2O)2(NO3)].H2O###CuC14H18N6O9S###509.94###72.0###Green###> 360

###(32.88)###(3.45)###(16.38)###(6.15)###(12.35)

###35.46###3.40###14.77###6.76###22.75

6###[AgL(H2O)2]###AgC14H16N5O5S###474.24###55.7###White###> 360

###(35.33)###(3.31)###(14.68)###(6.69)###(22.66)

###ZnC14H14N6O7S###269-###35.34###2.97###17.67###6.74###13.74

7###[ZnL(NO3)].H2O###475.74###65.7###White

###270###(35.22)###(2.85)###(17.58)###(6.62)###(13.62)

###Yellowish-###265-###57.79###3.70###12.84###7.35###-

8###ISC###C21H16N4O5S###436.44###64.4

###White###266###(57.68)###(3.63)###(12.72)###(7.22)###-

###270-###61.72###3.73###11.52###6.59###-

9###ISB###C25H18N4O5S###486.50###67.4###Yellow

###271###(61.61)###(3.60)###(11.43)###(6.50)###-

Table-2: The characteristic vibrational stretching frequencies of the H2L, APT and DAR.

###Frequencies (cm-1)

###Assignment

###ISE###CAH###H2L

###-###3347, 3193 (s)###-###

###3132###3126###3167, 3140 (2NH)###

###-###-###3109 (m)###Caromatic

###2974###2966###2974 (w)###Caliphatic

###1681###1687###1697###C

###-###2261###2260###CN

###1615###-###1629 (vs)###C

###1643###-###1649###CC

###1400, 1126###-###1388, 1161###SO2

Table-3: 1H and 13C-NMR chemical shifts of the ISE, H2L ligand, Zn(II), Cd(II), coumarine and benzocoumarine compounds in DMSO.

Compd.###Chemical Shift (ppm)

No.###1H NMR###Assignment###13C-NMR###Assignment

###2.17###(s, 3H, CH3)###27.26###(CH3)

###7.26###(s, 2H, CH=CH of imidazole)###119.71###(2C)

###8.86-8.91###(m, 4H, Ar-H)###126.05###(2C)

###ISE

###13.86###(br, 1H, NH)###128.53###(2C)

###134.59, 137.36, 151.84###(3C)

###198.33###(C=O)

###2.26###(s, 3H, CH3)###14.38###(CH3)

###4.24###(s, 2H, CH2)###25.25###(CH2)

###7.60-7.76###(m, 6H, Ar-H + CH=CH of imidazole)###116.71###(C)

###9.08###(s, 1H, NH)###120.32###(2C)

###H2L

###14.22###(br, 1H, NH of imidazole)###125.96###(2C)

###134.82###(2C)

###138.19, 149.24, 153.18, 159.90###(4C)

###166.36###(C=O)

###2.26###(s, 3H, CH3)###

###4.65###(s, 1H, CHCN)###

###7.58-7.91###(m, 6H, Ar-H + CH=CH of imidazole)###

###7

###9.58###(br, 2H, H2O)###

###10.68###(s, 1H, NH)###

###11.03###(br, 1H, NH of imidazole)###

###8###2.31###(s, 3H, CH3)###39.64###(C)

###7.25-7.86###(m, 11H, Ar-H + NH + CH=CH of imidazole)###115.00###(C)

###8.61###(s, 1H, H4 of coumarin)###118.42###(2C)

###13.59###(s, 1H, NH)###119.93###(C)

###121.28###(C)

###124.27###(C)

###125.56###(2C)

###125.96###(C)

###130.17###(C)

###133.37###(2C)

###137.61###(2C)

###142.12, 149.10, 152.16, 153.41###(4C)

###155.86###(C=O)

###158.04###(C=O)

###2.33###(s, 3H, CH3)###39.50###(C)

###7.46-8.49###(m, 13H, Ar-H + NH + CH=CH of imidazole)###119.92###(2C)

###9.20###(s, 1H, H4 of benzocoumarin)###115.81###(3C)

###13.67###(s, 1H, NH)###119.09, 121.93, 125.57###(3C)

###125.98###(2C)

###128.86, 128.97, 129.19###(3C)

###9

###129.75###(2C)

###134.68###(C)

###137.21###(2C)

###137.64, 149.16, 152.19, 153.39###(4C)

###155.91###(C=O)

###158.21###(C=O)

Characterization of the metal complexes

Interaction between the mono-basic bidentate Schiff base ligand (H2L ) having the coordination donors N and O, with Mn(II), Fe(III), Co(II), Ni(II), Cu(II), Ag(I) and Zn(II) ions affords a series of mononuclear transition metal complexes. The structural characterization of the present newly synthesized compounds were obtained by using elemental analyses and spectroscopic techniques IR, UV-visible, Ms, 1H- and 13C-NMR. Additionally, TGA, magnetic susceptibility and molar conductivity measurements were employed.

Infrared spectra

Table-4 lists the characteristic frequencies of the H2L ligand with its metal complexes and assignments. The spectral frequencies of the metal complexes were compared with the frequencies observed in the free ligand spectrum. The (-C=N-) band in the spectra of the complexes shifted to lower frequencies (1620-1576 cm-1) in comparison with the original band of free ligand at 1629 cm-1. This behavior is most likely attributable to the coordination of the (N-N=CH) group with metal ions [16, 33, 34].

The NO3 ions coordinated to the metal ions as unidentate for the Mn(II) complex (1) with C2v symmetry. The unidentate nitrate was comprised of a set of non-degenerated vibration models (s, s and as) that appeared at 1407, 1321 and 811 cm-1, respectively. The s(NO3 ) of the unidentate NO3 was shifted to a lower frequency in comparison to that of the free nitrate, which ranged between 1700 and 1800 cm-1 [35]. This information was then used to estimate the covalent binding that developed from a moment of electronegativity from NO3 to the metal ion. For complexes of iron(III) 2, cobalt(II) 3, nickel(II) 4, copper(II) 5 and zinc(II) 7, the NO3 group services served as bidentate ligand. The nitrate group thus possessed three non-degenerate modes of vibrations (, a and s) at 1548-1541, 1387-1375 and 1304-1302 cm-1.

The (OH) of the water molecules in the complexes was assigned by broad bands ranging from 3420-3291 cm-1. The weaker bands ranging from 579-530 cm-1 and 459-439 cm-1 were applied to the stretching frequency of the (M-O) and (M-N) bands, respectively. These results illustrate the behavior of bonds between the ligands and metal ions in the presence of an O atom and (N-N=CH) atoms Electronic, magnetic, mass, and molar conductance measurements

In electronic, magnetic, mass and molar conductance assessment, electronic constructions can be both assembled and predicted with the assistance of magnetic moments created by a majority of metal ions (Table-5). The I a I* and n a I* electronic transitions due to C=N and C=C groups that took place due to the presence of the H2L ligand in a metal complex were observed: 318-316 and 410-408 nm, respectively. The transitions were lower compared to subsequent absorption bands of the H2L ligand that occurred at 324 and 417 nm. This shift may have formed as a result of the development of coordinate bonding between O and N atoms of the ligand with the metal ions.

Furthermore, extra bands were observed while electronic spectrometry of the metal complexes was conducted. They appeared to persist at parallel points, which is dissimilar from the results of electronic spectrometry of the complimentary H2L ligand, which are a range of wavelengths from the transitions of the aromatic ring and was seen at 273-272 and 337-334 nm, respectively.

Only a single unsymmetrical band (712 nm) formed from an electronic spectrum of the green compound 5 and was given to Eg a 3T2g(G) transition in square planar geometry. The measured value of the magnetic moment for complex 5 was 2.11 B.M., which serves as a confirmation of the square planar structure [36].

Electron spin resonance (ESR) study of complex 5 shows a single wide-ranging band with geff = 2.03 and the structure of the spectrum served as a confirmation for square planar geometry [37, 38]. The mass spectrum of 5 (Fig. 2), revealed the parent peak at m/z 831 which matched well with the formula mass (831.78) of the compound.

Table-4: Infrared spectral bands (cm-1) and their assignments for H2L and its metal complexes.

###v(M-###v(M-###v(CH)###v(CH)###v(NH,

###Other band###I'(H2O)###v(C=O)###v(C=C)###v(C=N)###v(OH)###Cpd.

###N)###O)###Aliphatic###Aromatic###NH2)

###-###-###-###-###-###1649###1629###2974###3109###3167, 3140###3430 s, br###H2L

###1407 s, 1321 s, br, 811 m;

###455 w###538 w###1638 vs###-###1666###1632###2920###3032###3410, 3197###3428 s, br###1

###unidentate NO3 group.

###1542 vs, br, 1379 vs, 1304 vs,

###457 w###536 w###1645 vs###-###2983###3100###3344, 3317###3433 s, br###2

1053 m; bidentate NO3 group

###1546 vs, br, 1384 vs, 1302 vs,

###453 w###542 w###1688 vs###-###1666###1618###2985###3078###3194, 3136###3425 s, br###3

1058 m; bidentate NO3 group.

###1541 vs, br, 1387 vs, 1303 vs,

###453 w###542 w###1629 vs###-###1666###1618###2924###3074###3186, 3132###3300 s, br###4

1046 m; bidentate NO3 group.

###1548 vs, br, 1375 vs, 1304 vs,

###456 w###530 w###1636 vs###-###1645###2932###3022###3400, 3192###3298 s, br###5

1051 m; bidentate NO3 group.

###452 w###544 w###1662 v###-###1648###1643###2969###3027###3347, 3187###3431 s, br###6

###1547 vs, br, 1380 vs, 1311 vs,

###455 w###542 w###1662 vs###-###1666###1617###2987###3074###3259, 3236###3291 s, br###7

1055 m; bidentate NO3 group

###1678,

###-###-###-###-###-###1614###2970###3051###3313, 3113###-###8

###1659

###1685,

###-###-###-###-###1643###1617###2997###3039###3321, 3290###-###9

###1670

Table-5: Magnetic moments, molar conductance and electronic transition bands of the metal complexes of H2L.

Compound No.###nm ;Iumax(Electronic transition; I>>max)a###Ab###ueff(B.M.)c###Assignment###d-d transition

###n a n*

###H2L###417 (0.557) 335 (0.485) 324 (0.823) 272 (0.732) 14###---###---###---

###4A1 a 4T1 (G)###865 (0.000)

###1###410 (0.558) 334 (0.467) 316 (0.464) 272 (0.852) 112###5.51

###4A1 a 4T2 (G)###490 (0.000)

###Charge transfer tailing from the UV###830 (0.021),

###2###408 (0.392) 336 (0.511) 318 (0.842) 273 (0.784) 160###4.70

###region to the visible region.###519 (0.047)

###4T1g a 4T2g###1060 (0.089)

###3###410 (0.542) 337 (0.452) 318 (0.414) 272 (0.723) 117###4.90###4T1g(F) a 4A2g(F)###685 (0.060)

###4T1g(F) a 4T1g(P)###516 (0.022)

###3A2g a 3T2g###1000 (0.083)

###4###410 (0.523) 337 (0.462) 318 (0.423) 272 (0.741) 121###3.21

###3A2g a 3T1g(F)###625 (0.057)

###5###410 (0.561) 337 (0.43) 318 (0.443) 272 (0.752) 123###1.91###Eg a 3T2g(G)###712 (0.035)

###6###409 (0.566) 333 (0.451) 322 (0.476) 273 (0.741) 22###Diamagnetic###---###---

###7###409 (0.422) 335 (0.515) 318 (0.824) 272 (0.857) 111###Diamagnetic###---###---

###8###421 (0.573) 339 (0.574) 329 (0.924) 277 (0.713) 31###---###---###---

###9###423 (0.544) 341 (0.565) 327 (0.933) 279 (0.828) 28###---###---###---

Two electronic transitional bands were seen from the pale yellow Mn(II) complex (5). The bands are occurred at 664 and 525 cm-1 and may be assigned to the 6A1g a 4T1g(4G) (3) and the 6A1g a 4T2g(4G) (2) transitions, in order of rising energy.

The third band due to the 6A1g a 4A1g, 4Eg(4G) (1) transition exists in a range of the ligand transitions and it was hard to notice. Measurement of the magnetic moment gave 6.83 B.M. This value ascertains to the fact that complex (5) has high spin octahedral geometry [39].

The UV-Vis spectrum of the blue Co(II) complex (3) recorded in DMF showed that this compound has an octahedral environment. Two bands were observed in the visible region, one of them at 680 nm refers to 4T1g(F) a 4A2g(F) (2) transition, while, the other one at 511 nm corresponding to 4T1g(F) a 4T1g(P) (3) transition.

At room temperature, the magnetic moment of Co(II) complex was 4.90 B.M. [40].

The absorption spectrum in UV-Vis region of the green Ni(II) complex (4) displayed two bands at 1000 and 623 nm which are assigned to 3A2g a 3T () and 3A a 3T (F) () transitions, respectively, confirming octahedral structure of d8 configuration. The other indication which confirms the octahedral environment of (4) complex is the value of magnetic moment at 3.21 B.M. [41, 42]. The third 3A2g a 3T1g(P) 3 transition was covered by charge transfer of the organic ligand.

The high spin octahedral geometry of the brown Fe(III) complex (2) was proven only by magnetic measurement at 5.94 BM due to a strong charge transfer (CT) band tailing from the UV-region to the visible region, two bands at 562 and 483 nm not appeared and the identity of the type of the d-d transition are not possible [43, 44].

Table-6: Thermogravimetric analysis degradation of metal complexes.

Complex###Thermogravimetric analysis degradation###Kinetic and thermodynamic parameters

###Temp. AdegC###Weight loss###Species formed###TAdeg (K)###A###Ea###H###G###S

###Found (Calcd.)

###1###38.39 - 254.30###27.82 (27.10)###[MnL(0.94)]###0543###02.11###33.610###30.690###167.52###-0.2519

###294.82 - 428.48###40.19 (60.12)###Mn2O3###0773###02.94###52.610###49.690###244.62###-0.2521

###2###153.26 - 416.85###30.50 (28.70)###[FeL]###055###03.33###39.460###36.540###174.65###-0.2484

###442.83 - 551.88###33.69 (33.36)###[FeL(0.45)]###0693###07.43###72.490###69.570###238.35###-0.2435

###594.92 - 628.66###25.51 (27.29)###Fe2O3###0943###02.56###65.940###63.020###303.46###-0.2549

###3###156.60 - 219.10###67.92 (61.73)###[CoLNO3] [CoL(0.3)]###0568###0.150###25.490###22.570###178.40###-0.2743

###388.39 - 492.06###18.94 (19.67)###Co2O3###0833###03.08###60.750###57.890###268.09###-0.2524

###4###24.78 - 63.31###4.06 (3.56)###[NiLNO3]###0387###05.35###31.930###29.020###122.53###-0.2416

###82.31 - 276.31###45.77 (45.64)###[NiL(0.6)]###0568###0.150###25.490###22.570###178.40###-0.2743

###289.45 - 569.18###40.56 (39.36)###NiO###0833###03.08###60.750###57.890###268.09###-0.2524

###5###49.66 - 135.88###7.65 (7.06)###[CuLNO3]###0521###02.95###35.350###32.430###162.09###-0.2488

###136,66 - 370.40###6.16 (6.49)###[CuL(0.99)]###0650###07.07###65.860###62.940###221.17###-0.2434

###403.58 - 680.80###66.90 (64.32)###CuO###0993###0.720###43.860###40.940###324.04###-0.2850

###6###23.12 - 67.47###4.86 (3.79)###[AgL]###0335###0.160###15.380###12.470###046.38###-0.2506

###70.15 - 155.76###4.86 (3.79)###[AgL]###0573###0.187###28.210###25.290###181.48###-0.2725

###160.43 - 317.64###16.45 (16.07)###[AgL(0.77)]###0698###17.09###126.01###123.01###288.22###-0.2366

###353.10 - 419.67###15.04 (15.37)###[AgL(0.55)]###0823###11.27###111.94###109.02###307.79###-0.2415

###452.90 - 551.30###36.99 (38.42)###Ag2O

###7###22.85 - 100.43###3.60 (3.78)###[ZnLNO3]###0459###01.12###24.500###21.590###139.03###-0.2558

###100.43 - 216.65###12.75 (13.03)###[ZnL]###0631###05.21###53.640###50.730###205.78###-0.2457

###217.65 - 336.86###46.61 (46.66)###[ZnL(0.33)]###0833###03.42###62.860###59.950###253.51###-0.2323

###402.63 - 544.96###19.59 (22.98)###ZnO

The coordination occurred between the oxygen of the phenolic group and the Zn(II) and Ag(I) ions, where the proton of OH group of phenolic disappeared in 1H-NMR spectra (Table-3). When compared the MS of compounds 3 and 12 (Fig. 2) with the molecular weights (835.50 and 929.51) showed the base peaks m/z at 836 and 929; respectively.

The molar conductivities of the present nitrato metal chelates in DMF solution lie in the range 22.32 - 30.98 a|-1 mol-1 cm2 indicate partial dissociation because the observed values are much lower than that expected for 1:1 electrolytes indicating that they behave essentially as non-electrolytes in this solvent [45]. This non-electrolytic nature of these metal complexes also indicates that the associated counter anions remain coordinated to metal ion as mono anion in solution.

Thermal analysis (TGA and DTG)

The TGA and DTG date of the investigated complexes are listed in (Table-6). The thermogram of the hydrated manganese (II) complex, [Mn(HL)(H2O)3-(NO3)] (1), displayed two stages of decomposition within an ambient temperature range extending to 500 C. The initial weight loss of 27.82% is in a good agreement with the theoretical value (27.10%), which corresponds to volatilization of the coordinated water molecules, removal of the non-electrolytic nitrate ion and partial degradation of the organic ligand. The recorded mass loss in the range of 295 to 430 AdegC with DTG maximum at 400 AdegC can attribute to complete thermal decomposition of the remaining organic ligand in successive steps.

The final product of the thermal pyrolysis for the complex was Mn2O3.

The thermal decomposition of the hydrated iron(III) complex, [Fe(HL)(H2O)2(NO3)2] (2), occurs in three stages within the temperature range 35.0 - 628.66 C. The first decomposition stage starts at 153.26 C and ends at 280.22 C with DTG maximum at 245.47 C. The weight loss is corresponding to the volatilization of three molecules of the coordinated water. The observed mass loss (23.50 %) is due to the explosion of the coordinated nitrate molecules and the corresponding activation energy value is 39.46 kjmol-1. The final thermal degradation stage comprises several successive and unresolved steps within the temperature range 318.89 - 658.90 C, with maximum decomposition peaks (DTGmax) at 377.71, 551.88 and 628.66 C. The corresponding mass loses are due to the partial and complete decomposition and removal of the organic portion.

The mass loss in this stage is in an almost agreement with calculated mass loss and the final product is quantitatively proved to be anhydrous Fe2O3. The corresponding activation energy values are 72.49 and 65.94 kjmol-1.

The thermogram of the hydrated cobalt complex, [Co(HL)(H2O)3(NO3)] (3), exhibits three stages of thermal pyrolysis within the temperature range 21.30-534.39 C. (Table-6). The first thermal degradation step starts at 130.06 C and ends at 387.37 C with the maximum decomposition peak (DTGmax) at 219.10 C. The corresponding mass loses are due to the complete volatilization of the three coordinated water molecule and counter anion (NO3) in addition to partial removal of the organic portion with a mass loss of 61.69% in accordance with calculated values 61.73 %. The final thermal pyrolysis stage involves a significant mass loss extending from 388.39 to 550.00 C with DTG maximum at 492.06 C. The corresponding mass loses are due to the complete removal of the organic moiety. The final product is anhydrous metallic oxide (Co2O3).

The activation energy value of this thermal degradation process calculated from thermal analysis data is 60.75 kJmol-1. Further horizontal constant curve observed may be due to the remaining part from the metal oxides.

The thermogram of the nickel(II) complex, [Ni(HL)(H2O)3(NO3)] (4), represents the following features. The initial weight loss within the temperature range 124-213 AdegC is in a good agreement with the theoretical value which corresponds to the removal of the coordinated water molecule with DTGmax peak at 189 C. A complete removal of the non-electrolytic and coordinated nitrato counter anion in addition to partial degradation of the organic ligand were achieved at the second thermal decomposition stage in successive steps within the temperature range 82-276 AdegC. The experimental weight loss value 45.77 % agrees well with the expected value of 45.64 %. The third stage describes the complete releasing of the organic moiety in the temperature range of 289-569 AdegC leaving behind nickel(II) oxide as a final product. The observed activation energy values of the different thermal decompositions of this complex are low and reported within the range 43.360 - 32.040 kjmol-1.

The thermogram of the copper(II) complex, [Cu(HL)(H2O)3(NO3)] (5), represents the following features. The first stage of thermal decomposition starts at 156.60 C and is marked with a regular loss in mass up to 219.10 C. The observed weight loss is in good agreement with the theoretical value of 19.06 %, which is due to thermal removal of water content. Table-6 lists the activation energy of this thermal dehydration step (37.69 kjmol-1), which suggests the weak interaction of the water and no role in the lattice forces. The second stage was within the temperature range 219.10-370.40 C, with DTG maximum at 193.40 C corresponds to the volatilization of the axially coordinated non-electrolytically counter anion (NO3) in addition to a partial decomposition of the organic ligand. The activation energy value is 35.35 kjmol-1 (Table-6). The third stage in the region between 403.58 and 680.80 C with DTG maximum of 285.12-488.68 C describes the complete decomposition of the organic moiety.

The TG curve of the hydrated silver(I) complex, [Ag(HL)(H2O)2] (6), shows decomposition steps within the temperature range 40-552 AdegC. The first thermal pyrolysis steps at the temperature range of 155.76-235.56 AdegC represent the volatilization of two water molecule as inferred from observed mass loss values which are in a good agreement with expected theoretical value as shown in Table-6. The second thermal degradation stage within the temperature range 235.56-317.64 ascribed to the partial decomposition of the organic moiety with experimental mass loss of 27.37 % (calculated; 28.12 %). The final thermal decomposition process within the temperature range 353 - 551 AdegC with a DTGmax at 550 AdegC correspond to the loss of the residual organic portion of the complex in successive steps with a mass loss of 36.99 % (calcd. 38.24 %) giving the final residual Ag2O. The reported activation energy values of these thermal decompositions lye in the range 15.380 - 111.94 kjmol-1.

The data obtained from the TG curve for zinc(II) complex, [Zn(HL)(H2O)(NO3)] (7), (Table-6), displays mainly three regions of changes. The first stage of thermal degradation in the temperature range 122 - 210 C is corresponding to dehydration of the complex and the DTG curve displays maximum peak at 186 C. In the second stage the TG curve shows a mass loss in the 100 - 216 AdegC region representing thermal removal of the coordinated nitrate anion with DTGmax peak at 358 C. The third stage represents a partial decomposition of the coordinated organic ligand in the temperature range 217 - 336 AdegC with a DTGmax peak at 560 AdegC. The final decomposition stage within the temperature rang of 402 - 544 AdegC, represents the further decomposition of the remaining portion of complex leaving behind the final product ZnO. The value of the final product deviates slightly from that theoretically expected for ZnO.

On the other hand the experimental mass loss values for the other thermal pyrolysis stages agree well with the theoretical expected mass losses. The observed activation energies of these thermal degradation stages are 24.500, 53.640 and 62.860 kjmol-1 for the first, second and third stage respectively (Table-6).

Characterization of coumarin 8 and benzocoumarin 9 compounds

13C-NMR spectrum of ISE showed one signal at I' 27.26 ppm for the methyl group, six signals for nine aromatic and imidazole carbon atoms from I' 119. 71 to 151.84 ppm and one signal at I' 198.33 ppm corresponding to the carbonyl group. H NMR spectrum of coumarin 8 showed three singlet signals at I' 2.31, 8.61 and 13.59 ppm for the methyl group, H4 of coumarin and NH group, as well as multiple signals for aromatic protons and CH=CH of imidazole from I' 7.25 to 7.86 ppm. 13C-NMR spectrum of benzocoumarin, 9 showed one signal at I' 39.50 ppm for the methyl group, sixteen signals for twenty-two aromatic and imidazole carbon atoms from I' 119.92 to 153.39 ppm and showed two signals at I' 155.91 and 158.21 ppm corresponding to 2C=O. Table-4 lists the 13C-NMR spectra chemical shifts (ppm) of coumarin 8 and benzocoumarin 9 in DMSO. Table-3 lists the 1H NMR spectra chemical shifts (ppm) of ISE, H2L ligand, coumarin 8 and benzocoumarin 9 in DMSO.

On the basis of the above discussion the general structures have been proposed for the present metal complexes as shown in Fig. 3.

Antimicrobial activity

The inspected metal complexes and the reported organic compounds, namely coumarin and benzocoumarin, were assessed for antimicrobial activity against three strains of Gram-positive bacteria Staphylococcus aureus [RCMB 010027], Streptococcus pneumonia [RCMB 010010] and Bicillis subtilis [RCMB 010067]); four Gram-negative bacteria Klebsiella pneumoniae [RCMB 0010093], Escherichia coli [RCMB 010052], Mycobactrium tuberculosis [RCMB010120] and Sallmonella typhimurium [RCMB 010315] and the fungi Aspergillus fumigatus [RCMB 02568]. The results of the antimicrobial study of the present newly synthesized compounds are presented in Table-7.

The antibacterial assay of compounds 3 and 4 showed good activity against all strains of Gram negative bacteria except (P. aeruginosa); compounds 2, 6, 8 and 9 showed good activity against two strains of this classification of bacteria (K. pneumonia and M. tuberculosis). On the other hand, complexes 2 and 6 exhibited significant antimicrobial activity against gram-negative bacteria Sallmonella typhimurium [RCMB 010315]. In addition, silver metal complex 6 showed good activity against Escherichia coli (RCMB 010052).

However, all tested compounds showed no activity against any strains of Gram positive bacteria nor against the Gram negative bacterium P. aeruginosa. The values of the zones of inhibition (ZOI) obtained indicated that metal complexes 3 and 4 demonstrated better activity against strains of Gram negative bacteria when compared with organic compounds, such as coumarin and benzocoumarin derivatives 8 and 9.

It is generally expected that metal complexes would be more active against Gram-positive than Gram-negative bacteria as the outer membrane layer of Gram-negative bacteria do not allow complexes to enter the cell. However, some of the metal complexes observed in the present study were more active against Gram-negative than Gram-positive bacteria (Table-7). This behavior suggests that these complexes increased lipophilicity, which favors permeation of complexes through the lipid layer of the cell membrane. These results are consistent with some previous studies [46, 47].

Regarding the anti-fungal activity of the tested compounds, only 3, 4, 6 and 9 showed moderate activity against Aspergillus fumigatus fungi and the other compounds presented no activity against Candida albicans. According to the antimicrobial results, most of the compounds showed low activity against fungi compared to their activity against bacteria (Table-7). This difference might be due to the differences between the cell structure of fungi and bacteria. The cell wall of fungi contains chitin and ergosterol, while the cell wall of bacteria contains murein [48, 49].

In vitro anticancer studies

The results of the in vitro anticancer activity of the synthesized Schiff base ligand (H2L), metal complexes and coumarin compounds 8 and 9 against the human colon carcinoma (HCT-116) cell line and human breast carcinoma cells (MCF-7) are presented in Table-8 and 9. The cytotoxic effects of the Schiff base ligand (H2L), as well as its metal complexes and the investigated organic compounds, on normal cells in the Baby Hamister Kidney (BHK) cell line is presented in Table-10. In addition, SI values for the present inspected compounds against the (HCT-116), (MCF-7) and Baby Hamister Kidney (BHK) cell line are displayed in Tables-11 and 12. According to the MCF-7 cell line results, complexes 6, 2 and 4 with SI values 84.72, 9.23 and 7.30 exhibited more significant activity than the reference drug (DOXORUBICIN; DOX with S.I. value 4.00), while compound 6 with SI value 5.36 was found to be nearly as active as DOX (Table-11).

The results of the HCT-116 cell line revealed that compounds 6, 7, 9 and 4 with SI values 29.84, 8.95, 7.26 and 7.25 showed better activity against the cell line than the reference drug DOX with SI value 4.00 (Table-12).

These results revealed that minute amounts of copper(II) 5 can cause cytotoxicity in the HCT-116 cell line, as compared to the higher amounts required to cause the same effect in the control cells. The significant differences noted regarding the cytotoxicity of the synthesized compounds in both the experiment and control cells were supported by the ratio of the concentration that caused half of the BHK cell line to die to the concentration that caused a similar proportion of death in HCT-116 [50-52].

Docking studies

Docking of Doxorubicin into DHFR [53], Fig. 4.

Fig.s 4-15 illustrate the docking of DOX and compounds into DHFR. Docking score energy of the novel compounds are listed in Table-13.

Docking of ISE into DHFR. One H-bond interaction as one O atom of SO2 group acted as a H- bond acceptor with the amino acid; Thr 56 (2.68 A) (50.9%). However, it revealed the existence of hydrophobic interactions involving other atoms of ISE with various amino acid residues as drown on Fig. 5.

Docking of H2L into DHFR. N atom of CN function represented as a H-bond acceptor with the side chain; Arg 70 (71.6 A) (2.86%). Furthermore, it revealed the existence of hydrophobic interactions involving other atoms of H2L with various amino acid residues as drown on Fig. 6.

Docking of compound 1 into DHFR. The O of the SO2 acted as a H-bond receptor of hydrogen with the residues of the amino acid; Thr 56 in conjunction with Ser 59 (2.28 A as well as 3.41 A) with strength (63.2% and 3.4%, respectively). In addition, H2O molecule acted as the donor of the H- bond with the residue of the amino acid; Val 115 (2.94 A) with strength 2.9%. Thus, the N atom of the CN acted as a H-bond acceptor of the amino acid; Val 8 coupled with Thr 136, respectively (3.20 A as well as 3.03 A) with strength 3.9% and 36%, respectively. Additionally, it revealed the existence of hydrophobic interactions involving other atoms of 1 with various amino acid residues as drown on Fig. 7.

Docking of compound 2 into DHFR. One O atom of SO2 moiety acted as a H-bond acceptor with the amino acid; Asn 64 (2.43 A) (15.8%). Besides, N3 of imidazole group acted as a H-bond acceptor with the amino acid; Arg 70 (3.06 A) (14.2%). Furthermore, it revealed the existence of hydrophobic interactions involving other atoms of 2 with various amino acid residues as drown on Fig. 8.

Docking of compound 3 into DHFR. N atom of CN function acted as a H-bond acceptor with the amino acid, Ser 59 (3.74 A) (4.9%). Also, one O atom of SO2 moiety acted as a H-bond acceptor with the amino acid; Asn 64 (2.45A) (17.3%). Moreover, N3 of imidazole moiety acted as a H-bond acceptor with the amino acid; Arg 70 (3.09 A) (13.1%). Furthermore, the water molecule acted as both a H- bond donor with the side chain residue Val 115 (2.69 A) (22.3%). In addition to, it revealed the existence of hydrophobic interactions involving other atoms of 3 with various amino acid residues as drown on Fig. 9.

Table-7: Anti-bacterial and anti-Fungal activities of the examined compounds a.

Comp. No.###Gram-positive Bacteria###Gram-Negative Bacteria###Fungi

###S. aureus###S. pneumoniae###B. subtilis###K. pneumoniae###E. Coli###M .tuberculosis###S. typhimurium###A. fumigatus

###ISE###NAc###NA###NA###NA###NA###NA###NA###NAb

###H2L###NA###NA###NA###NA###NA###NA###NA###NA

###1###16.9 +- 0.58###16.5 +- 044###18.4 +- 0.44###16.8 +- 0.63###17.3 +- 0.58###15.1 +- 0.37###18.3 +- 0.58###14.2 +- 0.58

###2###17.2 +- 0.63###17.8 +- 0.37###19.9 +- 0.25###21.6 +- 0.25###19.7 +- 0.63###19.6 +- 0.25###24.3 +- 0.63###15.6 +- 0.25

###3###20.1 +- 0.58###20.9 +- 0.58###20.2 +- 0.25###20.2 +- 0.37###27.8 +- 0.58###18.2 +- 0.24###24.3 +- 0.58###19.7 +- 0.32

###4###20.9 +- 0.44###22.8 +- 0.25###21.9 +- 0.36###21.8 +- 0.19###28.3 +- 0.58###20.1 +- 0.19###25.4 +- 0.44###20.4 +- 0.58

###5###15.2 +- 0.44###15.9 +- 0.58###16.2 +- 0.32###13.4 +- 0.44###15.2 +- 0.63###12.4 +- 0.37###16.8 +- 0.25###14.3 +- 0.37

###6###17.4 +- 0.25###17.9 +- 0.58###19.2 +- 0.44###22.5 +- 0.37###26.6 +- 0.25###20.4 +- 0.37###25.7 +- 0.48###19.2 +- 0.63

###7###16.2 +- 0.37###17.4 +- 0.44###18.3 +- 0.37###15.8 +- 0.12###18.2 +- 0.63###14.1 +- 0.25###16.2 +- 0.44###18.2 +- 0.25

###8###18.2 +- 0.44###19.8 +- 0.58###21.2 +- 0.37###18.7 +- 0.44###20.3 +- 0.36###17.1 +- 0.37###21.3 +- 0.25###17.3 +- 0.58

###9###19.3 +- 0.44###19.8 +- 0.44###20.4 +- 0.36###17.8 +- 0.36###19.2 +- 0.72###16.5 +- 0.25###20.1 +- 0.63###20.4 +- 0.58

St. Controlb###28.9 +- 0.14###25.3 +- 0.58###26.3 +- 0.34###17.3 +- 0.12###27.3 +- 0.44###16.3 +- 0.58###23.8 +- 0.63###23.7 +- 0.10

Table-8: Anti-breast cancer activity (MCF-7) of novel compounds.

Comp. No.###Validity for sample Conc. (ug/mL)

###50###25###12.5###6.25###3.125###1.56###0###IC50

###ISE###15.39 +- 1.3###28.54 +- 1.7###67.38 +- 3.2###84.64 +- 1.5###91.05 +- 09###97.78 +- 0.7###100###18.1 +- 0.1

###H2L###47.26 +- 2.5###73.46 +- 4.3###87.51 +- 1.4###94.17 +- 1.###98.23 +- 0.7###100###100###47.4 +- 0.1

###1###12.33 +- 1.0###28.02 +- 1.9###45.80 +- 2.7###73.53 +- 4.3###84.69 +- 1.5###94.42 +- 0.1###100###11.6 +- 0.1

###2###16.57 +- 1.2###47.85 +- 2.6###84.12 +- 1.5###93.56 +- 0.9###98.71 +- 0.6###100###100###24.3 +- 0.

###3###71.84 +- 4.2###87.53 +- 1.3###94.42 +- 1.1###98.71 +- 0.9###100###100###100###> 50

###4###15.79 +- 1.3###25.91 +- 2.2###47.84 +- 2.8###81.07 +- 1.5###92.72 +- 0.###98.45 +- 0.6###100###12.1 +- 0.1

###5###6.36 +- 1.2###10.89 +- 0.14###19.53 +- 1.5###33.18 +- 2.###64.49 +- 3.7###83.84 +- 1.4###100###4.57 +- 0.1

###6###3.41 +- 1.3###6.88 +- 2.2###10.56 +- 2.8###16.72 +- 1.1###20.94 +- 1.3###28.41 +- 1.9###100###0.53 +- 0.1

###7###4.12 +- 1.2###7.98 +- 0.9###18.83 +- 1.5###64.36 +- 3.8###82.51 +- 1.4###91.48 +- 0.9###100###8.22 +- 0.1

###8###16.22 +- 1.2###37.47 +- 2.0###78.94 +- 3.0###89.12 +- 1.5###95.38 +- 0.9###99.08 +- 1.9###100###21.2 +- 0.2

###9###13.89 +- 1.1###35.62 +- 2.3###76.48 +- 4.5###85.69 +- 1.3###93.13 +- 0.9###98.24 +- 1.9###100###20.6 +- 0.2

###DOX###12.82 +- 0.1###21.89 +- 0.1###43.83 +- 0.1###78.17 +- 0.1###89.28 +- 0.1###93.64 +- 0.1###100###10.68 +- 2.10

Table-9: Anti-colon cancer activity (HCT-116) of novel compounds. a

Comp. No.###Validity for sample Conc. (ug/mL)

###50###25###12.5###6.25###3.125###1.56###0###IC50

###ISE###21.23 +- 1.9###38.94 +- 2.###71.56 +- 4.1###83.87 +- 1.4###92.45 +- 1.0###98.74 +- 0.1###100###20.8 +- 0.1

###H2L###43.59 +- 2.###71.82 +- 3.###86.78 +- 1.4###94.06 +- 1.1###98.43 +- 0.7###100###100###44.3 +- 0.1

###1###11.84 +- 0.9.###20.93 +- 1.8###31.42 +- 2.1###58.96 +- 3.###86.57 +- 1.5###98.13 +- 0.6###100###4.36 +- 0.1

###2###87.13 +- 1.5###96.37 +- 0.9###99.42 +- 0.6###100###100###100###100###> 50

###3###48.25 +- 2.6###60.58 +- 3.###79.27 +- 4.4###91.04 +- 1.2###98.46 +- 0.6###100###100###46.5 +- 2.1

###4###14.91 +- 1.2###30.87 +- 2.0###48.52 +- 2.###82.34 +- 1.5###91.78 +- 0.8###98.12 +- 0.6###100###12.2 +- 0.1

###5###6.57 +- 0.1###9.3 +- 0.1###14.08 +- 1.2###30.97 +- 2.3.###62.38 +- 3.9###86.74 +- 1.4###100###4.36 +- 0.1

###6###4.18 +- 1.3###8.23 +- 2.2###16.78 +- 1.3###28.19 +- 1.9###36.42 +- 2.7###49.27 +- 2.8###100###1.51 +- 0.1

###7###6.83 +- 0.4###11.36 +- 0.8###24.05 +- 1.###35.94 +- 2.41###78.12 +- 3.8###89.08 +- 0.1###100###5.21 +- 0.1

###8###15.69 +- 1.1###27.81 +- 1.###46.03 +- 2.3###80.94 +- 1.5###91.21 +- 1.1###98.04 +- 0.6###100###11.8 +- 0.3

###9###13.53 +- 1.1###25.49 +- 1.7###43.87 +- 2.8###78.19 +- 4.###89.63 +- 1.5###97.41 +- 0.6###100###11.4 +- 0.2

###DOX###11.82 +- 0.1###10.89 +- 0.1###20.83 +- 0.1###42.17 +- 0.1###69.28 +- 0.1###89.64 +- 0.1###100###6.55 +- 2.10

Table-10: Cytotoxicity of compounds against Normal fibroblasts of Baby Hamister Kidney (BHK) cell line.a

###Comp. No.###Validity for sample Conc. (ug/mL)

###50###25###12.5###6.25###3.125###0###CC50

###ISE###52.91 +- 3.2###64.17 +- 3.4###79.46 +- 1.5###88.92 +- 1.3###95.18 +- 0.9###100###71.9

###H2L###80.96 +- 1.5###91.22 +- 1.3###96.38 +- 0.9###98.26 +- 0.6###100###100###161

###1###51.23 +- 3.3###68.19 +- 4.1###81.22 +- 1.2###90.78 +- 0.9###96.62 +- 0.8###100###55.3

###2###95.28 +- 0.7###98.76 +- 0.6###100###100###100###100###>200

###3###91.28 +- 1.1###97.54 +- 0.8###99.08 +- 0.6###100###100###100###>200

###4###58.97 +- 4.2###72.86 +- 1.5###87.54 +- 1.1###93.28 +- 0.9###98.71 +- 0.6###100###88.4

###5###40.97 +- 3.1###49.56 +- 3.8###60.34 +- 4.5###76.13 +- 1.5###88.92 +- 1.4###100###24.5

###6###45.27 +- 3.1###68.92 +- 4.4###83.75 +- 1.5###90.24 +- 1.3###97.56 +- 0.7###100###45

###7###48.31 +- 3.4###60.67 +- 4.2###74.23 +- 1.5###89.16 +- 1.1###94.37 +- 0.8###100###46.6

###8###48.37 +- 3.1###63.89 +- 4.2###76.84 +- 1.5###89.02 +- 1.1###93.87 +- 0.9###100###47.4

###9###61.86 +- 4.1###78.12 +- 1.5###84.91 +- 1.2###91.23 +- 0.9###97.42 +- 0.7###100###82.7

###DOX###42.33 +- 0.590###77.54 +- 0.756###83.05 +- 0.328###90.75 +- 0.439###100.0 +- 0.489###100###42.70

Table 11: Cytotoxicity of breast carcinoma cell line (MCF-7) and mammalian cells of normal fibroblasts of Baby Hamister Kidney (BHK) cell line on compounds. a

###Comp. No.###CC50 (g/mL)###CC50 (M)###IC50 (g/mL)###IC50 (M)###S.I.

###ISE###71.9###287.29 +- 0.54###18.1###72.32 +- 0.76###3.97

###H2L###161###485.89 +- 0.34###47.4###143.05 +- 0.67###3.40

###1###55.3###110.31 +- 0.44###11.6###23.14 +- 0.45###4.77

###2###> 200###366.15 +- 0. 28###24.3###44.49 +- 0.76###9.23

###3###> 200###395.78 +- 0.32###>50###98.95 +- 0.27###3.99

###4###88.4###175.02 +- 0.33###12.1###23.96 +- 0.56###7.30

###5###24.5###48.04 +- 0.38###4.57###8.96 +- 0.84###5.36

###6###45###94.89 +- 0.54###0.53###1.12 +- 0.56###84.72

###7###46.6###97.95 +- 0.54###8.22###17.28 +- 0.67###5.67

###8###47.4###108.72 +- 0.75###21.2###48.57 +- 0.68###2.24

###9###82.7###169.99 +- 0. 43###20.6###42.34 +- 0.35###4.01

###DOX###42.70###94.05 +- 2.15###10.68 +- 1.88###23.51 +- 2.10###4.00

Table-12: Cytotoxicity of colon carcinoma cell line (HCT-116) and Normal fibroblasts of Baby Hamister Kidney (BHK) cell line compounds. a

###Comp. No.###CC50 (g/mL)###CC50 (M)###IC50 (g/mL)###IC50 (M)###S.I.

###ISE###71.9###287.29 +- 0.38###20.8 +- 2.18###83.11 +- 0.98###3.46

###H2L###161###485.89 +- 0.67###44.3 +- 5.67###133.70 +- 0.83###3.63

###1###55.3###110.31 +- 1.01###8.28 +- 1.37###16.52 +- 0.67###6.68

###2###> 200###366.15 +- 0.67###> 50###91.54 +- 1.23###3.99

###3###> 200###395.78 +- 0.45###46.5###92.02 +- 0.45###4.30

###4###88.4###175.02 +- 1.07###12.2###24.15 +- 2.01###7.25

###5###24.5###48.04 +- 1.56###4.36 +- 2.78###8.55 +- 0.45###5.62

###6###45###94.89 +- 0.39###1.51###3.18 +- 0.49###29.84

###7###46.6###97.95 +- 0.48###5.21 +- 10.35###10.95 +- 0.78###8.95

###8###47.4###108.72 +- 0.76###11.8###27.04 +- 0.57###4.02

###9###82.7###169.99 +- 0.84###11.4###23.43 +- 0.55###7.26

###DOX###42.70###94.05 +- 2.30###6.55 +- 1.88###14.42 +- 2.10###6.52

Table-13: Docking score energy of the novel compounds.

###Compound###Score###E_conf.###E_place###E_score 1###E_refine###E_score 2

###ISE###-13.797###37.67665###-78.5349###-9.36795###-10.8012###-13.797

###H2L###-17.3951###63.52516###-74.4478###-10.2521###3.222391###-17.3951

###1###-19.4795###88.66785###-97.5022###-11.657###19.06533###-19.4795

###2###-20.5666###82.78984###-114.965###0###2.673034###-20.5666

###3###-18.7949###102.6056###-112.595###0###60.43481###-18.7949

###4###-19.7473###85.98631###-110.93###0###17.28483###-19.7473

###5###-18.4784###97.39848###-114.707###-10.3926###27.39854###-18.4784

###6###-17.8914###97.77918###-123.532###-10.2685###31.77247###-17.8914

###7###-18.093###73.65321###-134.911###-12.3487###1.247415###-18.093

###8###-21.5886###105.2864###-116.845###-11.535###34.07844###-21.5886

###9###-25.1034###124.8704###-93.6111###-12.2335###45.90167###-25.1034

Docking of compound 4 into DHFR. H-bond interactions between two O atoms of SO2 group as they acted as a H-bond acceptor with the side chain residue Asn 64 (2.96 A and 2.82 A, respectively) (14.5% and 18.7%), respectively. Also, water molecule acted as both a H-bond donor with the amino acid; Val 115 (2.28 A) (15.6%). Moreover, it revealed the existence of hydrophobic interactions involving other atoms of 4 with various amino acid residues as drown on Fig. 10.

Docking of compound 5 into DHFR. O atom of SO2 moiety acted as a H-bond acceptor with the amino acid; Ser 59 (3.24 A) (3.4%). Furthermore, the OH molecule acted as a H-bond donor with the amino acids; Ile 7 and Val 115 (3.74 A and 2.02 A, respectively) (9.9% and 59.2%), respectively. Additionally, the H2O molecule acted as both a H-bond donor with the amino acids; Ile 7 and Tyr 121 (2.88 A and 2.59 A, respectively) (2.7% and 60.5%), respectively and as a H-bond acceptor with the amino acids; Ala 9 and Tyr 121 (3.74 A and 2.59 A, respectively) with a strength of 2.8% and 60.5%, respectively. Furthermore, it revealed the existence of hydrophobic interactions involving other atoms of 5 with various amino acid residues as drown on Fig. 11.

Docking of compound 6 into DHFR. N atom of CN function acted as a H-bond acceptor with the amino acid; Asn 64 (2.60 A) (50.5%). Furthermore, O atom of SO2 moiety acted as a H-bond acceptor with the side chain; Thr 56 (3.13 A) (8.5%). Also, N3 of imidazole moiety acted as a H-bond acceptor with the amino acid; Ala 9 (3.63 A) (1.6%). Also, it revealed the existence of hydrophobic interactions involving other atoms of 6 with various amino acid residues as drown on Fig. 12.

Docking of compound 7 into DHFR. Two O atoms of SO2 moiety acted as a H-bond acceptor with the amino acid, Thr 56 (3.07 A) (16.6%). Moreover, the N atom of CN group acted as a H-bond acceptor with the side chain, Asn 64 (3.06 A) (18.8%). In addition to, it revealed the existence of hydrophobic interactions involving other atoms of 7 with various amino acid residues as drown on Fig. 13.

Docking of compound 8 into DHFR. O atom of the SO2 acted the H-bond acceptor of the residue of the amino acid, Ser 59 (3.01 A) (23.3%). Also, there is an arene cation interaction between Arg 70 coupled with the cumarene ring. Additionally, it revealed the existence of hydrophobic interactions involving other atoms of 8 with various amino acid residues as drown on Fig. 14.

Docking of compound 9 into DHFR. O atom of SO2 moiety acted as a H-bond acceptor with the amino acid, Arg 70 (3.21 A) (12.9%). Also, N3 of imidazole moiety acted as a H-bond acceptor with the amino acid, Lys 68 (3.04 A) (11.3%). In addition to, it revealed the existence of hydrophobic interactions involving other atoms of 9 with various amino acid residues as drown on Fig. 15.

Conclusions

In the present study, Ag(I), Fe(III), Zn(II), Mn(II), Co(II), Ni(II) and Cu(II) complexes prepared with N'-(1-(4-(1H-imidazol-2-ylsulfonyl)-phenyl)ethylidene)-2-cyanoacetohydrazide showed promising antimicrobial and anticancer activity. The results obtained indicated that metal complexes 2 and 6 present significant activity against three Gram (-ve) bacteria; silver metal complex 6 exhibits good activity against Escherichia coli while complexes 4, 6 and 9 present significant activity against both cell lines (breast and colon) when compared with DOX as a reference drug. The observed activity from molecular docking analysis showed that complexes 2, 8 and 9 may exert their action through inhibition of the DHFR enzyme. It can be concluded that, coordination of the metal ions Ag(I), Fe(III), Zn(II), Mn(II), Co(II), Ni(II) and Cu(II) to the synthesized Schiff base ligand (H2L) enhances its anti-microbial and anti-cancer activities.

Moreover, it was observed that metal ion complexes Co(II), Ni(II) and Ag(I) were more active than the other metal complexes, Fe(III), Zn(II), Mn(II) and Cu(II).

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