Synthesis, characterization and biological screenings of 5-coordinated Organotin(IV) complexes based on carboxylate ligand

Journal Pre-proof Synthesis, characterization and biological screenings of 5-coordinated Organotin(IV) complexes based on carboxylate ligand Muhammad Sirajuddin, Saqib Ali, Vickie McKee, Abdul Matin PII:

S0022-2860(20)30006-5

DOI:

https://doi.org/10.1016/j.molstruc.2020.127683

Reference:

MOLSTR 127683

To appear in:

Journal of Molecular Structure

Received Date: 22 October 2019 Revised Date:

26 December 2019

Accepted Date: 2 January 2020

Please cite this article as: M. Sirajuddin, S. Ali, V. McKee, A. Matin, Synthesis, characterization and biological screenings of 5-coordinated Organotin(IV) complexes based on carboxylate ligand, Journal of Molecular Structure (2020), doi: https://doi.org/10.1016/j.molstruc.2020.127683. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2020 Published by Elsevier B.V.

Synthesis, Characterization and Biological Screenings of 5-Coordinated Organotin(IV) Complexes Based on Carboxylate Ligand Muhammad Sirajuddina,*, Saqib Alib,*, Vickie McKeec, Abdul Matind,e a

Department of Chemistry, University of Science and Technology Bannu, 28100, Pakistan

b

Department of Chemistry, Quaid-i-Azam University, Islamabad 45320, Pakistan

c

Department of Chemistry, Dublin City University, Ireland

d

Department of Medical Laboratory Sciences, College of Applied Medical Sciences, Majmaah

University, Majmaah 11952, Saudi Arabia e

Department of Medical Lab Technology, University of Haripur, Hattar Road, Haripur, Khyber

Pakhtunkhwa, 22620, Pakistan *For correspondence: [email protected] (M. Sirajuddin) and [email protected] (S. Ali). Abstract This article focus on the synthesis of three 5-coordinated Sn(IV) complexes based on carboxylate ligand: 4-((4-methoxy-2-nitrophenyl)amino)-4-oxobutanoic acid. The complexes were characterized in both solid state (FT-IR, single crystal XRD and micro elemental analysis) as well as in solution state (1H,

13

C and

119

Sn NMR) and their results demonstrate that the

carboxylate moiety of the ligand is linked with Sn atom. A 5-coordinated trigonal bipyramidal structure was obtained from crystallographic data for the reported complexes with little bit distortion from the ideal structure. Preliminary antifungal screening showed that the tested compounds have 100% fungal inhibition capability as shown by the well know antifungal drug, Terbinafine. Bacterial inhibition results showed that these compounds have outstanding inhibition capability against various pathogens as compared to the market available standard antibacterial drugs, Cefixime and Roxithromicine. The antiproliferative/anticancer result against H157 cancer cell line revealed that compounds 1 and 2 have shown the maximum activity (4969% cytotoxicity even at 2 µg/mL final concentration). The tested compounds were found active against human corneal epithelial cells (HCEC). The synthesized compounds were almost nontoxic towards HCEC. Compounds 1 and 2 have shown 6.9% and 7.9% antiproliferative activity, respectively when used at the 2.0 µg/mL final concentration while compound 3 is totally nontoxic towards HCEC like the standard drug, Methotrexate. The antileishmanial results of these compounds are good though their activity is lower than that of the Amphotericin B. An 1

intercalative mode of interaction was observed during the interaction of the titled compounds with DNA. Keyword: Sn(IV) complex; Structural peculiarity; Anticancer activity; DNA interaction; Antimicrobial activity; Antileishmanial activity 1.

Introduction

Metal complexes have widespread application in the treatment of various common human diseases such as gastric and duodenal ulcers, rheumatoid arthritis, cancer and so many others. Due to these beneficial applications, the medicinal bioinorganic chemistry research area is getting considerable attention to search new metal based drugs [1]. The most threatening disease for the mankind in the present time is the cancer and for this purpose more than half million synthetic compounds were screened for their antitumor application but only about 25 of them gave fruitful results and are used in market today [1, 2]. The anticancer effect of the cisplatin, discovered by Rosenberg, is limited by some serious side effect like neuro-, nephro-, and ototoxicity [1]. To overcome these side effects, investigation for new metal based compounds other than platinum with limited toxicity and better clinical efficiency is on peak nowadays. Sn compounds (called organotins or organotin(IV) compounds which contain at least one direct SnC covalent bond) have got the prime importance among non-platinum metal based compounds because in case of organotin(IV) compounds there is no resistance of the cells against them and they also exhibit less toxicity than cisplatin derivatives [3]. The activity of organotin(IV) compounds depend both on the nature and on the number of organic group (alkyl or aryl) as Gielen pointed out that the anticancer activity of many organotin(IV) compounds against colon carcinoma and breast carcinoma cell lines is dependent on the nature of organic group attached to Sn center [4]. Moreover, from SAR (Structure Activity Relationship) of organotin(IV) compounds it is observed that the these compounds possess the following properties: availability of coordination position on Sn, existence of stable ligand-Sn bond and slow hydrolytic decomposition [5]. In orgnaotin(IV) compounds, triorganotin(IV) derivatives exhibit good cytotoxic activity than diorganotin(IV) derivatives because of possessing more free coordination position [6]. Among the organotins, organotin(IV) carboxylates have caught the concentration of the scientists because of their wide range biological and non-biological application mainly due to structural 2

diversity [7-9] as compared to organotin(IV) with thiolato and dithiocarbamato ligands [6, 10]. The prime biological applications of organotin(IV) carboxylates include as antitumor, antiviral, anti proliferative/anticancer, antileishmanial and antimicrobial agents, etc [11-18]. Keeping in mind the anti proliferative properties of triorganotin(IV) carboxylates, here in this paper we are investigating the synthesis and some important medicinal applications especially anticancer potential of 4-(4-methoxy-2-nitrophenylamino)-4-oxobutanoic acid based three novel triorganotin(IV) carboxylates. FT-IR, multi NMR, mass spectrometry, micro elemental analysis and single crystal X-ray analysis were used for successful characterization of the synthesized compounds. 2.

Experimental section

2.1.

Materials and methods

Trimethyl-, triethyl- and tributyltin chlorides and succinic anhydride were purchased Aldrich while Sodium salt of Salmon sperm DNA (SS-DNA) was obtained from Arcos and were used as such as received. The solvent used in the experiments were purchased from E. Merck, and Fluka and dried prior to use in accordance to the reported method [19]. Gallenkamp electrothermal device was used for recording the melting point of the desired compounds. Thermo Nicolet-6700 FT-IR Spectrophotometer equipped with DTGS (deuterated triglycine sulphate) detector was used to record the spectra of sodium 4-((4-methoxy-2-nitrophenyl)amino)-4-oxobutanoic acid and its three triorganotin(IV) complexes in the range 4000–400 cm−1. CE-440 Elemental Analyzer (Exeter

Analytical, Inc) was used for the determination of percent compositions of C, N and H. A 400 MHz JEOL ECS NMR was used for recording the 1H,

13

C and

119

Sn NMR spectra in

DMSO. In the NMR data interpretation, s, d, t, q, dd and m are used to represent singlet, doublet, triplet, quartet, doublet of doublet and multiplet, respectively. The DNA-compound interaction study was performed by 1800 Shimadzu UV-Visible Spectrophotometer. Bruker Apex II CCD diffractometer was used to collect the crystal data of the compounds. SHELXS-97 [20] and SHELX2012 [21] were used to solve and refine the structures. The poor quality of the crystal of compound 3 is reflected by the comparatively high value of the Rint parameter. MAT-311A Finnigan (Germany) mass spectrometer was used to record the mass spectra of the desired compounds with m/z values by considering H = 1, C = 12, N = 14, O = 16, Cl = 35, and Sn = 120. 3

2.2.

Synthesis

Synthesis of sodium N-[(2-methoxyphenyl)]-4-oxo-4-[oxyl]butanamide (NaL) N-(4-methoxy-2-nitrophenylamino)-4-oxobutanoic acid was synthesized according to our previously reported method [22]. Sodium N-(4-methoxy-2-nitrophenylamino)-4-oxobutanamide was synthesized by adding the aqueous solution of sodium hydrogen carbonate (NaHCO3) to a suspended solution of N-(4-methoxy-2-nitrophenylamino)-4-oxobutanoic acid in distilled water (Scheme 1) [17]. After the addition, a clear solution was obtained which on rotary evaporation of the water gives the desired sodium salt. Synthesis of triorganotin(IV) complexes Organotin(IV) carboxylates were synthesized by refluxing a mixture of R3SnCl (5 mmol) and the sodium salt of ligand NaL (5 mmol) in dry toluene for 6-8 h (Scheme 1) [17]. The refluxed solution was kept for overnight at room temperature. The NaCl precipitate was removed by filtration and the solvent was removed under the reduced pressure. The product was purified by recrystallization from chloroform at room temperature.

Scheme 1: Synthesis of NaL and its triorganotin(IV) complexes 1-3 2.3.

Biological applications

2.3.1. DNA interaction Study The binding ability of the desired compounds was SS-DNA was studied UV-visible spectroscopy according to the procedure described in our previously reported paper [22]. 1 mM solutions of the compounds were prepared in 70% ethanol and their absorbance was measured in the absence 4

and presence of various concentrations of SS-DNA. The UV absorption titrations were performed by keeping the concentration of the compound fixed while that of the DNA was from varied 9 µM to 81 µM [23, 24]. 2.3.2. Antimicrobial studies The potential of the desired compounds for bacteria killing was studied against the following bacterial pathogens two Gram-Positive: Micrococcus luteus and Staphylococcus aureus and two Gram-negative: Escherichia coli, Bordetella bronchiseptica using agar well-diffusion method as reported earlier [18, 25]. A 100 µL of the compounds (concentration: 1 mg/mL) prepared in DMSO was added to the respective wells. DMSO was used as a negative control while Roxyithromycin (1 mg/mL) and Cefixime (1 mg/mL) as positive controls. Triplicate plates of each bacterial strain were prepared, which were incubated aerobically at 37 ºC for 24 h. The activity was determined by measuring the diameter of the zone showing complete inhibition (mm). Similarly the potential of the desired compounds for fungal growth was checked against four fungal strains (Helminthosporium solani, Aspergillus niger, Fusarium solani and Mucor sp.) using agar tube dilution method according to the procedure described previously [18, 25]. 4 mL Sabouraud dextrose agar (SDA) medium was added to screw caped test tubes and were autoclaved at 121 ºC for 15 min and then allowed to cool at 50 ºC. A 66.6 µL of compounds from the stock solution (1 mg/mL in DMSO) was loaded to non-solidified SDA. DMSO and Turbinafine were used as a negative and a positive control, respectively. The tubes were incubated at 28 ºC for 7 days and growth was determined by measuring the linear growth (mm) and growth inhibition was calculated with reference to growth in the control [18, 25]. 2.3.3. Anticancer study The desired compounds were checked for their anticancer activity in two ways according to the procedure described in our previously reported paper [22]. The activity was performed by Skehan et al., method [26] in one case Vincristine was used as a standard drug with H-157 and BHK-21 cell lines with compounds concentrations (100, 10, 1 and 0.1 µg/mL) of the while in other case Methotrexate was used as a standard drug with H-157 and HCEC cell lines and with compounds concentrations (0.5-2 µg/mL) [27]. 2.3.4. Antileishmanial study 5

The synthesized compounds were evaluated for their antileishmanial activity performed against the promastigote forms of leishmania major using MTT assay and Amphotericin B as standard drug [28]. 10 µL of MTT was added to each well and plates were incubated for 3 h at 25 ± 1°C. Enzyme reaction was then stopped by the addition of 100 µL of 50% isopropanol and 10% sodium dodecyl sulfate. The plates were incubated for an additional 30 min. under agitation at room temperature. Relative optical density (OD) was then measured at a wavelength of 570 nm using a 96-well microplate reader. The background absorbance of plates was measured at 690 nm and subtract from 570 nm measurement. The absorbance of the formazan produced by the action of mitochondrial dehydrogenases of metabolically active cells is shown to correlate with the number of viable cells. All experiments were repeated at least three times. Results reported are mean of three independent experiments (± SEM) and expressed as percent inhibitions calculated by the formula [22]:

3. Results and discussion The synthesis of the desired three new triorganotin(IV) derivatives (methyl, ethyl and butyl) were carried out in an easy way of refluxing the sodium 4-((4-methoxy-2-nitrophenyl)amino)-4oxobutanoate suspending in dried toluene with the corresponding triorganotin(IV) derivatives in 1:1 molar ratio for 6-8 h. Their purity were first checked by the sharp melting point that was quite different from that of the precursors (melting point of NaL = 198-200ºC) and then were subjected to micro elemental analysis for the determination of the percent compositions of C, N and H as shown in Table 1. The close matching between the calculated and found values of C, N and H confirm the successful formation of the desired products. Table 1 also gives the detail of the various physical properties like molecular formula and molecular weight of the descried compounds. The synthesized compounds were air stable and were obtained in good yield (8385%). 3.1.

FT-IR

Table 1 shows the detail of the main characteristics peaks necessary for the confirmation of the synthesis of the desired compounds. First of all the sodium 4-((4-methoxy-2-nitrophenyl)amino)4-oxobutanoate was confirmed by the replacement of the H at 3107 cm-1 of the OH group of the carboxylate moiety with Na. The sodium 4-((4-methoxy-2-nitrophenyl)amino)-4-oxobutanoate 6

was then used as starting material for getting the desired organotin(IV) complexes. In the Far region of the spectra of the complexes the existence of Sn–C and Sn–O in the range of 546–521 cm-1 and 452–441 cm-1, respectively proves the synthesis of organotin(IV) complexes [29-31]. The binding mode of the carboxylate ligand in the organotin(IV) carboxylate complexes was determined by calculating the value of ∆ν. The value of ∆ν was calculated as: ∆ν = ν (COOasym) – ν (COOsym). Upon interaction between O and Sn atoms the value of ν (COOasym decreases while that of ν (COOsym increases and as a result the value of ∆ν decreases [8, 32]. As shown in Table 1, the ν (COOasym) and ν (COOsym) in NaL appear at 1536 cm-1 and 1275 cm-1, respectively giving the ∆ν = 261. But for organotin(IV) complexes the value of ∆ν is less than that of NaL which reflect the chelating or asymmetric bridging bidentate nature of the carboxylate moiety with 5-coordinated geometry around the Sn atom [8, 33]. 3.2.

Mass Spectrometry

Table 1 describes the mass data of the desired compounds obtained by Electrospray ionization (ESI) method. The data shows that there is a close matching between the resultant and expected fragments of the compounds. Three fragmentation patterns were proposed, based on observed m/z in their spectra in which two patterns give the final end product in the form of [R']+ and other pattern give [Sn]+ as end product as shown in Scheme 2 [17, 34].

7

Table 1: Physical, micro elemental composition, FT-IR and Mass spectrometry data of the synthesized compounds* Comp. %age Melting point Formula Formula Carbon Hydrogen Nitrogen No Yield (°C) weight calculated/found calculated/found calculated/found 85 145-147 431.0 C14H20N2O6Sn 39.0 (39.3) 4.7 (4.5) 6.3 (6.3) 1 2

83

113-115

473.1

C17H26N2O6Sn

43.0 (42.9)

5.5 (5.5)

5.9 (5.8)

3

85

68-70

557.3

C23H38N2O6Sn

49.6 (49.2)

6.9 (6.9)

5.0 (4.8)

FT-IR data, ν (cm-1) NH

C=Oamide

COOasym

COOsym

∆ν

Sn-C

Sn-O

NaL

3365

1689

1536

1275

261

-

-

1

3373

1699

1507

1357

150

538

448

2

3378

1696

1511

1366

145

521

441

3

3358

1704

1509

1371

138

546

452

Mass spectrometry data, ESI-MS, m/z (%) 1

[C14H20N2O6SnNa]+ = 455 (80); [C14H20N2O6Sn]+ = 432 (2); [C11H11N2O6]+ = 267 (5); [C10H11N2O4]+ = 223 (8); [C3H9Sn]+ = 165 (100); [C2H6Sn]+ = 150 (5); [CH3Sn]+ = 135 (15); [Sn]+ = 120 (20); [C13H17N2O6Sn]+ = 417 (23); [C12H17N2O4Sn]+ = 373 (34); [C11H14N2O4Sn]+ = 358 (21); [C10H11N2O4Sn]+ = 343 (10).

2

[C17H26N2O6SnNa]+ = 497 (20); [C17H26N2O6Sn]+ = 474 (5); [C11H11N2O6]+ = 267 (4); [C10H11N2O6]+ = 223 (5); [C6H15Sn]+ = 207 (100); [C4H10Sn]+ = 178 (15); [C2H5Sn]+ = 149 (10); [Sn]+ = 120 (25); [C15H21O6N2Sn]+ = 445 (6); [C14H21O4N2Sn]+ = 401 (1); [C12H16N2O4Sn]+ = 372 (3); [C12H16N2O4]+ = 252 (5)

3

[C23H38N2O6SnNa]+ = 581 (21); [C23H38N2O6Sn]+ = 558 (25); [C11H11N2O6]+ = 267 (4); [C10H11N2O4]+ = 223 (5); [C12H27Sn]+ = 291 (100); [C8H18Sn]+ = 234 (5); [C4H9Sn]+ = 177 (5); [Sn]+ = 120 (10); [C19H29O6N2Sn]+ = 501 (1); [C18H29N2O4Sn]+ = 457 (4); [C14H20N2O4Sn]+ = 400 (10); [C14H20N2O4Sn]+ = 343 (4).

* See Scheme 1 for compound numbering.

8

Scheme 2: Mass fragmentation pattern for the synthesized compounds 3.3. NMR results The detailed data of 1H,

13

C and

119

Sn NMR of the NaL and its three organotin(IV) complexes

recorded in DMSO is given Table 2. The absence of the carboxylic proton in the spectrum of the NaL confirms its formation. The other proton such as NH, aliphatic protons (H2 and H3) and aromatic protons (H6, H7 and H9) appear in their respective regions. The protons of the organic moiety (especially methyl derivative) attached to Sn atoms give important information regarding the geometry. In complex 1, the methyl protons resonate at 0.37 ppm with clear satellite peaks due to coupling with

119

Sn,

117

Sn and

115

Sn isotopes. The

coupling constant values calculated for complex 1 for 2J[119/117/115Sn-1H] are 70, 68, 40 Hz. The coupling constant value of the most dominant isotope

119

Sn (70 Hz) falls in the range of 5-

coordinated trigonal bipyramidal geometry (TBPG) about Sn atom in solution state [23]. In 9

complex 2, the ethyl protons attached to Sn atom appear as a quartet at 1.04 ppm (Hα) and as a triplet at 1.21 ppm (Hβ). Similarly in complex 3, Hα and Hδ of the butyl protons give triplets Hβ and Hγ give multiplet. The value of the 2J[119Sn-1H] = 70 Hz observed in complex 1 was put into the Lockhart’s equation to calculate the C-Sn-C angle (θ) that was equal to 118º which also satisfy the range of 5-coordinated TBPG [35]. In

13

C NMR the most important phenomenon observed was the downfield shifting of the C=O

(C1 and C4) upon complexation with Sn atom which is due to the shifting of electron density from the ligand (in free ligand C1 was observed at 170.7 ppm while C4 at 174.1 ppm [28]) toward the Sn atom [36]. The other carbons peaks were observed in their respective regions. The value of 1J[119Sn–13C] observed in

13

C NMR spectrum of complex 1 was put in the

Lockhart’s equation to find the C-Sn-C bond angle which comes equal to 123º. Similarly for complexes 2 and 3 Howard et al. equation [36] was used to calculate the C-Sn-C bond angle from 1J[119Sn–13C] value. For complexes 2 and 3 the value of C-Sn-C bond angle was 128º and 121º, respectively (Table S1 of supplementary data). The value of C-Sn-C bond angle for all the three complexes fall in the range of 5-coordinated TBPG [35]. 119

Sn NMR was also done for the desired complexes and gives a sharp single (Fig. S1 of the

Supplementary data) in all the three complexes (Table 2) which an indication of the presence of one species in solution. However the 119Sn values appear in highly shielded region which may be attributed to solvent effects [37].

10

Table 2: 1H NMR, 13C and 119Sn NMR data of the synthesized compounds* Proton No. Comp. No NaL

1

2

3

H2

H3

NH

H6

H7

H9

2.40,

2.22,

11.62,

7.48, d

7.28, dd

7.42, d

t (6.8)

t(6.4)

s

(8.8)

(2.8)

(2.8)

2.44,

2.30, t

10.0,

7.51, d

7.31, dd

7.45, d

t (6.6)

(6.6)

s

(8.7)

(3)

(3)

2.51,

2.38, t

10.02,

7.53, d

7.31, dd

7.47, d

t (7.8)

(7.8)

s

(9.0)

(2.7)

(2.7)

2.47,

2.34, t

10.04,

7.56, d

7.29, dd

7.45, d

t (6.6)

(6.6)

s

(9.0)

(3)

(3)

H11

α

β

γ

δ

3.82, s

-

-

-

-

-

-

-

-

-

1.27, m

0.84, t (7.4)

0.37, s 3.82, s

[70, 68, 40]

3.82, s

3.81, s

1.21, t

1.04, q

(8.1)

1.04, t

1.54, q

(8.1)

119

Carbon No.

Sn

C11

Α

β

γ

δ

NaL

172.1 33.7 33.4 175.7 126.6 124.6 119.9 155.4 108.6 143.1 55.7

-

-

-

-

-

1

171.4 31.3 30.7 176.1 127.6 124.9 120.8 156.4 109.4 143.7 56.5

-

-

-

-9.7

2

171.4 32.8 31.3 176.3 127.5 125.0 120.8 156.3 109.4 143.6 56.5 11.3[581] 10.7[32]

-

-

3

171.3 32.9 31.5 176.1 127.3 125.1 120.8 156.3 109.4 143.3 56.5 19.5[464] 28.2[28]

C1

C2

C3

C4

C5

C6

C7

C8

C9

C10

0.67 [521,496]

27.0 [74,35]

14.1

19.2 17.4

nJ[119/117/115Sn-1H]and nJ[119/117/115Sn-13C] in Hz in square brackets. 3/4J[1H-1H] in Hz in parenthesis. * See Scheme 1 for compound numbering as well as for NMR numbering pattern

11

3.4. X-Ray Crystallography discussion Figures 1 and 2 describe the molecular structure along with atomic numbering scheme and packing diagram along with unit cell of complex 1 while Figures 3 and 4 describe that of complex 2 and Figures 5 and 6 describe that of complex 3, respectively. Tables 3-5 describe the crystal data along with structure refinement parameters, selected bond lengths, bond angles and H-bonding, respectively. The molecules exist in polymeric form because of the presence of the 4((4-methoxy-2-nitrophenyl)amino)-4-oxobutanoate units that asymmetrically binds two Sn atoms:

one through O1 and other through O2. The environment around the Sn atom comprise of two oxygen atoms of two carboxylate moieties and three alkyl groups of the organic moieties. The alkyl groups lie at the equatorial positions while the two oxygens lie at the axial positions resulting in trigonal bipyramidal geometry (TBPG). However, the geometry is slightly distorted from the ideal one as determined by τ value (τ = (β-α)/60, where β and α are 1st and 2nd largest basal angle around the tin atom, respectively) [38]. The τ value for the complexes 1, 2 and 3 are 0.83, 0.82 and 0.73, respectively that are typical for the distorted TBPG [17]. Intramolecular and intermolecular interactions play a vital role in the stabilization of crystal structure. One of the remarkable features of these compounds is the presence of C–H---O intramolecular interactions are observed and their bond distance C6-H6···O3 is 2.02-2.381Å as shown in Table 6. All the three complexes display beautiful 3D-packing diagram mainly due to the presence of intramolecular C–H---O interactions, C–H---π interactions and intramolecular hydrogen bonds (N1–H1---O3). These interactions stabilize the polymeric chains in zigzag manner which are linked into a three-dimensional network via C–H---π interactions. Moreover, extended networks of O-Sn-O, C–H--O and C–H---π contacts lead to aggregation and a supramolecular assembly.

12

Table 3: Crystal data and structure refinement parameters for complexes 1-3 Parameters

1

2

3

Chemical formula Molecular weight Temperature for data collection (K) Crystal system Space group a (Å) b (Å) c Å)

C14H20N2O6Sn 431.01

C17H26N2O6Sn 473.09

C23H38N2O6Sn 557.24

150(2)

150(2)

150(2)

Monoclinic P21/c 16.632(3) 10.7400(18) 9.869(3)

Monoclinic P21/c 12.6943(14) 10.2081(11) 16.5523(18)

α, β, γ (°)

90, 104.468(16), 90

90, 110.245(2), 90

V(Å3) Z ρ(mg/mm3) µ(mm-1) F(000) Crystal size/mm3 2θ range for data collection Radiation MoKα (λ) Reflections collected

1707.0(6) 4 1.677 1.527 864 0.50×0.26×0.06

2012.4(4) 4 1.561 1.303 960 0.55×0.24×0.09

Triclinic P-1 9.796(12) 10.261(14) 28.17(6) 90.46(19), 91.90(17), 94.58(10) 2821(8) 4 1.312 0.940 1152 0.33×0.26×0.05

4.6 to 61.0°

3.4 to 56.6°

1.5 to 55.1°

0.71073 19412 5162 [Rint = 0.195, Rsigma = 0.111]

0.71073 20153 5007 [Rint = 0.032, Rsigma = 0.028]

0.71073 7380 7084 [Rint = 0.108, Rsigma = 0.210]

5161/0/211

5007/0/242

7084/0/274

0.988

1.021

1.474

0.040, 0.100

0.026, 0.061

0.198, 0.497

0.047, 0.104 1018674

0.033, 0.063 1018675

0.257, 0.517 1018676

Independent reflections Data/restraints/ Parameters S on F2 Final R indexes [I>=2σ (I)] Final R indexes [all data] CCDC#

13

Table 4: Selected bond lengths (Å) for complexes 1-3 1 Sn1-O1

2.323(2)

Sn1-C12

2.125(3)

Sn1-O2

2.217(2)

Sn1-C13

2.125(3)

C2-C1

1.522(3)

Sn1-C14

2.122(3)

O1-C1

1.250(3)

O2-C1

1.278(3)

2 Sn1-O1

2.347(1)

Sn1-C12

2.140(3)

Sn1-O2

2.227(1)

Sn1-C14

2.138(3)

C2-C1

1.514(3)

Sn1-C16

2.136(2)

O1-C1

1.251(2)

O2-C1

1.276(2)

3 Sn1-O1

2.22 (2)

Sn1-C12

2.28 (3)

Sn1-O8

2.400 (2)

Sn1-C16

2.47 (3)

O1-C1

1.38 (4)

Sn1-C20

2.09 (4)

O2-C1

1.16 (3)

Sn2-C35

2.17 (5)

Sn2-O7

2.36 (3)

Sn2-C39

2.32 (3)

Sn2-O2

2.569 (2)

Sn2-C43

2.05 (4)

O7-C24

1.33 (4)

O8-C24

1.09 (3)

14

Table 5: Selected bond angles (º) for complexes 1-3 1 C12-Sn1-O1

87.4(1)

O2-Sn1-C12

93.94(9)

C13-Sn1-O1

86.9(1)

O2-Sn1-C13

88.9(1)

C14-Sn1-O1

90.4(1)

O2-Sn1-C14

92.08(9)

O2-Sn1-O1

175.69(7)

C13-Sn1-C12

118.0(1)

O2-C1-O1

122.3(2)

C14-Sn1-C12

126.0(1)

C2-C1-O1

120.8(2)

C14-Sn1-C13

115.7(1)

2 C12-Sn1-O1

87.71(8)

O2-Sn1-C12

94.34(9)

C14-Sn1-O1

89.68(9)

O2-Sn1-C14

94.21(9)

C16-Sn1-O1

86.01(8)

O2-Sn1-C16

87.64(8)

O2-Sn1-O1

173.50(6)

C14-Sn1-C12

124.5(1)

O2-C1-O1

122.6(2)

C16-Sn1-C12

120.6(1)

C2-C1-O1

120.9(2)

C16-Sn1-C14

114.5(1)

3 C12-Sn1-O1

90.4 (10)

O7-Sn2-C35

105.0 (15)

C16-Sn1-O1

85.8 (10)

O7-Sn2-C39

82.9 (10)

C20-Sn1-O1

101.3 (14)

O7-Sn2-C43

88.2 (14)

C16-Sn1-C12

120.7 (12)

C39-Sn2-C35

131.2 (15)

C20-Sn1-C16

134.4 (13)

C43-Sn2-C35

114.8 (19)

C20-Sn1-C12

104.4 (15)

C43-Sn2-C39

113.5 (16)

O8-Sn1-O1

175.8 (8)

O2-Sn2-O7

174.8 (7)

15

Table 6: Hydrogen-bond angles and bond lengths (Å,˚) for complexes 1-3 D-H····A

D-H

H····A

D····A

D-H····A

1 N1-H1···O5

0.880

2.052

2.635

122.76

C6-H6···O3

0.950

2.381

2.889

113.14

2 N1-H1···O6

0.877

1.938

2.627

134.42

C6-H6···O3

0.950

2.193

2.836

124.00

3 N1-H1···O5

0.88

2.37

3.01(4)

130.1

C6-H6···O3

0.95

2.02

2.78(5)

134.8

N3-H3···O11

0.88

2.01

2.73(4)

138.8

C29-H29···O9

0.95

2.44

3.04(5)

120.9

Symmetry transformations used to generate equivalent atoms for complex 3: (i) +X,+Y,1+Z

Fig. 1: Molecular structure of the 1 with atoms numbering scheme. For clarity purpose H atoms are removed. 16

Fig. 2: 3D-Packing digram of 1 viewed along b-axis along with unit cell.

Fig. 3: Molecular structure of the 2 with atoms numbering scheme. For clarity purpose H atoms are removed.

17

Fig. 4: 3D-Packing digram of 2 viewed along b-axis along with unit cell.

Fig. 5: Molecular structure of the 3 with atoms numbering scheme. For clarity purpose H atoms are removed. 18

Fig. 6: 3D-Packing digram of 3 viewed along b-axis along with unit cell. 3.5. DNA binding study by UV-Visible spectroscopy The interaction between the desired compounds and SS-DNA was performed on UV-Vis. absorption spectroscopy. During the experiment the conc. of the compound was kept constant while that of DNA was varied as shown in Figures 7, 8 and 9. The Figures show two strong bands at 235-236 nm and 361–363 nm. The band of 235–236 nm may due to the absorptions of n(N)→π*(CO) and n(O)→π*(CO) electron transition while the band at 360-363 nm may due to π–π* (aromatic group). During the interaction two phenomenons were observed: hypochromism along with bathochromic effect upto 4 nm. When these two phenomenons occur then the dominant mode of interaction is intercalation which involves strong stacking between the chromophore and the base pairs of DNA [39]. Considering the approximate planar structure of the compounds, it suggests that the main part of compound, especially the carbonyl group (CO), should have intercalated the neighboring base pairs of DNA, and the π orbital coupling between π* orbital of CO and π orbitals of DNA bases could exist. Therefore, based on this viewpoint, the 19

interaction between compounds and SS-DNA could be noncovalent intercalative binding [4043]. Benesi–Hildebrand equation [44] was used to calculate the binding constant value of the compound-DNA adduct as its values are shown in the inset graph of the Figures 7, 8 and 9. Similarly the value of Gibb's free energy (∆G) determined from the equation: ∆G = -RT lnK, are also given in the inset graphs. The negative value of ∆G shows the spontaneity of compoundDNA adduct.

Fig. 7: UV-visible graph of compound 1-DNA (Conc. of compound = 1 mM and conc. of DNA = 0 (a), 9 (b), 18 (c), 27 (d), 36 (e), 45 (f), 54 (g), 63 (h), 72 (i) and 81 (j) µM). Binding constant (K) and Gibb’s free energy (∆G) are obtained from the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1.

20

Fig. 8: UV-visible graph of compound 2-DNA (Conc. of compound = 1 mM and conc. of DNA = 0 (a), 9 (b), 18 (c), 27 (d), 36 (e), 45 (f), 54 (g), 63 (h), 72 (i) and 81 (j) µM). Binding constant (K) and Gibb’s free energy (∆G) are obtained from the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1.

21

Fig. 9: UV-visible graph of compound 3-DNA (Conc. of compound = 1 mM and conc. of DNA = 0 (a), 9 (b), 18 (c), 27 (d), 36 (e), 45 (f), 54 (g), 63 (h), 72 (i) and 81 (j) µM). Binding constant (K) and Gibb’s free energy (∆G) are obtained from the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1. Antimicrobial activity data Table 7 describes the in vitro antibacterial activity results of the desired compounds against four bacterial strains. It can be seen from the data that all the three compounds are highly active against the studied bacterial strains. The order of activity for the desired compounds is as: 2 > 1 > 3 > NaL. The activity of compound 2 is even higher that both the standard drugs: Cefixime and Roxithromicine. The bactericidal and bacteriostatic effects of the tested compounds may be responsible for their antibacterial activity [33]. Table 8 describes the antifungal activity of the tested compounds against four fungal strains. It can be seen from the data that all the three compounds show outstanding activity against the studied fungal strains. The order of activity for the desired compounds is as: 2 ~ 3 > 1 > NaL. Compounds 2 and 3 show 100% growth inhibition like the standard drug, Terbinafine. Compound 1 shows 100% growth inhibition against A. niger and F. solani while 90% against Mucor sp. and H. solani fungal strains. The antibacterial activity of the tested compounds may be due to their interference with the synthesis of cellular walls, causing damage that can lead to altered cell permeability characteristics or disorganized lipoprotein arrangements, ultimately resulting in cell death [24].

22

Table 7: Zone of inhibition (mm) of antibacterial activity of the synthesized compoundsa Comp. No

E. coli

S. aureus

M. luteus

B. bronchiseptica

NaL

0

0

12 ± 0.5

0

1

30 ± 1.3

41 ± 1.1

24 ± 0.2

28 ± 1.1

2

35 ± 1.5

45 ± 1.4

32 ± 1.3

30 ± 1.3

3

18 ± 1.0

15 ± 1.3

21 ± 1.1

24 ± 1.1

Cefixime

30 ± 1.2

42 ± 1.5

38 ± 1.5

35 ± 2.8

Roxithromicine

26 ± 1.0

27 ± 1.2

30 ± 1.2

24 ± 1.0

b b

MIC (µg/mL) 1

2.0

4.9

10

11.5

2

1.3

3.4

6.9

5.3

3

4

6

4.5

14.3

Cefixime

0.2

8

2

4

Roxithromicine

8

2

4

32

b b

a) See Scheme 1 for compound numbering. b) Reference drugs

Table 8: Zone of inhibition (mm) of antifungal activity of the synthesized compoundsa Comp. No

Mucor sp.

A. niger

F. solani

H. solani

NaL

80 ± 1.0

85 ± 1.1

95 ± 1.5

87 ± 1.3

1

90 ± 1.2

100

100

96 ± 1.4

2

100

100

100

100

3

100

100

100

100

DMSO

0

0

0

0

Terbinafine

100

100

100

100

b c

a) See Scheme 1 for compound numbering. b) Vehicle control. c) Reference drug

Anticancer activity results The anticancer potential of the desired compounds that was checked by two group of researchers: one with H-157 and HCEC cell lines while Methotrexate as a standard drug and second with H-

23

157 and BHK-21 cell lines while Vincristine as a standard drug. Their results are presented in Figure S2 (supplementary data) and Table 9, respectively. Figure 10 explores the cytotoxic effect evaluation of the investigated compounds on the growth and proliferation of cancerous (H157) as well as normal cell lines (HCEC) using the MTT assay [45]. It can be seen from Figure S1 that the anticancer potential of all the compounds is dose dependent. The activity of compounds 1 and 2 (49-69% cytotoxicity even at 2 µg/mL final concentration) against H157 cell line is much higher than that of the Methotrexate while that of compound 3 and NaL is comparable to that of the Methotrexate. The results also revealed that compounds 1 and 2 have little effect on normal cell line HCEC while compound 3 and NaL are totally non-toxic like the Methotrexate. Interestingly, lowering the compound concentration had little effect on their antiproliferative activity and all compounds maintained their antiproliferative activity near to 8% even at the 0.5 µg/mL final concentration Similarly the results of second group evaluated against BHK21 and H157 cell lines are shown in Table 9. The tabulated data reveal that the activity of the desired compounds against BHK-21 and H157 is slightly lower than that of Vincristine. But overall the anticancer potential is good and are able to be apply for cancer treatment after additional studies that are needed to verify this data in other normal cells and cancer cell types to assess the effectiveness of the synthesized compound in an in vivo model system.

24

Table 9: Anticancer activity of NaL and 1-3a against BHK21 and H157 cell lines Cytotoxicity (%) against BHK21

Cytotoxicity (%) against H157

Dose Conc. (µg/mL)

Dose Conc. (µg/mL)

Comp. No 100

10

1

0.1

100

10

1

0.1

NaL

62.7 ± 1.4

58.9 ± 1.3

49.9 ± 1.6

31.8 ± 2.1

62.7 ± 2.1

54.5 ± 2.5

49.9 ± 2.0

35.8 ± 1.4

1

69.7 ± 1.5

63.8 ± 2.4

51.7 ± 1.4

41.2 ± 1.6

65.6 ± 2.0

60.3 ± 0.8

54.2 ± 2.7

41.7 ± 1.2

2

71.8 ± 1.4

60.8 ± 1.6

55.6 ± 1.7

48.2 ± 1.9

73.2 ± 1.3

62.6 ± 1.5

51.9 ± 1.3

40.2 ± 2.8

3

69.3 ± 1.3

65.8 ± 2.1

62.4 ± 1.5

40.5 ± 1.4

63.7 ± 1.6

61.5 ± 1.3

50.4 ± 1.4

35.3 ± 2.3

Vincristineb

74.5 ± 2.9

72.6 ± 3.1

70.9 ±2.4

69.8 ± 1.9

74.5 ± 2.9

72.6 ± 3.1

70.9 ± 2.4

69.8 ± 1.9

a) See Scheme 1for compounds numbering. b) Standard drug

25

Antileishmanial activity results Table 10 describes the antileishmanial potential of the screened compounds in comparison with the market available standard drug, Amphotericin B. The tabulated data shows that the antileishmanial potential of the screened compounds was evaluated at four different concentration doses and all of them have shown good antileishmanial potential in comparison standard. Among the screened compounds, compound 2 has shown the maximum antileishmanial potential of about 70.5 at 100 µM concentration dose which is very much near to the activity of the standard 79.8. The antileishmanial activity of the tested compounds may be due to interference with the function of parasite mitochondria [28]. The antileishmanial data therefore demonstrate the potential use of these compounds for the treatment of leishmania. Table 10: Antileishmanial activity data of NaL and 1-3a Dose Conc. (µg/mL)

Comp. No 100

10

1

0.1

NaL

55.4 ± 1.5

51.3 ± 1.3

43.2 ± 2.2

36.4 ± 1.2

1

65.2 ± 1.2

57.9 ± 1.7

52.9 ± 1.5

38.6 ± 1.3

2

70.5 ± 1.7

60.2 ± 2.1

54.1 ± 1.6

38.2 ± 1.1

3

63.8 ± 2.1

59.7 ± 1.2

45.3 ± 1.3

39.3 ± 1.7

Amphotericin Bb

79.8 ± 1.8

76.3 ± 1.4

74.8 ± 2.7

69.8± 2.3

a) Numbering of compounds is according to the Scheme 1 b) Standard drug

Conclusions The synthesis of three new 5-coordinated triorganotin(IV) carboxylate derivatives were successfully performed and were analyzed by various spectroscopic techniques. The results of solid state FT-IR and single crystal X-ray crystallography as well as solution state multinuclear NMR suggest the coordination of the carboxylate ligand takes place via oxygen atom of the carboxylate moiety. The antimicrobial activity of the synthesized compounds is either comparable or even higher than the corresponding standard drugs. The possible mode of binding of the titled compounds (1-3) with DNA is the intercalative mode of interaction as their interaction with DNA cause hypochromic effect as well as bathochromic shift. The evaluation of the desired compounds for anticancer activity reveals that 1 and 2 have very little effect on normal cell line HCEC while 3 and NaL are totally non-toxic like the standard drug, Methotrexate. The 26

antileishmanial activity of these compounds is good although slightly lower than that of the standard drug, Amphotericin B.

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31

Highlights • Synthesis of new triorganotin(IV) carboxylates • Interaction with SS-DNA via intercalative mode of interaction • In vitro antimicrobial against various strains • Anticancer Activity against H157, BHK21 ad HCEC cell lines • Antileishmanial activity against leishmanial major strain kwh 23

Author Contribution Statement

Muhammad Sirajuddin: Conceptualization, Methodology, Software, Writing- Original draft preparation Saqib Ali: Supervision Vickie McKee: Crystal analysis ,Abdul Matin: Biological study

Conflict of interest There is no conflict of interest