Synthesis, spectroscopic characterization, antileishmanial, DNA binding, antioxidant, cytotoxicity, antifungal and antibacterial studies of organotin(IV) carboxylate complexes

2.2.1 Synthesis of ligand (NaL)

4-(aminomethyl)cyclohexanecarboxylic acid (0.5 g, 3.18 mmol) was suspended in 15 mL methanol in two necks round bottom flask fitted with a condenser. A clear solution was obtained after being stirred for 15 min. Equimolar solution of 5-chloro-2-hydroxyacetophenone (0.54 g, 3.18 mmol) was added to the reaction system. The general reaction is shown in Scheme 1. The reaction mixture was refluxed for 6 h. To this solution sodium bicarbonate (0.26 g, 3.18 mmol) was added and refluxed for 1 h. A bright yellow compound was obtained after evaporating the solvent at room temperature.

Close

Scheme 1

Synthesis of sodium salt of ligand (NaL).

Export to PPT

Yield 76%, m.p 174–176 °C; anal. (calc.) for C₁₆H₁₉ClNNaO₃: C, 56.13 (57.92); H, 5.11 (5.77); N, 3.96 (4.22);IR (cm⁻¹): 1686 υ(COO)_asym, 1438 υ(COO)_sym, 1586 υ(C = N), 3535 υ(OH); ¹H NMR (CDCl₃-d) δ (ppm):12.14 (s, 1H, OH), 7.22 (d, J = 8.8 Hz,1H, phenyl), 7.45 (s, 1H, phenyl), 7.18 (d, J = 8.9 Hz, 1H, phenyl), 3.39 (d, J = 2.5 Hz, 2H, N-CH₂), 2.62 (s, 3H, CH₃),2.10 (p, J = 6.4 Hz, 1H, cyclohexyl), 1.91–1.93 (m, 2H, cyclohexyl), 1.48–1.51 (m, 2H, cyclohexyl), 1.11–1.26 (m,1H, cyclohexyl). ¹³C NMR (CDCl₃-d), δ (ppm): 182.50 (C-1); 170.91 (C-7); 130.45 (C-9); 136.81 (C-11); 136.17 (C-13); 163.94 (C-12); 120.77 (C-8); 128.87 (C-10); 55.39 (C-6); 26.72 (C-2); 43.14 (C-5); 30.43 (C-4,4ʹ); 29.15 (C-3,3ʹ); 38.01 (C-14)carbon are assigned with numbers as presented in Numbering Scheme 3.

2.3.1 Dibutyltin(IV)bis[4-((5-chloro-2-hydroxyphenyl)ethylideneamino)methyl)cyclohexane carboxylate]; (1)

Yield: 68%, m.p. 205–207 °C: anal. calc. for C₄₀H₅₆C_l2N₂O₆Sn: C, 56.49 (55.84); H, 6.64 (5.33); N, 3.29 (3.54); IR (4000–400 cm⁻¹): 1632 υ(COO)_sym;1477 υ(COO)_asym; 155 $\mathrm{\Delta }$ υ; 1586 υ(C = N); 3373 υ(OH)_phenolic; 453 υ(Sn-O); 567 υ(Sn-C): ¹H NMR (CDCl₃-d) δ (ppm): 7.20 (d, J = 8.8 Hz, 1H, phenyl), 7.45 (s, 1H, phenyl), 6.84 (d, J = 8.9 Hz, 1H, phenyl), 3.42 (d, J = 5.6 Hz, 2H, –NCH₂), 2.3 (s, 3H, CH₃), 2.46–2.50 (m, 1H, cyclohexyl), 2.07–2.10 (m, J = 13.2, 7.7, 6.1, 4.9 Hz, 4H, cyclohexyl), 1.44–1.46 (m, J = 13.2, 4H, cyclohexyl), 1.43 (t, 4H, α-H), 1.12–1.12 (m, 4H, β,γ-H), 0.93 (t, 6.72, 6H, δ-H). ¹³C NMR (CDCl₃-d) δ (ppm): 192.26 (C-1), 168.20 (C-7), 156.80 (C-9), 148.70 (C-11), 118.69 (C-13), 127.23 (C-12), 123.08 (C-8), 110 (C-10), 57.98 (C-2), 41.45 (C-5), 34.82 (C-4,4ʹ),28.73 (C-3,3ʹ), 30.67 (C-γ), 26.13 (C- β),25.69 (C-α), 14.88 (C-14), 14.16 (C-δ).

2.3.2 Dimethyltin(IV)bis[4-((5-chloro-2-hydroxyphenyl)ethylideneamino)methyl)cyclohexane carboxylate]; (2)

Yield: 73%, m.p. 182–184 °C: 766.34: anal. calc. for C₃₄H₄₄C_l2N₂O₆Sn: C, 53.29 (52.88): H, 5.79 (5.63): N, 3.66 (3.56): IR (4000–400 cm⁻¹): 1652 υ(COO)_sym;1466 υ(COO)_asym; 186 $\mathrm{\Delta }$ υ; 1594 υ(C = N); 3387 υ(OH)_phenolic; 466 υ(Sn-O); 570 υ(Sn-C): ¹H NMR (CDCl₃-d) δ (ppm): 7.20 (d, 8.1 Hz, 1H, phenyl); 6.85 (d, J = 8.8 Hz, 1H, phenyl);3.41 (d, J = 5.5 Hz, 2H, –NCH₂); 2.33 (s, 3H, CH₃); 1.92–1.95 (m, 1H, cyclohexyl); 1.38–1.41 (m, 4H, cyclohexyl); 1.26–1.30 (m, 4H, cyclohexyl)1.09–1.11 (m, 1H, cyclohexyl); 0.93 (s, 6H, αH): ¹³C NMR (CDCl₃-d) δ (ppm): 172.26 (C-1); 161.6 (C-7); 159.5 (C-9); 132.69 (C-11); 129.08 (C-12); 122.50 (C-8); 116.16 (C-10); 57.95 (C-6); 42.45 (C-2); 37.25 (C-5); 36.82 (C-4,4ʹ); 28.67 (C-3,3ʹ);18.48 (C-α); 17.86 (C-14) (Numbering Scheme 3).

2.3.3 Trimethyltin(IV)bis[4-((5-chloro-2-hydroxyphenyl)ethylideneamino)methylcyclohexane carboxylate]; (3)

Yield: 71%, m.p. 215–217 °C: 472.59: anal. calc. for C₁₉H₂₈ClNO₃Sn: C, 48.29 (47.93); H, 5.97 (5.77); N, 2.96 (2.45): IR (4000–400 cm⁻¹): 1652 υ(COO)_sym;1474 υ(COO)_asym; 178 $\mathrm{\Delta }$ υ; 1584 υ(C = N); 3422 υ(OH)_phenolic; 453 υ(Sn-O); 561 υ(Sn-C): ¹H NMR (CDCl₃-d) δ (ppm): 7.45 (s, 1H, phenyl), 7.22 (d, J = 8.9 Hz, 1H, phenyl), 6.85 (d, J = 8.8 Hz, 1H, phenyl), 3.42 (d, J = 6.5 Hz, 2H, –NCH₂), 2.32 (s, 3H, CH₃), 2.06–2.09 (m, 1H, cyclohexyl), 1.43–1.46 (m, 4H, cyclohexyl), 1.27–1.30 (m, 4H, cyclohexyl), 1.12–1.15 (m, 1H, cyclohexyl), 0.53 (s, 9H, α-H).: ¹³C NMR (CDCl₃-d) δ (ppm): 183.81 (C-1), 173.30 (C-7), 166.44 (C-12), 135.02 (C-11), 129.79 (C-13), 123.37 (C-8), 121.78 (C-10), 57.16 (C-6), 46.16 (C-2), 40.50 (C-5), 32.96 (C-4,4ʹ), 31.61 (C-3,3ʹ), 16.76 (C-α), 2.44 (C-14).

2.3.4 Tributyltin(IV)bis[4-((5-chloro-2-hydroxyphenyl)ethylideneamino)methyl)cyclohexane carboxylate]; (4)

Yield: 76%, m.p. 232–234 °C: 598.84: anal. calc. for C₂₈H₄₆ClNO₃Sn: C, 56.16 (56.01); H, 7.74 (7.62); N, 2.34 (2.21): IR (4000–400 cm⁻¹): 1647 υ(COO)_sym;1467 υ(COO)_asym; 180 $\mathrm{\Delta }$ υ; 1543 υ(C = N); 3337 υ(OH)_phenolic; 432 υ(Sn-O); 554 υ(Sn-C): ¹H NMR (CDCl₃-d), δ (ppm): 7.63 (s,1H, phenyl), 7.26 (d, J = 6.3 Hz,1H, phenyl), 7.74 (d, J = 6.3 Hz,1H, phenyl), 3.43 (d, J = 6.5 Hz, 2H, –NCH₂), 2.37 (s, 3H, CH₃), 1.87–1.90 (m, 1H, cyclohexyl), 1.84 – 1.76 (m, 4H, cyclohexyl), 1.31–1.34 (m, 4H, cyclohexyl), 1.26 – 1.29 (m, 1H, αH), 1.30–1.33 (m, J = 7.2 Hz, 4H, γH), 1.06–1.09 (m, 4H, βH), 0.86 (t, J = 7.3 Hz, 6H, δH): ¹³C NMR (CDCl₃-d), δ (ppm): 183.44 (C-1), 172.77 (C-7), 165.15 (C-9), 132.57 (C-11), 128.38 (C-12), 121.29 (C-13), 119.47 (C-8), 119.33 (C-10), 54.63 (C-2), 44.97 (C-5), 39.59 (C-4,4ʹ), 28.23 (C-γ), 26.81 (C-α), 29.71 (C- β), 14.80 (C-δ), 14.13 (C-14).

2.3.5 Triphenyltin(IV)bis[4-((5-chloro-2-hydroxyphenyl)ethylideneamino)methyl)cyclohexane carboxylate]; (5)

Yield: 78%, m.p. 245–248 °C: 658.8: anal. calc. for C₃₄H₃₄ClNO₃Sn: C, 61.99 (61.54); H, 5.20 (5.10); N, 2.13 (2.01): IR (4000–400 cm⁻¹): 1625 υ(COO)_sym;1440 υ(COO)_asym; 185 $\mathrm{\Delta }$ υ; 1532 υ(C = N); 3336 υ(OH)_phenolic; 434 υ(Sn-O); 545 υ(Sn-C): ¹H NMR (CDCl₃-d) δ (ppm): 7.69 (m, 1H, phenyl), 7.47 (d, 1H, δH) 7.45 (s, 1H, phenyl), 7.43 (d, 1H, βH, βʹH), 7.25 – 7.14 (m, 1H, γH, γH), 6.83 (d, J = 8.9 Hz, 1H, phenyl), 3.40 (d, J = 5.7 Hz, 2H, –NCH₂), 2.62 (s, 3H, CH₃), 1.91–1.94 (m, 1H, cyclohexyl), 1.51–1.54 (m, 4H, cyclohexyl), 1.26–1.29 (m,4H, cyclohexyl), 1.11–1.14 (m, 1H, cyclohexyl).: ¹³C NMR (CDCl₃-d) δ (ppm): 182.50 (C-1), 170.89 (C-7), 163.90 (C-9), 160.91 (C-10), 137.53 (C-12), 137.26 (C-9), 130.36 (C-α), 128.66 (C-β), 127.0 (C-γ), 119.74 (C-δ), 120.12 (C-11), 129.16 (C-8),55.41 (C-6), 43.14(C-2), 38.01(C-5), 30.43 (C-4,4ʹ), 26.72 (C-3,3ʹ), 14.35 (C-14).

2.3.6 Dioctyltin(IV)bis[4-((5-chloro-2-hydroxyphenyl)ethylideneamino)methyl)cyclohexane carboxylate]; (6)

Yield: 73%, m.p. 192–194 °C: anal. calc. for C₄₈H₇₂C_l2N₂O₆Sn: C, 59.88 (59.56); H, 7.54 (7.34); N, 2.91(2.76): IR (4000–400 cm⁻¹): 1655 υ(COO)_sym;1482 υ(COO)_asym; 173 $\mathrm{\Delta }$ υ; 1563 υ(C = N); 3364 υ(OH)_phenolic; 473 υ(Sn-O); 581 υ(Sn-C): ¹H NMR (CDCl₃-d) δ (ppm): 8.16 (s, 1H, OH), 6.98 (d, J = 6.6 Hz, 1H, phenyl), 7.02 (s, 1H, phenyl), 6.95 (d, 1H, phenyl), 3.47 (d, J = 6.6 Hz, 2H, –NCH₂), 2.32–2.36 (m, 1H, cyclohexyl), 2.28 (s, 3H, –CH₃), 1.59–1.61 (m, 4H, cyclohexyl), 2.06 (t, J = 14.1 Hz, 4H, α´H), 1.94–1.97 (m, 4H, cyclohexyl), 1.30–1.33 (m, 4H, β-β´H), 1.251.30 (bs, γ--γ´H), 1.21 – 1.23 (m, 6H, δH), 0.90 (t, J = 6.2 Hz, 2H, αH).: ¹³C NMR (CDCl₃-d) δ (ppm): δ 173.30 (C-1), 166.44 (C-7), 135.03 (C-11), 129.80 (C-12), 123.20 (C-8), 121.79 (C-10), 57.89 (C-2), 46.16 (C-5), 40.51 (C-4,4ʹ), 25.12 (C-3,3ʹ), 32.97 (C-γ), 31.87 (C-β), 31.62 (C-α) 30.50 (C-γ), 27.97 (C-βʹ), 26.97 (C-β), 18.97 (C-δʹ), 16.76 (C-δ).

2.4.1 Antileishmanial assay

Antileishmanial activity of the synthesized compounds was evaluated in vitro against the promastigote form of leishmaniatropica using XTT (2, 3-bis-(2-methoxy-4-nitro-5-dulfophenyl)–2H-tetrazolium-5-carboxanilide) based assay as a marker of cell viability (Meyre-Silva et al., 2013). Stock solution (5 mg/mL) of XTT was prepared in phosphate buffer solution and stored in dark at 4 °C. RPMI-1640, fetal bovine serum (FBS), potassium chloride (KCl), sodium bicarbonate (NaHCO₃), phosphate buffer saline (PBS), antibiotic (Penicillin/Streptomycin) and distilled water were used in preparation of culture medium. For anti-leishmaniasis, each well of the 96-well plate was charged 100 μL culture media and 2.5 × 10⁴ cells/mL were seeded into each well. The seedling density was counted during log phase using a Neubauer chamber. The promastigotes were allowed to enter to their log phase until the count reach from 5 to 6 million cells per mL. Three different concentrations (1.25 µg/mL, 2.5 µg/mL, and 5 µg/mL) of each compound were added to respective wells in 96-well plate. Amphotericin B was used as a reference drug and DMSO as negative control. The 96- well plate was incubated for 72 h at 25 ± 1 °C. After incubation 10 μL of XTT was added to each well and was again incubated for 3 h at 25 °C. 40 µL of DMSO was then added into each well of the plate as stopping solution and relative optical density (OD) the plate was recorded at 450 nm using an ELISA plate reader. The number of viable cells is correlated by the absorbance of formazan produced by the mitochondrial dehydrogenase of active cells (C. Williams et al., 2003). All the experiments were performed in triplicate and the Percent inhibitions of the compounds were calculated using the following formula: $$\mathrm{P}\mathrm{e}\mathrm{r}\mathrm{c}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{I}\mathrm{n}\mathrm{h}\mathrm{i}\mathrm{b}\mathrm{i}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}=\frac{\mathrm{m}\mathrm{e}\mathrm{a}\mathrm{n}\mathrm{O}\mathrm{D}\mathrm{o}\mathrm{f}\mathrm{S}\mathrm{a}\mathrm{m}\mathrm{p}\mathrm{l}\mathrm{e}-\mathrm{m}\mathrm{e}\mathrm{a}\mathrm{n}\mathrm{O}\mathrm{D}\mathrm{o}\mathrm{f}\mathrm{B}\mathrm{l}\mathrm{a}\mathrm{n}\mathrm{k}}{\mathrm{m}\mathrm{e}\mathrm{a}\mathrm{n}\mathrm{O}\mathrm{D}\mathrm{o}\mathrm{f}\mathrm{p}\mathrm{o}\mathrm{s}\mathrm{i}\mathrm{t}\mathrm{i}\mathrm{v}\mathrm{e}\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l}-\mathrm{m}\mathrm{e}\mathrm{a}\mathrm{n}\mathrm{O}\mathrm{D}\mathrm{o}\mathrm{f}\mathrm{B}\mathrm{l}\mathrm{a}\mathrm{n}\mathrm{k}}\times 100$$

2.4.2 DNA interaction study

DNA interaction study was carried out by UV–visible spectroscopy. SS-DNA (20 mg) was dissolved in water by overnight stirring. 20 mM PBS was used to check the percent purity of DNA by a ratio of UV absorbance at 260 and 280 nm. The final concentration of the SS-DNA solution was determined by using molar extinction coefficient (6,600 M⁻¹cm⁻¹) and was found to be 180 mM. A series of DNA solution (5 μM, 10 μM, 15 μM, 20 μM and 25 μM) were treated with test compounds (3 mM, 3 mL). The solutions were incubated for 5 min and then subjected to UV–visible absorption. Absorption spectra were recorded using quartz cuvettes in the range 600–200 nm (Sirajuddin, Ali, & Badshah, 2013).

2.4.3 Antioxidant activity

DPPH standard protocol was applied for antioxidant study (Yabalak, Emire, Adıgüzel, Könen Adıgüzel, & Gizir, 2020). DPPH solution (160 μM) was prepared in ethanol. To a series of test compounds (25 μM, 50 μM, 75 μM, 100 μM and 125 μM) DPPH solution (3 mL, 160 μM) was added. UV–visible absorption of the solutions at 517 nm was recorded using quartz cuvettes after being incubated for 2 min(Adeyemi et al., 2019).

2.4.4 Cytotoxicity

Brine shrimp’s lethality bioassay was used to evaluate the cytotoxicity of synthesized compounds. Briefly, a vessel was filled with sterile simulated sea water. The sea water was prepared by adding sea salt (38 g/L) and pH was adjusted to 8.5, using NaOH solution. Brine shrimp’s eggs were added to this sea water at room temperature. The eggs were hatched into brine shrimps after two days continuous aeration. Thirty active nauplii were collected in a vial containing yeast suspension and 4.5 mL brine solution. a series of three vials containing 4.5 mL brine solution and yeast suspension were provided with 0.5 mL test compounds solution with different concentrations (5, 50 and 100 μg/mL). The experiment was conducted in triplicate. Etoppside was used as standard reference drug. LD ₅₀ was determined by using Finney’s probit analysis.

2.4.5 Antifungal activity

Antifungal activity of the compounds was done by using standard agar tube dilution method. Five fungal strains F. Solani, C. Albicans, T. Longifusus, A. Flavis and M. Canis were tested. Miconazole and amphotericin were used as standard drugs. Growth media sabouraud dextrose was prepared by mixing glucose agar (45%), sabouraud (32.5 g), and agar (20 g) in 500 mL distilled water at 90 °C. 4 mL media was added into respective capped tubes and autoclaved for 15 min at 121 °C. 100 μg/mL of each test compounds was added to respective tube at 50 °C and the media was then solidified at room temperature. The tubes were then provided with 4 mm inoculums derived from a seven day old fungal culture. The tubes were then incubated at 25 °C for 8 days. Each fungal strain growth was measured with linear growth in mm and growth inhibition was calculated with reference to respective standard (Nural, Gemili, Yabalak, De Coen, & Ulger, 2018).

2.4.6 Antibacterial activity

Antibacterial activity of the synthesized compounds was assisted by agar well diffusion essay. Five bacterial strains including E. Coli, B. Subtillus, S. Aureus P. Aeruginosa and S. Typhi were checked. Briefly, petri plates were provided with nutrient agar growth media at 50C and solidified at room temperature. Growth media nutrient agar was prepared by standard procedure. Wells were bored in the media with the help of 6 mm sterile cork borer. Bacterial strains of about five hours old bacterial inoculums were spread on the surface of media using sterile cotton swab. Test compounds (100 μg/mL), standard reference (Imipenem) and negative control (DMSO) was provided into their respective wells. The prepared plates were incubated at 37C for 20 h. The antibacterial activity was determined by measuring zone of inhibition in millimeter. Growth inhibition was calculated with reference to standard drug (Nural et al., 2018).

3.2.1 ¹HNMR

NMR spectra of the prepared ligand and its organotin(IV) carboxylates were recorded in deuterated chloroform using TMS as internal standard and the data is given in experimental section. The ¹H and ¹³C NMR spectra of ligand and complexes (1–6) are shown in supplementary data (Fig. S8-S20, Supplementary materials). In the ¹H NMR spectrum of ligand, resonance signals for protons of cyclohexyl OH, phenyl, methyl and methylene groups with characteristic multiplicity and integration confirmed the formation of ligand (Aman, Matela, Sharma, & Chaudhary, 2015). However, in spectra of complexes (1–6), in addition to ligand characteristic signal, the appearance of respective alkyl/aryl group resonance signals attached to tin atom indicted the formation of complexes. The protons attached to cyclohexyl appeared as multiples in the range of at 1.00–2.32 ppm and methylene group resonate as doublet around 3.44 ppm (Fig S8-S20, Supplementary materials) (Hussain et al., 2014). Whereas the ¹H resonances of methyl group attached to N of the azomethine appears as singlet around 2.35 ppm. The protons of the aromatic part of the ligand appeared as two set of dupletsin the range of 6.74–7.37 ppm. The protons of alkyl/aryl groups attached to tin showed signals as expected. Coupling constant²J(¹¹⁹Sn, ¹H) and Lockhart equation was used to determine the coordination around Sn atom. The calculated values for ²J(¹¹⁹Sn, ¹H) of compounds 2 and 3 were 83 and 79 Hz, agreeing with C-Sn-C angle of 132.87 and 130.15°, respectively, demonstrating five coordination around tin (Table 1) (Rabiee, Safarkhani, & Amini, 2019).

3.2.2 ¹³C NMR

The ¹³C NMR spectra displayed the carbon resonance of the ligand and coordinated Sn-R.The exact number of carbon resonance at their characteristics chemical shift values validated the formation of ligand and complexes (1–6). The carboxylate carbons resonance of free ligand at 182.50 ppm is shifted downfield to 192.26(1), 183.81 (3), 183.44 (4)and 185.2(6)ppm in complexes verifying COO-Sn coordination. These chemical shifts are consistent with that reported for butanoic acid (Tariq et al., 2014){Tariq, 2014 #1;Tariq, 2014 #1}. Similarly, the shifting of carboxylic carbon resonance to downfield compared to free ligand strongly recommending coordination via carboxylate group. No extra resonance signals were found in ¹³C NMR and thus suggesting the purity of compounds. The cyclohexyl carbons resonances are observed in the range 26–43 ppm. The other resonance lines in the spectra fall in the three regions, i.e., 14–26 ppm for aliphatic, 110–138 ppm for carbons of phenyl group and 172–192 ppm for carbonyl carbon of carboxyl group (Akintayo, Munzeiwa, Jonnalagadda, & Omondi, 2021).

3.2.3 XRD analysis

Powder XRD diffraction pattern of the compounds (1–6) were recorded by Oxford X-calibur Gemini diffractometer. The spectra exposed sharp peaks with prominent diffraction intensities and low background. Expert high score plus software was used to determine the indexing of powder diffraction pattern and searching space group of the crystal specimens (Fig. 1) (Akremi & Noubigh, 2019). The structural features and properties obtained are demonstrated in Fig.1 (Supplementary materials). The indexed 10 prominent diffraction lines of ligand demonstrated lattice parameter of orthorhombic crystal system with primitive bravais and P 21 am space group. Similarly the complexes 1 and 2 indexed lattice parameters fitted to orthorhombic crystal system with C face centered and primitive bravais (Masciocchi, Sironi, Chardon-Noblat, & Deronzier, 2002). Scherrer equation was used to calculate the crystallite size D of complexes 1, 2 and 3and found to be 142 nm, 76 nm and 70 nm respectively (Xue, Kamali, Yu, Appels, & Dewil, 2023). $$D=\frac{k\mathrm{\lambda }}{\mathrm{\beta }\mathrm{c}\mathrm{o}\mathrm{s}\theta }$$

Close

Fig. 1

Powder XRD pattern of complexes 1–3 and peaks indexation using X-pert high score plus software 10^3.

Export to PPT

Where D is the crystal size in the form of powder, K shape factor with value of unity, ʎ is wavelength of X-ray, β is FWHM and θ is Bragg angle.

PXRD data shows that all the prominent diffraction pattern of prepared compounds can be readily indexed by a set of lattice parameter. The relative deviations between experimental and calculated are about 1.12%, 0.45% and 0.36% for compounds 1, 2 and 3 correspondently, which could suggest that all the prepared compounds are single phase(Su et al., 2022).

3.3.1 Antileishmanial activity

In vitro antileishmanial activity of the prepared was done using XTT standard protocol against promestigotes form leishmaniatropica (Ozlem‐Caliskan, Ertabaklar, Bilgin, & Ertug, 2021). The examined compounds produced a significant reduction in viable promestigotes. The complexes showed high antileishmanial activity compared to the free ligand suggesting the charge distribution and compatibility of the compounds structure to cell membrane enzyme trypanothione synthetase. All the test concentrations showed significant antileishmanial activity (Fig. 2). The IC₅₀ of the compounds and standard is shown in Table 2. The compounds 1, 2 and 5 were most potent against promestigotes form of leishmaniatropica and exhibited IC₅₀ value 1.22 ± 0.014, 1.17 ± 0.007 and 1.22 ± 0.021 µg/mL as compared to the positive standard amphotericin B which exhibited 0.71 ± 0.05 μg/mL value. Compounds 3 and 6 also showed remarkable potency with IC₅₀ 1.68 ± 0.014 and 1.94 ± 0.03 respectively against leishmaniatropica as demonstrated in Table 2. The ligand and compound 4 were found inactive. The antileishmanial activity may be due to the interference with the functioning of leihmania mithocondria (Ozlem‐Caliskan et al., 2021). This investigation demonstrated the potential use of the prepared organotin(IV) carboxylate as a source of novel agents for the treatment of leishmaniasis.

Close

Fig. 2

Antileishmanial activity; Percentage inhibition and IC₅₀ of selected compounds.

Export to PPT

3.3.2 DNA interaction

The DNA-compounds interaction was examined by UV–Visible spectroscopy. The synthesized compounds (1–6) were interacted with different concentrations of DNA and UV–Visible spectra were taken (Fig. 3).The compounds showed two bands around 250 nm and 350 nm which are probably due to aromatic π - π̽ transition and 300–380 nm due to n - π̽ transition of CO or NC groups. As the DNA was interacted with increasing concentration, hypochromic effect, and red shift of about 5 nm in the bands were observed. The band shift of molecules after interacted with DNA could be evidence of binding mode of DNA with small molecules (Casini et al., 2001). In case of intercalative mode of binding, hypochromism effect coupled with red shift for the characteristic’s peaks will be observed due to strong stacking between DNA base pairs and chromophores of the compounds. Thus, the interaction between small molecules and DNA could be non-covalent intercalative binding. During intercalation of the base pairs of DNA, the π̽ orbital of the ligand could couple with π orbital of the DNA base pairs, thus resulting in bathochromic effect due to lowering π -π̽ transition energy. Similarly, the coupling of partially filled electrons with π orbital lowers the transition likelihood and hence result hyphochromism. The hypochromic effect may be observed due to π - π̽ (stacking interaction) in intercalative binding mode and bathochromic effect is observed when DNA duplex is stabilized (Aydinoglu et al., 2016). In case of compounds (2,3 and5) the DNA-compounds complex showed stability as revealed from red shift after 12 and 24 h. The intrinsic binding constant of compounds with DNA, based on variation in band shift, was calculated by using Benesi-Hildebrands equation. $$\frac{A_0}{\mathrm{A}-A_0}=\frac{{\epsilon }_G}{{\epsilon }_{H-G}-{\epsilon }_G}+\frac{{\epsilon }_G}{{\epsilon }_{H-G}-{\epsilon }_G}X\frac{1}{K[\mathrm{D}\mathrm{N}\mathrm{A}]}$$

Close

Fig. 3

UV–Vis spectra of DNA binding study and DNA binding constant.

Export to PPT

Where K is binding/association constant, A₀ and A are the absorbance of compound and DNA-compound complex, respectively. Ɛ_Gis the absorption coefficient of compound and Ɛ_H-_Gis the absorption coefficient of DNA-compound complex. The binding constants K were calculated from intercept to slop ratio of A₀/(A-A₀) vs 1/[DNA]. The intrinsic binding constants of compounds (2, 3, 4 and 5) are determined to 2.7 × 10³, 1.7 × 10³, 1.1 × 10²and 6.2 × 10³ respectively as demonstrated in Table 3 and Fig. 3. The compounds (2, 3, 4 and 5) showed significant binding constant whereas the ligand and compound (1 and6) were inactive. Gibbs free energy was calculated from equation: $\mathrm{\Delta }$ G = -RT lnK.

R is gas constant (8.314 JK⁻¹.mol⁻¹), T is temperature (298 K). The values of $\mathrm{\Delta }$ G and K are given in the inset graph (Fig. 3). The order of DNA-compound interaction is as 5˃2˃3˃4.The compounds might interact with nitrogenous base of the DNA and restrain cell growth by interfering the transcription and replication of DNA. The DNA interaction study revealed that these compounds may be used as anticancer drugs.

3.3.3 Antioxidant activity

Health pathologies such as diabetes, heart diseases, are all contributed by oxidative damage. The antioxidants agents can slow down or prevent oxidative damage to our body by converting the free radicals to stabile components. DPPH standard protocol was used to investigate the antioxidant activity of free ligand and compounds (1–6) (Sari, Qudus, & Hadi, 2020). The compounds showed significant percentage antioxidant activity range from 96% to 59% compared to free ligand (46%) (Fig. 4). The low IC₅₀ values (76.92, 56.31 and 42. 10 ΜM) of Compounds (1, 2 and 5) suggested significant antioxidant activity (Table 4 and Fig. 4). In the complexes, the phenolic group gives hydrogen radical and results the initiation of radical chain and they act as antioxidant agents. Similarly, the electronegative halogen substituent and conjugated system also served good antioxidant. Overall, the complexes showed comparable antioxidant activity reported in the previous work (Yabalak et al., 2020).

Close

Fig. 4

Antioxidant activity of HL, (3), (4), (5) and % RSA.

Export to PPT

3.3.4 Cytotoxicity

The ability of organotin(IV) complexes to interact with cell membrane, protein and DNA make them cytotoxic. The toxicity of the synthesized compound against Brine shrimps was tested. All the tested complexes revealed significant toxicity except ligand which showed moderate toxic effect (Table.5 and Fig. 5). Highest toxicity was observed for complex 3 (27 killed Brine shrimps 30/dilution) and 5 (28 killed Brine shrimps 30/dilution) which is almost similar to the standard drug (27 killed Brine shrimps 30/dilution). The lowest toxicity was found for ligand and complex 4 with LD₅₀ values of 14.22 μg/mL and 14.56 μg/mL. The LD₅₀ values of complexes 2 (6.26 μg/mL), 3 (6.79 μg/mL) and 5 (5.67 μg/mL) indicated comparable toxicity to standard drug (5.80 μg/mL). The high cytotoxicity of complexes (3 and 5) may be due to the presence of phenyl group. In addition to bioactive pharmacophore and bulky cyclohexyl, the phenyl group facilitates π-π interaction between biomolecules and complexes. The complexes showed potent cytotoxicity lower LD₅₀ value comparable to the previously reported work (Yabalak et al., 2017; Nural et al., 2018).

Close

Fig. 5

In vitro cytotoxicity of Organotin(IV) complexes with LD₅₀ against Brine Shrimps 30/dilution.

Export to PPT

3.3.5 Antifungal

Based on the provided data, different strains of fungi were tested for their susceptibility to various drugs, and the results are presented in terms of percentage inhibition and minimum inhibitory concentration (MIC) values (Table. 6 and Fig. 6). All the organotin(IV) derivatives of ligand revealed significant activity against tested fungal strains. Highest activity was found for complex 5 with % inhibition in the range 25–74%, where the complex was most effective against F. solani and M. canis with 72% and 74% inhibition respectively. Similarly, the complex (4) and (6) also showed significant activity, however the complexes revealed highest effectiveness against A. flayis and F. solani with growth inhibition of 55% and 68% respectively. Furthermore, the complexes (1, 2 and 3) were found most active against F. solani, A. falyis and F. solani respectively. All the complexes showed highest antifungal activity against F. solani. The ligand was found active against A. falyis and F. solani and inactive for other tested fungal strains. The lowest MIC (32 μg/mL and 43 μg/mL) was found for complex (5) and (6) respectively (Table. 6). The high antifungal activity of complexes may be due to the coordination and polarity of tin with oxygen of the ligand. The complexes (1, 2 and 3) showed high activity which may be due to the increase lipophilicity and permeability across the cell membrane (Hussain et al., 2015).

Close

Fig. 6

Comparative antifungal %inhibition of ligand and complexes with standard.

Export to PPT

3.3.6 Antibacterial

The synthesized ligand and its organotin(IV) complexes revealed antibacterial activity against the targeted bacterial strains (Table. 7 and Fig. 7). The ligand was found active against P. aeruginosa (18/25 mm) and S. typhi (16/22 mm), inactive against E. coli (5/30 mm) and moderately active against B. subtillus (15/32 mm) and S. aureus (10/25 mm). All the complexes showed significant activity against the tested bacterial strains compared to lignad. The highest activity was observed for complexes (5 & 6) against all tested bacterial strains among test complexes. The complexes (1, 2 and 3) showed moderate activity against B. subtillus, E. coli, S. aureus and S. typhi. The data revealed that all the complexes showed highest activity against S. typhi and P. aerruginosa. The antibacterial potential of these complexes revealed that the substituted bioactive pharmacophore containing complexes are of biological properties. The activity of these complexes can be attributed to the bulky cyclohexyl and active pharmacophore substituent that is good electron donating groups, enhancing the lipophilic property of complexes by strengthening ligand metal bond. The antibacterial result is corroborated to previously report tin complexes.

Close

Fig. 7

Antibacterial activity of ligand and its organotin(IV) complexes.

Export to PPT