Spectral, thermal, antimicrobial studies for silver(I) complexes of pyrazolone derivatives

Background Synthesize new complexes of Ag(I) to enhance efficacy or stability and also, pharmacological activities on the operation of pyrazolone's biological properties. Results Efficient and high yielding pathways starting from the versatile and readily available 3-methyl-1-phenyl-5-pyrazolone by Knoevenagel condensation of a sequence of 4-arylidene-3-methyl-1-phenyl-5-pyrazolone derivatives (2a-c) have been formed by the reaction of various substituted aromatic aldehydes Used as ligands to synthesize Ag(I) chelates. Synthesized compounds and their complexes have been characterized by elemental analysis, magnetic and spectroscopic methods (IR, 13C, 1HNMR, mass) and thermal analysis. The spectrophotometric determinations suggest distorted octaedral geometry for all complexes. Both ligands and their metal complexes have also been tested for their antibacterial and antifungal efficacy. Conclusions Newly synthesized compounds have shown potent antimicrobial activity. The results showed that the complex 's high activity was higher than its free ligands, and that Ag(I)-L3 had the highest activity.


Introduction
Pyrazolone chemistry began in 1883 when Ludwig Knorr first reacted to phenyl hydrazine with aceto-acetate ester. As pyrazolones were discovered as binding components for azo dyes in the late 1800s, they rapidly increased in importance. Today, pyrazolon is still an significant trade precursor to dyes and pharmaceuticals. Pyrazolone is a biologically important scaffold associated with different pharmacological activities such as antimicrobials [1][2][3][4][5], anti-inflammatory [6], analgesic [7], antidepressant [8], anticonvulsant [9], antidiabetic [10], antihyperlipidemic [11,12], antiviral [13,14], anti-tuberculosis [15,16], antioxidant [17,18] and anticancer [19,20]. For several years, the preparation of pyrazolone and its derivatives has attracted significant attention from organic and medicinal chemists, as they belong to a class of compounds with promising results in medicinal chemistry. The heterocycles condensed to the pyrazole ring are an important source of bioactive molecules [21,22]. Compounds containing both pyrazole and other essential heterocyclic active structural units usually demonstrate more remarkable biological activity. A number of condensed pyrazole derivatives have been reported as four-fold antibacterial agents against Gram-positive and Gram-negative bacteria compared to general pyrazole compounds [23,24]. A digit of antimicrobial active silver(I) complexes have the capacity to disrupt microbial transpiration as well as block tyrosinase synthesis and are extremely cytotoxic to cancer cells [24]. Massive attention in silver ions (Ag(I)) as a broad spectrum antimicrobial has upped the size and importance of in vitro biocompatibility research [25]. Silver ions are toxic to many bacteria, viruses, algae and fungi. Silver-based medicines have been widely used for this task for decades [26]. The objective of this study is to display the synthesis and characterization of three Ag(I) pyrazolone complexes in an attempt to verify the mode of coordination and the biological properties of the final complexes.

Infrared spectra
KBr disks registered mid-infrared spectra of L 1 , L 2 , L 3 and their metal complexes. As expected, with changes in band intensities and wave numbers, the absorption bands characteristic of L 1 , L 2 , L 3 acting as a monodentate unit are observed in the complexes. The proposed structures of the complexes must be considered prior to determining the assignments of the infrared spectra. Here, Ag(I) ion interacts with these monodentate ligands forming monomeric structure complexes in which the Ag(I) ion is four coordinated (Scheme 2) [27][28][29][30].
The complexes of three ligands with Ag(I) contain only one plane of symmetry and therefore the complexes that belong to C S symmetry and show 159 vibrational fundamentals, and all vibrations are distributed between movements of the types A \ and A 1 \\ , all of which are monodegenrate, infra-red and Raman active. The free ligand infrared spectrum shows bands at 1496, 1508 and 1550 cm −1 due to the stretching vibration of hydrazono (C = N) groups [31]. Comparing the Ag(I) IR spectrum with the free ligand spectrum, the transfer of (C = N) groups to lower frequency values (1512, 1515, 1523 and 1527 cm −1 ) and the change in strength of (C = N) from  [31];The stretching vibrations ν(C-H) of phenyl groups and −CH 3 units in these complexes are assigned as a number of bands in the region 3066-3100 cm −1 [11,12]. The ν(C = O) vibration appears in the region of 1666-1685 cm −1 . The spectra of the isolated solid complexes revealed a number of new bands of different intensities for ν(M-N). The ν(Ag-N) bands observed at 813, 837 cm −1 for Ag(I)-L 1 , at 748, 794 cm −1 for Ag(I)-L 2 and at 759, 779 cm −1 for Ag(I)-L 3 (Table 1) which are absent in the spectrum of free three ligands [30][31][32]. The coordinating water in the three complexes are characterized by the appearance of ν (Ag-O) at 577, 515, 544 cm −1 . Also the stretching vibrations at 813, 792, 779 cm −1 assigned to ν(Ag OH 2 ), sponsored coordinating water participation [32]. The suggested structural formulas are defined in Scheme 2 on the basis of the IR tests.

UV-Visible Spectra
The application of ultraviolet spectroscopy is more general and can be useful for all chelate structural determinations as they are all absorbed in this region [33]. Electronic absorption spectra confirmed the development of metal ligand complexes. Electronic absorption spectra L 1 for Ag(I), L 2 for Ag(I) and L 3 for Ag(I). Complexes within the spectrum of wavelengths between 200 and 800 nm are described in Additional file 1: Table S1 and Fig. 2. The free three-ligand UV spectrum (L 1 , L 2 and L 3 ) displays bands at 281, 297 and 297 nm that are assigned respectively to π-π * . And displays bands allocated to n-π * transitions at 330 nm. The modification of the reflectance band to higher (bathochromic shift) and lower values (hypochromic shift) and the appearance of new bands for complexes has resulted in the release of three ligands' complex actions towards metal ions. Complexes also present bands within the range 410-480 nm which can be due to the transition of ligand-metal charges for three ligands [34,36]. The molar absorptivity (ε) values of the prepared metal complexes under investigation were determined (Additional file 1: Table S1) using the relation: A = εcl, where, A = absorbance, c = 1.0 × 10 -3 M, l = length of cell (1 cm) [22]. The values of 10Dq (difference between t 2g and e g ) for the complexes were calculated by using the following Eq. 10Dq = E = hcν − where E = energy, h = blank constant = 6.626 × 10 −34 J.sec, c = 3 × 10 10 cm/sec, ν − = wave number cm −1 the data listed in Additional file 1: Table S1.

The 1 H NMR spectra
Suggested structure of the isolated Ag(I) complexes confirm about the efficiency of 1 H NMR spectra. Compared to the one of their complexes (Additional file 1: Table S2), the 1 H NMR spectra of new free three ligands in DMSOd 6 . The 1 H NMR spectra of L 1 and its metal complex shown in (Fig. 3a, b), the proton of (= CH-Ar) group observed in δ: 9.66 ppm and the protons of aromatic ring of (s, 9H, Aromatic-H) observed at δ: 7.14-7.97 ppm also the values of protons of -CH aliphatic observed in the range δ: 3.03-3.33 ppm (s, 6H, -N (CH 3 ) 2 ), the proton of (s, 3H, -CH 3 ) group observed in δ 2.28 ppm, no major differences were observed as opposed to the Ag(I) complex except that the signal is observed in 3.46 ppm due to H 2 O molecules [36]. This supports the hypothesis that L 1 interacts as a monodentate ligand bound to the Ag(I) ion through the hydrazono nitrogen group. [37]. The 1 H NMR spectra of L 2 and its Ag(I) complex shown in (Fig. 3c, d), the proton of (= CH-Ar) group observed in δ: 8.25 ppm and, the protons of aromatic ring of (s, 8H, Aromatic-H) observed at δ: 7.39-7.91 ppm [38]. The proton of (s, 3H, -CH 3 ) group observed in δ 2.30 ppm, simple differences are shown in comparison to the metal complex and the signal is observed in π: 3.47 ppm due to H 2 O molecules. This reinforces the hypothesis that L 2 reacts via the hydrazono nitrogen group as a monodentate ligand bound to the Ag(I) ion. The 1 H NMR spectra of L 3 and its Ag(I) complex shown in ( Fig. 3 (E, F)), the proton of (= CH-Ar) group observed in δ: 8.71 ppm and the protons of aromatic ring of (s, 9H, Aromatic-H) observed at δ: 7.18-7.46 ppm also the values of protons of -CH aliphatic observed in the range δ: 3.31 ppm (s, 3H, -O-CH 3 ), the proton of (s, 3H, -CH 3 ) group observed in δ 2.33 ppm, no major variations were noticed as opposed to the Ag(I) series. This supports the assumption that L 3 reacts as a monodentate ligand bound to the Ag(I) ion via the hydrazone nitrogen group.

Thermal studies
The thermal degradation of ligand (L 1 ) began at 190 °C and decay occurs at various temperatures at 310, 544 °C at one stage (Additional file 1: Fig. S1a). This step is accompanied by a net weight loss of 92. 36 Fig. S1d), the first phase occurs at 99 °C and is followed by a weight loss of 2.08 per cent relating to the removal of H 2 O, activation energy of 79.28 kJ mol −1 . The second step of decomposition occurs at temperature is 203, 528 and is accompanied by a weight loss of 75.90%; corresponding to the value of 10C 2 H 2 + 4HCN + 2H 2 O + NO 2 + SO + SO 2 theoretically, close to the calculated value 76.404%.

Table 1 Infrared frequencies (cm −1 ) a and tentative assignments b for (A) L 1 , (B) [Ag(L 1 ) 2 (H 2 O) 2 ]NO 3 , (C), L 2 (D) [Ag(L 2 ) 2 (H 2 O) 2 ]NO 3 .H 2 O, (E) L 3 and (F) [Ag(L 3 ) 2 (H 2 O) 2 ]NO 3
a s = strong, w = weak, sh = shoulder, v = very, br = broad, b ν = stretching and δ = bending The Residue value decomposition occurs at maximum 881 °C and the actual weight loss from this step is 23.35%, corresponding to Ag + 6C, close to the calculated value 23.596%. The thermal decay of L 3 happens in two phases of degradation (Additional file 1: Fig. S1e), the first step arises at 291 °C and is followed by a weight loss of 70.55 percent leading to a loss of 8C 2 H 2 similar to the measured value of 71.23 per cent with activation energy of 35.31 kJ mol −1 . The second step occurs at 518 ο C and is accompanied by a weight loss of 28.604%; corresponding to the value of 2CO + N 2 theoretically, close to the calculated value 28.67%. The [Ag(L 3 ) 2 (H 2 O) 2 ]NO 3 degradation takes place in two stages (Additional file 1: Fig. S1f ), the first occurs at 244 ο C and is accompained by a weight loss of 51.071% corresponding to loss of 14C 2 H 2 + 2H 2 O close to the calculated value 50.60% with an activation energy 15.31 kJ mol −1 . The second one begins at 543 ο C and is followed by a weight loss of 30.17%; corresponding to C 2 H 2 + CO + 2HCN + 3NO 2 theoretically, close to the calculated value 31.25%. The Residue remains at 677 °C and the actual weight loss is 17.76%, equal to Ag + 3C, close to the calculated value 18.15%.

Kinetic data
The kinetic parameters (activation energy, E*, entropy, ΔS*, enthalpy, ΔH*, and Gibbs free energy, ΔG*) have been evaluated by using the two mentioned methods in the literature [39,40] and shown in Additional file 1: Fig. S2 and listed in Table 3. The correlation coefficient for Arrhenius plots of thermal degradation stages were found to be in the range 0.943-0.985, revealing a good fit with linear function. The activation energies of decomposition were observed to be in the range 7.44-154.69 kJ mol −1 . The negative values of ΔS* indicate that the activation complex has a more ordered structure than the reactants or the reactions are slow. The positive ΔH* values postulate an endothermic nature of the formed complexes. The greater positive values of E* indicate that the processes involving in translational, rotational, vibrational states and a changes in mechanical potential

Mass spectra
The principle of a mass spectrometer focuses on the separation of fragments of ions based on the distribution of these ions with the mass to charge ratio (m/z). The L 1 , L 2 , L 3 fragmentation patterns and their complexes were obtained from the mass spectra, and were in good agreement with the structure suggested. The L 1 showed molecular ion peak (M +. ) with m/z = 305 (100%).  (Fig. 4), (Scheme 3). Fragmentation pattern of the complex [Ag(L 1 ) 2 (H 2 O) 2 ]NO 3 is given as an example in (Fig. 4), Additional file 1: Scheme S1. The molecular ion peak [a] appeared at m/z = 816 (20.5%) losses C 18 (Fig. 4), Scheme 4. Fragmentation pattern of the complex [Ag(L 2 ) 2 (H 2 O) 2 ] NO 3 .H 2 O is given as an example in (Fig. 4) (Fig. 4), Scheme 5. Fragmentation pattern of the complex [Ag(L 3 ) 2 (H 2 O) 2 ]NO 3 is given as an example in (Fig. 4) [42].

Biological activity studies Antimicrobial studies
The antimicrobial efficacy of L 1 , L 2 , L 3 and their free ligand complexes are explored in this experiment. Studies were conducted on E. Coli ATCC11229, Coliform ATCC8729, S. aureus ATCC6538, and Salmonella typhi ATCC14028 and fungal species as A. niger and P. expansum screening was tested against and examination and evaluation of the prepared complexes [42]. The same results were reported for E. Coli ATCC11229 of Ag(I)-L 2 and Ag (I)-L 1 followed by Ag(I)-L 3 considers that the lowest findings are equivalent to those of other complexes. The effect of free ligands on this strain has been shown to be below its complex and can be organized according to the sensitivity of the strains L 2 , L 3 and L 1 in the following ascending order. The effect of Ligands and their    complexes on Coliform ATCC8729 showed that Ag(I)-L 2 is highly important, giving 25.12 mm respectively. Although the remaining complexes showed lower results than the L 2 complexes. The results obtained in Table 4 and Fig. 5 showed that lower activity on the same strain and these results ensured that free ligand complexes were more active than free ligand complexes. In gram + ve bacteria, S. aureus ATCC6538, Highly important antibacterial activity of metal complexes with L 1 followed L 3 complex. The lesser activity from ligand L 2 and its complex. The antibacterial activity of metal complexes on Salmonella typhi ATCC14028 showed a good activity against (gram −ve), that recorded the best results Ag(I)-L 3 > Ag(I)-L 1 > Ag(I)-L 2 respectively. The action of the free ligands on gram -ve bacteria has yielded results lower than their complexes which give respectively 12.6, 11.43 and 7.8 mm, L 3 , L 1 , L 2 . The presence of different ligands and other complexes on both fungal strains of the testes, A. niger recorded that Ag(I)-L 3 showed a significant difference the highly results (20 ± 2.6) though free L 3 results showed less than its complex. Others did not show any activity against tested fungi (A. niger). The effect of various significant ligands and other complexes on P. expansum did not show any activity whereas the the highest broad spectrum of activity on the same test strain showed the best results on L 1 and its complexes [42]. Normal antibiotic efficacy of antimicrobials (AMC, CTX, NS, FU). The AMC mixture give the effective against E. coli, Coliform, S. aureus and NS high inhibitory activity on A. niger. Other antibiotics have shown no action on other microorganisms. Eventually, the bacterial strains showed a varied response to the three free ligands and their complex antimicrobial activity, but the results indicated that the high activity of ligand complexes was better than their free ligands. The two fungal strains are more resistant to synthesis ligands and their complexes than bacterial strains [42][43][44][45][46].

Determination of MIC for the most sensitive organisms
The artificial ligands and their complexes developed the biological efficacy towaeds the more resistant  Table 5E, F and Fig. 6 data showed that the lowest MIC for the two strains measured at conc. 0.02 mg/100 mL. Although MIC at complex L 3 was recorded by A.niger, the same result was recorded on Ag(I)-L 3 at conc. 0.02 mg/100 mL. Ligand L 1 and its complexes demonstrate the strongest MIC on P. expansum, although no behavior is displayed Table 4 The inhibitation diameters zone values (mm) for L 1, L 2 , L 3

Compounds
Microbial species

Chemistry
Analytical grade reagents, commercially available from multiple suppliers and used without further purification, were all the chemicals used in the complex preparation. Synthesized compounds and their complexes have been characterized by elemental analysis, magnetic and spectroscopic methods (IR, 13 C, 1 HNMR, mass) and thermal analysis using the known apparatuses [42].

Synthesis of the complexes
The brown solid complex [Ag(L 1 ) 2 (H 2 O) 2 ]NO 3 was prepared by adding 0.5 mmol (0.085 g) of AgNO 3 in 20 ml of acetone to a stirred suspended solution 1 mmol (0.305 g) of L 1 in 50 ml acetone. The reaction mixture was refluxed for 6 h, the precipitate was drained off, washed several times with acetone and dried under vacuum over anhydrous CaCl 2 . Dark brown [Ag(L 2 ) 2 (H 2 O) 2 ]NO 3 .H 2 O, [Ag(L 3 ) 2 (H 2 O) 2 ]NO 3 solid complexes were prepared in the same manner as mentioned above.