SONG Huan WANG Jia-Kai CHEN Xiao-Yan TIAN Xiao-Yan LI Ying-Lian LI Bing ②

a (State Key Laboratory of High-efficiency Utilization of Coal and Green Chemical Engineering, Ningxia University, Yinchuan 750021, China) b (Department of Chemistry and Chemical Engineering, Ningxia University, Yinchuan 750021, China)

ABSTRACT A new complex [Zn(3,4-APT)2(H2O)4]·8H2O (1, 3,4-HAPT = 4-(5-(pyridin-3-yl)-4H-1,2,4-triazol- 3-yl) benzoic acid) has been prepared and characterized by elemental analysis, X-ray single-crystal diffraction analysis, thermogravimetric analysis and infrared spectrum analysis. Theoretical calculation based on density functional theory (DFT) is also employed to explicate frontier orbitals of 3,4-HAPT. X-ray single-crystal diffraction analysis reveals that 1 belongs to the triclinic system, space group P with a = 7.5123(3), b = 8.6745(3), c = 15.2074(6) Å, α = 78.469(1), β = 87.387(1), γ = 65.448(1)°, V = 882.42(6) Å, Z = 1, Dc = 1.528 g·cm-3, μ = 0.780 mm-1, Mr = 812.09, F(000) = 424, the final R = 0.0401 and wR = 0.1136. Zn(II) ion is coordinated by two N atoms from two 3,4-HAPT as well as four O atoms from four coordinated water molecules, forming a 0D motif with distorted octahedral coordinated geometry. The adjacent 0D units are linked into a 3D supramolecular structure through hydrogen bonding interaction. In addition, complex 1 exhibits better antifungal activity against Colletotrichum gloeosporioides Penz than the ligand and metal salt by MIC, MBC tests and Kirby-Bauer disc diffusion method which exhibit potential application in the antifungal fields.

Keywords: 4-(5-(pyridin-3-yl)-4H-1,2,4-triazol-3-yl) benzoic acid, crystal structure, antifungal activity;

1 INTRODUCTION

Bacteria and fungus have resistance by the frequent use of antibacterial agents, leading to treatment drawbacks for a large number of drugs[1]. Therefore, it is necessary to find new efficiency and broad-spectrum antimicrobial agents which have different mechanisms of action aimed at a better understanding of antimicrobial resistance[2].

It is well know that triazole and their derivatives have extensive biological activity, including antitumor[3,4], antimi- crobial[5,6], insecticide[7,8], antivira[9], herbicide[10], etc. As a novel triazole derivative, 4-(5-(pyridin-3-yl)-4H-1,2,4-tria- zol-3-yl) benzoic acid (3,4-HAPT) has attracted much attention in recent years[11]. For one reason, triazole exists in numerous natural products and is extremely important in the chemistry of biological systems[12-14]. For another reason, the pyridyl and benzoic acid groups not only have strong coordination ability[15,16], but also exert potential antibacterial activity against the pathogenic bacteria[17].

In view of the above situations, a new complex [Zn(3,4- APT)2(H2O)4]·8H2O has been designed, synthesized and characterized by elemental analysis, X-ray single-crystal diffraction analysis, thermogravimetric analysis and IR spectrum analysis. The frontier orbitals of the ligand were searched by (DFT) with the 6-31G basis set to provide useful insights into the chemical reactivity and stability. Furthermore, the antifungal activity of 1 was also tested against

Colletotrichum gloeosporides Pens determined by minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) tests and Kirby-Bauer disc diffusion method.

2 EXPERIMENTAL

2. 1 Materials and measurements

All commercially available chemicals reagents were AR grade and used without further purification. Elemental analyses (C, H and N) were carried out at the Vario EL III analyzer. Infrared spectra were obtained from KBr pellets on a BEQ VZNDX 550 FTIR instrument within the 400～4000 cm-1region. Thermo- gravimetric analysis was carried out on a TA Instruments NETZSCH STA 449 C simultaneous TGA at a heating rate of 10 ℃·min-1under hydrostatic air.

2. 2 X-ray crystallography

Crystallographic data of complex 1 were collected on a Bruker/Siemens Smart Apex II CCD diffractometer with graphite-monochromated MoKα radiation (λ = 0.71073 Å) at 293(2) K. Cell parameters were retrieved using SMART software and refined using SAINTPLUS[18]for all observed reflections. Data reduction and correction for Lp and decay were performed using the SAINTPLUS software. Absorption corrections were applied using SADABS[19]. The structure was solved by direct methods and refined by full-matrix least-squares on F2using SHELXL-2017[20]and SHELXS- 2017[21]programs. All non-hydrogen atoms were refined anisotropically. Hydrogen atoms were placed in geometrically calculated positions. For 1, a total of 16877 reflections were collected in the range of 1.40≤θ≤27.60°, of which 3613 were independent (Rint= 0.0620). The final R = 0.0401 and wR = 0.1136 for all data with (Δρ)max= 0.370 and (Δρ)min= - 540 e·Å-3. Selected bond lengths and bond angles as well as hydrogen bonding geometry of 1 are shown in Tables 1 and 2.

Table 1. Selected Bond Lengths (Å) and Bond Angles (°) for Complex 1

Table 2. Hydrogen Bond Lengths (Å) and Bond Angles (°)

2. 3 Antifungal activity assays

Antifungal activities of the ligand, metal salt and complex 1 were tested against Colletotrichum gloeosporides Pens determined by MIC and MBC tests[22]and Kirby-Bauer disc diffusion method[23]. The complex was insoluble in water and the assay was carried out using DMF as solvent. A set of assay tubes containing only inoculated medium with DMF as negative control and prochloraz as positive control[5]were also done simultaneously. The diameters of filter paper were 5 mm. Flat plates were incubated at 25 ℃ for 12, 24, 36, 48, 60 and 72 h, respectively. Their inhibition diameter (including filter paper) was measured with a vernier caliper. All experiments were carried out in parallel for three times and the average diameter values were calculated.

2. 4 Synthesis of complex 1

A water solution (10 mL) of Zn(OAc)2·2H2O (0.0088 g, 0.04 mmol) and methanol (5 mL) was added to a solution of 3,4-HAPT (0.0053 g, 0.02 mmol) in DMF (1 mL). After 30 min of vigorous mixing, the resulting solution was filtered and kept at room temperature. Upon slow evaporation of the solvents, pale yellow crystals of 1 were obtained after ca. 14 days in a yield of 52% (based on 3,4-HAPT). Anal. Calcd. for (C28H42N8O16Zn) (%): C, 41.41; H, 5.21; N, 13.80. Found (%): C, 41.92; H, 5.65; N, 13.01. IR(cm-1): 3545s, 3489s, 3412s, 1616m, 1591m, 1552m, 1392m, 1132m, 989w, 748m, 702w, 478w.

3 RESULTS AND DISCUSSION

3. 1 Quantization calculation of the ligand

The density functional calculations were performed with the GAUSSIAN 09 program. The geometry optimization of the ligand has been performed by using B3LYP functional of DFT in conjunction with the 6-31G basis set.

The gap energies between the frontier molecular orbitals are often used to provide useful insights into the chemical reactivity and stability of chemical species[24]. The gap energy between the frontier molecular orbitals (ΔE = LUMO-HOMO) of the ligand is 0.15717 eV. The big absolute value of the gap energy shows that the 3,4-HAPT can keep thermodynamic stability in the experimental conditions and not convert to other products.

3. 2 Structural description

Single-crystal X-ray diffraction analysis reveals that 1 crystallizes in triclinic system with Pspace group. 1 is a 0D coordination pattern, including a Zn(II) center, two 3,4-HAPT, four coordinated water molecules and eight free water molecules. As shown in Fig. 1, the Zn(II) center is surrounded by four oxygen atoms in the equatorial plane from four coordinated water molecules and two nitrogen atoms in the axial position from two 3,4-HAPT, which form a distorted octahedral geometry. As shown in Table 1, the Zn-O distances range from 2.089(2) to 2.094(2) Å and Zn-N distances is 2.211(2) Å. The O-Zn-O bond angles range from 87.72(6)° to 180.00° and O-Zn-N bond angles change from 88.67(6)° to 91.33(6)°. In addition, the 0 motif can be linked into 1D chains via hydrogen bonds O(3)-H(3B)···O(2), O(4)- H(4A)···O(1) and O(8)-H(8A)···O(1). Further, the 1D chains are formed into a 2D pattern through O(3)-H(3A)···O(7), N(2)-H(2N)···O(5), O(5)-H(5B)···N(3), O(6)-H(6B)···O(5), O(7)-H(7B)···O(8) and O(8)-H(8B)···O(4). Finally, these plains are cross-linked by O(4)-H(4B)···O(7), O(5)- H(5A)···O(2), O(6)-H(6A)···N(1) and O(7)-H(7A)···O(5) to create a 3D supramolecular structure (Fig. 2).

Fig. 1. Coordination environment of complex 1 (Lattice water and H atoms are omitted for clarity)

Fig. 2. Hydrogen bonding interaction diagram of 1 (3D)

3. 3 IR spectra

The IR spectrum of 1 displays a medium intensity broad absorption band at 3489 cm-1, corresponding to the O-H stretching of water molecules in the complex. The sharp absorption band at 1392 cm-1corresponds to the C-N stretching of triazole. Asymmetric and symmetric C=O stretching modes of the ligated benzoate moieties were evidenced by very strong, slightly broadened bands at 1552 cm-1. The absence of any bands in the area of ～1706 cm-1indicates full deprotonation of carboxylate group in 1[25], which is consistent with the results of X-ray analysis.

3. 4 Thermogravimetric analysis

Thermogravimetric experiments were conducted to study the thermal stability of 1, which is an important parameter for complex. As shown in Fig. 3, the TGA curve of the title complex suggests the first weight loss of 26.6% (calcd.: 26.6%) in the range of 30 ～196 ℃, corresponding to complete loss of four coordinate water molecules and eight free water molecules with an exothermic peak at 119 ℃. The main framework remains intact until heated to 285 ℃, and then releases 3,4-HAPT completely from 285 to 843 ℃ with an exothermic peak at 301 ℃ and converts to ZnO with the residual amount of 9.9%, which is in agreement with the calculated value 10.0%. The residual sample was characterized by X-ray powder diffraction (XRPD) at room temperature. As shown in Fig. 4, all diffraction peaks are in good agreement with the standard diffraction data for ZnO (JCPDS card file No. 65-2880)[26].

Fig. 3. TGA curve for complex 1

Fig. 4. XPRD patterns of the residual and ZnO

3. 5 Antifungal activity

The antifungal activity qualitative determination for complex 1 is presented in Fig. 5. Complex 1 exhibits better antifungal activity against Colletotrichum gloeosporioides Penz than corresponding ligands and metal salt. What's more, with the increase of incubation time, the antifungal effect of 1 was gradually enhanced. The positive and negative control tests under the identical experimental condition were explored with prochloraz and DMF as substances, respectively. No antifungal activity was observed in DMF and the inhibition diameter of complex 1 was nearly 24 mm at 72 h, which is slightly less than of prochloraz (31 mm). Based on the concentration test, the inhibition diameter of 1 has also exhibited excellent antifungal effect on the tested microorganism (concentration = 1, 3, 5 mg/mL). In the same incubation time, the inhibition diameters of 1 are 12, 24 and 27 mm, respectively (Fig. 6), indicating that the title complex has concentration-dependent antifungal activity[27]. The MICs of 3,4-HAPT and 1 are 1500 and 93.75 μg/mL, respectively, while the MBCs of 3,4-HAPT and 1 are 3000 and 187.5 μg/mL, respectively. Compared with the 3,4-HAPT, the MIC and MBC of 1 were significantly lower, so the title complex has potential applications as antifungal agent[22].

From the results, it is realized that 1 has a higher activity than free ligand. According to Overtone’s concept of cell permeability[28], liposolubility is an important factor to control the antimicrobial activity. Complex 1 with good lipid- solubility favors to passage of the lipid membrane that surrounds the cell materials. Moreover, complex 1 may disturb the respiration process of the cell and thus blocks the synthesis of proteins that restricts the further growth of fungus[29].

Fig. 5. Mean inhibition diameters (mm) for the antifungal activity of DMF, ligand, metal salt and complex 1 with different incubation time

Fig. 6. Mean inhibition diameters (mm) of complex 1 against Colletotrichumgloeosporioides Penz in the range of 1～5 mg/mL

4 CONCLUSION

In summary, the title complex [Zn(3,4-APT)2(H2O)4]·8H2O has been successfully synthesized based on 4-(5-(pyridine- 3-yl)-4H-1,2,4-triazol-3-yl) benzoic acid. The structure was characterized by single-crystal X-ray diffraction, elemental analysis, IR spectroscopy and thermogravimetric analysis. The molecular frontier orbital of 3,4-HAPT through density functional theory (DFT) was obtained. Antifungal activity tests indicate that 1 exhibits better antifungal effect towards Colletotrichum gloeosporioides Penz than the ligand and metal salt which show potential applications for the control of fungal infections. Further investigation is currently being processed.

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