Synthesis, Crystal Structure, Dft Study and Antifungal Activity of 4-(5-((4-bromobenzyl) Thio)-4-phenyl-4h-1,2,4-triazol-3-yl)pyridine

The title compound

In view of these facts, and also as part of our work [22][23][24][25][26] on the synthesis of bioactive lead compounds for drug discovery, the title compound was designed by introducing pyridine pharmacophore into the 1,2,4-triazole scaffold.This new 1,2,4-triazole derivative characterized by 1 H NMR, MS, elemental analysis and single-crystal X-ray structure analysis as well as its antifungal activity has been tested.

Crystal Structure
The crystal crystallizes in the triclinic space group P-1.The molecular structure is shown in Figure 1, and the packing diagram is in Figure 2. The selected bond lengths and torsion angles are listed in Table 1.CCDC-1438378 contains the supplementary crystallographic data for this crystal.These data can be obtained free of charge at www.ccdc.cam.ac.uk/conts/retrieving.html or from the Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: +44-1223-336033; e-mail: deposit@ccdc.cam.ac.uk.

Crystal Structure
The crystal crystallizes in the triclinic space group P-1.The molecular structure is shown in Figure 1, and the packing diagram is in Figure 2. The selected bond lengths and torsion angles are listed in Table 1.CCDC-1438378 contains the supplementary crystallographic data for this crystal.These data can be obtained free of charge at www.ccdc.cam.ac.uk/conts/retrieving.html or from the Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: +44-1223-336033; e-mail: deposit@ccdc.cam.ac.uk.Generally, the average bond lengths and bond angles of ring systems (phenyl, pyridine and triazole) are in normal ranges [27][28][29].The C6=N4 bond (1.311(2)Å) in 1,2,4-triazole ring is longer than the general C=N double-bond (imine or Schiff Base) length of 1.27 Å.The bond angle of C7-S1-C14 is 98.36(7)°.The torsion angle of the thioether group C7-S1-C14-C15 is 174.13(10)°.From Table 1, calculated bond lengths and bond angles were found to differ from experimental values.For example,  Generally, the average bond lengths and bond angles of ring systems (phenyl, pyridine and triazole) are in normal ranges [27][28][29].The C6=N4 bond (1.311(2)Å) in 1,2,4-triazole ring is longer than the general C=N double-bond (imine or Schiff Base) length of 1.27 Å.The bond angle of C7-S1-C14 is 98.36 (7) ˝.The torsion angle of the thioether group C7-S1-C14-C15 is 174.13(10)˝.From Table 1, calculated bond lengths and bond angles were found to differ from experimental values.For example, the bond S1-C14 is 1.938 Å in our calculation result, but it is shorter in the crystal with the bond length of 1.8273(18) Å.
Crystals 2016, 6, 4 3 the bond S1-C14 is 1.938 Å in our calculation result, but it is shorter in the crystal with the bond length of 1.8273(18) Å.As shown in Figure 1, each of the four rings is planar.The interplanar angles between the triazole ring on the one hand and the pyridine, phenyl (C8-C13) and phenyl (C15-C20) rings on the other hand are 23.9°,72.9° and 37.5°, respectively.Meanwhile, the mean plane of the pyridine ring forms angles of 72.4° and 60.6°, respectively, with the phenyl rings (C8-C13) and (C15-C20).The planes of the two phenyl rings form an angle of 68.7° with one another.
An interesting feature is the intramolecular edge-to-face π-π stacking, which exists between the CH and the two phenyl rings (C8-C13 and C15-C20) (Figure 2); the distances between the CH and the centroids of the phenyl rings are 3.439 and 2.833 Å, and the angles of CH and the centroids of phenyl ring and pyridine ring are 88.05° and 87.42°, respectively.Between the CH of the phenyl ring and pyridine ring, the intramolecular edge-to-face π-π stacking also exists, with the distance between the CH and the centroid of the pyridine ring is 2.666 Å, and the angle of CH and the centroid of the pyridine ring is 85.38°, respectively.In addition, the intermolecular face-to-face π-π stackings exist between two phenyl rings: the centroid separation of them is 3.8744( 18) Å (C15-C20), and the dihedral angle between the two π planes is 0.00°.These interactions are estimated to play a role in stabilizing the crystal structure.

Frontier Orbital Energy Analysis and Molecular Total Energies
Molecular total energy and frontier orbital energy levels are listed in Table 2.The energy gap between Highest Occupied Molecular Orbital (HOMO) and Lowest Unoccupied Molecular Orbital (LUMO) was calculated by Becke's nonlocal three parameter exchange and correlation functional with Lee-Yang-Parr correlation functional (B3LYP).According to the frontier molecular orbital theory, HOMO and LUMO are the most important factors that affect the bioactivity.As shown in Figure 1, each of the four rings is planar.The interplanar angles between the triazole ring on the one hand and the pyridine, phenyl (C8-C13) and phenyl (C15-C20) rings on the other hand are 23.9 ˝, 72.9 ˝and 37.5 ˝, respectively.Meanwhile, the mean plane of the pyridine ring forms angles of 72.4 ˝and 60.6 ˝, respectively, with the phenyl rings (C8-C13) and (C15-C20).The planes of the two phenyl rings form an angle of 68.7 ˝with one another.
An interesting feature is the intramolecular edge-to-face π-π stacking, which exists between the CH and the two phenyl rings (C8-C13 and C15-C20) (Figure 2); the distances between the CH and the centroids of the phenyl rings are 3.439 and 2.833 Å, and the angles of CH and the centroids of phenyl ring and pyridine ring are 88.05 ˝and 87.42 ˝, respectively.Between the CH of the phenyl ring and pyridine ring, the intramolecular edge-to-face π-π stacking also exists, with the distance between the CH and the centroid of the pyridine ring is 2.666 Å, and the angle of CH and the centroid of the pyridine ring is 85.38 ˝, respectively.In addition, the intermolecular face-to-face π-π stackings exist between two phenyl rings: the centroid separation of them is 3.8744( 18) Å (C15-C20), and the dihedral angle between the two π planes is 0.00 ˝.These interactions are estimated to play a role in stabilizing the crystal structure.

Frontier Orbital Energy Analysis and Molecular Total Energies
Molecular total energy and frontier orbital energy levels are listed in Table 2.The energy gap between Highest Occupied Molecular Orbital (HOMO) and Lowest Unoccupied Molecular Orbital (LUMO) was calculated by Becke's nonlocal three parameter exchange and correlation functional with Lee-Yang-Parr correlation functional (B3LYP).According to the frontier molecular orbital theory, HOMO and LUMO are the most important factors that affect the bioactivity.HOMO has a priority to provide electrons, while LUMO can accept electrons firstly.Thus, study on the frontier orbital energy can provide useful information about the biological mechanism.From the Figure 3, the geometry of the title compound was optimized using the DFT method.The LUMO of the title compound is mainly located on the pyridine ring, 1,2,4-triazole ring, phenyl ring and SCH 2 group, while the HOMO of the title compound is located on the pyridine ring, 1,2,4-triazole ring and SCH 2 group.The fact is that the electron transitions from the pyridine ring, 1,2,4-triazole ring and SCH 2 group to the phenyl ring, while the energy gap between the HOMO and LUMO is 0.17499 Hartree.
Crystals 2016, 6, 4 4 HOMO has a priority to provide electrons, while LUMO can accept electrons firstly.Thus, study on the frontier orbital energy can provide useful information about the biological mechanism.From the Figure 3, the geometry of the title compound was optimized using the DFT method.The LUMO of the title compound is mainly located on the pyridine ring, 1,2,4-triazole ring, phenyl ring and SCH2 group, while the HOMO of the title compound is located on the pyridine ring, 1,2,4-triazole ring and SCH2 group.The fact is that the electron transitions from the pyridine ring, 1,2,4-triazole ring and SCH2 group to the phenyl ring, while the energy gap between the HOMO and LUMO is 0.17499 Hartree.

Mulliken Atomic Charges and Electrostatic Potential (ESP)
Table 3 exhibits the calculated mulliken atomic charges except for atoms H. Taking DFT, for example, again (Figure 4), all the nitrogen atoms (N1, N3, N4) are the most negatively charged ones, which can interact with the positively charged part of the receptor easily.Therefore, we supposed this compound can combine the amino-acid residue on its surface by the interaction of the pyridine ring or 1,2,4-triazole ring, which may be responsible for the bioactivity.

Mulliken Atomic Charges and Electrostatic Potential (ESP)
Table 3 exhibits the calculated mulliken atomic charges except for atoms H. Taking DFT, for example, again (Figure 4), all the nitrogen atoms (N1, N3, N4) are the most negatively charged ones, which can interact with the positively charged part of the receptor easily.Therefore, we supposed this compound can combine the amino-acid residue on its surface by the interaction of the pyridine ring or 1,2,4-triazole ring, which may be responsible for the bioactivity.

Instruments
Melting points were determined using an X-4 apparatus and uncorrected.The 1 H NMR spectra were measured on a Bruker AV-400 instrument (Fallanden, Switzerland) using TMS as an internal standard and CDCl3 as the solvent.Elemental analyses were performed on a Vario EL elemental analyzer (Hanau, Germany).Crystallographic data of the compound were collected on a rigaku saturn diffractometer (Tokyo, Japan).All the reagents are of analytical grade or freshly prepared before use.

General Procedure
The synthetic route of title compound was outlined in Scheme 1.The intermediate 1, 2 was synthesized according to the reference [30,31].A mixture of isonicotinyl hydrazine (1.37 g, 10 mmol) with isothiocyanatobenzene (1.35 g, 10 mmol) was refluxed for 5 h in ethanol.After cooling down to room temperature, the products were obtained and recrystallized from methanol to give 3, yield 95%.A mixture of compound 3 (10 mmol) in aqueous NaOH solution (5 mL, 2 N) was refluxed for 4 h.After cooling down to room temperature, HCl aqueous solution (4 N) was added to afford a large amount of precipitate.The solid was filtered, dried and recrystallized from methanol to give intermediate 4, yield 88%.A CEM designed 10 mL pressure-rated vial was charged with DMF (5 mL), 4 (0.25 g, 1 mmol), 4-bromo-1-(chloromethyl)benzene (1.1 mmol), and NaOH (0.05 g, 1.2 mmol).The mixture was irradiated in a CEM Discover Focused Synthesizer (Matthews, MO, USA) (150 W, 90 °C, 200 psi, 15 min).The mixture was cooled to room temperature by passing compressed air through the microwave cavity for 2 min.It was poured into cold ice (40 mL) and the formed precipitate was filtered.The crude solid was recrystallized from EtOH to give the title compound.

Evaluation of the Bioactivity
The in vivo fungicidal activities of the title compound against Stemphylium lycopersici (Enjoji) Yamamoto, Fusarium oxysporum.

Instruments
Melting points were determined using an X-4 apparatus and uncorrected.The 1 H NMR spectra were measured on a Bruker AV-400 instrument (Fallanden, Switzerland) using TMS as an internal standard and CDCl 3 as the solvent.Elemental analyses were performed on a Vario EL elemental analyzer (Hanau, Germany).Crystallographic data of the compound were collected on a rigaku saturn diffractometer (Tokyo, Japan).All the reagents are of analytical grade or freshly prepared before use.

General Procedure
The synthetic route of title compound was outlined in Scheme 1.The intermediate 1, 2 was synthesized according to the reference [30,31].A mixture of isonicotinyl hydrazine (1.37 g, 10 mmol) with isothiocyanatobenzene (1.35 g, 10 mmol) was refluxed for 5 h in ethanol.After cooling down to room temperature, the products were obtained and recrystallized from methanol to give 3, yield 95%.A mixture of compound 3 (10 mmol) in aqueous NaOH solution (5 mL, 2 N) was refluxed for 4 h.After cooling down to room temperature, HCl aqueous solution (4 N) was added to afford a large amount of precipitate.The solid was filtered, dried and recrystallized from methanol to give intermediate 4, yield 88%.A CEM designed 10 mL pressure-rated vial was charged with DMF (5 mL), 4 (0.25 g, 1 mmol), 4-bromo-1-(chloromethyl)benzene (1.1 mmol), and NaOH (0.05 g, 1.2 mmol).The mixture was irradiated in a CEM Discover Focused Synthesizer (Matthews, MO, USA) (150 W, 90 ˝C, 200 psi, 15 min).The mixture was cooled to room temperature by passing compressed air through the microwave cavity for 2 min.It was poured into cold ice (40 mL) and the formed precipitate was filtered.The crude solid was recrystallized from EtOH to give the title compound.

Structure Determination
The cube-shaped single crystal of the title compound was obtained by recrystallization from EtOH.The crystal size is 0.20 mm × 0.18 mm × 0.14 mm.A total of 9617 reflections were collected, 4313 of which were independent (Rint = 0.031) and 3145 were observed with I > 2σ(I).The calculations were performed with SHELXS-97 program [32] and the empirical absorption corrections were applied to all intensity data.The non-hydrogen atoms were refined anisotropically.The positions of H atoms were refined using riding models.A summary of the key crystallgraphic information was given in Table 4.

Therotical Calculations
According to the above crystal structure, a crystal unit was selected as the initial structure, while DFT-B3LYP/6-31G methods in Gaussian 03 package [33] were used to optimize the structure of the title compound.Vibration analysis showed that the optimized structures were in accordance with the minimum points on the potential energy surfaces, which means no virtual frequencies, proving that the obtained optimized structures were stable.All the convergent precisions were the system default values, and all the calculations were carried out on a DELL computer.

Structure Determination
The cube-shaped single crystal of the title compound was obtained by recrystallization from EtOH.The crystal size is 0.20 mm ˆ0.18 mm ˆ0.14 mm.A total of 9617 reflections were collected, 4313 of which were independent (R int = 0.031) and 3145 were observed with I > 2σ(I).The calculations were performed with SHELXS-97 program [32] and the empirical absorption corrections were applied to all intensity data.The non-hydrogen atoms were refined anisotropically.The positions of H atoms were refined using riding models.A summary of the key crystallgraphic information was given in Table 4.

Therotical Calculations
According to the above crystal structure, a crystal unit was selected as the initial structure, while DFT-B3LYP/6-31G methods in Gaussian 03 package [33] were used to optimize the structure of the title compound.Vibration analysis showed that the optimized structures were in accordance with the minimum points on the potential energy surfaces, which means no virtual frequencies, proving that the obtained optimized structures were stable.All the convergent precisions were the system default values, and all the calculations were carried out on a DELL computer.

Conclusions
In summary, a new crystal structure, 4-(5-((4-bromobenzyl)thio)-4-phenyl-4H-1,2,4triazol-3-yl)pyridine, has been prepared by multi-step reaction and characterized by 1 H NMR, MS and elemental analyses and single-crystal X-ray structure determination.The results show that the crystal structure exhibits intermolecular π-π stacking.The frontier orbital energy analysis, mulliken atomic charges and electrostatic potential were also studied by using the DFT method.The antifungal bioassay showed that it possessed moderate activity.

Figure 1 .
Figure 1.View of the title compound, with displacement ellipsoids drawn at the 30% probability level.

Figure 1 .
Figure 1.View of the title compound, with displacement ellipsoids drawn at the 30% probability level.

Figure 2 .
Figure 2. A view of pack title compound.

Figure 2 .
Figure 2. A view of pack title compound.

Scheme 1 .
Scheme 1.The synthetic route of title compound.

Scheme 1 .
Scheme 1.The synthetic route of title compound.

Table 1 .
Selected bond lengths (Å), angles ( ˝) and theoretical calculations for the title compound.

Table 2 .
Total energy and frontier orbital energy.

Table 2 .
Total energy and frontier orbital energy.

Table 3 .
Mulliken atomic charges except for atoms H (e).

Table 3 .
Mulliken atomic charges except for atoms H (e).

Table 4 .
Crystal data of the title compound 5.

Table 4 .
Crystal data of the title compound 5.