Crystal Structures, Vibrational Spectra, and Fungicidal Activity of 1,5-Diaryl-3-oxypyrazoles

The aryloxypyrazole structure is present in a number of bioactive molecules. Four 1,5-diaryl-3-oxypyrazoles containing benzoyl (I), thiazolidinethione (II and III) or per-O-acetylated glucopyranosyl (IV) moieties were characterized by single-crystal X-ray diffraction. Compounds I and II crystallize in a triclinic P-1 system, whereas III and IV crystallize in an orthorhombic Pbca and a monoclinic P21 space groups, respectively. The dihedral angles between the two benzene rings of the pyrazole are 61.33° (I), 62.87° (II), 57.09° (III) and 70.25° (IV). The structures were stabilized by classical intra- (C-H···S for II and III, C-H···O for IV) and intermolecular (C-H···O for I and IV) H-bonds, as well as intermolecular C-H···π stacking interactions. The theoretical FTIR results showed good agreement with the experimental data. Compounds IV, II and III showed moderate fungicidal activity against Sclerotinia sclerotiorum and Gibberella zeae. The structure-activity relationships were discussed.

In this paper, we report the crystal structures and FTIR spectra of four 1,5-diaryl-3-oxypyrazoles bearing benzoyl (I), thiazolidinethione (II and III) or per-O-acetylated glucopyranosyl (IV) moieties ( Figure 1). Meanwhile, their in vitro fungicidal activity against Sclerotinia sclerotiorum and Gibberella zeae has been investigated, and their structure-activity relationshipe were also discussed.

Experimental and Theoretical FTIR Results
The experimental and theoretical FTIR spectra for I-IV are shown in Figure 7, where the intensity is plotted against the vibrational frequencies. The primary vibrational frequencies with assignments are listed in Table 4. The main signals are grouped in three regions, including the ranges of 2800-3200 cm −1 , 1000-1800 cm −1 and 500-1000 cm −1 . The second region shows strongly mixed vibrational bands. The C=O stretching vibrations of I-IV were found at 1739 cm −1 , 1714 cm −1 , 1708 cm −1 , and 1755 cm −1 , whereas they were calculated at 1737 cm −1 , 1718 cm −1 , 1711 cm −1 , and 1752 cm −1 , respectively. The experimental and theoretical C=C stretching vibrations of the phenyl rings were found in the region of 1440-1630 cm −1 , which were consistent with the literature value of 1,430-1,625 cm −1 [16]. The bands observed in the region of 1000-1300 cm −1 corresponded to the symmetric and asymmetric C-O stretching vibrations. The region below 1000 cm −1 exhibited the out of plane bending C-H vibrations of the aromatic rings, and the region in 3000-3200 cm −1 was the characteristic absorption of the aromatic C-H stretching vibrations. The C-Cl stretching vibration in II was consistent with the aromatic C-Cl stretching vibration in III. The experimental C-F stretching vibration at 1330 cm −1 in IV was consistent with its theoretical value of 1332 cm −1 . These results indicated that the observed and calculated FTIR data were in good agreement with each other. Wavenumber (cm -1 ) Wavenumber (cm -1 ) Wavenumber (cm -1 ) Wavenumber (cm -1 )

Fungicidal Activity
Compounds I-IV were evaluated for in vitro fungicidal activity against Sclerotinia sclerotiorum and Gibberella zeae, at a dosage of 10 μg/mL. As can be seen in Table 5, for Sclerotinia sclerotiorum, Compound IV (29%) possessing a per-O-acetylated glucopyranosyl moiety, displayed better activity than I (0%), II (21%), and III (14%). The reason was speculated that the glucopyranosyl moiety could form more H-bonds ( Figure 2) to improve the hydrophilicity of molecule, which could balance the HLB value and then increase the systemic of molecule within plant. Within the series of thiazolidinethione derivatives, compound II (21%, 32%) with an electron-withdrawing chloro group on the pyrazole ring displayed better fungicidal activity against the two fungi than III (14%, 29%), and both compounds showed better inhibitory activity against Gibberella zeae than I (4%) and IV (11%). The results might imply that the introduction of the heterocycle moiety by full consideration of the electronic effects was important for improving its fungicidal activity. However, compound I containing a benzoyloxy moiety showed the worst activity (0, 4%), which indicated switching the C3-substituent of the pyrazole ring from thiazolidinethione to benzoyloxy moiety had no effective impact on the inhibition rates. The structure-activity relationship revealed that the improvement of bioactivity might require a reasonable design of molecules (e.g., considering H-bonds effect and heterocycle) and full consideration of the electronic effects of electron-withdrawing groups to balance the HLB value and enhance the systemic of the whole molecule.

General Information
Melting points were measured on an X-4 microscope electrothermal apparatus (Taike, Nanjing, China) and were uncorrected. 1 H-NMR spectra were recorded on a Bruker spectrometer (Bruker, Leipzig, Germany) at 300 or 500 MHz using CDCl 3 or DMSO-d 6 as solvent, with tetramethylsilane as an internal standard. Elemental analyses were performed on a Flash EA-1112 elemental analyzer (Thermo, Illinois, USA). FTIR spectra of compounds I-IV were recorded in the region of 4000-400 cm −1 on a Nicolet 380 FT-IR spectrophotometer (Thermo, Waltham, MA, USA) using the KBr pellet technique.

Preparation of Compound I
To a solution of 5-(3,4-dimethoxyphenyl)-1-phenyl-1H-pyrazol-3-ol (0.59 g, 2.0 mmol) in CHCl 3 (50 mL) was added Et 3 N (0.41 g, 4.0 mmol). The mixture was stirred for 10 min and benzoyl chloride (0.42 g, 3.0 mmol) was added. Then, the mixture was stirred at r.t. for 2 h. The solvent was removed under reduced pressure and the residue was recrystallized from ethanol to give the white solid product

X-ray Crystallography
Suitable crystals of I-IV were obtained by slow evaporation of ethyl acetate solutions at r.t. Crystal data were performed on a Nonius CAD-4 diffractometer (Enraf-Nonius, Rotterdam, The Netherlands) by using MoK α (λ = 0.71073 Å) irradiation. All of the structures were solved by direct methods using SHELXS-97 and refined by full-matrix least-squares on F 2 for all data using SHELXL-97 [17]. All non-H-atoms were refined anisotropically, and H-atoms were introduced at calculated positions. The isotropic temperature factors were fixed to 1.2 times (1.5 times for methyl group) the equivalent isotropic displacement parameters of the C-atom the H-atom is attached to CCDC-906894 (I), CCDC-819778 (II), CCDC-918082 (III) and CCDC-925316 (IV) contain the supplementary crystallographic data for this paper. These data can be obtained free of charge via http://www.ccdc.cam.ac.uk/conts/retrieving.html (or from the CCDC, 12 Union Road, Cambridge CB2 1EZ, UK; Fax: +44 1223 336033; E-mail: deposit@ccdc.cam.ac.uk).

FTIR Spectra
The structures in the ground state (in vacuo) were optimized by the Gaussian 03 program using the B3LYP (DFT) method with the 6-31G (d) basis set [18][19][20]. The initial configurations for calculation were constructed according to the X-ray data. Frequency calculations at the same levels of theory revealed no imaginary frequencies, indicating that the B3LYP/6-31G (d) method was the optimal one in our system.

Fungicidal Activity Assays
The in vitro fungicidal activity of compounds I-IV against Sclerotinia sclerotiorum and Gibberella zeae was investigated at a dosage of 10 μg/mL, according to a reported method [10]. The fungi were obtained from Jiangsu Pesticide Research Institute Co., Ltd., Nanjing, China. The tested compounds I-IV were dissolved in acetone and added to a sterile agarized Czapek-Dox medium at 45 °C. In preliminary screenings, the compounds were used in a concentration of 10 μg/mL. The control sample contained only one equivalent of acetone. The media were poured onto 8-cm Petri dishes (10 mL for each dish) and after 2 days inoculated with 5-mm PDA discs of overgrown mycelium. In the case of Sclerotinia sclerotiorum, the medium was inoculated by a prick of laboratory needle containing fungus spores. The Petri dishes were incubated at r.t. in the dark. After 4 days, the diameters of the inoculation of the cultures were measured. The percentage inhibition of fungal growth was determined by comparison between the development of fungi colonies on media containing compounds and on the control. Three replicates of each test were carried out.

Conclusions
Four 1,5-diaryl-3-oxypyrazoles containing benzoyl (I), thiazolidinethione (II and III) or per-O-acetylated glucopyranosyl (IV) moieties have been analyzed by X-ray diffraction. The molecules were stabilized by classical intra-and intermolecular H-bonds, as well as intermolecular C-H···π stacking interactions. Compound IV with more H-bonds in the crystal displayed better activity (29%) against Sclerotinia sclerotiorum than I (0%), II (21%), and III (14%). Compound II (21%, 32%) showed better fungicidal activity against the two fungi than III (14%, 29%), and both II and III exhibited better inhibitory activity against Gibberella zeae than I (4%) and IV (11%). The structure-activity relationship revealed that the improvement of bioactivity might require full consideration of H-bonds effect, heterocycle, and electronic effects of electron-withdrawing groups to balance the HLB value and enhance the systemic of molecule.