Synthesis, Density Functional Theory, Insecticidal Activity, and Molecular Docking of Some N-Heterocycles Derived from 2-((1,3-Diphenyl-1H-Pyrazol-4-yl)Methylene)Malonyl Diisothiocyanate

Abstract The extensive pharmacological applications of the pyrazole-based compounds stimulated us to synthesize the highly functionalized, pyrazolylmalonyl diisothiocyanate derivative (2) via treating 2-((1,3-diphenyl-1H-pyrazol-4-yl)methylene)malonyl dichloride (1) with ammonium thiocyanate in acetonitrile at room temperature. The diisothiocyanate derivative 2 was subsequently conducted with a double molar ratio of different nitrogen nucleophiles like 4-acetylaniline, 2-aminobenzoic acid, 6-aminothiouracil, 2-aminothiadiazole derivative, hydrazine, phenylhydrazine, thiophene-2-carbohydrazide, 2-hydroxybenzohydrazide, 2-cyanoacetohydrazide, thiourea, thiosemicarbazide, 2-aminoaniline, and thiocarbohydrazide, aiming to construct some valuable heterocyclic systems. The synthesized compounds were screened for their insecticidal activity against healthy late third instar larvae P. interpunctella and Nilaparvata lugens and the results showed that pyrimidinethione, thiadiazolotriazine, and tetrazinethione derivatives exhibited the highest insecticidal potency that are supported by DFT stimulation and their molecular docking.

In turn, hydrazinolysis of the diisothiocyanate derivative 2 using hydrazine hydrate acquired triazepinotriazepine derivative 7 (Scheme 2).The IR spectrum of 7 offered the absorption bands for NH, C¼O, and C¼S groups.The olefinic proton (CH¼) disappeared in its 1 H NMR spectrum.The formation of 7 can be explained via Scheme 3. The bistriazole derivative 8 was produced via treating 2 with phenylhydrazine.The IR spectrum of 8 was devoid of carbonyl absorption.Also, its 1 H NMR spectrum showed an exchangeable singlet signal at d 14.43 ppm corresponding to the SH proton.This reaction can proceed via first addition to the N ¼ C¼S moiety followed by 1,5-exo-trig cyclization to eliminate a water molecule.
On the other hand, the interaction of 2 with thiophene-2-carbohydrazide, 2-hydroxybenzohydrazide, and 2-cyanoethanohydrazide provided the bisthiosemicarbazide derivatives 9-11, respectively (Scheme 2).Their IR spectra retained the carbonyl absorption bands, as well as their 1 H NMR spectra conserved the singlet signal of the olefinic proton.The assigned structures were fully supported by their spectral data (cf.Experimental).
Noteworthy, the reaction of 2 with thiourea afforded the carbamothioyl derivative 12 (Scheme 4).On the other hand, the bistetrazepine 13 and bisbenzotriazepine 14 derivatives were obtained by treating 2 with thiosemicarbazide and 2-aminoaniline, respectively.The IR spectrum of 13 and 14 was devoid of the C¼O group.Finally, the tetrazinethione derivative 15 was provided by treating 2 with thiocarbohydrazide (Scheme 4).The IR spectrum of 15 offered the absorption bands for NH, C¼O, and C¼S groups.

Insecticidal activity
The insecticidal activities of compounds 3-15 were measured against healthy late third instar larvae P. interpunctella and Nilaparvata lugens according to the standard test 35,36 with a slight modification.The test analogs were dissolved in DMF and serially diluted with water containing Triton X-80 (0.1 mg/L) to obtain the required concentrations.The insects were reared at 25 (±1) C and groups were transferred to glass Petri dishes.All experiments were carried out in three replicates for statistical requirements.Assessments of mortality were calculated 48 h by the number and size of the live insects relative to those in the negative control.Evaluations were based on a percentage scale of (0 ¼ no activity and 100 ¼ complete eradication).The mortality rates were subjected to probity analysis. 37hiamethoxam was used as positive control while water containing Triton X-80 (0.1 mg/L) was used as a negative control.The results listed in Table 1 indicated that most of the compounds showed Scheme 1. Synthesis of compounds 3-6.
weak insecticidal activity against the two pests.However, some of the compounds displayed good insecticidal activities.For example, compounds 5, 6, and 15 showed 100% activities at 400 mg/mL and 50% (IC 50 ) activities at 100 and 50 mg/mL, respectively, against both P. interpunctella and Nilaparvata lugens.Meanwhile, the activities of compounds 2 and 4 against Nilaparvata lugens were similar to those of thiamethoxam at 200 mg/mL; compounds 5, 6, and 15 showed 60.0 and 83.3% activity, respectively, against Nilaparvata lugens at 200 mg/mL.In addition, compounds 6 and 15 also showed good insecticidal activities, their mortalities against P. interpunctella were 100% at (400 mg/mL) conc., while the activity of compound 6 at 200 mg/mL concentration, against P. interpunctella was still 53.3%.Moreover, compounds 11 and 13 processed 56.6 and 53.3% activities on P. interpunctella and 60 and 56.6% on Nilaparvata lugens at 400 mg/mL, respectively.The authors thought Trypsin enzymes of P. interpunctella larvae were inhibited by compound 15, and thiamethoxam control which also demonstrated the capacity to bind to chitin.

Molecular docking studies
Molecular docking can be used to study the binding of a drug to receptor enzyme.To understand the molecular basis of insecticidal activity of the potent compounds (5, 6, and 15) and to discover the binding mechanisms of these molecules, these molecules were docked into the active site of GABA-AT receptor (PDB ID: 1OHW) (Table 2).The reference drug, thiamethoxam showed two H-bonds with GLU270 and HIS44 (Table 3).The docking scores of the active compounds were good.The docking results of all molecules in GABA-AT active site are displayed in Table 2.Among them, compound 5 showed eleven hydrogen bonding (GLU419, GLU270, GLU270, HIS206, ARG422, ASN423, HIS206, GLN301, ARG192, ARG192, and ARG192), one pi-cation, and one pi-pi interactions.Compound 6 displayed three hydrogen bonds (TYR69, ASN39, and SER427) and pi-H interaction.In turn, triazinethione derivative 15 disclosed two hydrogen bonds (ARG192 and LYS329) and two pi-H interactions (Table 4).

Density functional theory (DFT)-based characterization
From the minimum inhibitory concentration LC 50 for the most potent compounds 5, 6, and 15, the quantum mechanical program was used for the molecular parameters for the most potent compounds are listed in Table 5, and the fully optimized minimum energy geometrical configuration of the most potent insecticidal compounds.Proceeds, DFT-based LC 50 of such compounds supported their high insecticidal activity.
It is well known that high E HOMO is likely to indicate a strong tendency of the molecules to donate electrons.9][40] The DE of a molecule is a measure of the hardness or softness of a molecule.Hard molecules are characterized by larger values of DE and vice versa.The linear correlation between E HOMO energy level and the insecticidal activity proved that the lower the HOMO energy (more negative values) of the thiacarbamoyl structure, the greater the trend of accepting electrons.The order of decreasing E HOMO , increasing E LUMO values and the energy gap (DE) are directly proportional to increasing the insecticidal efficiency.The tendency of an electron cloud to be distorted from its normal shape is referred to as its polarizability, the greater the polarizability the more inhibitor molecules of the Trypsin enzymes damage the chitin protective film of the larvae, we can consider the polarizability as a resultant of all intramolecular electron transfer interactions.This increased volume enhances the ease of the distortion of the electron cloud which will promote the adsorption of the compound.The HOMO energy (E HOMO ) is a parameter of direct relation to the ionization potential and its value expresses the susceptibility of organic molecules toward attacks by electrophiles.Unlikely, the LUMO energy (E LUMO ) refers to the electron affinity and its value expresses the vulnerability of the molecules toward a nucleophilic attack.To obtain these parameters, the molecules must be subjected first to geometry optimization, and then these parameters are calculated.The HOMO and LUMO distributions are displayed in Figures 2-4.
The listed results indicate that the values of gap energy (DE), where DE ¼ E LUMO À E HOMO , follow the order: 5 < 6 < 15.DE is indicative of the reactivity of organic compounds toward the interaction with Trypsin enzymes to damage the chitin surfaces of larvae.Compounds having  small DE values are generally referred to as soft compounds, while those having large values are called hard compounds.In general, soft compounds are more reactive toward the chitin surface of insect interactions; being capable of donating electrons easily to insect surfaces. 39Generally, for organic material to interact effectively with insect surfaces, it must contain heteroatoms rich in non-bonded electrons (free lone pairs) and/or aromatic rings having p-electrons.Heteroatoms and aromatic rings are the major adsorption centers at which electron transfer occurs between the additive and the microbial cell wall during operating conditions. 40The synthesized compounds under study are rich in O and N atoms containing free lone pairs of electrons beside double bonds and aromatic rings containing p-electrons.Low-gap-energy compounds generally provide good interaction with the chitin of the cell wall of the larvae because the energy required to remove an electron from the last occupied orbital (HOMO) of the most potent compound will be minimized and it will be easy to donate electrons.Several other various quantum chemical parameters can help to predict the molecular characteristics of the optimized structure of the most potent compounds such as ionization potential (I), electron affinity (A), absolute electronegativity (v), and absolute chemical hardness (g) be sure these arrange the binding according to 5 < 6 < 15. v and g are two quantities related to I and A, where v ¼ I þ A/2 and g 5 I À A/2, I and A are calculated in turn, from E HOMO and E LUMO , where I ¼ ÀE HOMO and A ¼ ÀE LUMO .For any two molecules in contact with each other, electrons will be partially transferred from the one of low value to that of higher value.For the studied compounds, v is generally lower than that of the chitin of larvae, which is 6 eV, and thus chitin-most potent compounds interaction may proceed via electron transfer from the former to the latter.Since soft compounds are preferred for such interactions, one can conclude that compound 15 is the best candidate for interacting with its surface, where it has the highest value of chemical softness (s), which is defined as the inverse of hardness (s ¼ 1/g). 39The electrophilicity x index is given by the simple expression: x ¼ E 2 /g.The electrophilicity x index encompasses the tendency of an electrophile to acquire an extra amount of electron density, given by binding energy (E), and the resistance of a molecule to exchange electron density with the environment, given by g.Thus, a good electrophile is a species  characterized by a high (E) value and a low (g) value.Besides, the maximum number of electrons that an electrophile can acquire is given by the expression: DN max ¼ E/g.Hence, compounds 6 and 15 are predicted to be the best powerful additive for insects.

Conclusion
New series of pyrazole-based heterocycles like pyrimidine, thiadiazolotriazine, triazepinotriazepine, triazole, benzotriazepine, and tetrazine derivatives as well as acyl thiourea skeletons have been synthesized using the highly functionalized malonyl diisothiocyanate derivative through reactions with some nitrogen nucleophiles.The insecticidal activity screening of these compounds against healthy late third instar larvae P. interpunctella and N. lugens offered that pyrimidinethione, thiadiazolotriazine, and tetrazinethione derivatives exhibited the highest insecticidal potency, which based on DFT optimization and supported with their molecular docking.

General
Melting points are uncorrected and were measured on a GALLENKAMP electric melting point apparatus.The IR spectra (, cm À1 ) were recorded by KBr disks on Fourier Transform Infrared Thermo Electron Nicolet iS10 Spectrometer (Thermo Fisher Scientific Inc., Waltham, MA) at Faculty of Science, Ain Shams University.The 1 H NMR spectra (d, ppm) were run at a Varian GEMINI 300 using tetramethyl silane as an internal standard in deuterated dimethyl sulfoxide (DMSO-d 6 ) at Faculty of Science, Cairo University.Elemental analyses were measured using Perkin-Elmer 2400 CHN elemental analyzer at Faculty of Science, Ain Shams University, and satisfactory analytical data (±0.4) were obtained.The reactions were monitored by thin-layer chromatography (TLC) using Merck Kiesel gel 60 F 254 obtained from Fluka, Switzerland.The key malonyl chloride derivative 1 was previously synthesized by us. 34General procedure for the synthesis of compounds 3 À 6 A solution of malonyl chloride derivative 1 (2 mmol, 0.7 g) and ammonium thiocyanate (5.2 mmol) in dry acetonitrile (10 mL) was stirred at room temperature for 20 min and then 4-acetylaniline, 2-aminobenzoic acid, 6-aminothiouracil, or 5-(4-nitrophenyl)-1,3,4-thiadiazol-2amine (4 mmol) was added.The reaction mixture was heated under reflux for 4 ⁓ 10 h.The precipitated solid while hot was collected and recrystallized from the appropriate solvent to afford compounds 3-6, respectively.

Scheme 3 .
Scheme 3. Plausible pathway for the formation of compound 7.

Figure 3 .
Figure 3. Optimized structure of compound 6 (left), HOMO (middle), and LUMO (right) for most potent insecticidal compounds.Color index: White H, Grey C, Blue N, Red O, and yellow S.

Figure 2 .
Figure 2. Optimized structure of compound 5 (left), HOMO (middle), and LUMO (right) for most potent insecticidal compounds.Color index: White H, Grey C, Blue N, Red O, and yellow S.

Figure 4 .
Figure 4. Optimized structure of compound 15 (left), HOMO (middle), and LUMO (right) for most potent insecticidal compounds.Color index: White H, Grey C, Blue N, Red O, and yellow S.

Table 5 .
Quantum chemical parameters calculated for the studied compounds.