The Synthesis, Antimicrobial Activity, and Molecular Docking of New 1, 2, 4-Triazole, 1, 2, 4-Triazepine, Quinoline, and Pyrimidine Scaffolds Condensed to Naturally Occurring Furochromones

This study aims to synthesize a new series of furochromone derivatives, evaluate their antimicrobial properties, and improve the permeability of potent compounds to inhibit different types of bacteria and fungi. Hence, Substituted furo[3,2-g]chromene-6-carbonitrile (3a,b) readily form 7-amino-5-methyl-furo [3,2-g]chromene-6-carbonitrile (4a,b) via reduction using sodium borohydride in methanol. The same compounds of (4a,b) were used as starting materials for the synthesis of new furochromone derivatives such as furochromeno [2,3-d]pyrimidines, N- (6-cyano- 5-methyl-furochromene) acetamide, N-(6-cyano-5-methyl-furo chromene)-2-phenyl acetamide, N- (6-cyano-5-methyl-furochromene) formimidate, furochromeno[1,2,4]triazepin-5-amine, furochrom ene-6-carboxamide, furochromeno[1,2,4]triazolopyrimidines, and furochromeno[2,3-b]quinolin- 6-amine. The structures of the new compounds were determined using spectroscopy: Nuclear Magnetic Resonance (1H, 13C), Mass spectra, Infrared, and elemental analysis. Molecular docking studies were conducted to investigate the binding patterns of the prepared compounds against DNA-gyrase (PDB 1HNJ). The results displayed that compounds furochromenotriazolopyrimidine (20a,b), furochromenoquinolin-6-amine (21a,b), furochromenotriazepin-amine (9a,b), and furo- chromenopyrimidine-amine (19a,b) were excellent antimicrobials.

The IR spectra of (17a,b) exhibited the absence of any absorption due to amino groups and the presence of a cyano function at ν 2220-2218 cm −1 ; the 1 HNMR spectrum of (17a) showed the presence of triplet and quartet signals at δ 1.25 and 3.66 ppm due to the ethoxy group.
The IR spectra of (17a,b) exhibited the absence of any absorption due to amino groups and the presence of a cyano function at ν 2220-2218 cm −1 ; the 1 HNMR spectrum of (17a) showed the presence of triplet and quartet signals at δ 1.25 and 3.66 ppm due to the ethoxy group.

Structural Activity Relationship (SAR)
The results show that some types of bacteria and fungi are more sensitive to the synthesized compounds; some compounds have better antimicrobial activity, such as furo [3 ,2 :6,7]  Based on previous studies and practical results, the structure activity relationship of the compounds, with results showing good antimicrobial activity have been discussed, and the following can be confirmed: The presence of functional groups linked with furochromones such as methyl, methoxy, amino, imino, hydroxyl, phenyl, thioxo, acetyl, 1,2,4-triazole, 1,2,4-triazepine, pyrimidine, quinoline, and fused rings; furochromenoquinoline, furochromenotriazolo-pyrimidine, furochromenotriazepine, and furochromenopyrimidine moieties; and heteroatoms such as oxygen, nitrogen, and sulfur.

Molecular Modeling
The structure of FabH (PDB 1HNJ) was obtained from the RCSB protein Data Bank [55]. Glide was used to analyze the interactions of the active compounds with the enzyme. All the heteroatoms were removed and isolated from the 1HNJ.pdb, to make complex receptors free of any ligand before docking. The water molecule of the enzyme was removed, and hydrogen atoms were added to the typical geometry before docking. The ligand file was submitted to the Chem3D Ultra Visualizing program to be reduced to the lowest energy and to obtain a standard 3D structure. A grid box was created with active residues of 1HNJ protein, using receptor grid generation in the glide tool of the Schrodinger suite to produce a good docking reaction at the formed binding domain. The engaged free energy of output docked complexes was studied using prime MMGBSA of the Schrodinger suite. The binding free energy demonstrated the consanguinity of H-bond and pi-sigma reactions between target 1HNJ protein and little ligand molecules. Table 3 shows eight docked complexes with an H-bond length below 3.2, suggesting that the docked complexes have steady conformation. The binding free energy of docked complexes was in the range of −38.8 to −49.84, with negative dG values designating the formation of steady complexes. Table 3. Glide score, glide energy, binding energies (MM/GBSA), and interaction of the synthesized molecules with amino acid residues of 1HNJ protein.     The FabH active site generally contains a catalytic triad tunnel involving Cys112, His244, and Asn274. A change in these amino acid resides may inhibit or even stop an enzyme's catalytic activity [57]. The direct outcome of this would be that fatty acid biosynthesis cannot carry on efficiently as the energy equipping the organism would not be enough, so the components of all cell membranes could not be formed, and antimicrobial activity would be revealed [58]. Subsequently, we carried out molecular docking studies of the prepared compounds with the crystal structure of E. coli FabH (entry 1HNJ in the Protein Data Bank) to discover their binding mode. From recent and previous scientific studies [59][60][61][62][63][64][65][66][67] The FabH active site generally contains a catalytic triad tunnel involving Cys112, His244, and Asn274. A change in these amino acid resides may inhibit or even stop an enzyme's catalytic activity [57]. The direct outcome of this would be that fatty acid biosynthesis cannot carry on efficiently as the energy equipping the organism would not be enough, so the components of all cell membranes could not be formed, and antimicrobial activity would be revealed [58]. Subsequently, we carried out molecular docking studies of the prepared compounds with the crystal structure of E. coli FabH (entry 1HNJ in the Protein Data Bank) to discover their binding mode. From recent and previous scientific studies [59][60][61][62][63][64][65][66][67], we know that the hydroxyl (OH) group contributes The FabH active site generally contains a catalytic triad tunnel involving Cys112, His244, and Asn274. A change in these amino acid resides may inhibit or even stop an enzyme's catalytic activity [57]. The direct outcome of this would be that fatty acid biosynthesis cannot carry on efficiently as the energy equipping the organism would not be enough, so the components of all cell membranes could not be formed, and antimicrobial activity would be revealed [58]. Subsequently, we carried out molecular docking studies of the prepared compounds with the crystal structure of E. coli FabH (entry 1HNJ in the Protein Data Bank) to discover their binding mode. From recent and previous scientific studies [59][60][61][62][63][64][65][66][67] The FabH active site generally contains a catalytic triad tunnel involving Cys112, His244, and Asn274. A change in these amino acid resides may inhibit or even stop an enzyme's catalytic activity [57]. The direct outcome of this would be that fatty acid biosynthesis cannot carry on efficiently as the energy equipping the organism would not be enough, so the components of all cell membranes could not be formed, and antimicrobial activity would be revealed [58]. Subsequently, we carried out molecular docking studies of the prepared compounds with the crystal structure of E. coli FabH (entry 1HNJ in the Protein Data Bank) to discover their binding mode. From recent and previous scientific studies [59][60][61][62][63][64][65][66][67] The FabH active site generally contains a catalytic triad tunnel involving Cys112, His244, and Asn274. A change in these amino acid resides may inhibit or even stop an enzyme's catalytic activity [57]. The direct outcome of this would be that fatty acid biosynthesis cannot carry on efficiently as the energy equipping the organism would not be enough, so the components of all cell membranes could not be formed, and antimicrobial activity would be revealed [58]. Subsequently, we carried out molecular docking studies of the prepared compounds with the crystal structure of E. coli FabH (entry 1HNJ in the Protein Data Bank) to discover their binding mode. From recent and previous scientific studies [59][60][61][62][63][64][65][66][67], we know that the hydroxyl (OH) group contributes

Binding Free Energy Calculation
The XP docked output molecules are used to calculate the binding free energy of protein-ligand complexes using prime MMGBSA (molecular mechanics generalized Born surface area) at force domain OPLS-2005 [56]. Free energy of binding describes the affinity of a ligand molecule with a protein. The binding free energy was calculated at binding poses of protein-ligand complexes as follows: where G Binding is the Minimized binding free energy; G complex, G protein, and G ligand represent the free energy of the protein-inhibitor complex, protein, and inhibitor, respectively.

Molecular Docking
The FabH active site generally contains a catalytic triad tunnel involving Cys112, His244, and Asn274. A change in these amino acid resides may inhibit or even stop an enzyme's catalytic activity [57]. The direct outcome of this would be that fatty acid biosynthesis cannot carry on efficiently as the energy equipping the organism would not be enough, so the components of all cell membranes could not be formed, and antimicrobial activity would be revealed [58]. Subsequently, we carried out molecular docking studies of the prepared compounds with the crystal structure of E. coli FabH (entry 1HNJ in the Protein Data Bank) to discover their binding mode. From recent and previous scientific studies [59][60][61][62][63][64][65][66][67], we know that the hydroxyl (OH) group contributes greater affinity in the interaction of the receptor and the ligand as compared to the methoxy (OCH 3 ) group by forming a hydrogen bond with the amino acid of the protein molecule, and the greater extent of hydrogen bonding leads to better interaction. In this study, molecular docking can offer worthwhile information on the action mechanism of our compounds. The docking  Table 3 and Figures 2-4. In silico studies discovered that most of the prepared molecules had a good binding free energy (kcal/mol) for the target protein, ranging from −38.8 to −49.84 kcal/mol Table 3. Moreover, the changes in MM-GBSA accorded well with the MIC values obtained for most of the compounds-specifically, compounds (20b) and (21b), with good activity, exhibited very low MM-GBSA values of −42.40 and −49.84 kcal/mol, respectively. The observations from the biological assay data and the molecular docking results ability suggest that the antibacterial activity of these compounds is derived from the reaction between the compounds and the enzyme FabH.

Materials and Methods
Through cooperation with other researchers, a research plan was made for the synthesis of new heterocyclic compounds. These new compounds were planned to study their antimicrobial activity, and the plan was successfully implemented.

General Information
All the melting points were assessed on an Electrothermal IA 9100 series digital melting point apparatus (Shimadzu, Tokyo, Japan). Elemental analyses were performed on Vario EL (Elementar, Langenselbold, Germany). Microanalytical data were processed at the microanalytical center of the Faculty of Science at Cairo University and National Research Centre. The IR spectra (KBr disc) were recorded using a Perkin-Elmer 1650 spectrometer (Waltham, MA, USA). NMR spectra were determined using JEOL 270 MHz and JEOL JMS-AX 500 MHz (JEOL, Tokyo, Japan) spectrometers with Me4Si as an internal standard. Mass spectra were recorded on an EI Ms-QP 1000 EX instrument (Shimadzu, Tokyo, Japan) at 70 eV. Biological evaluations were performed by the antimicrobial unit of Department of Chemistry of Natural and Microbial Products (National Research Centre, Giza 12622, Egypt). All starting materials and solvents were purchased from Sigma-Aldrich (St. Louis, MO, USA).
Method B. To a stirred solution of visnaginone (2a) (2.06 g, 0.01 mol) or khellinone (2b) (2.36 g, 0.01 mol) in ethanolic sodium ethoxide solution (0.5 g, 0.02-atom of sodium 35 mL of ethanol), malononitrile (0.66 g, 0.01 mol) was added and the mixture was heated under reflux for 2-4 h and checked by TLC. After cooling, the final solid product was collected and recrystallized from the proper solvent to give (3a) and (3b), respectively.
Method B. A stream of NH 3 gas was passed through (17a) (3.14 g, 0.01 mol) or (17b) (3.44 g, 0.01 mol) in a dioxane solution at room temperature for 2-4 h under control (TLC). The mixture was left in the refrigerator overnight, and the solid product that formed upon cooling was collected by filtration to give (5a) and (5b), respectively.
Method B. A mix of (4a) (2.58 g, 0.01 mol) or (4b) (2.88 g, 0.01 mol) and formic acid (10 mL) in formamide (35 mL) was refluxed for 5-8 h. After cooling, the solution was poured into cold water. The solid precipitate that formed was collected by filtration, washed with cold water/ethanol, and recrystallized from the proper solvent to give (6a) and (6b), respectively.
Method B. A solution of (7a) (2.98 g, 0.01 mol) or (7b) (3.28 g, 0.01 mol) in absolute ethanol (25 mL) with pyridine (5 mL) was heated and refluxed on water bath for 6-9 h under control (TLC), after cooling the solid precipitate was collected via filtration, washed with water, dried and recrystallized from appropriate solvent to give (8a) and (8b).

Human and Animal Rights
No human or animal subjects were used in the study. The research was conducted according to ethical standards in vitro.

Chemicals and Drugs
Types of Gram-positive bacteria Staphylococcus aureus and Streptococcus pyogenes, Gram-negative bacteria Escherichia coli and Klebsiella pneumoniae, and fungi Aspergillus niger, Alternaria alternate, Curvularia lunata, and Candida albicans were from the National Research Centre, Department of Chemistry of Natural and Microbial Products, Giza, Egypt, and cefotaxime sodium, nystatin, and DMSO were purchased from Sigma-Aldrich.