Targeting new N-furfurylated 4-chlorophenyl-1,2,4-triazolepropionamide hybrids as potential 15-lipoxygenase inhibitors supported with in vitro and in silico studies.

Lipoxygenases (LOXs) are a group of enzymes that catalyze the oxidation of polyunsaturated fatty acids and initiate the biosynthesis of secondary metabolites that are involved to control inflammation. In search of new and more potent LOX inhibitors, a series of new 3-(5-(4-chlorophenyl)-4-(2-furylmethyl)-1,2,4-triazole hybrids was prepared and screened for its LOX inhibitory potential. 4-Chlorobenzoic acid (a) was metamorphosed into N-furfuryl-5-(4-chlorophenyl)-4-(2-furylmethyl)-1,2,4-triazole (4) via intermediates like benzoate (1), hydrazide (2) and semicarbazide (3). Finally, triazole (4) was fused with propionamides (6a-o) and transformed it into the aimed derivatives (7a-o). The structural interpretations of the prepared derivatives (7a-o) were accomplished via FTIR, 1H-, 13C-NMR spectroscopy, EI-MS and HR-EI-MS spectrometry. The inhibitory potency of the compounds against soybean 15-LOX was determined by in vitro assay using chemiluminescence method. Compounds 7a and 7f exhibited potent LOX inhibitory profiles with IC50 21.83 ± 0.56 and 25.72 ± 0.51 µM, whereas 7d and 7e showed comparable inhibitory potential with IC50 values of 34.52 ± 0.39 and 39.12 ± 0.46 µM, respectively. Compounds 7a, 7f, 7d and 7e exhibited 65.58 ± 1.4%, 54.72 ± 1.3%, 58.52 ± 1.2% and 63.56 ± 1.4% blood mononuclear cells viability, respectively. Density functional theory and molecular docking studies further strengthened the studies of the synthesized compounds and these derivatives perceived to be potential 'lead' compounds in drug discovery as anti-LOX.Communicated by Ramaswamy H. Sarma.

LC-ESI-MS analysis, antioxidant, anti-diabetic and molecular docking studies on Corchorus depressus (L.) C.Chr.

The present study encompasses the ethnomedicinal consumption of Corchorus depressus (L.) C.Chr. (C. depressus) for diabetes. Samples were subjected to LC-ESI-MS analyses. The n-hexane, methanolic and water extracts were screened for α-glucosidase inhibition and in vivo anti-diabetic studies. Further, antioxidant (DPPH) and anti-inflammatory study was performed via luminol-enhanced chemi-luminescence assay. The identified compounds were docked against the target enzymes of diabetes. The n-hexane fraction (CD-J1) showed IC50 of 8.4 ± 0.1 µg/mL against α-glucosidase enzyme. The sub fractions CD-12 and CD-13 of CD-J1 obtained after flash column chromatography displayed further reduced IC50 values of 4.3 ± 0.1 and 6.3 ± 0.1, respectively, as compared with standard drug acarbose (IC50 values of 37.5 ± 0.2 µg/mL). Simultaneously, dereplication of most active sub-fraction CD-12 by LC-ESI-MS led to the identification of strophanthidin and some other active metabolites responsible for anti-diabetic activity. Molecular docking of strophanthidin with α-glucosidase and α-amylase revealed high affinity for these target enzymes.

4-Chlorophenyl-N-furfuryl-1,2,4-triazole Methylacetamides as Significant 15-Lipoxygenase Inhibitors: an Efficient Approach for Finding Lead Anti-inflammatory Compounds.

Lipoxygenases (LOXs) are a class of enzymes that catalyze the production of pro-inflammatory mediators, such as leukotrienes and lipoxins, via an arachidonic acid cascade as soon as they are released from the membrane phospholipids after tissue injury. In continuation of our efforts in search for new LOX inhibitors, a series of chlorophenyl-furfuryl-based 1,2,4-triazole derivatives were prepared and evaluated for their 15-LOX inhibitory activities. A simple precursor, 4-chlorobenzoic acid (a), was consecutively transformed into benzoate (1), hydrazide (2), semicarbazide (3), and N-furfuryl 5-(4-chlorobenzyl)-4H-1,2,4-triazole (4), which when further merged with electrophiles (6a-o) resulted in end products (7a-o). The structural elucidations of the newly synthesized compounds (7a-o) were carried out by Fourier transform infrared, 1H-, 13C NMR spectroscopy, EI-MS, and HR-EI-MS spectrometry. The inhibitive capability of compounds (7a-o) on soybean 15-LOX was performed in vitro using the chemiluminescence method. The compounds 7k, 7o, 7m, 7b, and 7i demonstrated potent activities (IC50 17.43 ± 0.38, 19.35 ± 0.71, 23.59 ± 0.68, 26.35 ± 0.62, and 27.53 ± 0.82 µM, respectively). These compounds revealed 79.5 to 98.8% cellular viability as measured by the MTT assay at 0.25 mM concentration. The structure-activity relationship (SAR) studies showed that the positions and the nature of substituents bonded to the phenyl ring are important in the determination of 15-LOX inhibitory activities. ADME, in silico, and density functional theory studies supported the evidence as yet another class of triazoles with potential lead properties in search for anti-LOX compounds with a safe gastrointestinal safety profile for various inflammatory diseases. Further work is in progress on the synthesis of more derivatives in search for anti-inflammatory agents.

Exploring ibuprofen derivatives as α-glucosidase and lipoxygenase inhibitors: Cytotoxicity and in silico studies.

This study reports the synthesis of a series of ibuprofen derivatives, including thiosemicarbazides 4a-f, 1,3,4-oxadiazoles 5a-f, 1,3,4-thiadiazoles 6a-f, 1,2,4-triazoles 7a-f, and their S-alkylated derivatives 8a-d. All of the newly synthesized derivatives were analyzed using 1 H NMR, 13 C NMR spectroscopy, and high-resolution mass spectra (electron ionization) spectrometry. These synthetic molecules were examined for their in vitro baking yeast α-glucosidase and soybean 15-lipoxygenase (15-LOX) inhibition and cell viability studies. The results revealed that the compounds N-(3,4-dichlorophenyl)-5-[1-(4-isobutylphenyl)ethyl]-1,3,4-oxadiazol-2-amine 5f (IC50 3.05 ± 1.23 µM) and N-(3-fluorophenyl)-5-[1-(4-isobutylphenyl)ethyl]-1,3,4-oxadiazol-2-amine 5b (IC50 3.12 ± 1.21 µM) were the most potent with respect to the α-glucosidase enzyme while in case of 15-LOX, the compound 4-(2,4-dichlorophenyl)-1-[2-(4-isobutylphenyl)propanoyl]thiosemicarbazide 4e showed potent inhibition with an IC50 value of 55.41 ± 0.41 µM. All these compounds were found least toxic by displaying a blood mononuclear cell viability value of 69.2%-97.8% by the MTT assay compared to the standards when assayed at 0.25 mM concentration. Molecular docking analyses were conducted to evaluate the inhibition profiles of these derivatives against the said enzymes and the data supported the in vitro profiles.

Identification of phenylcarbamoylazinane-1,3,4-oxadiazole amides as lipoxygenase inhibitors with expression analysis and in silico studies.

In search for new anti-inflammatory agents that inhibit the enzymes of arachidonic acid pathway as the drug targets, the present article describes the screening of 1,3,4-oxadiazole analogues against lipoxygenase (LOX) enzyme. The work is based on the synthesis of new N-alkyl/aralky/aryl derivatives (6a-o) of 2-(4-phenyl-5-(1-phenylcarbamoylpiperidine)-4H-1,3,4-oxadiazol-3-ylthio)acetamide which were obtained by the reaction of 1,3,4-oxadiazole (3) with various electrophiles (5a-o), in KOH. The synthesized analogues showed potent to moderate inhibitory activity against the soybean 15-LOX enzyme; especially 6g, 6b, 6a and 6l displayed the potent inhibitory potential with IC50 values 7.15 ± 0.26, 9.32 ± 0.42, 15.83 ± 0.45 & 18.37 ± 0.53 µM, respectively, while excellent to moderate inhibitory profiles with IC50 values in the range of 26.13-98.21 µM were observed from the compounds 6k, 6m, 6j, 6o, 6h, 6f, 6n and 6c. Most of the active compounds exhibited considerable cell viability against blood mononuclear cells (MNCs) at 0.25 mM by MTT assay except 6f, 6h, 6k and 6m which showed around 50% cell viability. Flow cytometry studies of the selected compounds 6a, 6j and 6n revealed that these caused 79.5-88.51% early apoptotic changes in MNCs compared with 4.26% for control quercetin at their respective IC50 values. The relative expression of 5-LOX gene was monitored in MNCs after treatment with these three molecules and all down-regulated the enzyme activity. In silico ADME and molecular docking studies further supported these studies of oxadiazole derivatives and considered it as potential 'lead' compounds in drug discovery and development.

Exploring phenylcarbamoylazinane-1,2,4-triazole thioethers as lipoxygenase inhibitors supported with in vitro, in silico and cytotoxic studies.

Searching small molecules as an auspicious approach to develop new anti-inflammatory drugs is a challenge for the researchers especially by modifying active pharmacophoric groups in the targeted molecules. In the current work, a series of new S-alkyl/aralky derivatives (8a-h; 9a-h) of 2-(4-ethyl/phenyl-5-(1-phenylcarbamoylpiperidine)-4H-1,2,4-triazol-3-ylthio)ether were synthesized and assessed for their inhibitory action against the 15-lipoxygenase from soybean (15-sLOX). The basic precursor ethyl piperidine-4-carboxylate (a) was consecutively changed into phenylcarbamoyl derivative (1), hydrazide (2), semicarbazides (3/4) and N-ethyl/phenyl-5-(1-phenylcarbamoylpiperidine)-1,2,4-triazoles (5/6), which further in association with electrophiles (7a-h) promoted to the final products (8a-h; 9a-h). The synthesized derivatives were characterized by FT-IR, 1H-, 13C NMR spectroscopy, EI-MS, and HR-EI-MS spectrometry. Amongst these, 8a, 8c, and 9c, expressed potent inhibitory profiles against the 15-sLOX enzyme with IC50 values of 12.52 ± 0.35 to 35.64 ± 0.29 µM, followed by the compounds 9b, 9g, 9d, 9a, 8b, 8e, 8d, 8g, 8h, 8f and 9h with IC50 values in the range of 43.78 ± 0.43 to 108.65 ± 0.38 µM. All compounds exhibited variable cellular viability levels by MTT assay. Flow cytometric data demonstrated that 8f, 8g, 8h have maximal lymphocyte cellular viability and all compounds affected cells in the late apoptosis phase. In silico ADMET studies supported the drug-likeness of most of the molecules. These studies were supported by molecular docking against 15-sLOX, human 5-LOX (5-hLOX) and human 15-LOX (5-hLOX); that inhibitors of 15-sLOX docked-in the active pocket of either 5-hLOX or 15-hLOX and docking score remained constant for all three enzymes within a narrow range (-6.8 to -9.7) as did it for standard quercetin (-8.4 to -9.0). The most dominant bonding interactions were π-π, π-anion, and π-alkyl type along with the hydrogen bonding. The data collected altogether demonstrates the better possibility of some of these compounds as good LOX inhibitors in search for 'lead' as anti-inflammatory agents in the process of drug discovery and development.

Identification of NSAIDs as lipoxygenase inhibitors through highly sensitive chemiluminescence method, expression analysis in mononuclear cells and computational studies.

Here we report the inhibitory effects of nine non-steroidal anti-inflammatory drugs (NSAIDs) on soybean 15-lipoxygenase (15-LOX) enzyme (EC 1.13.11.12) by three different methods; UV-absorbance, colorimetric and chemiluminescence methods. Only two drugs, Ibuprofen and Ketoprofen, exhibited enzyme inhibition by UV-absorbance method but none of the drug showed inhibition through colorimetric method. Chemiluminescence method was found highly sensitive for the identification of 15-LOX inhibitors and it was more sensitive and several fold faster than the other methods. All tested drugs showed 15-LOX-inhibition with IC50 values ranging from 3.52 ± 0.08 to 62.6 ± 2.15 µM by chemiluminescence method. Naproxen was the most active inhibitor (IC50 3.52 ± 0.08 µM) followed by Aspirin (IC50 4.62 ± 0.11 µM) and Acetaminophen (IC50 6.52 ± 0.14 µM). Ketoprofen, Diclofenac and Mefenamic acid showed moderate inhibitory profiles (IC50 24.8 ± 0.24 to 39.62 ± 0.27 µM). Piroxicam and Tenoxicam were the least active inhibitors with IC50 values of 62.6 ± 2.15 µM and 49.5 ± 1.13 µM, respectively. These findings are supported by expression analysis, molecular docking studies and density functional theory calculations. The expression analysis and flow cytometry apoptosis analysis were carried out using mononuclear cells (MNCs) which express both human 15-LOX and 5-LOX. Selected NSAIDs did not affect the cytotoxic activity of MNCs at IC50 concentrations and the cell death showed dose dependent effect. However, MNCs apoptosis increased only at the higher concentrations, demonstrating that these drugs may not induce loss of immunity in septic and other inflammatory conditions at the acceptable inhibitory concentrations. The data collectively suggests that NSAIDs not only inhibit COX enzymes as reported in the literature but soybean 15-LOX and MNCs LOXs are also inhibited at differential values. A comparison of the metabolomics studies of arachidonic acid pathway after inhibition of either COX or LOX enzymes may reconfirm these findings.

Synthesis of imidazole-pyrazole conjugates bearing aryl spacer and exploring their enzyme inhibition potentials.

Developing improved enzyme inhibitors is an effective therapy to counter various diseases. Aiming to build up biologically active templates, a new series of bis-diazoles conjugated with an aryl linker was designed and prepared through a convenient synthetic approach. Synthesized derivatives 6(a-m), having different substitutions at the 2nd position of the imidazole nucleus, depict the scope of present study. These compounds were characterized through spectroscopic methods and further examined for their in vitro enzyme inhibitory potentials against two selected enzymes: α-glucosidase and lipoxygenase (LOX). Overall, this series was found to be effective against α-glucosidase and moderately active against LOX enzyme. Compound 6k was the most potent α-glucosidase inhibitor with IC50 = 54.25 ± 0.67 µM as compared to reference drug acarbose (IC50 = 375.82 ± 1.76 µM). The docked conformation revealed the involvement of substituent's heteroatoms with amino acid residue Gly280 through hydrogen bonding. The most active LOX inhibitor was 6a with IC50 = 41.75 ± 0.04 µM as compared to standard baicalein (IC50 = 22.4 ± 1.3 µM). Docking model of 6a suggested the strong interaction of imidazole's nitrogen with iron atom of the active pocket of enzyme. Other features like lipophilicity, bulkiness of compounds, pi-pi interactions and/or pi-alkyl interactions also affected the inhibiting potentials of all prepared scaffolds.

Probing phenylcarbamoylazinane-1,2,4-triazole amides derivatives as lipoxygenase inhibitors along with cytotoxic, ADME and molecular docking studies.

Hunting small molecules as anti-inflammatory agents/drugs is an expanding and successful approach to treat several inflammatory diseases such as cancer, asthma, arthritis, and psoriasis. Besides other methods, inflammatory diseases can be treated by lipoxygenase inhibitors, which have a profound influence on the development and progression of inflammation. In the present study, a series of new N-alkyl/aralky/aryl derivatives (7a-o) of 2-(4-phenyl-5-(1-phenylcarbamoyl)piperidine-4H-1,2,4-triazol-3-ylthio)acetamide was synthesized and screened for their inhibitory potential against the enzyme 15-lipoxygenase. The simple precursor ethyl piperidine-4-carboxylate (a) was successively converted into phenylcarbamoyl derivative (1), hydrazide (2), semicarbazide (3) and N-phenylated 5-(1-phenylcarbamoyl)piperidine-1,2,4-triazole (4), then in combination with electrophiles (6a-o) through further multistep synthesis, final products (7a-o) were generated. All the synthesized compounds were characterized by FTIR, 1H, 13C NMR spectroscopy, EIMS, and HREIMS spectrometry. Almost all the synthesized compounds showed excellent inhibitory potential against the tested enzyme. Compounds 7c, 7f, 7d, and 7g displayed potent inhibitory potential (IC50 9.25 ± 0.26 to 21.82 ± 0.35 µM), followed by the compounds 7n, 7h, 7e, 7a, 7b, 7l, and 7o with IC50 values in the range of 24.56 ± 0.45 to 46.91 ± 0.57 µM. Compounds 7c, 7f, 7d exhibited 71.5 to 83.5% cellular viability by MTT assay compared with standard curcumin (76.9%) when assayed at 0.125 mM concentration. In silico ADME studies supported the drug-likeness of most of the molecules. In vitro inhibition studies were substantiated by molecular docking wherein the phenyl group attached to the triazole ring was making a π-δ interaction with Leu607. This work reveals the possibility of a synthetic approach of compounds in relation to lipoxygenase inhibition as potential lead compounds in drug discovery.

Conventional versus microwave assisted synthesis, molecular docking and enzyme inhibitory activities of new 3,4,5-trisubstituted-1,2,4-triazole analogues.

N-(Substituted)-5-(1-(4-methoxyphenylsulfonyl)piperidin-4-yl)-4H-1,2,4-triazol-3-ylthio) acetamide were synthesized by following conventional as well as microwave assisted protocol through five consecutive steps under the impact of various reaction conditions to control the reaction time and the yield of product. Starting from 4-methoxybenzenesulfonyl chloride and ethyl isonipecotate, product 3 was obtained which was converted into product 4 by treating with hydrazine hydrate. In step 3, the product 4 was refluxed with methyl isothiocyanate and KOH to yield compound 5 which was finally treated with variety of N-substituted acetamides to yield an array of different new compounds (8a-k). These synthesized compounds were evaluated for their inhibition potential against bovine carbonic anhydrase (bCA-II), acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes. Compound 8g demonstrated good activity against bCA-II, AChE and BChE with IC50 values of 8.69 ± 0.38 µM, 11.87±0.19 µM and 26.01±0.55 µM respectively. SAR studies assisted with molecular docking were carried out to explore the mode of binding of the compounds against the studied enzymes.