Skip to main content
NCBI home page
ACS Omega logoLink to ACS Omega
. 2023 Aug 23;8(35):31890–31898. doi: 10.1021/acsomega.3c03546

Triazolothiadiazoles and Triazolothiadiazines as New and Potent Urease Inhibitors: Insights from In Vitro Assay, Kinetics Data, and In Silico Assessment

Jalal Uddin †, Saeed Ullah ‡, Sobia Ahsan Halim ‡, Muhammad Waqas ‡, Aliya Ibrar §,*, Imtiaz Khan ∥, Abdullatif Bin Muhsinah †, Ajmal Khan ‡,*, Ahmed Al-Harrasi ‡,*
PMCID: PMC10483676  PMID: 37692208

Abstract

graphic file with name ao3c03546_0007.jpg

Hyperactivity of the urease enzyme induces the pathogenesis of peptic ulcers and gastritis. The identification of new urease inhibitors can reduce the activity of urease. Therefore, in the current study, we have evaluated 28 analogues of triazolothiadiazole and triazolothiadiazine heteroaromatics for their in vitro urease inhibitory efficacy. All the tested compounds displayed a remarkable inhibitory potential ranging from 3.33 to 46.83 μM. Among them, compounds 5k and 5e emerged as lead inhibitors with IC50 values of 3.33 ± 0.11 and 3.51 ± 0.49 μM, respectively. The potent inhibitory potential of these compounds was ∼6.5-fold higher than that of the marketed drug thiourea (IC50 = 22.45 ± 0.30 μM). The mechanistic insights from kinetics experiments of the highest potent inhibitors (4g, 5e, and 5k) revealed a competitive type of inhibition with ki values 2.25 ± 0.0028, 3.11 ± 0.0031, and 3.62 ± 0.0034 μM, respectively. In silico modeling was performed to investigate the binding interactions of potent inhibitors with the enzyme active site residues, which strongly supported our experimental results. Furthermore, ADME analysis also showed good druglikeness properties demonstrating the potential of these compounds to be developed as lead antiurease agents.

1. Introduction

Urease is a popular enzyme in living organisms, responsible for the hydrolysis of urea into carbon dioxide and ammonia.1,2 The high concentrations of ammonia are caused by urease hyperactivity, which leads to a rise in the pH of the stomach, thus resulting in complications like peptic and gastric ulcers,3 hepatic coma,4 pyelonephritis, and kidney stones.5,6 This condition of clinical complications demands such inhibitors that can regulate urease activity.2,6−8 Various studies have revealed the diverse variety of compounds such as Schiff base hydrazones, triazole-(thio) barbituric acids, N-thioacylated ciprofloxacin derivatives, and barbituric-hydrazine-phenoxy-1,2,3-triazole-acetamides which possess urease inhibitory effects.9−11,12,13 Urease inhibitors have revealed amplification of the urea N uptake in plants14 and reduction of environmental issues.6,15−17 Additionally, they play a role as strong antiulcer drugs.18,19

Nitrogen-containing heterocyclic compounds are the most common and important scaffolds for a wide range of synthetic medications, bioactive natural products, pharmaceuticals, and agricultural chemicals. These skeletons have gained attention due to their wide range of uses, and significant attempts have been made to develop synthetic techniques that could lead to the discovery of new bioactive molecules in medicinal chemistry.20 Conjugated heterocycles like triazolothiadiazoles and triazolothiadiazines have shown several biologically important properties, including antimicrobial,21,22 antitumor,21 antiviral,23 anticancer,24 central nervous system (CNS) depressant,25 anti-inflammatory, analgesic,26 antioxidant,27 anti-HIV,28 and antitubercular29 activities.

Several triazolothiadiazoles possess antiurease efficacy, phosphodiesterase, and c-Met protein inhibitory potential.30 The heteroaromatic (pyridine)-substituted triazolothiadiazole hybrids have displayed potent cytotoxic effects, anticholinesterase, antialkaline phosphatase, and antileishmanial activities.20,31,32 The excellent results of our previous findings on triazolothiadiazole and triazolothiadiazine derivatives33 and their inhibitory potential of cholinesterase and monoamine oxidases led us to explore their antiulcer potential by specifically targeting urease enzyme. This study initially scrutinized compounds 4a–l and 5a–p for urease inhibition, followed by a kinetic investigation to determine the probable mode of action. Later, their comprehensive structure–activity relationships were explained with molecular docking and pharmacokinetic analysis revealed the druglikeness properties.

2. Results and Discussion

2.1. Chemistry

The target compounds, triazolothiadiazoles (4a–l) and triazolothiadiazines (5a–p), were synthesized using the reaction sequences illustrated in Scheme 1.

Scheme 1. Designed Scheme for the Synthesis of Triazolothiadiazoles (4a–l) and Triazolothiadiazines (5a–p).

Scheme 1

Open in a new tab

Both 4-amino-1,2,4-triazole-3-thiols (3a,b) were accessed via a one-pot reaction of the corresponding benzoic acid (1a,b) with thiocarbohydrazide (2) at 190–200 °C. Compound 3 was a crucial intermediate for the formation of target structures.33 The thiol tautomeric form (3a,b) was found to be dominant over the thione form (3a,b*) in our investigation, which was characterized through FTIR and 1H NMR spectroscopic data. The absorption peaks in the FTIR spectra of compounds 3a and 3b indicated the presence of the −SH group at 2554 and 2563 cm–1, respectively, and the absence of the C=S group at 1300–1200 cm–1. Thiol isomer formation was also demonstrated by the disappearance of absorption bands for secondary N–H group in the specified range of 3350–3300 cm–1 and by the appearance of the primary NH2 group in the range of 3498–3495 cm–1. The formation of thiol tautomers in our work is clearly distinctive from the thione tautomer which is previously reported in the literature having N–H stretching near 3200 cm–1 and distinctive bands for thione (C=S) in the range of 1285–1250 cm–1 while lacking thiol (SH) absorption in the range of 2650–2500 cm–1.29,31−33,34,35

Similarly, 1H NMR spectra of both compounds (3a,b) showed resonances for the −NH2 protons as singlets at 5.77 and 5.80 ppm (integrating two protons) and the −SH proton as singlets at 13.83 and 14.03 ppm. The 1H NMR data of thione form (reported previously) indicate the presence of NH protons of triazole rings at 11.30–8.87 ppm and 13C NMR spectra showed the corresponding (C=S) resonance at ca. 170 ppm.34 These literature values for the thione tautomer are completely different from our thiol tautomer in the 1,2,4-triazole ring, giving confidence in the correct assignment of our proposed structure.36,37 Based on the comparison drawn using FTIR and NMR data, the preference for the thiol form is clearly determined for our compounds (3a,b) refuting the possible existence of the thione tautomer in the current study.

The reaction between aryloxy acids and 4-amino-1,2,4-triazole-3-thiol (3a,b) using phosphorus oxychloride produced 1,2,4-triazolo[3,4-b][1,3,4]thiadiazoles (4a–l).33 The aryloxy acids coupled with intermediate 3a,b to produce the corresponding conjugated products in high yield. The FTIR spectra of compounds 4a–l showed several absorption bands at 3064–3009, 2957–2803, 1612–1599, and 1578–1502 cm–1 for C–H aromatic, C–H aliphatic, C=N, and C=C vibration bands, respectively. These values were in close agreement with those reported in the literature such as 2991–2851, 1600–1541, and 1480–1471 cm–1.37 New peaks in the 1H NMR spectra were observed at 5.80 and 5.53 ppm (two proton integration) attributed to methylene protons and missing signals indicative of -SH and -NH2 protons. Electron-rich (OH, OMe, Me) and electron-deficient (F) substituents on the aryloxy ring were well tolerated.

Additionally, 4-amino-1,2,4-triazole-3-thiol (3a,b) was successfully coupled with substituted phenacyl bromides to give 1,2,4-triazolo[3,4-b][1,3,4]thiadiazines (5a–p).33 The FTIR spectra of compounds 5a–p showed several absorption bands at 3066–3000, 2975–2832, 1630–1591, and 1587–1494 cm–1 for C–H aromatic, C–H aliphatic, C=N, and C=C vibration bands, respectively. These values were in close agreement with those reported in the literature such as 3072–3030, 2985–2904, 1608–1537, and 1473–1448 cm–1.371H NMR spectra lacked the signals associated with the −SH and −NH2 protons, whereas a new singlet associated with the methylene protons was observed around 4.58–4.40 ppm (integration for two protons). The literature chemical shifts for this type of methylene were observed in the range of 4.37–3.95 ppm.37 The phenacyl bromide coupling partners were endowed with both electron-rich (Me, OMe) and electron-deficient (F, Cl) substituents showing functional group tolerance. Moreover, the bulky naphthyl and biphenyl ketones also proved successful coupling partners in the cyclocondensation reaction delivering triazolothiadiazines in good yields.

2.2. Biology

2.2.1. In Vitro Urease Inhibition

As the triazole heterocycle has already been reported for its antiurease activity, therefore, in the current investigation, 28 derivatives of triazolothiadiazole and triazolothiadiazine heterocycles proceeded with their medicinal use against stomach and peptic ulcers. Interestingly, all of them offered potent to significant ability to inhibit urease showing a potency range of 3.33–46 μM (Table 1) compared to the standard urease inhibitor, thiourea (IC50 = 22.36 ± 0.30 μM).

Table 1. Urease Inhibitory Activity of Triazolothiadiazoles (4a, l) and Triazolothiadiazines (5a–p).
compound urease inhibition
IC50 ± SEM (μM)
4a 32.91 ± 0.58
4b 25.24 ± 0.56
4c 11.95 ± 0.25
4d 38.99 ± 0.53
4e 8.14 ± 0.20
4f 6.65 ± 0.13
4g 4.50 ± 0.13
4h 6.79 ± 0.24
4i 9.63 ± 0.32
4j 4.67 ± 0.15
4k 28.44 ± 0.62
4l 46.83 ± 0.63
5a 5.91 ± 0.13
5b 8.37 ± 0.16
5c 6.93 ± 0.14
5d 5.16 ± 0.14
5e 3.51 ± 0.49
5f 10.51 ± 0.26
5g 6.52 ± 0.49
5h 5.30 ± 0.17
5i 9.55 ± 0.41
5j 11.48 ± 0.38
5k 3.33 ± 0.11
5l 5.56 ± 0.12
5m 5.79 ± 0.16
5n 7.20 ± 0.18
5o 7.72 ± 0.31
5p 12.95 ± 0.37
thiourea 22.36 ± 0.30
Open in a new tab

2.2.2. Structure–Activity Relationship Analyses

Triazolothiadiazoles and triazolothiadiazines were categorized into four subgroups according to variations in their R1 and R2 attached groups. Figure 1 presents the antiurease ability of compounds 4a–e, which comprise a similar R1 (OCH3) group and different R2-substituents. For instance, compound 4a with the 2-hydroxyl group at R2 exhibited significant antiurease potential (IC50 = 32.91 ± 0.58 μM). Changing the substituent from the 2 to 4-position slightly improved the urease inhibitory activity (4b; IC50 = 25.24 ± 0.56 μM). In compound 4c, the replacement of 2-OH with a methyl substituent surprisingly increased the antiulcer potential and displayed potent suppressive activity against urease (IC50 = 11.95 ± 0.25 μM) compared to compounds 4a and 4b. On the other hand, variation in the position of the methyl group (4-CH3) in compound 4d deteriorated the antiurease ability (IC50 = 38.99 ± 0.53 μM). Unlike 4d, compound 4e with a similar substituent position, however, with a different substituent (OCH3) showed an increase in the urease inhibitory activity (IC50 = 8.14 ± 0.20 μM). Similarly, compound 4f followed a similar pattern, and the 4-F substitution in 4f, further increased its antiurease activity (IC50 = 6.65 ± 0.13 μM) compared to compounds 4a–e.

Figure 1.

Figure 1

Open in a new tab

Structure–activity relationship of compounds 4a–f.

Compounds 4g–l contain a similar R1 substituent (Br) and different R2 substitutions (Figure 2). Variations in R2 showed a slight to high level of difference in antiurease activity. Compounds 4g and 4h with 2-OH and 4-OH displayed potent antiurease potential with IC50 values of 4.50 ± 0.13 and 6.79 ± 0.24 μM, respectively. In contrast, methyl group substitution at the 2-position in compound 4i (IC50 = 9.63 ± 0.32 μM) slightly decreased the activity compared to 4g. However, adding a methyl group at the 4-position in compound 4j (IC50 = 4.67 ± 0.15 μM) enhanced its activity over 4h. In contrast, substituting 4-OCH3 in compound 4k dropped the antiulcer activity of 4k (IC50 = 28.44 ± 0.62 μM) compared to compounds 4a–j. A further decline in the antiurease potential was observed when a 4-F substitution in compound 4l was made, making this molecule the least active antiulcer agent among 4a–l.

Figure 2.

Figure 2

Open in a new tab

Structure–activity relationship of compounds 4g–l.

Figure 3 presents a detailed structure–activity relationship of compounds 5a–h. These compounds have a similar R1 substituent (OMe) and a diverse R2 group. Compounds 5a and 5b with the addition of a 4-methoxyphenyl and 4-chlorophenyl as R2 position exhibited potent antiulcer activity with IC50 values of 5.91 ± 0.13 and 8.37 ± 0.16 μM respectively. Similar potent inhibitory activity against urease was observed for compounds 5c (R2 = 4-F-Ph, IC50 = 6.93 ± 0.14 μM) and 5d (R2 = 4-Me-Ph, IC50 = 5.16 ± 0.14 μM). Interestingly, adding a biphenyl moiety in 5e further increased the antiulcer activity (IC50 = 3.51 ± 0.49 μM). In contrast, the substitution of a naphthyl ring in 5f displayed an adverse effect and decreased the antiurease efficacy (IC50 = 10.51 ± 0.26 μM). However, in 5g and 5h, the addition of 4-nitrophenyl and 3,4-dichloro-phenyl as R2, exhibited almost similar potent suppressive effects against urease with IC50 values of 6.52 ± 0.19 and 5.30 ± 0.17 μM, respectively.

Figure 3.

Figure 3

Open in a new tab

Structure–activity relationship of compounds 5a–h.

The effect of similar R1 with varied R2 substituents was also evaluated to investigate the antiurease activity in compounds 5i–p (Figure 4), which contain Br as R1 and different R2 groups. Compound 5i with a 4-methoxyphenyl at R2 showed good inhibitory potential against urease (IC50 = 9.55 ± 0.41 μM) while compounds 5j (with 5-Cl-Ph, IC50 = 11.48 ± 0.38 μM) and 5p (R2 = 2,3-di-Cl2–Ph, IC50 = 12.95 ± 0.37 μM) displayed a decrease in antiulcer activity than 5i, reflecting the negative effect of the chlorine substituent. The substitution of 4-fluorophenyl in compound 5k demonstrated potent antiurease activity (IC50 = 3.33 ± 0.11 μM) and made it the most potent antiulcer agent in the series. Likewise, the addition of 4-methylphenyl, biphenyl, naphthyl, and 4-nitrophenyl groups in compounds 5l (IC50 = 5.56 ± 0.12 μM), 5m (IC50 = 5.79 ± 0.16 μM), 5n (IC50 = 7.20 ± 0.18 μM), and 5o (IC50 = 7.72 ± 0.31 μM), respectively, has produced good effects on the antiulcer potential of these compounds. Collectively, various structure–activity relationships revealed that both R1 and R2 groups are responsible for the significant antiulcer activity; however, the R2 group plays a key role in modulating the antiurease potential in combination with a specified R1 group. Based on the inhibition profile, these molecules might be developed as drug candidates for treating stomach and peptic ulcers.

Figure 4.

Figure 4

Open in a new tab

Structure–activity relationship of compounds 5i–p.

2.3. Kinetics Studies

Compounds with the highest urease inhibitory efficacy (4g, 5e, and 5k) were selected for their in vitro mechanistic studies to evaluate the mechanism of action. Kinetics studies of these compounds exhibited a competitive type of inhibition with Ki values of 2.25 ± 0.0028, 3.11 ± 0.0031, and 3.62 ± 0.0034 μM, respectively. In such inhibition, Vmax remains constant, while Km always increases (Table 2, Figure S1).

Table 2. Results of Kinetic Studies of Compounds 4g, 5e, and 5k against Urease.

compound IC50 ± SEM (μM) Ki ± SEM (μM) type of inhibition
4g 4.50 ± 0.13 2.25 ± 0.0028 competitive
5e 3.51 ± 0.49 3.11 ± 0.0031 competitive
5k 3.33 ± 0.11 3.62 ± 0.0034 competitive
Open in a new tab

2.4. Molecular Docking

The crystal structure of the Jack bean (Canavalia ensiformis) urease enzyme having a phosphate ion and covalently bonded with two nickel (Ni2+) ions was retrieved from the protein data bank server (PDB ID: 4GY7). HIS492 forms a hydrogen bond in the complex with the phosphate ion, while LYS490 and ASP633 form ionic interactions with the phosphate ion. Similarly, two Ni2+ ions form metal interactions with LYS490 and ASP633 (Figure 5). The detailed interactions of phosphate and Ni2+ ions are listed in Table 3.

Figure 5.

Figure 5

Open in a new tab

(A) Cartoon representation of the urease enzyme (PDB-ID 4GY7) having a phosphate ion in the active pocket interacting with the two Ni2+ ions. (B) Interaction of phosphate and Ni2+ ions with the active pocket residues. (C) Interactive residues of the urease active pocket mediate interactions with the selected compounds. (D) Urease active site shows the compounds’ binding mode (presented in the green stick model). H-bonds are presented in black dotted lines.

Table 3. Detailed Interaction of Jack Bean (C. ensiformis) Urease Active Site Residues with Phosphate and Two Ni2+ Ions (PDB-ID: 4GY7).

PDB code ligand atoms receptor atoms residues bond type distance (Å)
4GY7 O1 NE2 HIS492 HBA 2.93
O1 NE2 HIS492 HBA 2.93
O1 NE2 HIS492 HBA 2.93
O3 OQ2 LYS490 Ionic 3.03
O3 OQ1 LYS490 Ionic 3.01
O3 OD2 ASP633 Ionic 2.94
O3 OD1 ASP633 Ionic 2.63
NI901 OQ1 LYS490 Metal 2.05
NI902 OQ2 LYS490 Metal 2.03
NI902 OD2 ASP633 Metal 2.05
Open in a new tab

All of the compounds displayed a high efficiency in inhibiting the urease enzyme. Consequently, each of the compounds underwent a process of docking at the binding site of the urease. The docking results revealed that these molecules acquired nearly the same position with only slight variation in their conformation. The molecules were inserted deeply into the active site core, enabling interaction with any active site and metal binding residues (HIS492, LYS490, ASP633). The substituted groups were oriented toward the entrance of the active site, where the R group has the opportunity to rotate toward the active site and metal-interacting residues (HIS409, LYS490, HIS492, HIS519, HIS545, HIS593, and ASP633). Compound 5k, the most effective inhibitor, formed a strong hydrogen bond (H-bond) with ARG439 (2×) and ARG609 of urease. In addition, two critical residues in the active site (GLU493, HIS593) formed H-bonds and π–π interactions with compound 5e. Compound 4g, ranked third in terms of effectiveness, is positioned in such a way that it interacts with the side chains of GLY638 and ARG439 through H-bonds. On the other hand, compound 4j formed an H-bond with GLU493 and ARG609 side chains, and HIS593 formed π–π interaction with the R1 moiety of 4j.

In the docking simulation, several compounds were observed to interact with the active pocket of the urease enzyme in various ways. Compound 5d formed two H-bonds with the ARG439 via its R1 group and showed π-H interaction with MET637 that helped stabilize the ligand. Similarly, compounds 4c, 4f, 4l, 5c, 5h, 5l, and 5m showed the same interaction pattern with ARG439 by the formation of two H-bonds. The nitrogen of 5a formed a H-bond with the side chain of ARG609 and π–H interaction with MET637 while following the same interaction pattern with ARG439. Compounds 5g and 5o displayed a metal interaction with the Ni2+ ion and H-bonds with HIS492 and ARG609. In contrast, compounds 5b and 5f showed an H-bond with the side chain of ARG439, and nitrogen in the backbone of 5b also interacted with CYS592.

Compounds 5i, 4h, 4i, and 4b also adopted a similar interaction pattern with the ARG439 residue, mediating H-bonds. Additionally, 5i showed strong H-bonding with ARG609. Compound 4b showed an extra hydrogen bond interaction with ALA636. Compounds 4d, 4e, and 4k displayed a H-bond with ARG439, while HIS593 formed a π–π interaction with 4e and 4k. ARG609 also formed an π-cation interaction with 4k. Similarly, 5j and 5p showed H-bonds with ARG439, and the R2 group (Cl) of the 5j and 5p compounds formed halogen bonds with GLU493. Finally, compounds 5n and 4a showed almost similar interaction patterns with ARG439. The nitrogen of 5n interacted with the side chain of CYS592, while 4a mediated an H-bond with GLY638. These docking analyses suggest that the changes in the shape of a molecule can impact how it binds, leading to changes in its inhibitory effectiveness. Table S1 provides information about the docking scores and specific interactions of each molecule observed during docking. The compounds displayed docking scores between −6.82 and −4.60 kcal/mol, indicating a strong potential for binding with the active site of the urease enzyme. Figure 5 shows various binding modes of the compounds. Our docking results complement the experimental findings.

2.5. In Silico Prediction of ADMET Profile

In drug discovery, in silico predictions of various parameters like molecular weight, physicochemical features, pharmacokinetics, and bioavailability are vital to determine the potency of a drug molecule.38,39 The 28 compounds in the current study were subjected to in silico prediction of various parameters, and the results are summarized in Tables S2 and S4 (Supporting Information). The analysis revealed that the molecular weight of antiurease molecules varied from 354.38 to 447.35 g/mol. The predicted polarity (TPSA) and the number of rotatable bonds (NRB) range from 74.22 to 129.27 Å2 (values ≤140 Å2 indicate good oral bioavailability) and 2 to 5, respectively. These molecules have minimal hydrogen bond acceptors ranging from 2 to 6, and all adhere to Lipinski’s criteria.40 The molar refractivity (MR) of the compounds, a crucial factor in determining oral bioavailability, falls within the acceptable range according to Lipinski’s rule of five (except 5j, 5k, 5m, 5n, and 5p) and Ghose’s rules.41 The drug-likeness of compounds was evaluated using several rules, and all the compounds showed no violation of the Ghose filter (WLOGP < −0.4), except 5m, 5n, and 5p. However, eight compounds violated Muegge’s rules (XLOGP3 < −2).42

The lipophilicity score of compounds, which indicates their solubility in nonpolar solvents, ranged from 1.89 to 5.01. The molecules were also predicted to be moderately soluble in water, except for 5g and 5o. Most compounds showed high gastrointestinal absorption and no ability to penetrate the blood-brain barrier (except 5e, 5f, 5m, and 5n). Compounds were also unlikely to be the P-glycoprotein (P-gp) substrates except for the aforementioned four molecules (5e, 5f, 5m, and 5n). Furthermore, most compounds were predicted not to inhibit cytochrome P450 enzymes (CYP), a positive indication for safe human clearance. All the compounds had negative skin permeability coefficient values (Log Kp (cm/s) = −4.42 to −6.30), indicating that they are not very permeable through the skin. The estimated bioavailability scores of all compounds were favorable at 0.55, indicating their potential for good pharmacokinetic features. SwissADME analysis did not flag any PAINS alerts for any of the molecules, and all compounds showed one or two violations in the lead-likeness criteria (Mw < 250). The estimated synthetic accessibility scores of all compounds were between 3.14 and 3.56, demonstrating their easy synthetic accessibility.

3. Conclusions

In summary, the present work demonstrates the significant importance of nitrogen-containing heteroaromatics in medicinal chemistry while inhibiting the urease enzyme to the micromolar range. The excessive urease activity can cause peptic ulcers, gastritis, and environmental problems; therefore, we herein demonstrated the potential of triazolothiadiazoles and triazolothiadiazines as potent antiurease inhibitors. All the tested compounds showed excellent inhibitory data whereas 5k emerged as the most potent and lead inhibitor with IC50 and Ki values of 3.33 ± 0.11 and 3.62 ± 0.0034 μM, respectively. Kinetics analysis revealed the competitive mode of inhibition for compound 5k. Various structure–activity relationships were observed that indicate an intricate balance of two substituents at different positions is vital for achieving high inhibitory efficacy. Molecular docking revealed the types of interacting residues involved in establishing key contacts with potent inhibitors. Assessment of in silico ADME profile demonstrated good pharmacokinetic and physicochemical properties and followed the drug-likeness rules. Collectively, the findings reported in this work suggest that the identified heteroaromatic inhibitors could be developed as useful candidates in medicinal chemistry to treat peptic and gastric ulcers.

4. Experimental Section

4.1. Chemicals and Instrumentation

The chemicals used in this study were purchased from Aldrich and Alfa Aesar. For thin layer chromatography (TLC), Merck DF-Alufoilien 60F254 0.2 mm precoated plates were used. Product spots were visualized by using UV light at 254 and 365 nm. Stuart melting point apparatus was used to measure the melting points (SMP3). IR spectra were recorded using the FTS 3000 MX, Bio-Rad Merlin (Excalibur type) spectrophotometer. 1H and 13C NMR spectral data were obtained by using a Bruker Avance (300 MHz) spectrometer. Chemical shifts (δ) are quoted in parts per million (ppm), with the residual solvent serving as an internal standard (DMSO-d6 at 2.50 ppm for 1H NMR and 39.52 ppm for 13C NMR). Resonances are denoted by the letters s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), and Ar (aromatic).

4.2. Synthesis of 4-Amino-1,2,4-triazole-3-thiol (3a,b)

Benzoic acid (1.0 mmol) and thiocarbohydrazide (1.5 mmol) were heated in a sand bath for 1 h at 190–200 °C. The reaction mixture was cooled to room temperature, and the corresponding product was precipitated with hot water which was filtered, dried, and recrystallized from ethanol.33 The spectroscopic data were consistent with those we reported previously.33

4.2.1. 4-Amino-5-(4-methoxyphenyl)-4H-1,2,4-triazole-3-thiol (3a)

White solid (80%): mp 210–211 °C; Rf: 0.62 (10% MeOH/CHCl3); IR (ATR, cm–1): 3495 (N–H), 3015 (ArH), 2937, 2824 (CH3), 2554 (SH), 1602 (C=N), 1546, 1511 (C=C); 1H NMR (300 MHz, DMSO-d6): δ 13.83 (s, 1H, SH), 7.99 (d, 2H, J = 8.7 Hz, ArH), 7.07 (d, 2H, J = 8.7 Hz, ArH), 5.77 (s, 2H, NH2), 3.81 (s, 3H, OCH3); 13C NMR (75 MHz, DMSO-d6): δ 167.03, 161.28, 149.76, 130.05, 118.53, 114.40, 55.81.

4.2.2. 4-Amino-5-(3-bromophenyl)-4H-1,2,4-triazole-3-thiol (3b)

White solid (81%): mp 226–227 °C; Rf: 0.65 (10% MeOH/CHCl3); IR (ATR, cm–1): 3498 (N–H), 3035 (ArH), 2942, 2857 (CH2), 2563 (SH), 1605 (C=N), 1567, 1521 (C=C); 1H NMR (300 MHz, DMSO-d6): δ 14.03 (s, 1H, SH), 8.28–8.27 (m, 1H, ArH), 8.01 (d, 1H, J = 8.1 Hz, ArH), 7.76–7.73 (m, 1H, ArH), 7.49 (t, 1H, J = 7.8 Hz, ArH), 5.80 (s, 2H, NH2); 13C NMR (75 MHz, DMSO-d6): δ 167.72, 148.49, 133.66, 131.22, 130.84, 128.33, 127.39, 122.08.

4.3. Synthesis of 1,2,4-Triazolo[3,4-b][1,3,4]thiadiazoles (4a–l)

4-Amino-1,2,4-triazole-3-thiol (3a,b) (1.0 mmol) and the corresponding aryloxy acid (1.1 mmol) were refluxed in POCl3 (5 mL) for 6 h. The excess POCl3 was evaporated, and the resulting residue was neutralized with sodium bicarbonate. The precipitated solid was filtered, washed with cold water, dried, and recrystallized from ethanol to give 1,2,4-triazolo[3,4-b][1,3,4]thiadiazoles (4a–l).33 The spectroscopic data were consistent with those we reported previously.33

4.4. Synthesis of 1,2,4-Triazolo[3,4-b][1,3,4]thiadiazines (5a–p)

4-Amino-1,2,4-triazole-3-thiol (3a,b) (1.0 mmol) and the corresponding phenacyl bromide (1.2 mmol) were refluxed in absolute ethanol (10 mL) for 7 h. The reaction mixture was poured onto crushed ice and neutralized with sodium bicarbonate. The precipitated solid was filtered, washed with water, dried, and recrystallized from ethanol to give the conjugated products (5a–p). The spectroscopic data were consistent with those we reported previously.33 The detailed experimental and characterization data are provided in the Supporting Information.

4.5. In Vitro Urease Assay

The urease assay used 200 μL of the reaction mixture, 25 μL of the urease enzyme from the Jack bean (Canavalia ensiformis), and 5 μL of samples of various compounds. The plate was incubated for 15 min at 30 °C and added 55 μL of urea (100 mM). Phenolic and alkali reagents (A), (B) 45 > μL/well, and 70 μL were employed into a 96-well plate. Here, phenolic reagent comprising 1% w/v phenol and 0.005% w/v sodium nitroprusside constituents and alkali reagent is the combination of 0.5% w/v NaOH and 0.1% w/v NaOCl. The Weatherburn approach, which considers ammonia release upon hydrolysis, was used to assess urease inhibition of triazolothiadiazole and triazolothiadiazine derivatives. After 50 min, absorbance was measured using a microplate reader (xMark Microplate spectrophotometer, BIO-RAD).43 With a final volume of 200 μL, each reaction was run in triplicate. We have employed thiourea as a conventional urease inhibitor.44 The formula in eq 1 was used to compute the percent inhibition:

4.5. 1

4.6. Statistical Analysis

The IC50 values were determined by using the EZ-Fit Enzyme Kinetics tool (Perrella Scientific Inc., Amherst, USA).

4.7. Molecular Docking Analysis

4.7.1. Protein Coordinate Retrieval

The X-ray diffraction structure of the urease enzyme (Jack bean) was retrieved from the protein data bank server (RCSB-PDB) (https://www.rcsb.org/) with PDB-ID: 4GY7. The phosphate ion is attached to the active pocket of the protein with two Ni2+ also interacting with the active site residues. The Molecular Operating Environment version 2022.02 (MOE) was utilized to refine the protein’s crystal structure using all atoms Amber14:EHT45,46 force field. The protein structure’s missing residues were built using the MOE Loop modeler. The start and end of the protein (C–N terminals) were charged, and missing hydrogens were added. The force field missing parameters including atom type missing, bond strength parameters, bond angles missing parameters, and van der Waals missing parameters were added.

4.7.2. Molecular Docking of the Selected Compounds

The MOE Dock application was used to dock the selected compounds in the active site of the urease enzyme (PDB-ID:4GY7). Before docking, the protein and ligand were prepared using the MOE-QuickPrep module to add partial charges and missing hydrogens using AMBER14: EHT force field. The docking was performed using the triangle matcher method of ligand placement and scored by the London dG algorithm. One hundred poses were selected for ligand placement using the rotate bond conformations in the active pocket. The refinement was performed through the GBVI/WSA dG algorithm. Thirty poses were retrieved from the refinement using the receptor as a rigid body. The MOE protein–ligand interaction fingerprinting (PLIF) was used to analyze the interactions. The docking protocols were used to dock all of the selected compounds in the urease active site. Based on the docking score and good interactions in the protein–ligand complexes, the final pose for each compound was selected.

4.8. Estimation of Pharmacokinetics, Drug-likeness, and Physicochemical Profile

The SwissADME program was utilized to predict the drug-like properties, pharmacokinetics, medicinal properties, and physicochemical characteristics of triazolothiadiazoles and triazolothiadiazines.47,48

Acknowledgments

The authors extend their appreciation to the Deanship of Scientific Research at King Khalid University for funding this work through the Large Groups Project under grant number (RGP. 2/100/44).

Supporting Information Available

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.3c03546.

  • Characterization data, 1H and 13C NMR spectra for compounds, kinetics graphs, docking scores, and ADMET properties (PDF)

Author Contributions

⊥ J.U. and S.U. have contributed equally to this work.

The authors declare no competing financial interest.

Supplementary Material

ao3c03546_si_001.pdf (1.6MB, pdf)

References

  1. Kafarski P.; Talma M. Recent advances in design of new urease inhibitors: A review. J. Adv. Res. 2018, 13, 101–112. 10.1016/j.jare.2018.01.007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Kosikowska P.; Berlicki Ł. Urease inhibitors as potential drugs for gastric and urinary tract infections: a patent review. Expert Opin. Ther. Pat. 2011, 21 (6), 945–957. 10.1517/13543776.2011.574615. [DOI] [PubMed] [Google Scholar]
  3. Kusters J. G.; van Vliet A. H. M.; Kuipers E. J. Pathogenesis of Helicobacter pylori infection. Clin. Microbiol. Rev. 2006, 19 (3), 449–490. 10.1128/CMR.00054-05. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Rai R.; Saraswat V. A.; Dhiman R. K. Gut microbiota: its role in hepatic encephalopathy. J. Clin. Exp. Hepatol. 2015, 5 (Suppl 1), S29–S36. 10.1016/j.jceh.2014.12.003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Rosenstein I. J.; Hamilton-Miller J. M.; Brumfitt W. Role of urease in the formation of infection stones: comparison of ureases from different sources. Infect. Immun. 1981, 32 (1), 32–37. 10.1128/iai.32.1.32-37.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Krajewska B.; Ureases I. Functional, catalytic and kinetic properties: A review. J. Mol. Catal. B: Enzym. 2009, 59 (1–3), 9–21. 10.1016/j.molcatb.2009.01.003. [DOI] [Google Scholar]
  7. Ibrar A.; Khan I.; Abbas N. Structurally diversified heterocycles and related privileged scaffolds as potential urease inhibitors: A brief overview. Arch. Pharm. Chem. Life Sci. 2013, 346, 423–446. 10.1002/ardp.201300041. [DOI] [PubMed] [Google Scholar]
  8. Zahra U.; Zaib S.; Saeed A.; Ur Rehman M.; Shabir G.; Alsaab H. O.; Khan I. New acetylphenol-based acyl thioureas broaden the scope of drug candidates for urease inhibition: synthesis, in vitro screening and in silico analysis. Int. J. Biol. Macromol. 2022, 198, 157–167. 10.1016/j.ijbiomac.2021.12.064. [DOI] [PubMed] [Google Scholar]
  9. Hameed A.; Shafiq Z.; Yaqub M.; Hussain M.; Hussain M. A.; Afzal M.; Tahir M. N.; Naseer M. M. Me3N-promoted synthesis of 2,3,4,4a-tetrahydroxanthen-1-one: preparation of thiosemicarbazone derivatives, their solid state self-assembly and antimicrobial properties. New J. Chem. 2015, 39 (12), 9351–9357. 10.1039/C5NJ01879J. [DOI] [Google Scholar]
  10. Asgari M. S.; Azizian H.; Montazer M. N.; Mohammadi-Khanaposhtani M.; Asadi M.; Sepehri S.; Ranjbar P. R.; Rahimi R.; Biglar M.; Larijani B.; Amanlou M.; Mahdavi M. New 1, 2, 3-triazole–(thio) barbituric acid hybrids as urease inhibitors: design, synthesis, in vitro urease inhibition, docking study, and molecular dynamic simulation. Arch. Pharm. 2020, 353, e2000023 10.1002/ardp.202000023. [DOI] [PubMed] [Google Scholar]
  11. Rezaei E. B.; Abedinifar F.; Azizian H.; Montazer M. N.; Asadi M.; Hosseini S.; Sepehri S.; Mohammadi-Khanaposhtani M.; Biglar M.; Larijani B.; Amanlou M.; Mahdavi M. Design, synthesis, and evaluation of metronidazole-1,2,3-triazole derivatives as potent urease inhibitors. Chem. Pap. 2021, 75, 4217–4226. 10.1007/s11696-021-01653-4. [DOI] [Google Scholar]
  12. Pedrood K.; Azizian H.; Montazer M. N.; Moazzam A.; Asadi M.; Montazeri H.; Biglar M.; Zamani M.; Larijani B.; Zomorodian K.; Mohammadi-Khanaposhtani M.; Irajie C.; Amanlou M.; Iraji A.; Mahdavi M. Design and synthesis of new N-thioacylated ciprofloxacin derivatives as urease inhibitors with potential antibacterial activity. Sci. Rep. 2022, 12, 13827. 10.1038/s41598-022-17993-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hosseinzadeh N.; Montazer M. N.; Mohammadi-Khanaposhtani M.; Valizadeh Y.; Amanlou M.; Mahdavi M. Rational design, synthesis, docking simulation, and ADMET prediction of novel barbituric-hydrazine-phenoxy-1,2,3-triazole-acetamide derivatives as potent urease inhibitors. ChemistrySelect 2023, 8, e202203297 10.1002/slct.202203297. [DOI] [Google Scholar]
  14. Rego Y. F.; Queiroz M. P.; Brito T. O.; Carvalho P. G.; de Queiroz V. T.; de Fátima Â.; Macedo F. Jr A review on the development of urease inhibitors as antimicrobial agents against pathogenic bacteria. J. Adv. Res. 2018, 13, 69–100. 10.1016/j.jare.2018.05.003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Zaib S.; Younas M. T.; Zaraei S. O.; Khan I.; Anbar H. S.; El-Gamal M. I. Discovery of urease inhibitory effect of sulfamate derivatives: Biological and computational studies. Bioorg. Chem. 2022, 119, 105545 10.1016/j.bioorg.2021.105545. [DOI] [PubMed] [Google Scholar]
  16. McCarty G.; Bremner J.; Lee J. Inhibition of plant and microbial ureases by phosphoroamides. Plant Soil 1990, 127 (2), 269–283. 10.1007/BF00014435. [DOI] [Google Scholar]
  17. Khan K. M.; Iqbal S.; Lodhi M. A.; Maharvi G. M.; Choudhary M. I.; Perveen S. Biscoumarin: new class of urease inhibitors; economical synthesis and activity. Bioorg. Med. Chem. 2004, 12 (8), 1963–1968. 10.1016/j.bmc.2004.01.010. [DOI] [PubMed] [Google Scholar]
  18. Aslam M. A. S.; Mahmood S.-U.; Shahid M.; Saeed A.; Iqbal J. Synthesis, biological assay in vitro and molecular docking studies of new Schiff base derivatives as potential urease inhibitors. Eur. J. Med. Chem. 2011, 46 (11), 5473–5479. 10.1016/j.ejmech.2011.09.009. [DOI] [PubMed] [Google Scholar]
  19. Onoda Y.; Iwasaki H.; Magaribuchi T.; Tamaki H. Effects of the new anti-ulcer agent 12-sulfodehydroabietic acid monosodium salt on healing of acetic acid-induced gastric ulcers in rats. Arzneimittelforschung 1991, 41 (5), 546–548. [PubMed] [Google Scholar]
  20. Mermer A.; Keles T.; Sirin Y. Recent studies of nitrogen containing heterocyclic compounds as novel antiviral agents: A review. Bioorg. Chem. 2021, 114, 105076 10.1016/j.bioorg.2021.105076. [DOI] [PubMed] [Google Scholar]
  21. Bhardwaj N.; Pathania A.; Kumar P. Naturally available nitrogen-containing fused heterocyclics as prospective lead molecules in medicinal chemistry. Curr. Tradit. Med. 2021, 7 (1), 5–27. 10.2174/2215083805666190613125700. [DOI] [Google Scholar]
  22. Tejaswini D. P. Design, in silico analysis, synthesis, and evaluation of novel benzofused nitrogen-containing heterocyclic N-substituted mercaptobenzimidazole derivatives as potential antimicrobial agent. Int. J. Green Pharm. 2021, 15 (3), 226–237. 10.14233/ajomc.2021.AJOMC-P325. [DOI] [Google Scholar]
  23. Hosseinzadeh Z.; Ramazani A.; Razzaghi-Asl N. Anti-cancer nitrogen-containing heterocyclic compounds. Curr. Org. Chem. 2018, 22 (23), 2256–2279. 10.2174/1385272822666181008142138. [DOI] [Google Scholar]
  24. Lang D. K.; Kaur R.; Arora R.; Saini B.; Arora S. Nitrogen-containing heterocycles as anticancer agents: An overview. Anti-Cancer Agents Med. Chem. 2020, 20 (18), 2150–2168. 10.2174/1871520620666200705214917. [DOI] [PubMed] [Google Scholar]
  25. Singh K.; Pal R.; Khan S. A.; Kumar B.; Akhtar M. J. Insights into the structure activity relationship of nitrogen-containing heterocyclics for the development of antidepressant compounds: An updated review. J. Mol. Struct. 2021, 1237, 130369 10.1016/j.molstruc.2021.130369. [DOI] [Google Scholar]
  26. Wang W.; Xiong L.; Li Y.; Song Z.; Sun D.; Li H.; Chen L. Synthesis of lathyrane diterpenoid nitrogen-containing heterocyclic derivatives and evaluation of their anti-inflammatory activities. Bioorg. Med. Chem. 2022, 56, 116627 10.1016/j.bmc.2022.116627. [DOI] [PubMed] [Google Scholar]
  27. Kamboj V. K.; Kapoor A.; Jain S. Synthesis, antimicrobial, and antioxidant screening of aryl acetic acid incorporated 1,2,4-triazolo-1,3,4-thiadiazole derivatives. J. Heterocycl. Chem. 2019, 56, 1376–1382. 10.1002/jhet.3513. [DOI] [Google Scholar]
  28. Khan I.; Ibrar A.; Abbas N. Triazolothiadiazoles and triazolothiadiazines - Biologically attractive scaffolds. Eur. J. Med. Chem. 2013, 63, 854–868. 10.1016/j.ejmech.2013.01.060. [DOI] [PubMed] [Google Scholar]
  29. Karthikeyan M. S.; Holla B. S.; Kalluraya B.; Kumar N. S. Biological studies of some 2,4-dichloro-5-fluorophenyl containing triazolothiadiazoles. Monatsh. Chem. 2007, 138, 1309–1316. 10.1007/s00706-007-0718-y. [DOI] [Google Scholar]
  30. Khan I.; Ali S.; Hameed S.; Rama N. H.; Hussain M. T.; Wadood A.; Uddin R.; Ul-Haq Z.; Khan A.; Ali S.; Choudhary M. I. Synthesis, antioxidant activities and urease inhibition of some new 1,2,4-triazole and 1,3,4-thiadiazole derivatives. Eur. J. Med. Chem. 2010, 45, 5200–5207. 10.1016/j.ejmech.2010.08.034. [DOI] [PubMed] [Google Scholar]
  31. Khan I.; Zaib S.; Ibrar A.; Rama N. H.; Simpson J.; Iqbal J. Synthesis, crystal structure and biological evaluation of some novel 1,2,4-triazolo[3,4-b]1,3,4-thiadiazoles and 1,2,4-triazolo[3,4-b]1,3,4-thiadiazines. Eur. J. Med. Chem. 2014, 78, 167–177. 10.1016/j.ejmech.2014.03.046. [DOI] [PubMed] [Google Scholar]
  32. Khan I.; Ibrar A.; Zaib S.; Ahmad S.; Furtmann N.; Hameed S.; Simpson J.; Bajorath J.; Iqbal J. Active compounds from a diverse library of triazolothiadiazole and triazolothiadiazine scaffolds: Synthesis, crystal structure determination, cytotoxicity, cholinesterase inhibitory activity, and binding mode analysis. Bioorg. Med. Chem. 2014, 22, 6163–6173. 10.1016/j.bmc.2014.08.026. [DOI] [PubMed] [Google Scholar]
  33. Khan I.; Bakht S. M.; Ibrar A.; Abbas S.; Hameed S.; White J. M.; Rana U. A.; Zaib S.; Shahid M.; Iqbal J. Exploration of a library of triazolothiadiazole and triazolothiadiazine compounds as a highly potent and selective family of cholinesterase and monoamine oxidase inhibitors: design, synthesis, X-ray diffraction analysis and molecular docking studies. RSC Adv. 2015, 5, 21249–21267. 10.1039/C5RA00906E. [DOI] [Google Scholar]
  34. Saeed A.; Ashraf Z.; Nadeem H.; Simpson J.; Pérez H.; Erben M. F. An investigation of supramolecular synthons in 1,2,4-triazole-3(4H)-thione compounds. X-ray crystal structures, energetic and Hirshfeld surface analysis. J. Mol. Struct. 2019, 1195, 796–806. 10.1016/j.molstruc.2019.06.049. [DOI] [Google Scholar]
  35. Khan K.; Hameed S.; Al-Masoudi N. A.; Abdul-Reda N. A.; Simpson J. New triazolothiadiazole and triazolothiadiazine derivatives as kinesin Eg5 and HIV inhibitors: synthesis, QSAR and modeling studies. Z. Naturforsch. 2015, 70 (1 b), 47–58. 10.1515/znb-2014-0162. [DOI] [Google Scholar]
  36. Gholivand K.; Farshadian S.; Erben M. F.; Della Védova C. O. Synthesis and characterization of the first phosphonic diamide containing thiazolyl groups: Structural properties and tautomeric equilibrium. J. Mol. Struct. 2010, 978, 67–73. 10.1016/j.molstruc.2009.12.018. [DOI] [Google Scholar]
  37. Hozien Z. A.; EL-Mahdy A. F. M.; Ali L. S. A.; Abo Markeb A.; El-Sherief H. A. H One-pot synthesis of some new s-triazole derivatives and their potential application for water decontamination. ACS Omega 2021, 6, 25574–25584. 10.1021/acsomega.1c03675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Krämer S. D.; Wunderli-Allenspach H. Physicochemical properties in pharmacokinetic lead optimization. Il Farmaco 2001, 56, 145–148. 10.1016/S0014-827X(01)01028-X. [DOI] [PubMed] [Google Scholar]
  39. Neervannan S. Preclinical formulations for discovery and toxicology: physicochemical challenges. Expert Opin. Drug Metab. Toxicol. 2006, 2 (5), 715–731. 10.1517/17425255.2.5.715. [DOI] [PubMed] [Google Scholar]
  40. Lipinski C. A.; Lombardo F.; Dominy B. W.; Feeney P. J. Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv. Drug Deliv. Rev. 1997, 23, 3–25. 10.1016/S0169-409X(96)00423-1. [DOI] [PubMed] [Google Scholar]
  41. Ghose A. K.; Viswanadhan V. N.; Wendoloski J. J. A knowledge-based approach in designing combinatorial or medicinal chemistry libraries for drug discovery. 1. A qualitative and quantitative characterization of known drug databases. J. Comb. Chem. 1999, 1, 55–68. 10.1021/cc9800071. [DOI] [PubMed] [Google Scholar]
  42. Muegge I. Selection criteria for drug-like compounds. Med. Res. Rev. 2003, 23, 302–321. 10.1002/med.10041. [DOI] [PubMed] [Google Scholar]
  43. Weatherburn M. Phenol-hypochlorite reaction for determination of ammonia. Anal. Chem. 1967, 39, 971–974. 10.1021/ac60252a045. [DOI] [Google Scholar]
  44. Islam M.; Khan A.; Shehzad M. T.; Hameed A.; Ahmed N.; Halim S. A.; Khiat M.; Anwar M. U.; Hussain J.; Csuk R. Synthesis and characterization of new thiosemicarbazones, as potent urease inhibitors: In vitro and in silico studies. Bioorg. Chem. 2019, 87, 155–162. 10.1016/j.bioorg.2019.03.008. [DOI] [PubMed] [Google Scholar]
  45. Gerber P. R.; Müller K. MAB, a generally applicable molecular force field for structure modelling in medicinal chemistry. J. Comput. Aided Mol. Des. 1995, 9, 251–268. 10.1007/BF00124456. [DOI] [PubMed] [Google Scholar]
  46. Maier J. A.; Martinez C.; Kasavajhala K.; Wickstrom L.; Hauser K. E.; Simmerling C. ff14SB: improving the accuracy of protein side chain and backbone parameters from ff99SB. J. Chem. Theory Comput. 2015, 11, 3696–3713. 10.1021/acs.jctc.5b00255. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Daina A.; Michielin O.; Zoete V. SwissADME: a free web tool to evaluate pharmacokinetics, drug-likeness and medicinal chemistry friendliness of small molecules. Sci. Rep. 2017, 7, 42717. 10.1038/srep42717. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Aziz S.; Waqas M.; Iqbal A.; Halim S. A.; Abdellattif M. H.; Khan A.; Al-Harrasi A. Structure-based identification of potential substrate antagonists for isethionate sulfite-lyase enzyme of Bilophila wadsworthia: Towards novel therapeutic intervention to curb gut-associated illness. Int. J. Biol. Macromol. 2023, 240, 124428 10.1016/j.ijbiomac.2023.124428. [DOI] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

ao3c03546_si_001.pdf (1.6MB, pdf)

Articles from ACS Omega are provided here courtesy of American Chemical Society

RESOURCES