Investigating the potential of novel thiazolidinone derivatives as anti-alzheimer agents: A comprehensive study through kinetics, in silico, and in vitro approaches

2.1. General procedure for the synthesis of 4-thiazolidinone derivatives based on 2-mercaptobenzimidazol (2-MBI)

Synthesis of starting material hydrazone: 2-MBI derivatives were synthesized via multiple sequence reactions. Initially, 2-MBI was refluxed with bromoethane in EtOH using potassium hydroxide (KOH) as a base for 10 h. Following EtOH evaporation, 2-ethylthio-benzimidazole was produced as bright white needle-like crystals. In the second stage, 2-(2-(ethylthio)benzimidazolyl) acetate was produced in the semisolid form, which involved reacting 2-ethylthio-benzimidazole with ethyl chloroacetate in the presence of anhydrous potassium carbonate using dimethyl formamide (DMF) as a solvent under reflux for 15 h. After refluxing this intermediate with hydrazine hydrate in MeOH for 10 h, 2-((ethylthio)benzimidazolyl)acetohydrazide was formed. This precipitated when added to cold water and was filtered out. The last stage involved dissolving this hydrazide in methanol using a catalytic quantity of glacial acetic acid and then adding an aldehyde. The mixture was refluxed for 5 to 6 h, and when it cooled, the product was obtained as a precipitate. It was then filtered, cleaned, and dried in an open atmosphere.

Synthesis of final derivatives ( 1-12): For synthesis of thiazolidinone derivatives, 20 mL MeOH and 5 mL acetic acid were taken in a 100 mL round bottom flask, to which 0.733 mmol (0.09 g) of zinc chloride in molten form was added and stirred for about 5 min on a magnetic hot plate stirrer. Then 1.00 mmol (0.430 g) of the 2- mercaptobenzimidazole hydrazone derivative was added, and the mixture was stirred until completely dissolved on a hot plate. After 5 min, we added 1.00 mmol (0.14 mL) of mercapto-acetic acid (thioglycolic acid), and the reaction mixture was refluxed for about 8-10 h. We employed Thin Layer Chromatography to monitor the progress of the reaction. After completion of the reaction, the product was precipitated in freezing water, filtered, and washed with Na₂CO₃ to remove excess thioglycolic acid. The product was dried and preserved. The structures of novel derivatives were expanded upon using spectroscopic techniques like nuclear magnetic resonance (NMR), i.e., ¹H NMR and ¹³C NMR [19].

2.2. General

The current study utilized analytical-grade chemical reagents. AChE (EC; 3.1.1.7) (C3389-2KU), 5-dithiobis 2-nitrobenzoic acid (DTNB) (D8130-10G), Galantamine (Cat. No. 69353-21-5), and ACh (A7000-25G) were used. In addition, Sigma Aldrich sourced ethanol (No. 64-17-5), monobasic sodium phosphate (1342-35-0), and Sodium phosphate dibasic (558-79-4). The thiazolidinone derivatives (1-12) were previously synthesized and described earlier [19].

2.3. Inhibitory evaluation of AChE

We carried out the AChE inhibition assay following the method described earlier [20], where galantamine served as a control. Initially, 140 µL of sodium phosphate buffer (100 mM) pH 8, 10 µL DTNB (0.25 mM), 0.5 mM of thiazolidinone compound dissolved in 20 µL, and 20 µL AChE enzyme were mixed and incubated for 15-20 min at room temperature. Finally, 0.4 mM of substrate ACh (10 µL) was added to the mixture and incubated for a further af 5 min. The reaction started, and ACh was hydrolyzed by AChE enzyme in the presence of DTNB; a yellow-colored 5-thio-2-nitrobenzoate anion was generated, which indicates the reaction completion and was read at 412 nm wavelength. We performed each experiment in biological triplicate and analyzed the data using SoftMax Pro v. 6.3, using the following Eq. (1) to compute the inhibition as a percentage:

2.4. Kinetic study

The amount of compound required for 50% inhibition is known as IC₅₀, which was calculated using EZ-Fit (Perrella Scientific Inc; Amherst, USA) at various concentrations of the test compounds.

In the enzyme-substrate (ES) reaction, the AChE enzyme reacts with substrate ACh to form AChE-ACh complex, forming the product (P). The dissociation constant values were calculated using Lineweaver-Burk, Dixon plots, and their secondary replots [21]. The values of K_m, k_i, and V_max were calculated from Lineweaver-Burk and Dixon plots through a non-linear regression equation. The Lineweaver-Burk plot was employed to determine the K_i values; initially, values of 1/V _maxapp were found at each line intersection point of each concentration of the test compound on the y-axis. The slope of each line generated by the compound was plotted against various test compound concentrations on the Lineweaver-Burk plot.

2.5. Analysis of statistics

GraFit software [22] was used to evaluate correlation coefficients, slope, intercept, and standard error values.

2.6. AChE and ligand docking

The Molecular Operating Environment (MOE) package (www.chemcomp.com) was used to explore various binding conformations of AChE with different compounds. The 3D structure of Homo sapiens AChE (hAChE) (PDB code: 4EY6, resolution: 2.40 Å) was obtained from PDB (Protein Data Bank) (https://www.rcsb.org).

The online server Play-molecule was used to prepare the target protein, involving chain completion, protonation, and correction of bonds [23]. Crystalized H₂O molecules were removed from the retrieved crystal structure using MOE. Subsequently, 3D protonation was applied, and energy minimization was performed using the default parameters of the MOE program. The structures of synthesized thiazolidinone derivatives were built in the MOE program and protonated, and then energy was minimized through MMFP94x force fields with a 0.05 gradient. Each tested compound was docked in the active sites of the target AChE enzyme using the default parameters of the MOE-Dock program. Ten conformations were generated for each compound, and the top-ranked conformations based on docking scores were selected for further in silico studies.

To validate the reliability of the docking procedure, a re-docking experiment was performed using the AChE co-crystalized ligand as a reference (Figure S1). This step ensures the accuracy and consistency of the molecular docking method employed in this study.

2.7. Protein-ligand interaction fingerprint (PLIF)

The accuracy of the molecular docking data was confirmed using PLIF. The PLIF tool was linked with MOE and applied to display the principal interaction efficiency [24]. A docking protocol was employed to create a database. Bonding interactions, including hydrogen bonding, ionic bonding, and surface contacts, were analyzed per residue. The PLIF histogram was generated, which illustrates a database of the complex. The MOE PLIF was utilized to analyze all the molecular docking postures, and the software was executed using the default parameters. All the molecular docking poses were submitted to the MOE PLIF, and the program was run using default parameters.

2.8. MD simulations

A total of two top-ranked compounds (12 and 3) were short-listed based on IC₅₀ and docking scores for further MD simulations (GROMACS 5.1.2) [25]. Initially, a protein ligand complex was generated using the pdb 2 gmx module. Parametrization and topology of protein were generated by the OPLS-AA/L force field [26], and ligand topology was prepared using the Swissparam online web server (http://www.swissparam.ch/). Subsequently, each protein-ligand complex was solvated in a cubic box (10 × 10 × 10) surrounded by approximately 33,439 TiP3P water molecules. In the next step, 9 sodium ions were added to neutralize the complex. Using positional restraint and the steepest descent algorithm, energy minimization was carried out up to a maximum force of 10 KJ/mol [27], eliminating the atomic location mistake and structural variations such as bond length, bond angle, and structural conflicts between the ions, water molecules, and protein complex. The solvent around the protein was equilibrated under the number of atoms, volume, and temperature (NVT) ensemble, and then using the NPT ensemble. NVT and NPT would both perform 50,000 steps for 0.1 ns at 300 K and 1 bar of pressure. Each complex was then put through MD simulations utilizing a leapfrog algorithm for 10,000,000 ps at 300,000 atmospheric NPT ensemble pressure and periodic boundary conditions for 0.002 ps. All hydrogen bonds were contained throughout the procedure using the LINCS algorithm [28]. However, the particle MeshEwald module was functionalized with a 0.16 Å Fourier grid spacing [21].

2.9. Antispasmodic and Ca²⁺ antagonistic activities

The antispasmodic potentials of thiazolidinone compounds (1-12) were evaluated by the examination of spontaneous contracting rabbit jejunum [20]. The local breed rabbits of both sexes (1.5-2.0 kg) used in the current study were generously supplied by Agha Khan University, Pakistan. Animals were allowed free access to water, but their food was stopped 24 h before the experiment started. The animals were sacrificed with a blow to the back of the head, the abdomen was cut open, and a piece of their jejunum was removed. The 2 cm long segments were suspended in Tyrode’s solution (KCl 80 mM, MgCl₂ 1.05 mM, NaHCO₃ 11.87mM, NaCl 91.04 mM, NaH₂PO₄ 0.41mM, CaCl₂ 1.8 mM, and glucose 5.55 mM), which was aerated with a mixture of 95% O₂ and 5% CO₂, and preserved at 37°C. The spontaneous intestinal movements were observed and recorded isotonically using Harvard transducers and the Harvard Student Oscillograph. Each tissue was permitted to equilibrate for at least 30 min before any drug addition. The jejunum displayed spontaneous rhythmic contractions under these experimental conditions, enabling the study of spasmolytic activity directly without the use of an agonist. In addition, a Ca²⁺ antagonistic activity was conducted to authenticate the capability of the thiazolidinone compound (1-12) to relax the contractions induced by 80 mM K⁺. [29]

2.10. Cytotoxicity activity

Human neutrophil isolation: Human neutrophils were used to assess the cytotoxic effects of synthesized thiazolidine derivatives. For this purpose, whole blood (heparinized) was obtained from a local blood bank, and the required neutrophils were isolated [30]. The whole blood was then mixed with Ficoll-Paque, and after sedimentation, the unwanted RBCs were layered with the buffy coat on the Ficoll cushion (3 mL). The mixture was centrifuged for 30 min at 1500 rmp, and the supernatant was disposed of. The preparation was then centrifuged, and the neutrophils were cleaned with modified Hanks solution (MHS) and resuspended at a concentration of 1× 10⁷ cells/mL.

Assay procedure: The isolated neutrophils were incubated with thiazolidinone compounds for 30 min, followed by the addition of 0.25 mM WST-1 in a water bath shaker at 37°C [20]. The absorbance change was then measured at 450 nm in a 96-well plate using a Spectra Max, CA, United States. The OD (optical density) is the mean of five experimental replicates.

Percent viability was determined by Eq. (2).

3.1. Biology

Assessment of thiazolidinone inhibitory efficacy was performed against AChE. All compounds exhibited varying degrees of potential at various concentrations, with IC₅₀ values ranging from 209.53 ± 1.0 µM to 1656.01 ± 1.60 µM, as illustrated in Table 1. The current results suggest that these compounds possess significant potential as inhibitors of AChE, laying the foundation for further investigation. Compound 12 was the most potent inhibitor, with the lowest IC₅₀ (209.53 ± 1.01 µM) among the tested series, but had lower efficacy as compared to standard Galantamine (IC₅₀ = 92.49 ± 0.69 µM). The effectiveness of this compound may be due to the presence of an octyl group in its structure. Compound 3 (IC₅₀ = 249.52 ± 1.25 µM) with a hydroxyl moiety on the para position was the second most potent compound in this category. Compound 11 was the third most potent compound with an IC₅₀ of 345.15 ± 1.47 µM; the effectiveness of this compound is likely attributed to the existence of the 2-methyl furan ring. Compound 4 (IC₅₀ = 372.09 ± 1.04 µM) was ranked fourth in potency against AChE; the efficacy of this compound may be due to the high electronegative -Cl on ortho and para positions of a benzene ring. Compound 8 (IC₅₀ = 528.10 ± 1.45 µM) contains a hydroxyl group at the ortho and an ethoxy group at the meta positions. The reduced activity, compared to compound 3, suggests that the meta-positioned ethoxy group may hinder the inhibitory effect.

Table 1. Thiazolidinone inhibitory efficacy against AChE.

Compound no.	R	IC50 ± SEM (µM)	Docking Score
1		1172.64 ± 0.49	-9.529
2		1656.01 ± 1.60	-9.129
3		249.52 ± 1.25	-12.380
4		372.09 ± 1.04	-11.371
5		808.40 ± 0.5	-10.485
6		697.19 ± 0.78	-10.987
7		65.86 ± 0.91	-10.794
8		528.10 ± 1.45	-11.123
9		1227.97 ± 0.86	-10.321
10		801.74 ± 0.89	-10.699
11		345.15 ± 1.47	-11.842
12		209.53 ± 1.01	-13.511
Galantamine		92.49 ± 0.69	-14.542

Similarly, compared with compound 8, compound 6, which contains the hydroxyl group at the ortho position, exhibited a significant loss in efficacy (IC₅₀ = 697.19 ± 0.78 µM). This was observed when the ethoxy group was replaced with a methoxy group at the meta position of the compound. Compound 7 bearing an electron-withdrawing chloro group at the ortho position was the 7^th active compound of the series with IC₅₀ = 765.86 ± 0.91 µM. Compounds 10 and 5 displayed slightly different IC₅₀ values, IC₅₀ = 801.74 ± 0.89 µM, and IC₅₀ = 808.40 ± 0.5 µM. Compound 10, which has hydroxyl groups at the ortho and para positions, demonstrated greater efficacy than compound 5, substituted with an electron-withdrawing nitro group. Compounds 1, 9, and 2 were less potent with IC₅₀ values of 1172.64 ± 0.49, 1227.97 ± 0.86, and 1656.01 ± 1.60, respectively, as shown in Table 1. These results indicate that the activities of the compound may be associated with the electron-withdrawing hydroxyl and chloro groups or the electron-donating alkyl group.

3.2. Kinetics of AChE by thiazolidinone derivatives

Kinetic studies were employed to study the inhibitory efficacy of thiazolinone derivatives (1-12) on AChE. As a result, the tested compounds inhibited our enzyme with K_i values ranging from 41.74 ± 0.8 to 578.21 ± 0.4 µM. Notably, Compound 12 with Ki = 41.74 ± 0.8 µM established strong binding, whereas Compound 3 demonstrated weak interaction (Ki = 578.21 ± 0.4 µM) with AChE. Consequently, the IC₅₀ value for compound 12 (209.53 ± 1.01 µM) is lower and confirms its inhibitory potential compared to compound 3, which demonstrated a weak inhibitory efficacy.

According to kinetic parameters, as shown in Table 2, all compounds of Thiazolidinones acted as noncompetitive inhibitors of AChE. The Km values were consistent, and V_max decreased, which indicated a noncompetitive inhibition mechanism (Table 2). The steady-state inhibitory data of compound 12 against AChE have been illustrated in Figure 1.

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Figure 1.

Compound 12’s anti-AChE activity is represented in three plots. (a) Is the first plot of, Lineweaver-Burk, which shows the relationship between initial velocities and four constants conc of ACh: 100 µM (□), 150 µM (●), 175 µM (○), in the lack of compound 12 (■). (b) Is the second Dixon plot, which demonstrates the reciprocal relationship between initial velocities and variable concentration of compound 12 at constant conc. of ACh 100 µM (■), 150 µM (□), 175 µM (●) and 200 µM (○). (c) Is the third plot, which displays the correlation between various concentrations of compound 12 and the slopes.

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3.3. PLIF analysis

In the present study, all compounds of thiazolidinone were docked in the pocket of 4EY6 to generate barcodes and population plots Figures 2(a) and (b). Among all residues, TYR124 was found to be the most active residue in binding interactions with the target protein (4EY6) active site and followed by some other important interacting residues revealed by fingerprint bits such as TRP86, TYR341, SER125, GLY121, TYR337, TYR72, SER203, ASN74, ASN87, PHE295, HIS447, and TYR449.

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Figure 2.

PLIF between thiazolidinone compounds and 4EY6 (a) barcode (b) population.

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3.4. Molecular docking analysis

Molecular docking was conducted to validate in vitro results. The results revealed that all the compounds exhibited good binding affinity towards the 4EY6 active site. The top three compounds (12, 3, and 11) among the series with higher docking scores were evaluated for their interactions with the active site of AChE.

Compound 12 was the most potent compound of the series with IC₅₀ = 209.53± + 1.01 µM and docking score = - 13.511. The carbonyl oxygen of this compound acts as an acceptor and forms two hydrogen bonds, with TYR124 and SER125 having bond lengths of 3.1 and 2.8 Å, respectively. Moreover, the amido group of the same compound mediated hydrogen bond interaction with the side chain TYR337 with a bond length of 1.7 Å, as illustrated in Figures 3(a) and (b). Nevertheless, this compound revealed lower effectiveness than Galantamine (IC₅₀ = 92.49 ± 0.69 µM, docking score = ₋ 14.542). Galantamine was found to mediate two interactions with side chains TYR337 and TYR341, with 3.1 Å and 2.9 Å, respectively. The galantamine tetrahydrofuran moiety acted as an acceptor, making two other interactions with TYR124 and SER125 residues of the AChE (Figures 4a and b).

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Figure 3.

AChE active site interactions with compound 12 (a, b), Green sticks: amino acid residues; Yellow sticks: ligand; Dashed lines: hydrogen bonds. Red, white, blue, and green colors show oxygen, hydrogen, nitrogen, and carbon moieties respectively.

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Figure 4.

AChE active site interactions with galantamine (a, b), Green sticks: amino acid residues; Yellow sticks: ligand; Dashed lines: hydrogen bonds, Red, white, blue and green colors show oxygen, hydrogen, nitrogen and carbon moieties respectively.

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Compound 3 was the second most active compound against AChE, having an IC₅₀ = 249.52 ± 1.25 µM and a docking score of ₋12.380. This compound established a strong hydrogen bond with TYR124, having a bond length of 2.0 Å. In addition, the carbonyl oxygen of compound 3 forms a secondary hydrogen bond with SER125 (3.5 Å) and three π-π stacking interactions with TYR337 and TRP86 residues of the anionic subsite (Figures 5a and b). However, compound 3 exhibited lower potency than compound 12 and galantamine.

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Figure 5.

AChE active site interactions with compound 3 (a, b), Green sticks: amino acid residues; Yellow sticks: ligand; Dashed lines: hydrogen bonds, Red, white, blue, and green colors show oxygen, hydrogen, nitrogen, and carbon moieties respectively.

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Compound 11 (IC₅₀ value of 345.15 ± 1.47 µM, docking score of -11.842) emerged as the third most active compound among the series of thiazolidinone. Like compounds 12 and 3, the carbonyl oxygen of compound 11 established a hydrogen interaction with SER125 (3.5 Å). Additionally, it established a second strong hydrogen interaction with the TYR124 residue of the anionic subsite, with 2.8 Å (Figures 6a and b). Despite these interactions, compound 11 established lower potency than galantamine, compounds 12 and 3.

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Figure 6.

AChE active site interactions with compound 11 (a, b), Green sticks: amino acid residues; Yellow sticks: ligand; Dashed lines: hydrogen bonds, Red, white, blue and green colors show oxygen, hydrogen, nitrogen and carbon moieties, respectively.

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As a result, our compounds, despite interacting with all four subsites of the AChE active site, precisely interacted with the peripheral anionic site (PAS), anionic subsites, and allosteric subsites by hydrogen bonding, π-π bonding, and hydrophobic interactions. Therefore, docking analysis and kinetic studies are aligned, indicating a non-competitive behavior of our compounds.

Top-ranked compounds (12 and 3) based on high docking scores and lower IC₅₀ values, as well as control (galantamine), were chosen for further 100 ns simulation. The assessment of protein-ligand complex stability was calculated by means of root mean square deviation (RMSD). The RMSD curve of compound 12 was initially unstable and was followed by stabilization from 50-100 ns, shown by values from 0.20 to 0.3 nm. Similarly, the compound 3 complexes demonstrated a comparable trend, with instability up to 50 ns and sustaining stability till 100 ns. Our findings show that the stability and reliability of the compound 12-AChE complex were slightly better than those of the compound 3-AChE complex. The galantamine maintained the lowest value of RMSD, around 0.20-0.25nm, indicating the highest stability with minimal conformational drift when compared to the other two compounds, 12 and 3 (Figure 7(a)).

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Figure 7.

(a) RMSD, (b) RMSF (c) Rg, and (d) Hydrogen bonding plots of compound 12 (Black), compound 3 (Red), and Galantamine (Green) 100 ns in MD simulations.

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The root mean square fluctuation (RSMF) analysis was performed to calculate the flexibility of each residue in the AChE when complex with compounds 12 and 3 during MD simulations. The current investigation revealed that both complexes exhibited higher fluctuations in the highly flexible regions of the AChE, specifically in the N-terminal, C-terminal, and loop regions. In addition, residues contributing to α, β sheets and active sites establish decreased RMSF, as represented in Figure 7(b). These results are in line with the inherent mobility of these regions, which is known to support the structural flexibility of the enzyme during ligand binding and catalysis [31].

The radius of gyration (R_g) predicts the conformational changes and compactness values of the chosen protein-ligand complex. R_g values for the compound 12-AChE complex were observed in the range of 2.28 nm to 2.32 nm, which displayed the compactness of our protein and remained unchanged upon compound 12 binding. The compound 3-AChE complex exhibited identical values, which may show natural movement or rotations contained by the molecular structure. The galantamine curve at the end showed some conformational change, but overall, the Rg remains stable, suggesting that the structure retains its compactness without major unfolding (Figure 7(c)). Rg results indicate that the overall protein structure remains stable and compact during the MD simulation process, with minor fluctuations in the dimension relating to dynamic flexibility.

We further calculated hydrogen bonds formed between our compounds and AChE; compound 12 established a maximum of 3 H-bonds, and compound 3 formed 2 H-bonds with AChE during 100 ns MD simulations; galantamine exhibited 4 H-bonds, overall the results indicate that compound 12 formed a more stable complex with AChE than compound 3 but less stable complex as compared to galantamine as represented in Figure 7(d). H-bonds play a significant role in the stabilization of protein-ligand complexes by providing specific interactions that improve the binding affinity and structural integrity. The larger number of H-bonds in the compound 12-AChE complex shows stronger and more sustained interactions between the active residues of AChE and the compound. This may help explain its enhanced binding stability and inhibitory effectiveness. On the other hand, the compound 3-AChE complex smaller number of H-bonds, indicating comparatively weaker interactions, which could lead to reduced stability and efficiency.

3.5. Antispasmodic and Ca²⁺ antagonistic activities

Antispasmodic drugs are frequently used to treat spasms in the smooth muscles of the gastrointestinal tract [32]. Among these, the most commonly used drugs are those with anticholinergic properties, which act by blocking the activity of ACh, a neurotransmitter involved in muscle spasms. This mechanism efficiently reduces muscle contraction and eases the discomfort they cause [33]. Therefore, to analyze the anticholinergic characteristics of thiazolidinone derivatives, we also studied the potential of these compounds for putative muscle anti-contractility effect using the animal jejunum. This approach was adopted because the jejunum has a natural contraction ability, exclusive of utilizing any agonist. This helps explicitly investigate new drugs with smooth muscle relaxant activity [34]. Nevertheless, stimulants of smooth muscles can also be used, and their effects are compared with standard spasmogenic agents responsible for spasmogenicity. Current study results of the thiazolidinone series also exhibited suppression of spontaneous jejunal contractility (Table S1). Compounds, specifically 12 and 3 showed good spasmolytic activity with an ED₅₀ of 0.13±0.5 mg/mL (Mean ± SEM; n=3) among all compounds (Figures 8a and b).

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Figure 8.

Dose-response graphs illustrating the antispasmodic impact of (a) compound 12 and (b) 3 on both spontaneous/natural and K⁺ induced contractions in jejunal tissues (Mean ± SEM, n=3).

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The spasm of the rabbit jejunum and other smooth muscles is an indication of the increased level of Ca²⁺in the cytoplasm [35], which plays a key role in the interactions of actin and myosin leading to muscle contraction [36]. The initial elevated level of Ca²⁺is the direct consequence of Ca²⁺influx via voltage-gated channels (VDCs) into a cell or its release from the intracellular stores of Ca⁺². This depolarization and repolarization, consistent with the influx of Ca²⁺by VDCs, bring forth the intestine’s spontaneous contractile behavior [37]. In the present study, all Thiazolidinone derivatives were able to relax this contractile behavior of rabbit jejunal smooth muscles, probably by the inhibition of Ca²⁺entry via VDCs or by affecting the Ca²⁺release from intracellular Ca²⁺stores (Table S1). In conclusion, among all tested compounds, compounds 12 and 3 exhibited the most potent activities with median ED₅₀ values of 0.25±0.03 µg/mL and 0.26 ± 0.03 µg/mL (Mean ± SEM; n = 3) on K⁺ (80 mM) (Figures 8a and b).

3.6. Cytotoxicity evaluation of thiazolidinone (1-12)

Thiazolidinone compounds were subjected to cytotoxicity using a human neutrophil viability assay. All compounds of thiazolidinone (1-12) were nontoxic, like standard galantamine (Table S2). Among the whole series, compounds 12 and 3 showed the highest (94.07 ± 1.0 and 92.07 ± 1.4) percent cell viability at conc 200 µg/mL, as displayed in Figure 9.

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Figure 9.

A bar chart shows the percentage viability of human neutrophils upon exposure to the thiazolidinone derivatives (1-12). The error bars represent the standard deviation resulting from the triplicate experiments.

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Scheme 1.

Synthesis of 4-thiazolidinone derivatives. R= Aromatic or Aliphatic aldehyde.

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