Introduction of benzyloxy pharmacophore into aryl/heteroaryl chalcone motifs as a new class of monoamine oxidase B inhibitors

The inhibitory action of fifteen benzyloxy ortho/para-substituted chalcones (B1-B15) was evaluated against human monoamine oxidases (hMAOs). All the molecules inhibited hMAO-B isoform more potently than hMAO-A. Furthermore, the majority of the molecules showed strong inhibitory actions against hMAO-B at 10 μM level with residual activities of less than 50%. Compound B10 has an IC50 value of 0.067 μM, making it the most potent inhibitor of hMAO-B, trailed by compound B15 (IC50 = 0.12 μM). The thiophene substituent (B10) in the A-ring exhibited the strongest hMAO-B inhibition structurally, however, increased residue synthesis did not result in a rise in hMAO-B inhibition. In contrast, the benzyl group at the para position of the B-ring displayed more hMAO-B inhibition than the other positions. Compounds B10 and B15 had relatively high selectivity index (SI) values for hMAO-B (504.791 and 287.600, respectively). Ki values of B10 and B15 were 0.030 ± 0.001 and 0.033 ± 0.001 μM, respectively. The reversibility study showed that B10 and B15 were reversible inhibitors of hMAO-B. PAMPA assay manifested that the benzyloxy chalcones (B10 and B15) had a significant permeability and CNS bioavailability with Pe value higher than 4.0 × 10–6 cm/s. Both compounds were stabilized in protein–ligand complexes by the π-π stacking, which enabled them to bind to the hMAO-B enzyme's active site incredibly effectively. The hMAO-B was stabilized by B10- and B15-hMAO-B complexes, with binding energies of − 74.57 and − 87.72 kcal/mol, respectively. Using a genetic algorithm and multiple linear regression, the QSAR model was created. Based on the best 2D and 3D descriptor-based QSAR model, the following statistics were displayed: R2 = 0.9125, Q2loo = 0.8347. These findings imply that B10 and B15 are effective, selective, and reversible hMAO-B inhibitors.


Scientific Reports
| (2022) 12:22404 | https://doi.org/10.1038/s41598-022-26929-x www.nature.com/scientificreports/ the thermodynamically highly persistent trans form is much more prevalent than the cis form. Additionally, this unique conjugated kind of ketone performs as a Michael acceptor in numerous physiological signal transduction cascades in cells 48 . The electrophilic propensity of the enone's entity is driven by the charge distribution discrepancy of the aromatic core induced by the inclusion of diverse substituents 49 . These molecules are shown to possess a broad variety of physiological functions, such as the aptitude to suppress human MAOs (hMAOs). One of the two hMAO isotypes can be inhibited potently as well as preferentially by amending the scaffolding and meticulously considering the substituents inserted on the A and/or B rings. Relying on the linkers appended to the A/B rings, the conventional chalcones having these two aromatic rings as heads are inhibitors of hMAO-B in a low molar range. When compounds like furan and thiophene are incorporated into the A ring, such molecules often benefit from electron-donating lipophilic units coupled to the B ring. The halogens are frequently extensively endured, to varying degrees, and might improve the lipophilicity aiding in encounters in the entry chamber. The nitro and hydroxyl groups on B ring often contribute to a decline in interaction towards hMAO-B and the congestion of hydroxy or methoxy units on the corresponding ring. The inclusion of the hydroxyl group on ring A, on either hand, is effectively sustained, notably if halogens were bound to B ring. This molecule might create hydrogen bonds with tyrosine hydroxyl groups or water molecules within the enzyme cavities, strengthening the enzyme interactions [50][51][52][53] .
In light of these observations, the current work outlines the synthesis of a series of chalcones coupled to benzyloxy moieties and explores the compound's in vitro hMAO-A as well as hMAO-B inhibitory characteristics. The principal compounds were also put through extensive testing in the areas of kinetics, reversibility, blood-brain barrier (BBB) permeation, and molecular dynamics (MD) simulation.

Materials and methods
Synthesis. An equimolar mixture of 0.01 M of various substituted acetophenones, para/ortho-substituted benzyloxy benzaldehyde, 25 mL ethanol, and 7.5 mL 40% KOH was stirred for 20 to 24 h to synthesize the benzyloxy chalcones. Upon pouring the obtained mixture to crushed ice, the yellow solid product was obtained as precipitates which was then filtered by suction. It was dried after being washed with water. Employing methanol or ethanol, the dried products were recrystallized (Scheme 1). To monitor all of the reactions, thin layer chromatography was carried out on pre-coated TLC plates (Silica gel 60-120#) using the solvent system hexane:ethyl acetate (9:1).

Inhibition studies of hMAO-A and hMAO-B. Recombinant hMAO-A and hMAO-B were used in the
hMAO inhibitory activity assay with substrates kynuramine and benzylamine of 0.06 mM and 0.3 mM concentrations, respectively 54,55 . It was determined that the benzylamine-MAO-B Km ranged from 0.25 to 0.33 mM for kinetics. Toloxatone and clorgyline were served as reference hMAO-A inhibitors, while pargyline and Lazabemide had been used as reference hMAO-B inhibitors. From Sigma-Aldrich (St. Louis, MO, USA), enzymes, substrates, and reference chemicals were purchased.
Enzyme inhibition and kinetic studies. GraphPad Prism software 5 was employed to assess the activity at various doses of the compounds and to calculate the IC 50 value for the compound exhibiting a residual activity of less than 50% 56,57 . The inhibition effect was initially tested at a concentration of 10 μM. IC 50 of hMAO-A/IC 50 of hMAO-B was used to obtain the SI value of hMAO-B 58 . The enzyme kinetics of compounds B10 and B15 were assessed using hMAO-B at five different substrate dosages (0.0375-0.6 μM) 59 . The Lineweaver-Burk plots as well as their secondary plots were compared in order to investigate and ascertain the kinetic patterns.
Reversibility analysis of B10 and B15. After preincubation for 30 min at ~ 2 × IC 50 , as previously described, the reversibility of B10 and B15's hMAO-B inhibitions was assessed 60 . The reference for reversible hMAO-B inhibitor, lazabemide, and the reference for irreversible hMAO-B inhibitor, pargyline, were preincubated at ~ 2 × IC 50 (0.22 and 0.28 μM, respectively) for comparing with lead molecules. By contrasting the activities of dialyzed (A D ) and undialyzed (A U ) samples, reversibility patterns were investigated and determined.
Blood-brain barrier (BBB) permeability study. Initial drug research utilized the parallel artificial membrane permeation assay (PAMPA) method to forecast drugs' passive, transcellular permeation via the BBB. A "sandwich" structure was developed in PAMPA using a microtiter plate with 96 wells and a Millipore filter plate with 96 wells (ipvh, 125 μm thick filter, 0.45 μm pore), which was then drenched in 0.1 mL of n-dodecane. Drug sample stock solutions in DMSO were made at 10 mM concentration and retained at 0 °C until utilization. To accomplish a final sample concentration (0.01, 0.1, and 1 mM) and limit the DMSO concentration to 1% (v/v), the stock solution was diluted in buffer at pH 7.4, prior being incorporated into a 96-well filter plate. The donor wells received 270 μL of the final dilutions, while the acceptor well received 200 μL of pH 7.4 buffer. To create a sandwich, the acceptor filter plate was properly positioned atop of the donor plate (comprising of a synthetic lipid membrane in the center, an aqueous receiver on atop, and an aqueous donor carrying an analyte on the bottom). Accessing the acceptor well from across the lipid membrane, the test substance diffuses from the donor well. The penetration supposedly happened with the sandwich intact. The UV spectrometry was employed to quantify the drug concentration in the reference, donor, and receiver wells. The following expression was employed to figure out the rate of penetration 61 .  62,63 . The centroid box was identified using the grid generation technique based on the co-crystallized ligand to define the binding site. The 2D structure of the synthesized B10 and B15 compounds, and low energy 3D conformers with appropriate bond lengths and angles were obtained. At a physiological pH of 7.2 ± 0.2, the potential ionization states for each ligand structure were generated. The Glide module of the Schrödinger module was used to accomplish the docking, while all other parameters were left at their default settings 64,65 .

Molecular dynamic (MD) simulation.
MD studies were performed for the docking poses of synthesized compounds B10 and B15 with the lowest negative scores, i.e., top-docking poses, with an NVIDIA Quadro 6000 graphics processing unit, using the Desmond MD simulation program. More information for MD investigations (box type, thermostat and barometer settings, short-and long-range interactions calculations, etc.) can be found in previous studies because the same settings were used for the examined systems here., 100 ns of MD

Results and discussion
Chemistry. The benzyloxy chalcones were synthesized by the overnight stirring of various substituted acetophenones and para/ortho-substituted benzyloxy benzaldehyde via the Claisen-Schimdt reaction (Scheme 1) [73][74][75][76] . All final derivatives were characterized using 1 H NMR, 13  Inhibition studies of hMAO-A and hMAO-B. All fifteen chalcone derivatives (B1-B15) showed more effective inhibitory activity against hMAO-B than hMAO-A. Experimentally, most of the compounds showed residual activity of < 50% for hMAO-B at a concentration of 10 μM (Table 1). Compound B10 most potently inhibited hMAO-B with an IC 50 value of 0.067 μM, followed by B15 (IC 50 = 0.120 μM). Structurally, the benzylated chalcone in which the A-ring was substituted with thiophene showed the highest hMAO-B inhibition. Compared with the ethoxy group (B15) at the para position of the A-ring, hMAO-B inhibition of B10 was 1.79 times higher than B15. This means that thiophene substitution in the A-ring increases hMAO-B inhibition. In addition, for all compounds, regardless of the A-ring, the benzyloxy group at the para position of the B-ring showed higher hMAO-B inhibition than the ortho position. These results indicated that the benzyloxy group at the para position of the B-ring enhanced MAO-B inhibition. Selectivity index (SI) values for hMAO-B of B10 and B15 were calculated as 504.791 and 287.600, respectively, suggesting that B10 and B15 are potential selective hMAO-B inhibitors.  , respectively, in reversibility tests (Fig. 3). The recovery values of the samples were comparable to lazabemide, a reference inhibitor for hMAO-B that is reversible (from 45.13% to 80.78%), and distinct from pargyline, a reference inhibitor for hMAO-B that is irreversible (from 45.26% to 43.96%). These findings suggested that B10 and B15 were hMAO-B reversible inhibitors.

Structure-activity relationships (SAR).
In the SAR investigation, variously substituted chalcones with phenyl and heterocyclic systems were employed. The study focused primarily on the effects of placing the benzyloxy and different electron-donating and withdrawing groups. When chalcones with a heterocyclic system (B8-B11) were compared to those with a phenyl system in terms of their inhibitory profile, the heterocyclic chalcones exhibited relatively higher hMAO-B inhibition. Contrasting the inhibitory values of methylenedioxy (B1 and B2) and benzodioxane (B3) chalcones, the latter exhibited greater hMAO-B inhibition than the former, www.nature.com/scientificreports/ suggesting that increasing the number of alkyl groups between the two oxygen atoms enhanced hMAO-B inhibition as well as selectivity. Furthermore, the hMAO-B inhibitory ranges of the methylenedioxy chalcones (B1 and B2) displayed that compounds with identical attachments appeared to have vastly differing hMAO-B inhibitory potentials, depending on the position of the benzyloxy group, with the relocation of the benzyloxy group from the ortho (B1) to para position (B2), reducing the hMAO-B inhibitory value to half and intensifying the SI 1.17 times. The para placement of the benzyloxy group in benzodioxane chalcone (B3) raised the hMAO-B inhibitory range thrice higher than that of the methylenedioxy chalcone (B2) with 2.49 times greater selectivity in conjunction to the increased number of alkyl groups. Analogues with bromothiophene ring presented the strong hMAO-B inhibiting heterocyclic chalcone, when the benzyloxy group tethered to the para position (B8). Shifting the benzyloxy group to the ortho position (B9) exhibited the lowest inhibiting chalcone of the heterocyclic series with an SI difference of 45.39-fold. The greatest hMAO-B inhibitory chalcone (B10) of the entire series, with an SI value of 504.791 on anchoring the benzyloxy on para position, was shown by analogues with thiophene rings in comparison. In contrast, its ortho derivative (B11) showed a poor inhibition of hMAO-B and its SI value was 72.99 times lower than that of the para derivative. The inhibitory profiles of both bromothiophene (B8 and B9) and thiophene (B10 and B11) analogues showed how the bromine molecule affects the inhibition value of both para and ortho benzyloxy tethered chalcones, with the para analogues (B8 and B10), showing a variation of 4.97-fold and the ortho analogues (B9 and B11), expressing a difference of 3.09-fold in SI. Contrary to the benchmarks clorgyline (IC 50 = 0.110 µM) and pargyline (IC 50 = 0.140 µM), the highest activity compound (B10) had a low IC 50 value of 0.067 µM.
Similar to this, chalcones with a phenyl system demonstrate a change in IC 50 value when the position of the benzyloxy group was swapped from ortho to para. With an SI variation of 1.41-fold, methyl sulfonyl chalcones showed a lower IC 50 value for para benzyloxy linked molecule (B5) than ortho (B4). The same property was also disclosed by thiophenyl chalcones, but with a greater SI variation of 13.84 times for para (B7) over ortho (B6). The folder SI for trifluoromethyl chalcones differed by 147.05 for the para (B13) vs the ortho (B12) analogues, with the para analogue having a lower IC 50 value of 0.255 µM. In a similar vein, ethoxy chalcone (B14 and B15) had an IC 50 value for its para counterpart (B15) of 0.120 µM, which was 81.58 times more selective than its ortho analogue (B14).
All these inhibitory profiles of chalcones comprising the phenyl system accentuated that an electron-withdrawing and donating group also influenced the molecules along with the para positioning benzyloxy moiety. This highlighted the fact that chalcones with electron-donating groups, such as thiomethyl and ethoxy (B7), displayed stronger hMAO-B inhibitory values than chalcones with electron-withdrawing groups, such as methylsulfonyl and trifluoromethyl (B5 and B13). Especially contrasted to the heterocyclic series, ethoxy and thiomethyl chalcones (B15 and B7) exhibited the greatest hMAO-B IC 50 values. However, the former gave the second-highest IC 50 value of the whole series (IC 50 = 0.120 µM), which was greater than that of the reference standard pargyline. According to the SAR, chalcones with para versus ortho positioned benzyloxy groups, the former had a greater influence. Additionally, the electron-donating group had a more beneficial impact on chalcone activity than the electron-withdrawing group (Fig. 4). The summarized SAR of the present study is depicted in Fig. 4.
Parallel artificial membrane permeability assay (PAMPA) for blood-brain barrier (BBB) permeation study. PAMPA manifested that the benzyloxy chalcones (B10 and B15) had a significant permeability and CNS bioavailability with Pe value higher than 4.0 × 10 -6 cm/s ( Table 2). Brain penetration is acritical need for effective CNS medication delivery. In this study, PAMPA-BBB was used to assess the brain penetration of all derivatives. The rate of permeation was calculated using the equation and the compound's effective permeability (Log Pe). Molecules exhibiting a Pe value of lower than 2.0 × 10 -6 cm/s were categorized as possibly non-BBB permeable (CNS-), whereas compounds exhibiting a Pe value of more than 4.0 × 10 -6 cm/s were designated as potentially permeable (CNS+). www.nature.com/scientificreports/ Absorption, distribution, metabolism, and excretion (ADME) properties. A molecule must exhibit significant biological activity at minimal therapeutic doses, being low in toxic effects, and be effective till the intended result is achieved in order to be considered an efficacious medication. For a superior pharmacokinetic profile, the ADME features of drug prospects are taken into accounts during the process of drug discovery. Using online databases like SwissADME (http:// www. swiss adme. ch/) 78 and pkCSM (http:// biosig. unime lb. edu. au/ pkcsm/), the pharmacokinetic properties were estimated in silico 79 . The benzyloxy chalcones ADME characteristics were listed (Table 3). Gastrointestinal permeability and dissolution measurements were used to assess the absorption of the drug. The solubility of proposed molecules spanned from − 5.90 to − 7.165 in aqueous system and was expressed as the logarithm of molar concentration. The percentage absorption of the compounds was evaluated based on the majority of a drug's absorption by the small intestine when taken orally. Since Caco-2 from sentient colon cancer mimics intestine epithelial cells, it is often possible to anticipate the consumption of oral medicines based on Caco-2 permeability. To achieve superior permeability, the compound must have a P app value greater than 8 × 10 -6 cm/s. Oddly, all of the substances had high permeability. Each of the molecules exhibited substantial gastrointestinal absorption, ranging from 90 to 95%. Using a volume of distribution (VDss), fraction unbound, and BBB permeability, the drug's distribution profile was projected. If log VDss > 0.45, it suggests that the drug is distributed more widely in the tissues than in the plasma. In the tissues, each component is dispersed at a moderate to low level. Drug effectiveness measured by fraction bound suggests that it is less bound to blood proteins and is hence free to distribute. Both SwissADME and pkSCM were used to evaluate the BBB permeability. For neurodegenerative therapies, BBB permeability is crucial. The lead chemical B10 had a log BB value of 0.46, indicating that it may easily pass through the BBB. Molecules with a log BB value of 1 are poorly  www.nature.com/scientificreports/ distributed in the brain. The CNS is thought to be penetrable by compounds with a log PS > − 2, whereas those with a log PS < − 3 are thought to be ineffective. All of the substances tested in this study exhibited CNS penetration, therefore. Every molecule has some sort of interaction with cytochromes, whether it be as an inhibitor or a substrate. The study revealed that all benzyloxy chalcones had a reduced total clearance of − 0.196 to 0.755 logml/min/kg. B10 and B15 both exhibited total clearances of − 0.053 and 0.251 logml/min/kg, respectively. All of the compounds had favorable ADME characteristics, rendering them all plausible contender.

Molecular docking.
Compounds B10 and B15 were identified to be the most active derivatives in the hMAO-B enzyme inhibition series, as seen in the MAO inhibition assay studies. Accordingly, docking studies were conducted to assess the molecular interactions between the compounds and MAO-B as well as their potential to inhibit the enzyme in silico. The interactions between the compounds and hMAO-B binding pocket are shown in Fig. 5. The SP docking scores for compounds B10 and B15 were − 9.954 and − 10.852 kcal/mol, respectively, which were comparable to the cognate ligand (− 10.167 kcal/mol) present in the crystal structure. A detailed analysis of the docking poses of compounds B10 and B15 at the active site of hMAO-B revealed that they were located in the 'aromatic enclosure' delineated by Leu64, Leu171, Gly434, Tyr60, Tyr326, Leu328, Pro104, Phe103, Pro102, and Phe99, as illustrated in Fig. 5. The π-π stacking interaction via the phenyl rings of Tyr326 (3.75 Å) and Tyr398 (3.69 Å) with the thiophene and chalcone aromatic rings of B10, which is the most important interaction in the putative orientation of the hMAO-B inhibitor. In the case of B15, two hydrogen bond interactions with Tyr188 and Ser59 were visible at 3.65 Å and 3.62 Å. The B15 chalcone aromatic ring unit was buried in a large hydrophobic pocket surrounded by Val173, Cys172, Leu171, Phe168, Leu167, Leu164, Pro104, Phe103, Pro102, Phe99, Tyr326, and Leu328. Through π-π stacking and van der Waals interaction, the benzene ring of both promising compounds interacts with the hydrophobic residue Tyr398 phenyl ring. Interaction with the Tyr398 was required for catalytic activity, and binding of inhibitor candidates in the hMAO-B enzyme's substrate cavity facilitates hMAO-B enzyme inhibition 20 . These data suggest that compounds B10 and B15 bind extremely and efficiently to the active site of hMAO-B enzyme.

MD simulation.
The MD simulation is a prominent and popularly used method implemented in recent days in drug development research for enabling the comprehension of energetic details about protein and ligand interactions in a time-affordable fashion. It does this by reproducing the nearly precise or realistic dynamic behavior of a protein-ligand complex. Here, all-atoms classical MD simulations were run for 100 ns on each complex to examine the binding stability at the atomic level and clarify the dynamic properties of the promising hit inhibitors inside the hMAO-B binding cavity. A variety of characteristics from the MD simulation trajectories, including protein backbone RMSD, RMSF, radius of gyration (RGyr), PCA analysis, and binding free energy, were examined in order to assess the stability and flexibility of each protein-ligand complex.
Root-mean-square deviation (RMSD). One of the essential metrics that describes fluctuations in structural conformation of the protein backbone over time during system equilibration is the RMSD value acquired from the MD simulation trajectory, and low and consistent RMSD values show the protein structure's stability. To analyze the ligand-protein interaction, the protein should approach equilibrium, that is, the RMSD value should remain steady [80][81][82] . Figure 6 shows the RMSD computed for the two complexes based on the protein backbone using the Simulation interaction diagram tool. The average RMSD values were: (a) hMAO-B Apo protein = 2.232 ± 0.29 Å,    Principle component analysis (PCA). The PCA method was used to understand conformational distribution during the simulation time and investigate large-scale collective motions of the protein in protein-ligand complexes on the trajectories generated by simulations. The Essential dynamics (ED) analysis script of the Desmond program (trj_essential_dynamics.py) was used through the command line for predicting the dynamic behaviors of a protein 88 . This script calculates the principal components of the protein Cα atoms. The complex that occupies less phase space with a stable cluster was assumed to be more stable, whereas the complex that takes up more space with a nonstable cluster was assumed to be less stable 89 . First two principal components (PC1 and PC2) were selected to analyze their projection of trajectories during the simulations of compounds bound to hMAO-B protein in the phase space. The results clearly showed that the drug-protein complexes, B10-and B15-hMAO-B, occupied smaller regions of phase space (Fig. 9). The trajectories' centering inside a single cluster suggested Binding free energy analysis through MM-GBSA approach. A binding free energy analysis using the MM-GBSA approach was performed on both the protein-ligand complexes to analyze binding affinities of compounds B10 and B15 to hMAO-B protein. The MM-GBSA-based binding free energy (∆G Bind ) computations were performed on the100 ns long MDS trajectories. The binding energies assessed by this method were more efficient and precise, when compared to the binding energies determined in the molecular docking study 90 . The entire trajectories for 100 ns were used for the study, and the average ∆G Bind results are shown in Table 4. The main energy   The QSARINS 3 parametric model is currently in development. The descriptor VE3_DzE stands for the logarithmic coefficient sum of the final eigenvector from the Barysz matrix/weighted by Sanderson electronegativities. This description and the activity have negative correlations. The descriptor TPSA stands for the sum of solvent accessible surface areas of atoms having absolute values of partial charges greater than or equal to 0.2. A strong relationship exists between this description and the action. Number of para-hydroxylation sites is represented by the RDKit abbreviation fr_para_hydroxylation. The association between this descriptor and bioactivity is unfavorable.
For model 9, Fig. 10 shows graphs of experimental vs. projected pIC 50 values, an Insubria plot, a William's plot, a Y-scrambling plot, and an Insubria plot for model 9. In Table 5, there is additional information on the whole statistical analysis. Additional proof for the GA-MLR QSAR model's statistical robustness was supplied by its various cross-validation qualities (R2 cv, RMSE cv , MAE cv , CCC cv , and Q2 LMO). Greater results for the Tropsha and Golbraikh criterion, Q 2 -F 1 , Q 2 -F 2 , CCC ex , and Q 2 -F 3 demonstrated the external predictive power of the suggested models 91,92 . All models for statics and included descriptors are in Supporting information, Table S3.
It would be able to identify the reasons for variations in the MAO inhibitory effect of chalcones inhibitors based on benzyloxy pharmacophore by developing QSAR models using a variety of molecular descriptors. Although the current QSAR models have their limitations, more descriptor computation data, accurate modelling, and less statistical artefacts could lead to the development of better models. As a result, each model created here demonstrates the integration of all chosen chemical characteristics and forecasts future pIC 50 values for the aforementioned analogues.

Conclusions
Fifteen benzyloxy chalcones (B1-B15) were synthesized and their effectiveness to inhibit hMAO was evaluated in this study. Notably, contrasted to the reference drugs, the majority of the compounds had a significant selective hMAO-B inhibitory activity. With an IC 50 value of 0.067 µM, B10 demonstrated the strongest inhibitory action against hMAO-B, trailed by B15 (IC 50 = 0.120 µM). B10 and B15 were demonstrated to be competitive and reversible inhibitors of hMAO-B by kinetic and reversibility experiments. In a permeation investigation, B10 and B15 exhibited great BBB penetration. Novel insights into the binding modalities of the hMAO-B inhibitor-binding cavity were revealed by MD experiments. In aspects of binding to the hMAO-B enzyme's catalytic domain, both compounds were incredibly potent. Thus, the hMAO-B enzyme was stabilized by B10-and B15-hMAO-B complexes of higher binding affinities. Additionally, using the descriptors from RDKit and PaDEL, we created a QSAR model. A good balance of external predictive ability was present in the developed QSAR model. The developed model was successful in revealing not only the obvious correlation between structural features but www.nature.com/scientificreports/ also the hidden correlation between structural features and biological activity. The research also anticipated that introducing halogens to the chalcone framework's benzyloxy pharmacophore could augment MAO-B inhibition. This study, therefore, implies that B10 and B15 have therapeutic promise for the treatment of different neurodegenerative illnesses, such as AD and PD. For the lead molecules, in vivo experiments such as hMAO inhibitory activity in cell system including cytotoxicity and neuroprotective activity using OHDA-induced model for PD should be needed in the future study.