## 4 Discussion
To validate the enzymatic potential, each compound should be tested by in vitro analysis; however, this could not be possible due to the very limited concentration of these compounds in the plant extract and the commercial unavailability of some compounds. For this reason, these compounds can be evaluated virtually by using in silico molecular docking studies, which will predict the bioactive compounds responsible for enzymatic activities [83]. Nowadays, computational methods such as molecular docking have an important role in drug discovery as they help to predict and recognize low binding energy and high affinity, as well as better frameworks of the protein-ligand interactions. In our study, in silico molecular docking was performed in which hydrogen bonds, in addition to other interactions such as alkyl, pi-alkyl, pi-sigma, amide-pi, etc., play an important role in protein-ligand interactions and give stable binding of ligands and proteins [84]. A total of 29 compounds identified from GC-MS were docked against the \(\alpha\)-glucosidase (PDB: 3 wy1), tyrosinase (PDB: 3 nq1), and acetylcholinesterase- (PDB: 1 gqr) based on binding affinity and the energy level. All 29 compounds from GC-MS analysis along with the standards (Acarbose, Eserine, and Kojic acid) were docked against \(\alpha\)-glucosidase, acetylcholinesteraseerase, and tyrosinase enzymes, respectively. The results showed that 1,2-benzenedicarboxylic acid, 2-hydroxy-n-(2-phenylethyl)benzamide, \(\gamma\)-sitosterol, and lactose were the best-docked compounds with the least binding energy as compared to standards. Among all of these compounds, 1,2-benzenedicarboxylic acid showed the best \(\alpha\)-glucosidase inhibition activity due to the presence of hydrogen, alkyl, pi-pi, and pi-alkyl interactions with a \(-\)8.3 kcal/mol binding affinity as compared to Acarbose (\(-\)8.1 kcal/mol). For acetylcholinesterase-inbibition, \(\gamma\)-sitosterol was the best-docked compound among all with \(-\)11.6 kcal/mol and pi-sigma, pi-pi, and pi-alkyl bonds, while no hydrogen bond was present. The maximum hydrogen bond was observed by lactose with seven hydrogen bonds and \(-\)7.9 kcal/mol binding affinity as compared to the standard Eserin, having \(-\)8.1 kcal/mol binding affinity with only one H-bond. In the case of tyrosinase inhibition, 2-hydroxy-n-(2-phenylethyl)benzamide showed the lowest binding affinity (\(-\)7.9 kcal/mol) with hydrogen, amide-pi, pi-alkyl, pi-pi, and pi-sigma interactions; lactose has the highest number of hydrogen bonds (five), with \(-\)7.1 kcal/mol binding affinity as compared to standard Kojic acid (6.5 kcal/mol). These best-docked compounds were further subjected to ADMET studies to predict their drug-like behavior and toxicity. The SwissADME tool was used to investigate the physiochemical properties, drug-like behavior, and pharmacokinetics of the best-docked compounds [26]. These compounds were checked whether they follow the five rules described by [PERSON]. If a compound follows the rules it can be considered a therapeutic agent and if a compound does not follow more than one rule it is considered an orally unavailable drug [85]. Overall, lactose showed two violations while all the others showed one and even zero violations, rendering them as orally available drugs. A compound is said to have high bioavailability, absorption, and distribution if it has a lower molecular weight, lower hydrogen bond capacity, and lipophilicity. All compounds had high GIT absorption except Phthalic acid, bis-7-methylacetyl ester, lactose, and \(\gamma\)-sitosterol; however, tricylcb[4.3.1.1(3,8)]undecane-1-carboxylic acid, 1,2-benzenedicarboxylic, and _cis_-(\(-\))-carvone-5,6-oxide possessed blood-brain barrier penetration. Lipophilicity is measured in terms of LogP and its optimum value is between \(-\)0.7 to +5.0, and the optimum molecular weight ranges from 150 to 500 g/mol. The TPSA represents the size and polarity of compounds with an optimum range (20 to 130 A\({}^{2}\)). LogS (ESOL) ranges from 0 to 6 and measures solubility.