Nowadays, health professionals are seriously concerned regarding the escalating prevalence of diabetes mellitus in both developing and developed countries. World Health Organization (WHO) stats reveal that 30 million people were diagnosed with diabetes in the year 1985, which increased to 135 million by 1995. It has been further projected that nearly 300 million population will be suffering from diabetes by the year 2025 [1]. This rise in the prevalence of diabetes is causing substantial financial problems in healthcare settings of underdeveloped countries [2, 3]. There are two main categories of diabetes: type-I accounts for nearly 5 to 10% of cases, while type-II diabetes accounts for approximately 90% of cases [4]. Impaired secretion of insulin results in type-I diabetes, whereas insulin resistance causes type-II diabetes [5, 6]. Hyperglycemia is a state associated with uncontrolled diabetes that results in elevation of blood sugar levels. Prolonged hyperglycemic conditions may result in significant deterioration of different body organs, specifically blood vessels and the central nervous system. Presently, diversified synthetic drugs have been developed for the management of diabetes, such as meglitinides, biguanides, incretin mimetics, thiazolidinediones, sodium/glucose co-transporter 2 (SGLT2) inhibitors, and dipeptidyl peptidase-IV inhibitors [7]. Even though various developed anti-diabetic drugs have demonstrated significant results, they are associated with different side effects [8]. These adverse effects include visual complications, headache, diarrhea, impotency, and hypoglycemia [9]. Therefore, research and development of innovative anti-diabetic medications that have significant effects against diabetes along with the least adverse effects remains a vital opportunity for scientists [10, 11]. For this purpose, investigations on alternative natural medications have attained significant importance owing to their safe nature. Various folk medications, such as administration of different herbal plants and their resultant extracts, have been efficient alternatives to synthetic drugs in managing diabetes. Herbal plants have been a potential source of novel drug development, as nearly 90% of pharmaceuticals are directly or indirectly derived from them [12, 13].

Alzheimer’s disease (AD) is characterized as a neurodegenerative ailment known for its deteriorating effect on the central nervous system [14]. AD is related to different factors such as neuroinflammation, oxidative stress, aggregation of amyloids, and cholinergic deficiency [15, 16, 17, 18]. To date, the underlying origin and optimum treatment of AD is still unclear and unidentified, while managing AD might include targeting its associated conditions. Effective management of the progression of AD can be possible by significantly dealing with oxidative stress, cholinergic deficiency, aggregation of amyloids, and neuroinflammation [19]. Mostly, the treatments of AD focus on easing the symptoms associated with its progression rather than providing an ultimate cure. These therapies help in improving cognitive functionality, managing behavioral symptoms, and improving overall quality of life [19]. Even though these treatments are of significant benefits, continual research to hunt for a definitive cure is needed [20]. As a result, recent investigations focus on exploring improved and alternative therapies for effective treatment of neurodegenerative ailments. Acetylcholine-a, neurotransmitter that is consumed by cholinergic neurons via the central nervous system, plays a vital role in normal functionality [21]. However, acetylcholinesterase (AChE), an enzyme, is considered to be responsible for the reduction of acetylcholine (ACh), hence causing problems in neurotransmission. Recently, AChE inhibitors have been used as an effective alternative approach for the management of AD as they elevate the levels of acetylcholine in the brain [22]. Various investigations have shown the anti-cholinesterase potential of different plant extracts. In spite of recent advancements in the field of modern medication, plant constituents play a significant part in the health and well-being of the community. Pharmaceutical companies are increasingly interested in exploring higher plants as potential sources for discovering new lead compounds and developing effective drugs. This renewed interest highlights the potential therapeutic value of plant-based medicines in addressing various health conditions [23].

Inflammation is a complex biological process that plays a significant role in the development and progression of various disorders, including arthritis and cardiovascular disease [24]. The current approach for symptomatic treatment of inflammation primarily relies on nonsteroidal anti-inflammatory drugs (NSAIDs), which exert their anti-inflammatory effects by inhibiting cyclooxygenase (COX). COX exists in two isoforms: cyclooxygenase-1 (COX-1), which is constitutively expressed in most cells under normal conditions, and cyclooxygenase-2 (COX-2), which is induced by pro-inflammatory agents like tumor necrosis factor-$\alpha$ (TNF-$\alpha$), Lipopolysaccharide (LPS), and tumor-promoting factors [25]. As a consequence, novel dual COX-2/5-LOX inhibitors with anti-inflammatory properties have been identified through investigations involving both natural and synthetic sources. In an effort to contribute to advancements in this area, studies have been conducted to provide a comprehensive summary of the structural characteristics and pharmacological activities of heterocyclic scaffolds and natural products that have been investigated as dual COX-2/5-LOX inhibitors [26]. Another limitation of COX inhibition is that it can lead to an increased production of leukotrienes (LT) by 5-lipoxygenase (5-LOX). While 5-lipoxygenase (5-LOX) inhibitors have demonstrated a protective effect, their usage is hindered by several limitations. These limitations include poor bioavailability, lack of inhibitor specificity, potential hepatic and renal abnormalities, myelosuppression, gastrointestinal disturbances, and an inability to provide protection in chronic inflammation models [27]. The complex and potentially adverse side effects associated with COX and 5-LOX inhibitors have indeed restricted their long-term use as treatments for inflammatory disorders [28]. The limitations and risks associated with these inhibitors highlight the need for alternative therapeutic approaches that can effectively manage inflammation with a more favorable side effect profile.

Ficus benghalensis, a member of the Moraceae family, is widely recognized as the Banyan tree in English. It is native to a vast region of Asia. This plant is grown in several botanical gardens throughout different tropical areas across the world [29]. Studies have been performed to assess the phytochemistry and health-promoting benefits of F. benghalensis. Nevertheless, very few investigations have been conducted on leaves of F. benghalensis, which is of great importance owing to its medicinal applications [30]. F. benghalensis was reported to own different health-promoting benefits such as antibacterial [31], anti-diabetic, anticancer, and anti-inflammatory [32]. Moreover, it has demonstrated beneficial effects against skin ailments, gastric ulcers, and other gastrointestinal problems [33]. F. benghalensis methanolic extract was found to be effective against inhibiting acetylcholinesterase activity [21]. The composition of the leaves of F. benghalensis reveals the presence of crude protein (9.63%), Crude fiber (2.53%), phosphorous (0.4%), and calcium oxalate (2.53%). Phytochemical screening of leaves shows the presence of tannins, sterols, phenolic acids, saponins, and flavonoids. On the other hand, compounds like triterpenoids, volatile oils, and aromatic acids were not present in this plant [30]. In Ayurveda medications, this plant has been consumed for treating piles, dysentery, and diarrhea. Additionally, it is also employed as a hypoglycemic agent, indicating its potential in managing blood sugar levels [34]. Different extracts of Ficus bengalensis were assessed for their potential anti-allergic and anti-stress effects in asthma using milk-induced leucocytosis and milk-induced eosinophilia as indicators [35]. Literature reveals the potential of different Ficus species as anti-diabetic, antimicrobial, anticancer, and anti-inflammatory activities [36, 37]. The current study is unique as we have compared the in-vitro anti-diabetic, anti-cholinesterase, and anti-inflammatory potential of extracts and sub-fractions of different parts of F. bengalensis. In light of this, current study was designed to evaluate the in-vitro anti-diabetic [$\alpha$-amylase and $\beta$-glucosidase inhibition], anti-inflammatory (5-LOX and COX-2 inhibition), and anti-cholinesterase (AChE and butyrylcholinesterase [BChE] inhibition) activity of different extracts and subfractions from stem, leaves, and roots of F. bengalensis.

The global population’s reliance on plant-based remedies is on the rise, primarily attributed to their easy accessibility and affordability [43, 44]. Consequently, researchers are persistently engaged in the pursuit of significant natural-based remedies that can be employed in the treatment, diagnosis, mitigation, or prevention of various disorders [45]. Validating the therapeutic potential of these readily accessible natural products holds the promise of offering more cost-effective treatment alternatives, particularly in light of the prevailing high inflation rates globally. It is worth noting that the pharmacodynamics of many drugs are associated with the inhibition of enzymes present in various biological compartments. Local physicians have recognized the therapeutic properties of the Ficus plant. The local physicians have long recognized the therapeutic properties of various parts of the F. benghalensis plant. The milky fluid obtained from the plant is known for its external application in alleviating pain, rheumatism, bruises, backaches, and swollen soles of the feet. In India, the roots of F. benghalensis are traditionally used to treat conditions such as dysentery, biliousness, gonorrhea, and liver swelling. Additionally, the aerial roots and tips of the plant are employed for their medicinal benefits in the treatment of dysentery and vomiting [46]. The infusion of small branches is consumed to alleviate hemoptysis, while the bark is believed to possess potent tonic properties and is used as a cure for diabetes [47]. Different components of F. benghalensis are employed in the treatment of diarrhea, leucorrhea, wound healing, and skin diseases [48]. According to folkloric practices, the aerial parts of F. benghalensis are employed to alleviate persistent vomiting and as an anti-asthmatic remedy [49]. Additionally, F. benghalensis leaves and stems have been documented for their therapeutic applications in various ailments [50]. However, there is a limited exploration of the phytochemical composition of this plant. Some compounds identified in its leaves include $\beta$-sitosterol, psoralen, $\beta$-amyrin, quercetin-3-galactoside, leucopelargonon, rutin, and leucodelphinidin derivatives [51].

The present study was performed to assess the anti-cholinesterase, anti-inflammatory, and anti-diabetic properties of extracts/fractions of F. benghalensis. In-vitro anti-diabetic assay revealed a significant [p 0.05] concentration-dependent reduction in the activity of $\alpha$-amylase and $\alpha$-glucosidase due to the application of different extracts and fractions of F. benghalensis. In $\alpha$-glucosidase and $\alpha$-amylase assays, as compared to standard (Acarbose, IC${}_{50}$: 26.58 and 21.30 µg mL${}^{-1}$), the IC${}_{50}$ values of different extracts and fractions were in the range of 50.50–474.83 and 91.85–337.4 µg mL${}^{-1}$, respectively. The results of our study are in accordance with the findings of Blickle in 2008 [51]. They reported that the bark of F. benghalensis significantly inhibited the activity of both $\alpha$-glucosidase and $\alpha$-amylase [52]. Glucosidases play a vital role in various biological processes, including the breakdown of dietary carbohydrates [53]. $\alpha$-glucosidase is one of several glucosidases found on the brush-border surface membrane of intestinal cells, and it plays a crucial role in the digestion of carbohydrates [54]. Daniel et al. [55] in 1998 reported that the presence of phenolics and flavonoids in F. benghalensis bark contributes to its inhibitory effect on $\alpha$-glucosidase. These compounds possess potential antioxidant activity, and various investigations have shown that phenolic-enriched extracts of F. benghalensis demonstrate moderate free radical scavenging ability and strong inhibitory activity against $\alpha$-glucosidase. In a study conducted by Madiwalar et al. [56], it was reported that 17 phytoconstituents derived from F. benghalensis demonstrated blood glucose-lowering effects. Notably, the highest drug-likeness score has been shown by 4-methoxybenzoic acid, while maximum (–8.02 kcal/mol) binding affinity was demonstrated by lupeol acetate. Lupeol acetate formed nine pi-interactions with Ala451, Ile319, Phe323, Phe24, Ile200, Tyr324, and Ile28. Additionally, the extract displayed the most significant glucose uptake efficacy in yeast cells at a concentration of 500 µg/mL [56]. Aqueous extract of F. bengalensis bark had inhibitory (IC${}_{50}$: 4.4 µg/mL) effects of $\alpha$-amylase as experimented by Ponnusamy et al. [57].

In this study, the extracts and fractions of F. benghalensis were also examined for their potent anti-inflammatory properties. These two assays, namely COX-2 and 5-LOX inhibitory assays, were performed in this study. Results of both assays demonstrated a significant [p 0.05] reduction in the activity of COX-2 and 5-LOX activity. According to a study conducted by Kothapalli et al. [58], extracts of F. benghalensis leaves showed potential anti-inflammatory activity [58]. Moreover, an in-vivo study further investigated the anti-inflammatory potential of F. bengalensis bark extract in a rat model. They administrated varied content of extract [50–200 mg/kg] of F. bengalensis bark in carrageenan-induced paw edema. Results showed that administration of this extract reduced the inflammation in a concentration-dependent manner. They were of the view that tannins and flavonoids present in the bark extract of F. bengalensis were responsible for this anti-inflammatory potential. Oxidative stress induced by induction of carrageenan injection was also reduced on the application of experimented extracts [59]. Likewise, bark and leaves extract of F. curtipes—another species of genus Ficus-have been reported to downregulate the activity of 5-LOX in a dose-dependent manner. The bark extract of F. curtipes significantly inhibited 5-LOX activity with an IC${}_{50}$ value of 10.75 µg/mL. The inhibitory potential of bark was reported to be more owing to the presence of more phenolic compounds in stem (5374.1 mg/kg dry extract) as compared to leaves (611.5 mg/kg dry extract) [60]. In general, phenolic constituents are recognized as predominant inhibitors of 5-LOX activity [61]. The results of these studies are in accordance with the findings of the current study, which revealed the inhibitory potential of F. benghalensis extracts and fractions against the activity of 5-LOX and COX-2. Bengalensinone and benganoic acid have been isolated from roots of F. benghalensis and possessed inhibitory (164.5 and 154.5 µM) action against acetylcholinesterase, respectively. Both these compounds inhibited the activity of butyrylcholinesterase with an IC${}_{50}$ value of 224.9 and 120 µM, respectively [62]. Moreover, Hassan et al. [21] have shown the anti-acetylcholinesterase (IC${}_{50}$: 194.6 µg mL${}^{-1}$) activity of F. benghalensis.

Phytochemical profiling of F. benghalensis extracts have indicated the presence of diverse phytomolecules like steroids, tannins, flavonoids, alkaloids, anthraquinones, and glycosides [63]. Roots extract of F. benghalensis inhibited the activity of acetylcholinesterase in an in-vitro model. The ethyl acetate extract of roots showed inhibitory action with an IC${}_{50}$ value of 67 µg/mL as compared to the control (Donepezil), i.e., 33 µg/mL. The results of this study were comparable to the outcomes reported by Ramasamy et al. [63]. Moreover, Hassan et al. [21] 2020 also assessed the in-vitro AChE inhibitory potential of extracts (methanolic), fractions (ethyl acetate), and isolated compounds of F. benghalensis.

The findings of this study revealed the in-vitro potential of Ficus benghalensis extracts and fractions in various pharmacological activities. The leaf extract exhibited the highest alpha-glucosidase and alpha-amylase inhibitory activities among all the tested samples, indicating its potential as an anti-diabetic agent. Furthermore, the roots, leaves, and stem extracts, as well as the derived fractions, exhibited significant anti-cholinesterase activity, highlighting their potential for the management of neurodegenerative diseases. Additionally, the F-B-3 C fraction showed the highest COX-2 inhibitory activity, suggesting its potential as an anti-inflammatory agent. Overall, this study provides valuable insights into the medicinal potential of F. benghalensis and supports its traditional use in the treatment of diabetes, neurodegenerative diseases, and inflammatory conditions. Further studies are warranted to identify and characterize the active compounds responsible for these observed bioactivities and to evaluate their in-vivo efficacy and safety profiles.

The data generated in the present study are included in the figures and/or tables of this article.

AR was responsible for conceptualization, visualization, supervision, project administration and writing-original draft. MIbr and MIri contributed by conducting formal analysis, interpreting data, investigation and writing-original draft. AAK, NA, TSA, TK and MUK contributed by writing-original draft, analyzing, data curation, reviewing and editing of manuscript. KB, MSJ, and RS made significant revisions, visualization, and editorial changes. All authors read and approved the final manuscript. All authors have participated sufficiently in the work and agreed to be accountable for all aspects of the work.

Plant samples of F. benghalensis were gathered from Anbar Swabi. The collected specimen was taken to the Department of Botany, where it was examined and identified by Dr. Muhammad Ilyas, a member of the Botany Department at the University of Swabi. The voucher specimen number UOS-BOT/103 was then placed in the herbarium of the aforementioned department.

Authors would like to acknowledge Princess Nourah bint Abdulrahman University, Riyadh, Saudi Arabia for supporting this study.

Princess Nourah bint Abdulrahman University Researchers Supporting Project number [PNURSP2024R18], Princess Nourah bint Abdulrahman University, Riyadh, Saudi Arabia.

The authors declare no conflict of interest. Given his role as Guest Editor and Editorial Board Member of Frontiers in Bioscience-Landmark, Marcello Iriti had no involvement in the peer-review of this article and has no access to information regarding its peer review. Full responsibility for the editorial process for this article was delegated to Jen-Tsung Chen.