Antidiabetic Potential of Phytochemicals Found in Vernonia amygdalina

. Type 2 diabetes mellitus (T2DM), or “insulin-independent diabetes mellitus,” is a worldwide health concern. Diabetes afects roughly 415 million individuals worldwide, with 193 million undiagnosed cases. Te number of people aficted in the following decades is predicted to double. Although various synthetic medications are currently available to treat/manage T2DM, their side efects compel researchers to seek novel treatment options. Because of their afnity for biological receptors and broad bioactivity, nature has long been a source of innovative medication. V. amygdalina is one of the numerous natural productswith antidiabetic properties. Several studies have shown that the extracts have antidiabetic efects in vitro and in vivo. Tis review examined the antidiabetic and pharmacokinetic characteristics of phytoconstituents found in V. amygdalina .


Introduction
Diabetes, which is characterized by the body's difculty in efectively regulating blood sugar and insulin levels, is one of the major silent killers worldwide.In 2021, more than 537 million people between the ages of 20 and 79 were diagnosed with diabetes mellitus globally.It is estimated that 643 million people are expected to have diabetes by 2030 and a staggering 783 million by 2045 [1].Te expenditure on diabetes is rapidly increasing worldwide, and it varies signifcantly from one country to another.In India, the number of diabetic cases has seen a signifcant upsurge (7.1% in 2009 to 8.9% in 2019) in the last few decades, which is predicted to rise in the coming years, and will hugely enhance the burden on fnance as well as the healthcare sector [2].-Type 2 diabetes mellitus (T2DM) is brought on by the interaction of a genetic predisposition and a wide range of modifable and nonmodifable environmental risk factors, including obesity, a poor diet, and inactivity [3][4][5][6].Additionally, diabetes does not attack end-organ damage alone.For instance, cardiovascular disease (CVD) progresses, and eyesight loss and kidney failure might result from the vascular and nerve damage brought on by chronic hyperglycemia.Te main factor in preventable blindness is diabetes-induced retinopathy, while the main factor in kidney disease is diabetic nephropathy.Terefore, diabetes must be managed appropriately to reduce the emergence of chronic problems.Accordingly, the current therapeutic strategy includes a variety of medication classes.However, their adverse interactions and inadequate efcacy compelling researchers to develop safe alternatives.Cragg-Newman found that out of the 63 antidiabetic genre drugs approved between January 1, 1981, and September 30, 2019, biological macromolecules occupied the largest portion ( 38%), followed by synthetic drugs inspired by natural products(25.4%),and unmodifed natural products and their derivatives (14.3%).Tis data strongly highlights the importance of natural products [7].
Vernonia amygdalina (V.amygdalina) is a member of the Asteraceae family with dark green leaves and tiny, thistle-shaped fowers.Tis plant grows primarily in tropical Africa and is used as an herb and vegetable.Earlier studies suggest that this vegetable demonstrates a strong anticancer efect against breast cancer cell lines via inhibiting the DNA synthesis [8].Today, various classes of phytochemicals (e.g., sesquiterpene lactones, steroid glycosides, and saponins) are known from this plant, making it useful for the management of human and animal diseases [9][10][11].Tis review makes an earnest efort to put together its promise as a diabetes treatment, the gaps in existing research, and its potential future possibilities.Te survey of the existing literature has been conducted using the PubMed and SCOPUS databases.Te selected search period is from 2005 to 2022.

Type 2 Diabetes Mellitus (T2DM): Molecular Pathology and Medications
Although T2DM has been known to humans for ages, its complex nature and the lack of clarity in understanding the mechanistic aspects pose signifcant challenge.Initially thought to be a chromosomal polygene recessive condition with abnormal insulin secretion rates, the inheritance mode of type 2 diabetes (T2DM) remains uncertain.It is now understood that a complex interplay of genetic factors and external infuences, such as stress and chemicals, contributes to its development.Disruptions in glucose metabolism, incretin production, and insulin sensitivity involving key enzymes like α-glucosidase, α-amylase, DPP-4 (dipeptidyl peptidase 4), PPAR (peroxisome proliferator-activated receptor), PTP1B (protein tyrosine phosphatase 1B), and GLUT4 (glucose transporter type 4) play a central role in T2DM pathophysiology.α-amylase breaks down starch into oligosaccharides known as limit dextrins [12], which are further hydrolyzed into absorbable glucose in the small intestine by α-glucosidase.Elevated blood glucose levels stimulate the synthesis of glucagon-like peptide-1 (GLP-1), which aids in insulin secretion and pancreatic cell protection.However, GLP-1 is rapidly degraded by DPP-4, disrupting glucose metabolism [13].Insulin also infuences lipid metabolism and glycogen production in the liver and muscles, necessitating regulation through the GLUT4, PTP1B, and PPAR pathways [13].GLUT4 is crucial for postmeal glucose clearance, but altering its expression can disturb glucose homeostasis and promote insulin resistance [14].Insulin-dependent GLUT4 translocation involves a complex cascade of biochemical signals, including tyrosine kinase activation, IRS1 substrate phosphorylation, and PI3K/Akt, CAP/Cb1/Tc10, or Ras-MAPK pathway activation.Simultaneously, GLUT4 vesicles move from intracellular pools to the cell membrane to facilitate glucose uptake [15].Leptin enhances glucose absorption and energy metabolism by binding to its receptors and phosphorylating the JAK2/STAT pathway.PTP1B, found in the endoplasmic reticulum, skeletal muscle, and liver [16], dephosphorylates and inactivates these signal transduction cascades, making it a potential target against T2DM [17].Non-insulin dependent GLUT4 translocation can be induced by factors like severe exercise, increased Ca 2+ , and bradykinin [18], which stimulate GLUT4 translocation to the plasma membrane, aiding in efcient glucose control [19].Peroxisome proliferator-activated receptor gamma (PPAR) plays a critical role in adipocyte development and is involved in lipid and glucose metabolism regulation [20].Activating PPAR improves GLUT-4 and GLUT-1 mobility, facilitating glucose uptake in skeletal muscles and the liver.Infammation is closely linked to obesity and T2DM [21,22], with PPAR agonists showing promise in improving insulin sensitivity by reducing TNF-α [23] and increasing adiponectin expression [24].Infammatory substances like TNF-α impact insulin resistance through NF-κB signaling pathways, making this pathway a potential target for diabetes control [25].Diabetes can increase tissue sensitivity to damage from reactive oxygen species (ROS) and free radicals, leading to oxidative liver damage and decreased antioxidant enzyme function [25].Tis exacerbates insulin resistance and pancreatic cell dysfunction.Epigenetic alterations in DNA and histones are being explored through omics, including proteomics, genomics, and transcriptomics, to gain a better understanding of T2DM into pathogenesis [26,27].Despite these advancements, insights into the pathological mechanisms of T2DM continue to drive the discovery of novel drugs.
To treat T2DM, synthetic agents of various classes are available.Tis includes, but is not limited to, glimepiride (class: sulfonylurea), rosiglitazone (class: thiazolidinedione), metformin (class: biguanide), dapaglifozin (SGLT2 inhibitor) [28].However, side efects associated with the available synthetic agents are a major concern.For example, rosiglitazone is a thiazolidinedione-based compound that enhances insulin's efectiveness by promoting hepatic function.Nevertheless, there have been reports of cardiac failure as an adverse outcome.Similarly, metformin is a biguanide that acts by decreasing hepatic glucose output [28].In addition to synthetic agents, natural product-based antidiabetic drugs are also used globally [29].
Numerous studies have indicated that the organic and aqueous extracts of diferent parts of V. amygdalina plant possess unique bioactivities [49].For example, the ethanolic extract of the leaves has been found to suppress infammatory factors like TNF, IL8, and IL6 [50].Te saponins found in V. amygdalina exhibit selective cytotoxicity against cancerous cell lines [47].Following this work, Ejiofor et al. [51] studied the antidiabetic, anthelminthic, and antioxidant properties of the methanolic extract V. amygdalina stem bark, which contains phytochemicals responsible for the bitter taste [31].Sesquiterpene vernodalin and vernomygdin have been reported to show anticancer activity against nasopharynx cell lines [42].Anh et al. [8] reported the antidiabetic properties of V. amygdalina leaves.Tey found that the stigmastane type saponin vernoamyoside E shows remarkable antidiabetic property.Similarly, vernoniacum B was found to be mildly active against α amylase.[8].

Antidiabetic Potential of V. amygdalina Extracts
In 1992, Akah and Okafor [52] reported the antidiabetic efect of V. amygdalina extract.Tey noted that the intraperitoneal injection of aqueous leaf extract in rabbits leads to insulin secretion-independent hypoglycemia.However, later studies found that V. amygdalina extracts assist in β-cell regrowth.It was noted that when V. amygdalina was given intragastrically to alloxanized rats for two weeks, its blood and serum glucose levels reduced signifcantly as compared to the control [53,54].Histomorphological analysis revealed the regeneration β-cell regrowth, which is quite an interesting fnding.Another clinical study revealed that V. amygdalina produced signifcantly higher hypoglycaemic efects than the other vegetables at the majority of postprandial time points [55].Saliu et al. [56] investigated the inhibitory impact of free and bound phenol extracts of it on glucosidase and amylase activities in vitro.Tey found a signifcant inhibition in amylase and glucosidase activities in a dose-dependent manner (4−16 g/ml).It was noted that glucosidase had a more signifcant inhibitory efect than amylase [56].To understand how the combined extracts of V. amygdalina and Gongronema latifolium afect the pancreatic β-cells, Akpaso et al. [54] conducted a study on streptozotocin-induced diabetic Wistar rats.Tey noted that, among other changes, there was a decrease in blood glucose by 12.49% and 14.96% during a 28-day treatment period compared to diabetes control.Te synergistic efect of V. amygdalina extract on the antidiabetic activity of a synthetic drug has also been studied.Michael et al. [53] investigated the antidiabetic activity of an admix of metformin and an aqueous extract of V. amygdalina leaves.Tey found that a mixture in a ratio of 1 : 2 signifcantly reduced the blood sugar levels, while a 2 : 1 mixture was found to be better when taking drug safety into account [53].Following this study, Atangwho et al. [57] studied the efect of a herbal combination of V. amygdalina and Azadirachta indica and concluded that the use of combined extract is a useful way to control the glucose level [57].Ethanolic extract of V. amygdalina was found to improve glucose tolerance in animal models, and an improvement of 32.1% in fasting blood glucose was seen after a 28-day treatment.Surprisingly, V. amygdalina reduced triglyceride and total cholesterol levels by 18.2% and 41%, respectively, and shielded pancreatic β-cells from STZ-related harm.In addition, it was determined that polyphenols were the main candidates for mediating V. amygdalina's antihyperglycemic efect by boosting GLUT 4 translocation and inhibiting hepatic G6Pase.[58].[57] Erasto et al. [59] examined how leaf extract of V. amygdalina afects glucose utilization in different cells (chang-liver cells, C 2 C 12 muscles, and 3T3-L1).Tey found that the extracts, particularly aqueous ones, greatly enhanced glucose utilization in chang-liver cells and C 2 C 12 muscles.Onyibe et al. [60] found that glutathione S transferase (GSR) activity in alloxan-induced diabetic rats was similar to that of rats given metformin [60].Te importance of hepatic gluconeogenesis in glycogen metabolism is well known [61].Wu et al. observed reduced expression of key enzymes involved in gluconeogenesis post-V.amygdalina extract administration by activating the AMPK pathway in palmitic acid-induced HepG2 cells [62].However, they were unable to identify the key mechanism potentiating V. amygdalina's glucose-lowering efect.Ejiofor et al. [51] isolated various active compounds from the methanolic extract of V. amygdalina stem bark and noted that only vernoniaolide-glucoside showed an antidiabetic efect, while others were inactive.Similarly, vernonioside E, a saponin, was recently identifed by Uti et al. as having the antidiabetic component of V. amygdalina [63].
Recently, Djeujo et al. [64] found that polyphenolic chemicals in aqueous extracts of V. amygdalina roots and leaves made them less cytotoxic than ethanolic extracts, which primarily inhibited glucosidase activity.Although those made from leaves showed less efcacy, aqueous root extracts displayed a concentration-dependent activity.In addition, 10 g/ml V. amygdalina extracts were also found to inhibit the production of advanced glycation end products (AGEs) [64].Te fact that phytochemicals, namely polyphenols, are thermally degraded into a combination of distinct isomers and derivatives, which afects the potency of extracts, is one factor that is also implicitly implied by the diferential IC 50 values from diferent extraction procedures.Terefore, it is also essential to consider how to build an efective extraction technique.Hepatoprotective, anticancer, antioxidant, anti-infammatory, analgesic, antibacterial, anti-nociceptive, [30,45] Reproduced under the terms of the CC-BY creative commons attribution 4.0 International license (https://creativecommons.org/licenses/by-sa/4.0/)[35].
One of the important factors controlling insulin secretion is the MAFA protein, whose expression is negatively impacted by uric acid [70].Matsuoka et al. [71].It has been reported that luteolin restores the expression of MafA and suppresses the (NF)-κB-iNOS-NO pathway, preventing the pancreatic β cells from becoming dysfunctional [71].When used in combination with acarbose, luteolin was found to show synergistic inhibition of α-glucosidase [72].Besides, this study also found that luteolin shows respectable activity when compared to other well-known favonoids [72,73].Flavonoids are known to undergo substantial frst-pass phase II metabolism in the liver and epithelial lining of the small intestines to generate methyl, gluconorate, and sulphate-conjugated metabolites [74].Tey are mostly absorbed by the small intestines.Yasuda et al. [75] found that oral administration of Chrysanthemum morifolium extract to rats resulted in relatively rapid absorption and slow elimination of luteolin.Chen et al. [76] reported low excretion of luteolin (6.6%), suggesting probable metabolism of it into simple compounds or in vivo accumulation.However, the biosynthesis of luteolin involves the conversion of apigenin into luteolin.According to a study, luteolin and luteolin glucoside show an oral bioavailability of 10.6-26.6%, and luteolin-7-O-glucoside is primarily hydrolyzed to luteolin in the GI before being absorbed into the systemic circulation [77].Kure et al. [78] reported that the metabolites luteolin 3′O glucuronide, luteolin-7-O-glucoside, and luteolin-4′-O found in the liver, kidney, and small intestine might be the bioactive component.A recent structure-activity relationship (SAR) study on this class of compound revealed that the presence of an OH group at the 3′-4′ location of ring B in luteolin type 1 favone is an important element for greater glucosidase inhibition.Similar to hydrogenation at positions 2 and 3 of ring C, glycosylation at positions 7 and 6 of ring A, and methoxylation at positions 3′ and 4′ of ring B, these modifcations have been found to reduce glucosidase inhibitory activity [79].

Chlorogenic Acid
Derivatives.Chlorogenic acid and related compounds are abundantly found in phenolic acid in V. amygdalina [58].It was found that the chlorogenic acid derivatives reduces the postprandial glucose level [80,81].During a glucose tolerance test, Bassoli et al. [82] observed a decrease in plasma glucose levels due to chlorogenic acid, pointing to its potential involvement as a glycemic indexlowering drug.Chlorogenic acid has been found to improve lipid and glucose metabolism byactivating the AMPK pathway [83].Zheng et al. [84] reported the mixed-type inhibitory activity of chlorogenic acid on swine pancreatic amylase.Oboh et al. suggested that chlorogenic acid inhibited α-glucosidase with an IC 50 value of 9.24 g/ml in an in vitro model [85].In addition, reports have shown that chlorogenic acid inhibits the activity of the enzymes maltase and sucrase, with IC 50 values of 2.99 mM and 2.18 mM, respectively [85,86].Besides, 4,5-dicafeoyl quinic acid and 3,5-dicafeoyl quinic acid are also recognized for their ability to inhibit DPPIV and glucosidase, respectively [87].Some work also reported that 1,5 dicafeoylquinic acid had a dosedependent protective efect against glucotoxicity in RIN-m5F cells [88][89][90].In 2007, Ren et al. [91] administered chlorogenic acid to rats and observed rapid absorption and a relatively slow distribution followed by a slower elimination phase.In 2011, Xie et al. discovered chlorogenic acid  Journal of Chemistry metabolites such as O-methyl CA, hydrolyzed chlorogenic acid, and glucuronide conjugates in rats after intravenous treatment [91].It has been shown that the bioavailability of chlorogenic acid depends on gut microfora metabolism [92].
Keeping in mind the signifcance of the interplay of chemical dynamics among ligands and enzymes, Hemmerle et al. reported chlorogenic acid derivatives and studied their glucose -6-phosphate translocase inhibitory activity, which is one of the key enzymes for glucose homeostasis.Tey established that only one phenolic -OH moiety is enough for the action and increasing lipophilicity at position 1 enhanced the activity [93].Amylase and glucosidase are more strongly inhibited by the acylated derivatives of chlorogenic acid [94].It was observed that their inhibitory actions on enzymes spiraled upward with a positive shift in lipophilicity (mostly) with the highest activity in the C 12 acylated group.Tis could be attributed to a stronger bifurcated hydrophobic interaction within the microenvironment of amino acid residues and secondary structures [94].Te topological polar surface area for cafeic acid is 77.8 Å2, while that for quinic acid is 118 Å2 ; therefore, as the degree of esterifcation of quinic acid with cafeic acid increases, its lipophilicity increases [95].Terefore, chlorogenic acid is inherently less polar than one of its parents, quinic acid.Recently, Song et al. reported that cafeoyl substitution downturned the α amylase inhibitory activity of quinic acid mediated via reduced binding afnity postsubstitution [96].Te efectiveness of these compounds was not as strong as acarbose, but it outperformed another widely recognized glucosidase inhibitor, 1-deoxynojirimycin hydrochloride.Cardullo et al. [97,98] synthesized 11 amide derivatives of chlorogenic acid and found that one particular derivative, which featured a tertiary amine group on an alkyl chain and a benzothiazole scafold, exhibited signifcantly lower IC50 values compared to chlorogenic acid (45.5 µM for α-Glu; 105.2 µM for α-Amy).Tese derivatives were notably more potent as α-glucosidase inhibitors than the antidiabetic medication acarbose, which had an IC 50 = 268.4µM.Besides, amongst its derivatives, the inhibitory efect is a function of the number and position of the cafeoyl group.

Vernonioside E.
Vernonioside E, a stigmastane-type steroid, is a potential antidiabetic candidate.It was found that it imparts hepatoprotective efects, lowers blood glucose levels, improves the lipid profle, and decreases cardiovascular risks.[63] Tis hypoglycemic efect is likely caused by the activation of glucose-6-phosphate dehydrogenase via the shunt pathway, which enhances glucose oxidation, stimulation of insulin, regeneration of pancreatic cells, upregulation of enzymes involved in glucose metabolism, reduction of gluconeogenesis, and inhibition of glucose-6-phosphate and fructose-1, 6-bisphosphatases.However, whether the oxirane moiety, donor H-atoms, or acetoxy group could have a signifcant impact remains unclear.
5.5.Vernoamyoside E. Anh et al. [8] isolated vernonioside B 1 , vernonioside B 2 , vernoniacums B and vernoamyoside E. Vernonioside B 1 , vernoniacums B, and vernoamyoside E have similar skeletons with a diference at C 16 .Vernonioside B 1 , vernoniacum B, and vernoamyoside E have -OH, -OAc, and -H at C 16 of the cyclopentane ring, respectively.Vernoamyoside E was found to outperform the other derivatives and demonstrated a potent inhibitory action against glucosidase at 100 and 500 g/mL concentrations.All of them demonstrated inhibitory action against amylase, though less efectively than acarbose.5.6.11β,13-Dihydrovernolide. Okoduwa et al. [49] observed that 11β,13-dihydrovernolide possessed a hypoglycemic efect, which was attributed to the oxirane ring.However, additional studies are required to determine the mechanism.

Toxicological Studies
Te assessment of the toxicological and safety profles of a natural product is essential, especially when it is prescribed as herbal medicines.In 2016, Jamil et al. [100] reported the positive toxicity of ethanol and hexane extracts of V. anthelmintica in the brine shrimp lethality (BSL) test.Perera and coworkers [101] reported a dose-dependent toxicity of a formulation that contains V. anthelmintica as one of the components.Te BSL assay revealed that the aqueous extract of the formulation shows moderate lethality (>85% at a concentration over 900 μg/mL) with LD 50 = 807.6 ± 221.0 μg/mL.Methanolic and dichloromethane extracts have been found to show positive micronucleus tests, suggesting their clastogenic and or aneuploidic activity [102].However, an independent study found that the intraperitoneal LD 50 of the crude methanolic V. glaberrima extract was 1265 mg/kg, indicating its fairly toxic nature [103].V. amygdalina leaf extract exhibited no observable clinical signs of toxicity or adverse toxicological properties [104,105].It has also been found that the aqueous extract of V. amygdalina was more potent and less cytotoxic than the alcoholic extract [64].Similarly, Autamashih et al. [106] reported that the crude extract of V. galamensis is relatively safe for oral use [106].Table 2 collects the biological profle and mechanism of activity of diferent species of Veronia.
. Future Perspective V. amygdalina extracts, characterized by their remarkable dose-dependent antidiabetic efects, likely mediated by compounds such as favonoids, stigmastane-type steroids, and saponins, have exhibited promising potential.Notably, 6 Journal of Chemistry properties.In addition, synthetic chemists are designing derivatives with improved pharmacodynamic and pharmacokinetic profles [95].While some compounds such as vernoniosides and vernoamyosides may not adhere to Lipinski's rule [112], recent approvals of drugs exceeding these constraints ofer hope for their druggability.Challenges in drug discovery include sourcing natural products in sufcient quantities and ensuring their stability against degradation, which is being addressed through innovative extraction techniques.In summary, V. amygdalina extracts and their phytochemicals hold promise for managing T2DM and its complications.However, further research is needed to bridge the gap between their potential and practical application as therapeutic agents.

Conclusion
Diabetes, in which a human body is unable to deal with an upregulated sugar level, is a global health crisis.Diabetes not only leads to complications like cardiovascular disease, retinopathy, and kidney failure but also poses a signifcant risk to overall health and well-being.Te prevalence of diabetes is on the rise, with a projected increase in cases, leading to substantial economic burdens on healthcare systems worldwide.Among others, major factors that contribute to the increasing incidence of diabetes include the destruction of pancreatic β cells, a poor lifestyle, increased physical activity, and others.Efective diabetes management is crucial to prevent chronic complications and reduce the associated healthcare costs.However, although various therapeutic approaches involving various classes of medications are available, there is a growing interest in the development of natural product-based alternatives.Vernonia amygdalina (V.amygdalina), a plant found primarily in tropical Africa, has garnered attention for its potential medicinal properties.VA is rich in bioactive compounds like sesquiterpene lactones, steroid glycosides, and saponins, which have shown promise for various health benefts, including antimicrobial, anti-infammatory, and blood sugar regulation.However, despite the potential of as a diabetes treatment, its use for clinical applications has not been achieved yet.Tere are various factors in research that need to be addressed.For instance, studies related to mechanisms of action, safety, and efcacy are lacking.In addition, the exploration of natural products like V. amygdalina as potential therapeutic options opens up exciting possibilities for the future of diabetes management.In summary, diabetes is a growing global concern with signifcant economic implications.Vernonia amygdalina, with its rich bioactive compounds, presents a promising avenue for research into alternative diabetes treatments.Addressing these research gaps in research could lead to more efective and safer options for diabetes management in the future.

Table 1 :
Phytoconstituents of V. amygdalina and their medicinal properties.

Table 2 :
Antidiabetic activity of plant parts of Vernonia species.