Myricetin: A comprehensive review on its biological potentials

Abstract Myricetin is a critical nutritive component of diet providing immunological protection and beneficial for maintaining good health. It is found in fruits, vegetables, tea, and wine. The families Myricaceae, Polygonaceae, Primulaceae, Pinaceae, and Anacardiaceae are the richest sources of myricetin. Different researchers explored the therapeutic potential of this valuable constituent such as anticancer, antidiabetic, antiobesity, cardiovascular protection, osteoporosis protection, anti‐inflammatory, and hepatoprotective. In addition to these, the compound has been tested for cancer and diabetic mellitus during clinical trials. Health benefits of myricetin are related to its impact on different cell processes, such as apoptosis, glycolysis, cell cycle, energy balance, lipid level, serum protein concentrations, and osteoclastogenesis. This review explored the potential health benefits of myricetin with a specific emphasis on its mechanism of action, considering the most updated and novel findings in the field.


| INTRODUC TI ON
Natural products might be effective, new, and safe therapeutic agents if properly tested. In the current era, ample of drug molecules have their roots in natural products. The exploration of natural products against various preclinical cell or animal models can afford the emergence of novel drug candidates. Myricetin is found in many plant families including Myricaceae, Polygonaceae, Primulaceae, Pinaceae, and Anacardiaceae (Abd El-kader et al., 2013;Borck et al., 2018).
It is predominantly present in fruits, vegetables, berries, teas, and wine. Myricetin has been found as free molecule or glycosidically bound such as myricetin-3-Oβd-galactopyranoside, myricetin-  . Myricetin is less hydrophilic (16.6 µg/ml) but has good solubility in basic aqueous and in some organic media such as tetrahydrofuran, dimethylacetamide, acetone, and acetone dimethylformamide (Chang et al., 2012). Moreover, study on the degradation of this compound showed that it is quite stable even at pH 2, but it can differ with temperature. In the late eighteenth century, this compound was isolated from the bark of the Myrica nagi Thunb. A light yellow-colored crystals namely Myricaceae were harvested in India in the past (Perki, 1911). At first, the basic reason behind isolation was its coloring properties. Previously, Perkin (1902) carried out structural and physical properties elucidation and observed it contain various analogs such as ethyl, methyl, bromo, and potassium with melting point of 357°C. Myricetin glycoside (myricetin-3-O-rhamnoside) was also found for the first time in this study (Perkin, 1911). Hydrolysis of myricetin resulted in phloroglucinol and gallic acid production, this also resulted to confirm its chemical structure. The structure of the myricetin (1) is related to many other phenolic compounds such as quercetin (3), morin (4), kaempferol (5), and fisetin (6). Because of its structural similarity, myricetin is also named as hydroxyquercetin (3). Literature provides strong evidence about the nutraceuticals and antioxidant properties of myricetin (Lin & Huang, 2012). It also showed many pharmacological activities such as hepatoprotective, antitumour, anti-inflammatory, analgesic, and antidiabetic. The significant antioxidant activity of myricetin is attributed to the presence of three hydroxyl groups on ring B as compared to other flavonoids. The mineral chelation is attributed to the double-bonded oxygen group along with the two hydroxyl groups as shown in in vitro studies. Prooxidative effect of the myricetin is due to the catechol groups in its structure which forms semi-quinone radicals. This radical is oxidized when both 4-hydroxyl on the B and 4-hydroxyl group on the C ring form a quinine. There are many health benefits attributed to the myricetin such as inhibited hyperglycemia, decreased hepatic triglyceride, reduced oxidative stress and cholesterol contents, and protected liver injury (Chang et al., 2012;Choi et al., 2014;Guo et al., 2015;Semwal et al., 2016).
A study conducted by Dang explored the bioavailability and pharmacokinetic properties of myricetin using ultraperformance liquid chromatography-tandem mass spectrometry method. Myricetin was administrated orally and intravenous to the rats at dose-dependent manner, later on their bioavailability in blood was also determined (Dang et al., 2014). The results showed that the bioavailability of the myricetin by oral route was less due to the scarce absorption (9.62 and 9.74% at oral doses of 50 and 100 mg/kg, respectively).
Maximum concentrations (C max) and area under the cure (AUC) of myricetin increased after oral administration which was proportional to the dose which shows that the myricetin is absorbed by the passive diffusion in vivo. Longer time of achieving maximum concentration (T max ) (6.4 hr) indicated its low aqueous solubility.
Pharmacokinetic properties and bioavailability of co-administered drugs along with myricetin have been studied. The inhibition of the cytochrome P450s (drug metabolizing cytochrome) and P glycoproteins (drug efflux pumps) is a recognized mechanism of myricetin (Li et al., 2011). However, absorption of the myricetin was increased by 41.8%-74.4% and 48.4%-81.7%, respectively, by the combination of myricetin with tamoxifen as compared to the control group in which only tamoxifen was used. Co-administration of myricetin with losartan was also found to induce metabolism (Choi et al., 2010) ( Figure 1).

| HE ALTH PROPERTIE S
Myricetin possesses interesting pharmacological potentials such as anticancer, antidiabetic, and anti-inflammatory activities. This paragraph summarizes the existing literature on myricetin offering a comprehensive review on health effects of such molecule. Table 1 reports the fundamental works and the potential mechanisms of action of myricetin (Table 1).

| Anticancer effect
Myricetin has been found a significant inhibitor of migration, invasion, and adhesion and could reduce the matrix metalloproteinase (MMP-2/9) activities and mRNA levels of ST6 N-Acetylgalactosaminide Alpha-2,6-Sialyltransferase 5 genes (ST6GALNAC5) and MDA-Mb-231Br cells in concentration-dependent manner and in animal model when treated with 50 mg/kg dose (Ci et al., 2018). In recent study by  . Similarly, myricetin also had a role in lowering the proliferation rate and apoptotic cell death, in modulating cell cycle, invasion and pro-angiogenic properties via mitogen-activated protein kinase (MAPK) and PI3K/AKT signaling pathways. In addition, myricetin decreased free radicals, peroxidation of lipids, prevented from depletion of glutathione, and loss of membrane potentials in mitochondria in choriocarcinoma cell models (JEG-3 and JAR). Moreover, myricetin augmented the cytosolic Ca 2+ release from the endoplasmic reticulum (ER) linked with ER stress modulation . Human anaplastic thyroid cancer cell (SNU-80 HATC) proliferation has also been significantly reduced by the myricetin up to approximately 70%. A concentration-dependent cell death was observed sub-G 1 . The myricetin (100 μM) increased in the ratio of the Bax:Bcl-2 and caspase cascades . It was also found that myricetin activated apoptosis through apoptosis-inducing factors found in the cytosol and mitochondria and by disturbing the membrane potential (Jo et al., 2017). Earlier study reported by Jose pathways, reduction of IL-6 and PGE2 (pro-inflammatory cytokines) in blood, and downregulation of the p38 MAPK/Akt/mTOR pathways . Anti-leukemia activity of myricetin was reported by interfering with biosynthetic pathway of purine nucleotides by inhibiting inosine 5'-monophosphate dehydrogenase (hIMPDH) 1/2 catalytic activity (Mondal et al., 2016).
Researchers determined the anticancer role of myricetin against human MCF-7 cells by applying different doses at (0-80 µM) for 12, 24 and 48 hr (Pujari & Mishra, 2021). Multiple cell pathways were involved with consequent cell viability reduction, apoptosis induction, suppression of protein expression of p21-activated kinase 1 and phosphorylated extracellular MAPK and activation of Bax protein expression, GSK3β, promotion of caspase-3 activity, and suppression of β-catenin and cyclin D1, respectively (Jiao & Zhang, 2016). In another study, myricetin with methyl eugenol (MEG) and cisplatin (CP) in human cervical cancer cells have been found to hinder the growth of cancer cells by inducing apoptosis, dramatically as compared to single drug treatment. The combined treatment showed higher mitochondrial membrane potential loss (ΛΨm) and caspase-3 activity as compared to single drug treatment (Yi et al., 2015). Myricetin (60 μM) has been tested in human glioma U251 cells, inducing apoptosis, enhancing expressions of Bax and Bad levels, lowering levels of Bcl-xl and Bcl-2, and inducing cell cycle arrest in G2/M phase. All of these mechanisms were time-and dose-dependent (Fu et al., 2019). Wang and their colleagues investigated myricetin along with chitosanfunctionalized pluronic P123/F68 micelles. They showed alteration in levels of apoptotic proteins, such as bad, Bcl-2, and bax, in mice (Rajendran et al., 2021). In another study, myricetin exhibited antiproliferative activity in human cancer cells. In this study, their anti-angiogenic effects were investigated with in vitro (HUVEC) and in vivo (CAM) models, which showed myricetin inhibited an- factor-1α (HIF-1α), p-Akt and p-70S6K protein levels . In another study by Feng and colleagues, cell cycle arrest, induction of apoptosis and reduction of cell proliferation as well as regulation of related proteins GC HGC-27 and SGC7901 cells were observed after myricetin treatment. The phytochemical also exhibited strong binding affinity for RSK2 that resulted in a prompt expression of the Mad1 (Feng et al., 2015). In hepatocellular carcinoma cell line, myricetin suppressed the p21-activated kinase 1 (PAK1) in Ras signaling pathway and activated intrinsic caspase-mediated apoptosis.
This resulted in reduced expression of survivin and anti-apoptotic Bcl-2, as well as pro-apoptotic Bax increase. MAPK/ERK and PI3K/ AKT signaling pathways were blocked along with downstream Wnt/ β-catenin pathway (Iyer et al., 2015).

Myricetin
Ong and Khoo (2000) In vivo Lowered glycemia up to 50% after 2 days of treatment at 3 mg/12 hr

| Antidiabetic effects
Since it is an antidiabetic agent, myricetin (200 mg/kg/day) was found to be effective in the treatment of mice having dilated cardiomyopathy (DCM)-associated cardiac injury. In this trial, significant alleviation in apoptosis interstitial fibrosis and cardiac hypertrophy was observed during 6-month treatment. Also, significant stimulation of Nrf2/HO-1 pathway was demonstrated. Similarly, GPx and superoxide dismutase (SOD) activities were reversed, and malondialdehyde (MDA) production diminished resulting in increased antioxidative stress capacity. In addition, inflammation process was modulated, given the inhibition of IκBα/NF-κB pathway that resulted in cytokines (TNFα, IL-6, and IL-1β) decrement. Consistently, TGFβ/ Smad3 pathway was suppressed in DCM mice treated with myricetin. Bax and caspase-3 decrements showed the advantageous effects of myricetin treatment on cardiomyocytes. Neonatal rat cardiomyocytes (NRCM) treated with high glucose have also been tested using myricetin with similar results. It was also noted that regulation of IκBα/NFκB by myricetin was independent by suppressing Nrf2 in NRCM (Liao et al., 2017). Another work explored the potential effects of myricetin as a natural GPCR (G protein-coupled receptor) agonist to treat T2DM (type 2 diabetes mellitus). The possible mechanism associated with GPCR agonistic effect of myricetin is the stimulation of secondary messenger (cAMP), which further

Biological effects Mechanisms of action References
Antiobesity effects Improve hepatic steatosis and systemic insulin resistance along with body weight reduction Modulated thermogenic regulation proteins Enhanced the thermogenic protein expression and beige formation (Hu et al., 2018).
Lowered levels of protein expression phosphorylated AKT serine/threonine kinase 1 (Akt) Yao et al. (2018) Up regulated the β-endorphin and adropin levels Chao et al. (2017) Cardiovascular effects Reduced inflammatory cytokines and inhibited cellular apoptosis Degraded IκBα and nuclear translocation of p65 Prevented from the iNOS overexpression and oxidoreductase activity Zhang et al. (2018) In earlier study, myricetin also has been known to suppress the activator of transcription 1 (STAT1) activation and signal transducer Scarabelli et al. (2009) Normalized the nitric oxide, endothelin nitric oxide synthase, serum high-density lipoprotein cholesterol, and prostaglandin I2 levels (Guo et al., 2015).
Lowered the lactate dehydrogenase and creatine kinase levels Reduced the infarct size and cardiomyocyte apoptosis levels Reduced levels of MDA and increased levels of GSH/GSSG ratio Downregulated the p38, cytochrome P450, and cyclooxygenase−2 Upregulated fatty acid synthase and 6-phosphogluconate dehydrogenase Qiu et al. (2017) Anti-inflammatory effects Decreased the keratinocyte death, and COX2, IκB/NFκB expressions Xie and Zheng (2017) Suppressed the NF-κB, IL−6-12, NO, iNOS, TNFα (inflammatory mediators), binding activity NF-κB DNA Altered the NF-κB, IκBα, phosphorylation of STAT1 and production Cho et al. (2016); Latief et al. (2015) Suppressed the pro-inflammatory mediators, that is, TNFα-stimulated creation, Akt, mTOR and NF-κB Lee and Lee (2016) Hepatoprotective effects Lowered the AST and ALT levels Regulated by the myricetin such as caspase−3/9 and P53 protein, mitogen-activated protein kinase, nuclear factor-kappa B (NF-κB) activation inhibition of toll-like receptor 4 (TLR4), heme oxygenase−1 (HO−1), increase and enhanced expression of Nrf2 (nuclear factor-erythroid 2-related factor 2) Lowered AMPK/ACC signaling and activated Keap1-Nrf2/HO−1 Enhanced the HO−1 and Nrf2 protein expressions Lv et al. (2020) Hampered phosphorylation of Smad2, type I deposition by suppression of α-smooth muscle actin and collagen Geng et al. (2017) Lowered miR−146b expression to elevate TRb levels Xia et al. (2019) Osteoporosis prevention Increased body weight gain, upregulated osteocalcin Improved alkaline phosphate activity, and inhibited reduction Lowered tartrate-resistant acid phosphatase and C-terminal telopeptide of type I collagen levels Enhanced the osteopontin (OPN) levels, collagen type I alpha 1, COL1A1, ALP, Runx2, BMP2 and OCN Fan et al. (2018) Prevented from bone resorption Increased alveolar rest height Huang et al. (2016) TA B L E 3 (Continued) triggers protein kinase A/C (PKA/PKC) and then transcriptional factors (TF) are activated (TFa) the TFa then internalized to the nucleus and producing special mRNA with special protein coding. Once the mRNA is released to the cytoplasm, initiating the protein synthesis, these special proteins are transplanted act the cell surface in the form of glucose transporters (Glut) which stimulate the glucose uptake. This glucose uptake is responsible for hypoglycemic actions.
Glucagon-like peptide-1(7-36) amide (GLP-1) is a secreted peptide that acts as a key determinant of blood glucose homeostasis by virtue of its abilities to slow gastric emptying, to enhance pancreatic insulin secretion, and to suppress pancreatic glucagon secretion.
Glucagon-pike peptide-1 (GLP-1) is secreted from enteroendocrine cells (L cells) of the gastrointestinal mucosa in response to a meal, and the blood glucose-lowering action of GLP-1 is terminated due to its enzymatic degradation by dipeptidyl-peptidase-IV (DPP-IV).
Released GLP-1 activates enteric and autonomic reflexes while also circulating as an incretin hormone to control endocrine pancreas function . The supplementation of myricetin (770 μg/ ml) prevented the postprandial hyperglycemia through inhibiting the α-amylase and α-glucosidase activities (Meng et al., 2016).
Choline also lowered NO, eNOS, serum HDL cholesterol and prostaglandin I2 levels whereas administration of myricetin (400 and 800 mg per kg bw) to experimental mice normalized these changes (Guo et al., 2015). Subcutaneously isoproterenol given two time a day at a dose of 85 mg/kg along remarkably enhanced the concentration of cardiac marker enzymes, that is, lactate dehydrogenase, creatine kinase and aspartate aminotransferase in serum, and lowered the levels of catalase and dismutase: when myricetin was given at a dose ranging from 100 to 300 mg/kg p.o., it reverted these changes in animals (Tiwari et al., 2009)

| Anti-inflammatory effects
The COX-I is distributed throughout the body tissues as compared to COX-II, moreover the COX-II is over expressed at the site of inflammation as compared to COX-I. The suppression or block- Production of the reactive oxygen is actually inhibited by the induction of the myricetin. Moreover, myricetin also proved to change skin inflammatory disease by suppressing the pro-inflammatory mediators. Another study showed that myricetin could inhibit NF-κB, IL-6, IL-12, NO, iNOS, TNFα (inflammatory mediators), could modulate the binding activity of NF-κB to DNA and degradation of the p65 NF-κB subunit, phosphorylation of STAT1 in LPS-and IFNβ-stimulated RAW264.7 macrophages. Similarly, Nrf2/HO-1 system resulted modified by the use of myricetin (Cho et al., 2016;Latief et al., 2015).
A study on the iNOS in C57B16/J knockout male Swiss mice Mouse bone marrow (LPS-stimulated)-derived dendritic cells (DCs) treated with 10 µg/ml myricetin resulted in a significant reduction in TNFα secretion, IL-12 and IL-6. Histocompatibility CD40, CD86 and class II was also blocked by the flavonoid as well as migratory and endocytic capacity of DC cells. Moreover, T-cell proliferation by LPS-stimulated DC-elicited allogene was abolished by myricetin (Fu et al., 2019;Hagenacker, Hillebrand, Wissmann, et al., 2010).

Different works on IL-1β-stimulated SW982 synovial cells proved
anti-inflammatory role of myricetin along with the reduction in production of MMP-1 and IL-6, weakening the phosphorylation of p38 MAPK and Jun NH2-terminal kinase (Lee & Lee, 2016).

| Hepatoprotective effects
Hepatoprotective effect of myricetin is well documented by different researchers all over the world. D-GalN (D-galactosamine) and LPS-induced fulminant hepatitis: myricetin as proved by the lower mortality rate in animals could reduce such inflammation process.
Other markers such as AST and ALT levels were also decreased along with reduced oxidative stress, hepatic apoptosis, histopathological changes and inflammation. In addition, several metabolic key factors were mediated and regulated by myricetin, including caspase-

| Osteoporosis prevention
The beneficial effect of myricetin against osteoporosis could be due to its inhibitory effect on osteoclastogenesis and elevation of osteo-

| Other properties
Several therapeutic effects can be associated with the use of myricetin. One of them is its beneficial effect against cataract development thank to the strong aldose reductase inhibition, as proved in a study conducted on the galactosemic rats by using 1% dose of the myricetin (Mohan et al., 1988). Hodges and coworkers showed that in an in vivo study a dose of 1 mg/kg, i.v. could the intraocular pressure in normotensive rabbits, suggesting as myricetin was noticeably beneficial for treatment of glaucoma (Hodges et al., 1999). Similarly, in human retinal pigment epithelial cells, myricetin at different levels (10, 20, 50 and 100 µM) reduced cell proliferation and migration, as well as the Vascular endothelial growth factor (VEGF) secretion . At low concentration, myricetin reduced VEGF expression whereas at high concentration increased VEGF. Similarly, high doses affected cell viability inducing necrosis. Myricetin could also trigger caspase-3 independent retinal pigment epithelial cell necrosis by the activation of phospholipase A2 and calpain as well as by producing free radicals. Myricetin seems to have a role also in coagulation cascade and platelet aggregation. In rabbit platelets, Zang et al. (2003) found inhibitory effect of myricetin on specific receptors bindings of PAF (platelet activating factor). Myricetin con- found for the rabbit platelet adhesion. The polyphenol was also effective on neutrophil elastase and thrombin activity, with IC50 values of 7 and 28 µM, respectively (Melzig & Henke, 2005). Different IC50 values such as 17.5-64.1 µM inhibited the agglomeration and release of PAF-induced serotonin. Contrarily, lower concentration (7.9 µM) possessed no effect on the release of serotonin from platelets (Chen et al., 2001). Another research showed that myricetin could hamper thrombin production which made it very useful in the thrombotic disease treatment (Liu et al., 2010). Increased levels of platelet adenosine 3',5'-cyclic monophosphate (cyclic-AMP) stimulated by prostacyclin was triggered by myricetin. The detailed mechanism behind the anti-aggregating activity was the modification in the metabolism of platelet cyclic-AMP followed the phosphodiesterase activity inhibition.
A study conducted on cat blood showed that platelet aggregation was stopped by the intravenous administration of the myricetin at 3.6 µg/kg body weight. Moreover in in vitro experiment 60 nM of myricetin could disaggregate platelet thrombi. Myricetin could bind to platelet membranes and inhibit the formation of prostacyclin synthase reducing radicals of the oxygen, for example, singlet oxygen, hydroxyl radical (¨OH), superoxide anion radical (O ⋅ ⋅ 2 ) and perhydroxyl radical (HO 2 ). In addition, PGE2 levels in peritoneal fluid (reduced by myricetin) were decreased, eliciting less platelet aggregation (Tong et al., 2009).
Also, neuroprotective potential of myricetin was explored. A rat model was used to investigate the effect of 0.1-10 mg/kg i.p. myricetin for its remarkable effect on the neuropathic pain. Thermal hyperalgesia and mechanical allodynia were induced by reducing spinal nerve ligation for several hours Hagenacker, Hillebrand, Wissmann, et al., 2010). They showed 18%-78% decrease of IK(V) in an in vitro study by using nerve cell at concentration pf 1-75 µM. Reduction in the IK(V) was found to be dependent on p38 modulation. Significant analgesic effect was produced by myricetin as proved by the writhing test induced by acetic acid and time to lick during the late phase of formalin test. A 10-100 mg/kg i.p. dose of myricetin in the mice's hind paws was experimented in bradykinin-induced nociception assay. The dose of 100 mg/kg provoked a 57% reduction in the cinnamaldehyde-induced nociception. Significant reduction in acidified saline-induced nociceptive responses by 30-100 mg/kg, i.p. dose of myricetin (Córdova et al., 2011).
Not only for antinociceptive or neurological protection was tested myricetin, but also for its possible role as antimicrobial com-  (Pasetto et al., 2014;Yuan et al., 2012). Similarly, studies on its antibacterial property revealed that it was effective against multidrug-resistant Burkholderia cepacia, vancomycin-resistant Enterococci, methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis (Xu et al., 2016). DnaB helicase, a bacterial replicative factor, was found to be the main target for antibacterial activity of myricetin. DnaB helicase along with primase is the main part during DNA replication and elongation in Escherichia coli was noncompetitively inhibited (IC50 = 11.3 lM) by myricetin (Griep et al., 2007;Lin & Huang, 2012). Stimulation of the epithelial Cl-secretion by myricetin was found usefulness in the prevention of viral/bacterial infection. Its effect was different from quercetin as this last could trigger epithelial Cl-secretion merely under basal environment in epithelial A6 cells whereas myricetin could trigger it under basal as well as under cAMP-stimulated conditions (Sun et al., 2014).

| CON CLUS I ON S AND FUTURE PER S PEC TIVE S
Myricetin shows great therapeutic potential especially against cancer, T2DM, liver injury, cardiovascular diseases, obesity and osteoporosis. These health benefits are proved by several in vitro and in vivo studies as previously reported. Convincingly, myricetin can induce apoptosis and can inhibit invasion, migration, adhesion along.
It also interacts with several intracellular pathways, such as those related to insulin signaling, energy production. Moreover, infarct size and cardiomyocyte apoptosis levels were also found to be reduced by myricetin. Similarly, cytoprotective and inflammatory mediators have also been increased by myricetin, providing health benefits related to heart disorders and inflammation. Other studies suggested that hepatic biomarkers were modulated together with reduced oxidative stress and histopathological changes of the liver. Bone health was also demonstrated to be improved by myricetin, which was able to fight osteoporosis. In addition to these pharmacological activities, further studies especially based on a mechanistic approach are essential. These structure activity relationship (SAR) might be helpful to find a significant derivative of myricetin. Further, acute and chronic toxicological data on vital organs are of extreme importance.
The clinical trial studies on myricetin are limited. According to a clinical survey, the consumption of myricetin can lead to low incidence of prostate cancer risk. Another clinical trial on lung cancer reported that the regular consumption of myricetin was associated with lung cancer decreased incidence. The consumption of myricetin along with other flavonoids by menopausal women resulted in a reduced risk of CHD (coronary heart diseases). The study also reported a dropping in sera TG, LDL and apolipoproteins. Moreover hyperglycemic levels were significantly normalized in T2DM patients. However, these clinical trials cannot be considered sufficient. Keeping in view the broad potential of this valuable compound, it is strongly recommended that other randomized double blinded clinical trials could be urgently conducted. Only after such research and deepen analysis, myricetin will be ready for market and we can look forward to a new tool for fight human diseases.

ACK N OWLED G EM ENT
The authors are grateful to Higher Education Commission Islamabad, Pakistan, and Universities for providing facilities to carry out this activity.

CO N FLI C T O F I NTE R E S T
There is no conflict of interest among authors.

DATA AVA I L A B I L I T Y S TAT E M E N T
Data available on request from the authors.