Product name: Eupatilin
Synonym name:
Catalogue No.: BP0576
Cas No.: 22368-21-4
Formula: C₁₈H₁₆O₇
Mol Weight: 344.319
Botanical Source: Eupatorium semiserratum, Tanacetum vulgare (tansy) and Liatris punctata
Physical Description: Yellow powder
Type of Compound: Flavonoids

Purity: 95%~99%
Analysis Method: HPLC-DAD or/and HPLC-ELSD
Identification Method: Mass, NMR
Packing: Brown vial or HDPE plastic bottle

Storage: Store in a well closed container, protected from air and light. Put into refrigerate or freeze for long term storage.
Whenever possible, you should prepare and use solutions on the same day. However, if you need to make up stock solutions in advance, we recommend that you store the solution as aliquots in tightly sealed vials at -20℃. Generally, these will be useable for up to two weeks.

The product could be supplied from milligrams to grams, up to kilograms
Inquire for bulk scale.

Descriptions:
Eupatilinthe has anti-proliferative effect in MCF10A- ras cells, is associated with its blockade of cell cycle progression which appears to be attributable in part to inhibition of ERK1/2 activation.[1]
Eupatilin attenuates bile acid-induced hepatocyte apoptosis by suppressing bile acid-induced kinase activation, it might be therapeutically efficacious in a variety of human liver diseases associated with cholestasis.[2]
Eupatilin and jaceosidin have effects on cytochrome p450 enzyme activities in human liver microsomes, have potential pharmacokinetic drug interactions in vivo due to inhibition of CYP1A2 and CYP2C9.[3]
Eupatilin is a potent anti-atherogenic agent that inhibits PDGF-BB-induced proliferation and migration in HASMCs as well as aortic sprouting, which is likely mediated through the attenuation of PI3K, MKK3/6, and MKK4 activation.[4]
Eupatilin suppresses oxidative damage and reciprocally enhances extracellular matrix production in articular chondrocytes, making eupatilin a promising therapeutic option for the treatment.[5]
Eupatilin improves the acute hepatic IRI by reducing inflammation and apoptosis, it is a promising therapeutic agent against acute IR-induced hepatic damage.[6]
Eupatilin protects against tumor necrosis factor-α-mediated inflammation inhuman umbilical vein endothelial cells.[7]
Eupatilin induces Sestrin2-dependent autophagy to prevent oxidative stress.[8]

References:
[1] Kim D H, Na H K, Oh T Y, et al. Biochem Pharmacol, 2004, 68(6):1081-7.
[2] Su C P, Yoon J H, Kim W, et al. J Gastroenterol, 2006, 41(8):772-8.
[3] Ji H Y, Kim S Y, Kim D K, et al. Molecules, 2010, 15(9):6466-75.
[4] Son J E, Lee E, Seo S G, et al. Planta Med, 2013, 79(12):1009-16.
[5] Jeong J H, Moon S J, Jhun J Y, et al. Plos One, 2015, 10(6):e0130882.
[6] Lee H M, Jang H J, Kim S S, et al. Transpl P, 2016, 48(4):1226-33.
[7] Yu K, Li X M, Xu X L, et al. Int J Clin Exp Med, 2016, 8(12):22191-7.
[8] Jegal K H, Ko H L, Sang M P, et al. Apoptosis, 2016, 21(5):1-15.
[9] Zhou Q, Sun L L, Jiang B, et al. China Pharm, 2013, 24(47):4464-6.

HPLC of Eupatilin

Storage conditions：Short-term storage at 2~8℃, long-term storage at -20 ~ -80℃

Introduction/Overview

Natural products have always been an important treasure trove for innovative drug discovery, among which flavonoids have attracted much attention due to their broad biological activity and low toxicity. Eupatilin, also known as 5,7-dihydroxy-3 ', 4', 6-trimethoxyflavone, is a lipophilic flavonoid with a unique trimethoxy substitution pattern. Its CAS number is 22368-21-4, mainly isolated from Artemisia argyi, a plant in the Asteraceae family, as well as various plants in the Citrus and Salvia genera. Since its discovery, isoflavones have gradually become a hot molecule in natural product pharmacology research due to their significant multiple pharmacological activities such as anti-inflammatory, antioxidant, anti ulcer, and anti-tumor.

Traditionally, Artemisia argyi has been commonly used in East Asian traditional medicine to treat inflammation, bleeding, and abdominal pain, and some of its medicinal effects are believed to be closely related to active ingredients such as isoflavones. Modern pharmacological research has confirmed that isoquercetin is not only an agonist of peroxisome proliferator activated receptor alpha (PPAR alpha), but also exerts a wide range of cellular protective and anti disease effects by regulating multiple key signaling pathways and molecular targets. Especially in the field of cancer, its multi-target mechanism of action shows promising therapeutic potential. This article aims to systematically review the chemical properties, plant sources, pharmacological activities, molecular mechanisms of action, pharmacological evaluation, and clinical application prospects of isoquercetin, in order to provide comprehensive scientific references for the in-depth research and future drug development of this compound.

Chemical structure and physicochemical properties

Isocyanin belongs to the flavonoid class, and its basic parent nucleus is 2-phenylchromenone. The specific chemical structural feature is that the C-5 and C-7 positions of the flavonoid skeleton are replaced by hydroxyl groups (- OH), while the C-6, C-3 ', and C-4' positions are respectively replaced by methoxy groups (- OCH3), forming a 5,7-dihydroxy-3 ', 4', 6-trimethoxyflavonoid structure. This specific substitution pattern has a decisive impact on its physicochemical properties and biological activity.

From the analysis of physical and chemical properties, the molecular weight of isoquercetin is 344.3190 g/mol. Its lipophilicity is strong, and the calculated lipid water partition coefficient (LogP) is 2.5236, indicating that the compound is more soluble in organic solvents and belongs to lipophilic molecules. This characteristic is consistent with its form of existence in plants (often related to lipids) and its transmembrane transport capacity. Its topological polar surface area (TPSA) is 98.36 Å ², which is relatively moderate. The water solubility is poor, about 0.0085 mg/mL, which to some extent limits its direct application in aqueous systems, but also suggests that it may have good membrane permeability.

From the spectroscopic characteristics, isoquercetin exhibits typical flavonoid absorption peaks in the UV visible region. The crystal structure analysis shows that there are intramolecular hydrogen bonds (such as between C5-OH and C4 carbonyl groups) within the molecule, which helps to stabilize the molecular conformation. These clear chemical structural information provide a solid foundation for studying structure-activity relationships, structural modifications, and quality control.

Plant sources and extraction methods

Isocyanin is relatively widely distributed in nature, but mainly enriched in a few types of plants.
1. Main plant sources：
* Asteraceae plants：Artemisia argyi It is the most important and famous source. Artemisia argyi has a long history of medicinal use in Chinese folk culture, and isoflavones are considered one of its key active ingredients.
* Rutaceae plants: Multiple types Citrus spp Isocyanin is also present in the skin or leaves of the plant.
* Lamiaceae plants As follows:Salvia pubescens Waiting for plants of the Salvia genus.
*In addition, it has also been detected in some other plants of the Artemisia genus in the Asteraceae family.

1. Extraction and Separation Methods：
   Due to its lipophilicity, isoquercetin is often extracted using organic solvent methods.
   • Solvent extraction The most commonly used method is to use methanol, ethanol, acetone or their mixed solvents with water for reflux extraction or ultrasound assisted extraction of dried and crushed plant materials. Ethanol is the preferred choice for laboratory and industrial scales due to its safety, low cost, and high extraction efficiency.
   • Extraction enrichment After vacuum concentration, the crude extract was subjected to liquid-liquid extraction to remove highly polar impurities such as sugars and proteins, taking advantage of the good solubility of isocyanine in moderately polar organic solvents such as ethyl acetate and chloroform.
   • Separation and purification Further purification relies on chromatographic techniques. Silica gel column chromatography is commonly used, with petroleum ether ethyl acetate or chloroform methanol gradient elution. High performance liquid chromatography (HPLC), especially preparative HPLC, is a key step in obtaining high-purity isoquercetin (for pharmacological research), commonly using C18 reverse phase chromatography columns with methanol water or acetonitrile water as the mobile phase.
   • appraisal The identification of pure compounds comprehensively utilizes mass spectrometry (MS), nuclear magnetic resonance (NMR, including 1H and 13C NMR), ultraviolet spectroscopy (UV), and HPLC method compared with standard samples.

In recent years, some green extraction technologies such as supercritical CO2 extraction have also been explored, aiming to improve efficiency and reduce the use of organic solvents.

Pharmacological activity research

A large number of in vitro and in vivo studies have confirmed that isoflavones have diverse and significant pharmacological activities.

1. Anti inflammatory and antioxidant activity：
   This is the core activity of isoquercetin that was first recognized. In various acute and chronic inflammation models (such as lipopolysaccharide induced macrophage inflammation, mouse ear swelling model, colitis model), isoflavones can effectively inhibit the production of pro-inflammatory mediators (such as tumor necrosis factor - α, interleukin-6, nitric oxide, prostaglandin E2). Its antioxidant effect is manifested by its strong free radical scavenging ability, which can enhance the activity of endogenous antioxidant enzymes such as superoxide dismutase and glutathione peroxidase, and reduce the levels of oxidative stress markers such as malondialdehyde. These functions are the basis for its application in fields such as gastric mucosal protection (anti ulcer) and neuroprotection.

2. Anti ulcer and gastrointestinal protective effects：
   Isocyanine is a Korean marketed gastric medicine called Stillen ®” The main active ingredient is used to treat gastritis and gastric ulcers. Its function not only stems from anti-inflammatory and antioxidant effects, but also from increasing gastric mucosal blood flow, promoting mucus secretion, inhibiting gastric acid secretion, and regulating the balance of mucosal cell apoptosis and proliferation, thereby protecting the gastric mucosa from damage caused by ethanol, stress, or nonsteroidal anti-inflammatory drugs through multiple pathways.

3. Antitumor activity：
   This is currently the most active field of research. Isoeupatorium flavin has growth inhibition and apoptosis induction effects on a variety of human cancer cell lines, including gastric cancer, liver cancer, colorectal cancer, breast cancer, lung cancer, prostate cancer, ovarian cancer, etc. Its anti-tumor effect is multifaceted:

   • Inhibit cell proliferation Block the cell cycle at G1/S or G2/M phase.
   • Inducing cell apoptosis Activation of Caspase cascade reaction through mitochondrial pathway and death receptor pathway.
   • Inhibit invasion and metastasis Reduce the migration and invasion ability of cancer cells.
   • Angiogenesis inhibition Inhibit tumor angiogenesis.
   • Sensitization chemotherapy Combination with conventional chemotherapy drugs can produce synergistic effects and reverse multidrug resistance.

4. Other pharmacological activities：

   • neuroprotection It has a protective effect in models such as cerebral ischemia and Alzheimer's disease, with mechanisms involving anti-inflammatory, antioxidant, and anti apoptotic effects.
   • metabolic regulation As a PPAR α agonist, it may be involved in regulating lipid metabolism.
   • anti-fibrotic Has an improving effect on liver fibrosis and pulmonary fibrosis.
   • Anti-allergy Plays a role by stabilizing mast cells and inhibiting histamine release.

Mechanism of action and molecular targets

The pharmacological effects of isoflavones, especially their anti-tumor activity, are achieved by regulating a complex molecular network involving multiple key signaling pathways and specific targets.

1. Regulation of core signaling pathways：

   • MAPK/ERK pathway Isocyanine can inhibit the phosphorylation activation of MAPK1 (ERK2), thereby affecting cell proliferation and survival signals.
   • JAK/STAT pathway Of particular importance is the STAT3 Inhibition of signaling pathways. Isocyanine can inhibit the phosphorylation and nuclear translocation of STAT3, as well as the expression of downstream target genes (such as Bcl-2, Cyclin D1, MMP-2), which is one of its core mechanisms for inducing apoptosis and inhibiting metastasis.
   • PI3K/Akt pathway By inhibiting the activation of Akt, promoting the expression of pro apoptotic proteins and inhibiting their downstream survival signals.
   • NF - κ B pathway Inhibiting the degradation of I κ B α and nuclear translocation of NF - κ B p65 subunit, thereby downregulating the expression of various inflammatory factors and anti apoptotic genes.
   • HIF-1 α pathway: Inhibition HIF1A The stability and transcriptional activity of the tumor interfere with its hypoxic adaptation and angiogenesis.

2. Key molecular targets：

   • Apoptosis regulatory targets Upregulation of pro apoptotic protein Bax and downregulation of anti apoptotic protein BCL2 and MCL1 Inducing mitochondrial membrane potential collapse and releasing cytochrome C.
   • Cell cycle targets Downregulation of Cyclin D1 and CDK4/6, upregulation of p21 and p27, leading to cell cycle arrest.
   • Invasion and metastasis related targets Inhibition of matrix metalloproteinases MMP2 The expression and activity of MMP9 decrease the ability to degrade extracellular matrix.
   • Enzyme and receptor targets：
     • TOP1 and TOP2A As a topoisomerase inhibitor, it interferes with DNA replication and transcription.
     • CYP19A1(Aromatase): inhibiting its activity may interfere with the growth of estrogen dependent tumors (such as breast cancer).
     • ESR1(Estrogen receptor alpha): By antagonizing or regulating its activity, it affects related signals.
     • 5-Lipoxygenase As an inhibitor, it reduces the production of inflammatory mediators such as leukotrienes.
   • Nuclear receptor target As:PPARαExcitants that participate in the regulation of metabolism and inflammation.

In summary, isoflavones exert their effects through a "multi-target, multi pathway" approach, which is not only the advantage of their therapeutic potential, but also brings complexity to their mechanism research. The core of its functional network is to synergistically induce apoptosis, inhibit proliferation, and block invasion and metastasis.

Evaluation of drug properties and pharmacokinetics

Although isoflavones have significant biological activity, their development as drug candidate molecules requires systematic pharmacological evaluation.

1. Analysis of drug properties parameters：

   • Permeability and Distribution Moderate LogP values and moderate TPSA suggest good membrane permeability. Predict it Low blood-brain barrier permeability Although this limits the direct application of the central nervous system, it may also reduce the risk of central side effects. Its binding rate with plasma proteins is expected to be high, which will affect its free blood drug concentration and distribution volume.
   • Preliminary safety warning According to the data, it affects the hERG potassium channel No significant inhibition The potential risk of arrhythmia is low. The Ames test result is 0.6 (usually expressed as a mutagenicity index, less than 2 is considered negative), indicating that it has no significant genetic toxicity. But this still requires more comprehensive in vitro and in vivo toxicology experiments to confirm.
   • Water solubility Poor water solubility (0.0085 mg/mL) is one of its main pharmaceutical deficiencies, which may lead to irregular oral absorption and low bioavailability.

2. Pharmacokinetic study：
   Animal and partial human studies have revealed the pharmacokinetic characteristics of isoflavones:

   • absorb After oral administration, it can be absorbed in the gastrointestinal tract, but the degree and speed of absorption are greatly affected by the dosage form. Its lipophilicity is conducive to passive diffusion absorption.
   • distribution After absorption, it is widely distributed in various tissues, with higher concentrations in the liver, kidneys, and gastrointestinal tract.
   • Metabolism Isocyanin undergoes extensive metabolism in the body, mainly II combined reaction Enzymes in liver microsomes and gut microbiota, such as UDP glucuronosyltransferase and sulfotransferase, catalyze their glucuronidation and sulfation, producing more polar metabolites. In addition, its methoxy group may undergo demethylation reaction. The CYP450 enzyme system may be involved in its minor phase I metabolism.
   • excretion Metabolites are mainly excreted through the kidneys and urine, and some prototype drugs and metabolites can also be excreted through bile and feces. Its elimination half-life is relatively short in animal models, suggesting that multiple administrations may be necessary to maintain effective blood drug concentrations.
   • bioavailability Existing data shows that its absolute oral bioavailability is not high, mainly due to first pass effects (intestinal and hepatic metabolism) and solubility limitations.

3. Formulation strategy：
   To improve its bioavailability, researchers are exploring various advanced drug delivery systems, such as:

   • Solid dispersion Disperse the drug in an amorphous form in a water-soluble carrier, greatly improving the dissolution rate.
   • Cyclodextrin inclusion complex Increase the solubility and stability of drugs in water.
   • nano-formulation Including nanocrystals, liposomes, polymer nanoparticles, etc., by reducing particle size or changing delivery methods, their solubility, targeting, and pharmacokinetic behavior can be improved.
   • Phospholipid complex Enhance its lipophilicity, promote lymphatic absorption, and may bypass some liver first pass effects.

Clinical application prospects and prospects

Isocyanin has evolved from a traditional medicinal plant to modern clinical practice, demonstrating multiple potential applications, but also facing challenges.

1. Current and potential clinical applications：

   • Gastrointestinal diseases This is the most mature field. As a prescription drug or OTC medication, it is used to treat acute and chronic gastritis, gastric and duodenal ulcers, with definite therapeutic effects. In the future, its application in inflammatory bowel diseases such as ulcerative colitis can be explored.
   • Tumor adjuvant therapy and chemoprevention Based on its multi-target anti-tumor properties and the effect of sensitizing chemotherapy, isoflavones are expected to be developed as Tumor adjuvant therapy drugs or Chemical preventive agent Especially for digestive tract tumors such as gastric cancer and liver cancer, they may have advantages due to their high local concentration in the gastrointestinal tract. Combined with conventional therapies, it can reduce chemotherapy dosage, alleviate side effects, and overcome drug resistance.
   • Inflammatory related diseases Such as dermatitis, arthritis, asthma, etc., can be used as a supplement or alternative to anti-inflammatory treatment.
   • Metabolic diseases Its PPAR α activation activity suggests that it may have application value in metabolic diseases such as non-alcoholic fatty liver and hyperlipidemia.

2. challenges faced：

   • Bioavailability bottleneck Low water solubility and significant first pass effect are the main obstacles to its oral administration.
   • Complexity of mechanism of action The multi-target characteristic is both advantageous and complex in determining precise mechanisms of action, potential off target effects, and optimal indications.
   • Clinical Evidence Level At present, most research is still in the preclinical stage (cell and animal experiments), lacking large-scale, multicenter randomized controlled clinical trial data to confirm its effectiveness and safety in humans (especially in tumor treatment).
   • Intellectual Property and Standardization As a natural product, its compound patents may have expired or face challenges. Strict quality control standards need to be established from raw materials to finished products.

3. Future research directions and prospects：

   • Structural modification and derivative development By chemically modifying the parent nucleus of isoquercetin (such as modifying methoxy and hydroxyl groups, introducing other functional groups), the aim is to improve its activity, selectivity, solubility, and metabolic stability, thereby obtaining candidate molecules with better drug properties.
   • Innovative formulation research and development Vigorously developing the above-mentioned new drug delivery systems and conducting systematic pharmacokinetic studies of formulations is the key path to achieving their clinical translation.
   • In depth mechanism exploration Using omics technologies (proteomics, metabolomics) and gene editing tools to more systematically and accurately elucidate its functional network and key node targets.
   • Conduct clinical research Design rigorous clinical trials, first explore optimized solutions in the already marketed indications (gastritis), and then gradually advance towards new indications such as tumor adjuvant therapy, accumulating high-level evidence-based medicine evidence.
   • Explore combination therapy Systematically study its synergistic effect with existing standard therapeutic drugs (chemotherapy drugs, targeted drugs, immune checkpoint inhibitors) to find the optimal combination therapy.

Conclusion

Isocyanin, as a natural flavonoid compound derived from traditional medicinal plants, has shown great potential in anti-inflammatory, antioxidant, anti ulcer, and especially anti-tumor fields due to its unique chemical structure and multi-target, multi pathway pharmacological mechanisms. From Artemisia argyi leaves to the laboratory, from studying its mechanism of action to exploring its medicinal properties, our understanding of isoflavones is constantly deepening. Although it still faces challenges in terms of bioavailability and clinical translation, these challenges are gradually being overcome through the cross fusion of modern medicinal chemistry, pharmacy, and pharmacology methods. In the future, with deeper mechanism research, more optimized formulation development, and more rigorous clinical validation, isoflavones and their derivatives are expected to develop from potential lead compounds into innovative drugs for treating gastrointestinal diseases, tumors, and other inflammation related diseases, fully demonstrating the sustained value of natural products in modern medicine. Its research process has also provided valuable paradigms for the development of other natural active ingredients.