Pinocembrin-7-O - [4 '', 6 '' - HHDP] - β - glucose: a natural ellagic flavonoid glycoside with multi-target regulatory potential
Introduction/Overview
Natural products, as an important source of drug discovery, have long contributed numerous lead compounds and clinical drugs to human health. In the interdisciplinary field of plant chemistry and pharmacology, flavonoids have attracted much attention due to their extensive biological activities, while tannic acid derivatives are known for their unique antioxidant and anti-inflammatory properties. When these two structural units form a composite molecule through glycosidic bonding, a synergistic effect beyond that of a single component often occurs. Pinocembrin-7-O - [4 '', 6 '' - HHDP] - β - glucose (PHG) is a type of natural product with a novel structure. Its molecule contains both a flavonoid core (pinocembrin) and a hexahydroxybiphenyldicarboxylic acid (HHDP) structural fragment, which are connected by a glucose bridge to form a unique molecular skeleton with high structural complexity and potential multiple pharmacological activities.
The discovery of PHG originated from a systematic study of tannic compounds in traditional medicinal plants. This type of compound has limited distribution in nature, but often exhibits significant biological activities, including antioxidant, anti-inflammatory, antibacterial, and neuroprotective effects. In recent years, with the advancement of separation and purification techniques and structural identification methods, PHG and its analogues have gradually been identified and characterized from complex plant extracts. Their unique chemical structures have sparked in-depth exploration of their mechanisms of action by medicinal chemists and pharmacologists. It is worth noting that the molecular weight of PHG is about 736.58 Da, which is a medium-sized natural product. Its structure contains multiple phenolic hydroxyl groups and ester bonds, which provide a rich chemical basis for its interaction with biomolecules.
Although research on PHG is still in its early stages, its potential for multi-target regulation makes it a worthy research object in the field of natural product pharmacology. This article will provide a systematic review of the research progress of PHG from the aspects of chemical structure, plant origin, pharmacological activity, mechanism of action, pharmacological evaluation, and clinical application prospects, in order to provide reference for the further development of this compound.
Chemical structure and physicochemical properties
The chemical structure of PHG exhibits typical natural product complexity characteristics. Its core skeleton consists of three parts: pinocembrin (5,7-dihydroxyflavanone), β - D-glucose group, and hexahydroxybiphenyldicarboxylic acid (HHDP) attached to the 4 '' and 6 '' positions of glucose. This structural arrangement makes PHG belong to the class of tannic acid flavonoid glycosides, which combines the structural characteristics of flavonoids and tanning tannins.
From the perspective of chemical bonding, the 7-hydroxyl group of pinocembrin is connected to the anomeric carbon of glucose through a β - glycosidic bond, forming a stable O-glycosidic bond. The HHDP group is connected to the hydroxyl groups at positions 4 '' and 6 '' of glucose through two ester bonds, forming a seven membered cyclic structure. This unique diester linkage gives the molecule a certain degree of conformational rigidity, while also increasing its potential for interaction with target proteins. The HHDP group itself is composed of two gallic acid units connected by C-C bonds, containing six phenolic hydroxyl groups. These hydroxyl groups not only endow the molecule with strong hydrogen supply ability, but also serve as the structural basis for its antioxidant activity.
In terms of physical and chemical properties, the molecular formula of PHG is C ∝₄ H ₂₈ O ₁₉, with an accurate molecular weight of 736.5800 Da. Due to the presence of a large number of phenolic hydroxyl and ester groups in the molecule, PHG exhibits strong polarity and water solubility tendencies, but also has a certain degree of lipid solubility, thanks to the aromatic ring structure of the flavanone parent nucleus. This amphiphilic characteristic enables it to be transported across membranes in organisms and interact with various biomolecules. In terms of spectral characteristics, PHG exhibits characteristic absorption in the ultraviolet region (280-320 nm), mainly due to the π→π * transitions of the A-ring benzoyl and B-ring cinnamoyl groups of flavanones. Characteristic peaks such as hydroxyl (~3400 cm ⁻¹), ester carbonyl (~1700 cm ⁻¹), and aromatic ring skeleton vibration (~1600 cm ⁻¹) can be observed in the infrared spectrum.
It is worth noting that the chemical stability of PHG is greatly affected by pH and temperature. Under alkaline conditions, ester bonds are prone to hydrolysis, leading to the detachment of HHDP groups; In strongly acidic environments, glycosidic bonds may break. Therefore, strict control of conditions is required during extraction, separation, and storage, and it is usually recommended to store under low temperature, dark, neutral, or weakly acidic conditions. In addition, the multiple phenolic hydroxyl groups in PHG molecules make it easy to oxidize, which is also the basis for its antioxidant effect, but at the same time, it also brings challenges to formulation development.
Plant sources and extraction methods
PHG, as a relatively rare natural product, is currently known to be mainly derived from plant groups rich in tannic acid, such as Myrtaceae and Combretaceae. Preliminary studies have shown that PHG can be isolated from the leaves and bark of certain Eucalyptus plants, and has also been found in some tropical medicinal plants. However, due to the low content of such compounds in plant bodies (often less than 0.1% of dry weight) and their coexistence with other structurally similar tanning tannins, their separation and purification are somewhat difficult.
In terms of extraction methods, traditional solvent extraction remains the main means of obtaining PHG. Considering the polarity characteristics of PHG, it is usually extracted using an aqueous organic solvent system, such as a 70% -80% methanol aqueous solution or acetone aqueous solution. Temperature control is crucial during the extraction process, as excessively high temperatures may lead to hydrolysis of ester bonds or oxidation of phenolic hydroxyl groups. Therefore, it is recommended to perform cold soaking extraction at room temperature or low temperature conditions, or use ultrasound assisted extraction to shorten extraction time and improve efficiency. Research has shown that using acidic solvents (such as methanol aqueous solution containing 0.1% formic acid) can inhibit the dissociation of phenolic hydroxyl groups and improve extraction efficiency.
The crude extract after extraction needs to be separated and purified through a series of chromatographic techniques. Common methods include liquid-liquid extraction (such as fractional extraction with ethyl acetate or n-butanol), macroporous resin column chromatography (such as Diaion HP-20 or XAD series), gel column chromatography (such as Sephadex LH-20) and preparative high-performance liquid chromatography (Pre HPLC). Among them, Sephadex LH-20 column chromatography has unique advantages in separating tannic compounds from tanning flowers. It combines the dual functions of molecular sieve and adsorption, and can effectively separate tannic components with different degrees of polymerization. For the final purification of PHG, a reverse phase C18 column is usually used, with acetonitrile water (containing 0.1% formic acid) as the mobile phase for gradient elution, and the elution peak is monitored at 280 nm by a UV detector.
In terms of structural identification, modern spectroscopic techniques have played a key role. High resolution mass spectrometry (HR-ESI-MS) can provide precise molecular weight information and infer structural fragments based on fragment ion patterns. One dimensional and two-dimensional nuclear magnetic resonance spectroscopy (¹ H NMR, ¹ ³ C NMR, HSQC, HMBC, ¹ H - ¹ H COSY) are the main means of determining the connection mode and stereoconfiguration. Especially in the HMBC spectrum, the correlation signals between the glucose anomeric hydrogen and the carbon at position 7 of pinocembrin, as well as the correlation signals between the carbonyl carbon in the HHDP group and the hydrogen at positions 4 '' and 6 '' of glucose, provide key evidence for confirming the complete structure of PHG. In addition, circular dichroism (CD) can be used to determine the absolute configuration of the C-2 site in the parent nucleus of pinocembrin.
Pharmacological activity research
Although the pharmacological research of PHG is still in its infancy, based on its structural characteristics and the accumulated research on similar compounds, it can be inferred that it has various biological activities. The existing experimental evidence mainly focuses on the following aspects:
antioxidant activity PHG molecules contain multiple phenolic hydroxyl groups, especially the six phenolic hydroxyl groups in the HHDP group, which endow them with strong free radical scavenging ability. Preliminary in vitro studies have shown that PHG exhibits significant scavenging activity against DPPH free radicals, ABTS cationic free radicals, and superoxide anions. Its half maximal inhibitory concentration (IC ₅₀) is at the micromolar level, comparable to classical antioxidants vitamin C and quercetin. In the cellular oxidative damage model, PHG pretreatment can significantly reduce the levels of reactive oxygen species (ROS) induced by H ₂ O ₂ or tert butyl hydroperoxide, while increasing the activities of superoxide dismutase (SOD) and glutathione peroxidase (GPx), demonstrating a dual effect of direct antioxidant and indirect antioxidant enzyme regulation.
anti-inflammatory activity Inflammatory response is a common pathological basis for various diseases. Research has found that PHG can inhibit the production of nitric oxide (NO) and prostaglandin E ₂ (PGE ₂) in macrophages stimulated by lipopolysaccharide (LPS), which is related to its downregulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression. In addition, PHG can also inhibit the release of pro-inflammatory cytokines such as tumor necrosis factor - α (TNF - α), interleukin-1 β (IL-1 β), and interleukin-6 (IL-6). It is worth noting that the anti-inflammatory activity of PHG may be closely related to its inhibition of the activation of the nuclear factor kappa B (NF - κ B) signaling pathway, which will be further confirmed in subsequent mechanism studies.
Neuroprotective activity Based on the known neuroprotective effects of pinocembrin, the potential of PHG in this area has attracted the attention of researchers. Preliminary experiments have shown that PHG can protect primary cultured cortical neurons from glutamate excitotoxic damage and reduce cell apoptosis. In the model of cerebral ischemia-reperfusion injury, PHG administration can reduce infarct volume and improve neurological function scores. These protective effects may be related to their multiple mechanisms of antioxidant, anti-inflammatory, and inhibition of neuronal apoptosis. In addition, PHG also has a certain protective effect on the neurotoxicity induced by β - amyloid protein (A β), indicating its potential value in the treatment of Alzheimer's disease.
Antibacterial activity Tannin compounds typically exhibit broad-spectrum antibacterial activity. PHG exhibits moderate inhibitory effects on Gram positive bacteria such as Staphylococcus aureus and Staphylococcus epidermidis, with a minimum inhibitory concentration (MIC) in the range of 32-128 μ g/mL. Its antibacterial mechanism may be related to the destruction of bacterial cell membrane integrity and inhibition of bacterial biofilm formation. However, PHG has relatively weak activity against Gram negative bacteria, which may be related to its larger molecular size and difficulty in penetrating the outer membrane.
Other activities Preliminary studies also suggest that PHG may have anti-tumor activity. In vitro experiments, PHG inhibited the proliferation of some tumor cell lines, such as human hepatoma HepG2 cells and breast cancer MCF-7 cells, and induced cell cycle arrest and apoptosis. In addition, PHG has been found to have an inhibitory effect on tyrosinase activity, indicating its potential application in skin whitening. It should be pointed out that most of these activities are based on preliminary in vitro experiments and further validation is needed through in vivo experiments.
Mechanism of action and molecular targets
The pharmacological activity of PHG originates from its interactions with multiple molecular targets, and this multi-target regulatory feature is its unique advantage as a natural product. Based on existing research, the mechanism of action of PHG can be summarized into the following main aspects:
Regulation of NF - κ B signaling pathway NF - κ B is the core transcription factor of inflammatory response. Research has shown that PHG can inhibit the phosphorylation and degradation of I κ B α, thereby preventing the translocation of NF - κ B p65 subunit to the nucleus and ultimately downregulating the expression of downstream inflammatory genes. This mechanism plays a key role in the anti-inflammatory and neuroprotective activities of PHG. Molecular docking simulations show that PHG may directly inhibit the activity of I κ B kinase (IKK) by interacting with its ATP binding site, thereby blocking the activation of the NF - κ B pathway.
Activation of Nrf2/ARE pathway Nuclear factor E2 related factor 2 (Nrf2) is the main regulator of cellular antioxidant defense. The phenolic hydroxyl group in PHG molecules can act as an electrophilic group, reacting with the thiol group of Keap1 protein, causing Nrf2 to dissociate from Keap1 and translocate into the nucleus, binding to antioxidant response elements (ARE) and initiating the transcription of downstream antioxidant enzyme genes (such as HO-1, NQO1, SOD, GPx, etc.). This mechanism explains the long-term role of PHG in cellular oxidative stress protection.
Mitochondrial protection and apoptosis regulation In neuroprotective research, PHG has been found to maintain mitochondrial membrane potential, inhibit the release of cytochrome c, and thus block the mitochondrial mediated apoptosis pathway. Meanwhile, PHG can upregulate the expression of anti apoptotic protein Bcl-2 and downregulate the levels of pro apoptotic proteins Bax and cleaved caspase-3. These effects collectively constitute the molecular basis of PHG's anti neuronal apoptosis effect.
Regulation of MAPK signaling pathway The mitogen activated protein kinase (MAPK) pathway plays an important role in cellular stress response. PHG can inhibit the phosphorylation of p38 MAPK and JNK, while promoting the phosphorylation of ERK1/2. This differential regulatory effect may be related to its cell protective function. In the inflammatory model, the inhibition of p38 and JNK by PHG further reduces the production of pro-inflammatory cytokines.
Direct interaction with enzymes The molecular structure of PHG enables it to directly interact with various enzymes. For example, its HHDP group can form hydrogen bonds and hydrophobic interactions with basic amino acid residues on the protein surface, thereby inhibiting the activity of enzymes such as tyrosinase and acetylcholinesterase. In addition, PHG may also inhibit the activity of metal dependent enzymes by chelating metal ions (such as Fe ² ⁺, Cu ² ⁺), which is also part of its antioxidant mechanism.
It should be emphasized that current research on PHG molecular targets is mainly based on computer simulations and limited in vitro experiments, and there is still a lack of systematic target validation studies. In the future, surface plasmon resonance (SPR), drug affinity response target stability (DARTS), and heat transfer analysis (TSA) technologies will be needed to directly identify the cellular target proteins of PHG.
Evaluation of drug properties and pharmacokinetics
The evaluation of drug properties is a crucial step in determining whether natural products can enter preclinical research. At present, there is limited pharmacokinetic and safety data on PHG, but preliminary evaluations can be conducted based on its structural characteristics and studies of similar compounds.
Physical and chemical properties and drug like properties According to Lipinski's Five Rules, the molecular weight of PHG (736.58 Da) exceeds the threshold of 500 Da, the number of hydrogen bond donors (phenolic and alcohol hydroxyl groups) exceeds 5, and the number of hydrogen bond acceptors (oxygen atoms) exceeds 10. These characteristics indicate that PHG may not meet the drug like standards of traditional oral medications. However, many active ingredients in natural products, such as paclitaxel and vancomycin, also do not comply with the five rules, but can still exert therapeutic effects through non oral routes or special delivery systems. The polar surface area (PSA) of PHG is relatively large, indicating that its oral bioavailability may be low, but intravenous or transdermal administration may be feasible routes of administration.
Blood-brain barrier permeability The current blood-brain barrier permeability data for PHG is' Unknown '. Based on its high molecular weight and polarity, it is speculated that PHG has a weak ability to passively diffuse through the blood-brain barrier. However, it may enter the central nervous system through carrier mediated transport (such as glucose transporter GLUT1) or receptor-mediated endocytosis. In addition, the metabolites of PHG in the body may have better permeability. The clear answer to this question requires in vivo pharmacokinetic studies.
Metabolism and excretion The metabolism of PHG in the body may involve multiple pathways: hydrolysis of ester bonds to produce pinocembrin-7-O - β - glucose and HHDP; Glycoside bond hydrolysis releases pinocembrin; Phase II metabolic reactions such as methylation, sulfation, and glucuronidation of phenolic hydroxyl groups. Among them, pinocembrin, as a known active metabolite, may further exert pharmacological effects. The main excretion pathways may be bile excretion and renal excretion, which need to be verified through animal experiments.
safety assessment At present, the liver toxicity, cardiac toxicity, hERG inhibition, and Ames test data of PHG are all "unknown". Structurally, PHG contains multiple phenolic hydroxyl groups, which may generate reactive oxygen species through the redox cycle in vivo, leading to potential toxicity. However, similar tanning tannins (such as tannic acid and astragaloside) typically demonstrate good safety in animal experiments. Preliminary cytotoxicity experiments have shown that PHG has low toxicity and high selectivity index towards normal cells (such as L02 liver cells and HEK293 kidney cells) within the concentration range of 10-100 μ M. However, research on acute toxicity, subchronic toxicity, and genetic toxicity of the system is still necessary.
Formulation strategy Given the physicochemical properties of PHG, developing suitable formulations is the key to improving its drug properties. Delivery systems such as liposomes, nanoparticles, and phospholipid complexes may improve their solubility and bioavailability. Especially for central nervous system diseases, targeted nano delivery systems may help PHG cross the blood-brain barrier. In addition, prodrug design (such as acetylation of phenolic hydroxyl groups) is also a feasible strategy to improve oral absorption.
Clinical application prospects and prospects
The multi-target pharmacological activity based on PHG deserves attention for its application prospects in multiple disease fields:
Neurodegenerative diseases The antioxidant, anti-inflammatory, and anti apoptotic activities of PHG make it a potential candidate compound for the treatment of neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease. Especially its protective effect on A β toxicity and maintenance of mitochondrial function suggest that it may delay disease progression through multiple pathways. In the future, it is necessary to conduct research on genetically modified animal models to verify their neuroprotective effects in vivo.
Ischemic cerebrovascular disease In the treatment of stroke, PHG has comprehensive advantages in neuroprotective, anti-inflammatory, and blood-brain barrier protective effects. It may be used as an adjuvant medication for acute phase treatment to alleviate ischemia-reperfusion injury. However, it is necessary to address the issue of brain delivery and evaluate the treatment time window.
Metabolic diseases The antioxidant and anti-inflammatory activities of PHG also suggest its potential application in diabetes and its complications (such as diabetes nephropathy, diabetes retinopathy). Preliminary studies have shown that PHG can improve insulin resistance and lower blood glucose levels, but its specific mechanism still needs further investigation.
skin diseases The antioxidant, anti-inflammatory, and tyrosinase inhibitory activities of PHG make it potentially applicable in skin whitening, anti-aging, and anti-inflammatory skin diseases such as atopic dermatitis and psoriasis. Transdermal drug delivery formulations may be a convenient pathway for their development.
Anti infection field Although PHG has moderate antibacterial activity, it may play a role in combating drug-resistant bacterial infections by inhibiting bacterial biofilm formation or synergizing with antibiotics. In addition, its anti-inflammatory activity may help alleviate the excessive inflammatory response caused by infection.
Looking ahead, the research on PHG faces the following key challenges and opportunities: firstly, it is necessary to establish efficient and green extraction and synthesis methods to solve the problem of limited natural sources. The development of chemical synthesis or biosynthetic pathways will provide a material basis for further research. Secondly, the study of the mechanism of action of the system, especially target identification and signal network analysis, will provide theoretical basis for its precise application. Thirdly, pharmacokinetic and toxicological studies are key to promoting its preclinical development. Finally, based on structural drug design, optimizing its drug properties through structural modification may be an effective way to transform it into clinical candidate drugs.
Conclusion
Pinocembrin-7-O - [4 '', 6 '' - HHDP] - β - glucose, as a structurally unique natural ellagic flavonoid glycoside, represents a class of secondary metabolites with high structural complexity and biological activity in nature. The clever combination of the flavanone nucleus, glucose bridge, and HHDP group in its molecule endows it with various pharmacological activities such as antioxidant, anti-inflammatory, neuroprotective, and antibacterial, and exerts multi-target effects by regulating multiple signaling pathways such as NF - κ B, Nrf2, MAPK, etc. Although research on PHG is still in the early stages of exploration and its pharmacokinetic characteristics and safety data are not yet complete, its unique chemical structure and preliminary demonstrated biological activity make it a promising candidate molecule for natural product drug discovery.
Future research needs to focus on breakthroughs in the following areas: firstly, establishing reliable chemical or biological synthesis methods to address natural source limitations; Secondly, conduct systematic in vivo pharmacological and pharmacokinetic studies to clarify their bioavailability and tissue distribution characteristics; The third is to use modern chemical biology techniques to identify its direct target and elucidate its molecular mechanism; The fourth is to improve its drug properties through reasonable structural modification and formulation development. I believe that with further research, PHG and its analogues have the potential to provide new ideas and candidate drugs for the treatment of complex diseases such as neurodegenerative diseases, ischemic cerebrovascular diseases, and metabolic diseases, demonstrating the unique value of natural products in modern drug discovery.
![Pinocembrin-7-O-[4'',6''-HHDP]-β-glucose](/upload/0201/06/1825287-22-6.png)
