Proanthocyanidins isolated from lotus seed skin mitigate glycolipid metabolism disorder through the p38/Nrf2/NF-κB signaling pathway.

Lotus seed skin extract is rich in flavonoids, making it a promising candidate for developing health products. In a previous study, we found that proanthocyanidins from lotus seed skin, particularly proanthocyanidin B1 (PB1), can indirectly activate the Nrf2 signaling pathway, exerting an antioxidant effect. In this study, we isolate proanthocyanidins from lotus seed skin (PLS) using ethanol extraction and RP-HPLC identification, and investigate its effects on glycolipid metabolism both in vivo and in vitro. Our results demonstrate that PLS reduces body weight in high-fat diet (HFD) mice by decreasing feed efficiency. PLS also normalizes serum glucose, insulin secretion, glycosylated hemoglobin (HbA1c), and intraperitoneal glucose tolerance (IPGTT). Furthermore, PLS significantly improves blood lipid parameters and inhibits the expressions of six proinflammatory factors, including IL-1α, IL-1ß, IL-3, IL-6, IFN-γ and TNF-α in HFD mice. Additionally, analysis of fresh liver tissues reveals that PLS and PB1 induce the expressions of antioxidant proteins such as HO-1 and NQO1 by activating the p38-Nrf2 signaling pathway and inhibiting the NF-κB signaling pathway. In conclusion, proanthocyanidins from lotus seed skin regulate glycolipid metabolism disorders by targeting the p38/Nrf2/NF-κB signaling pathway. Our study offers a new approach for the high-value comprehensive utilization of lotus seed skin by-products and precise dietary intervention for metabolic syndrome.

Extract of Gardenia jasminoides Ellis Attenuates High-Fat Diet-Induced Glycolipid Metabolism Disorder in Rats by Targeting Gut Microbiota and TLR4/Myd88/NF-κB Pathway.

Gardenia jasminoides Ellis is abundant in crocin and has a longstanding historical usage both as a dietary and natural ethnic medicine. Enhanced studies have increasingly revealed the intricate interplay between glycolipid metabolism and gut microbiota, wherein their imbalance is regarded as a pivotal indicator of metabolic disorders. Currently, the precise molecular mechanism of the crude extract of crocin from Gardenia jasminoides Ellis (GC) targeting gut microbiota to regulate glycolipid metabolism disorder is still unclear. Firstly, we explored the effect of GC on digestive enzymes (α-amylase and α-glucosidase) in vitro. Secondly, we investigated the effect of GC on the physical and chemical parameters of high-fat diet (HFD) rats, such as body weight change, fasting blood glucose and lipid levels, and liver oxidative stress and injury. Then, 16S rDNA sequencing was used to analyze the effects of GC on the composition and structure of gut microbiota. Finally, the impact of GC on the TLR4/Myd88/NF-κB signaling pathway in the intestine was assessed by Western Blotting. In the present study, GC was found to exhibit a hypoglycemic effect in vitro, by inhibition of digestive enzymes. In animal experiments, we observed that GC significantly reduced fasting blood glucose, TC, and TG levels while increasing HDL-C levels. Additionally, GC demonstrated hepatoprotective properties by enhancing liver antioxidative capacity through the upregulation of SOD, CAT, and GSH-Px, while reducing ROS. 16S rDNA sequencing results showed that GC had a significant effect on the gut microbiota of HFD rats, mainly by reducing the ratio of Firmicutes/Bateroidota, and significantly affected the genera related to glycolipid metabolism, such as Akkermansia, Ligilactobacillus, Lactobacillus, Bacteroides, Prevotellaceae, etc. The Western Blotting results demonstrated that GC effectively downregulated the protein expressions of TLR4, Myd88, and NF-κB in the intestine of HFD rats, indicating that GC could target the TLR4/Myd88/NF-κB pathway to interfere with glycolipid metabolism disorder. Correlation analysis revealed that GC could target the Akkermansia-TLR4/Myd88/NF-κB pathway axis which attenuates glycolipid metabolism disorder. Therefore, this study establishes the foundation for GC as a novel therapeutic agent for glycolipid metabolism disorder chemoprevention, and it introduces a novel methodology for harnessing the potential of natural botanical extracts in the prevention and treatment of metabolic syndrome.

Theoretical and experimental study on the radiative properties of Parachlorella kessleri: considering the effect of internal microstructure.

A microalgal cell model with multiple organelles considering both the irregular overall shape and internal microstructure was proposed. The radiative properties of Parachlorella kessleri during the normal phase, starch-rich phase, and lipid-rich phase were calculated by the discrete dipole approximation method in the visible wavelengths. The accuracy of the model is verified with experimental measurements. The results showed that the theoretical calculation of the established microalgal cell model is more accurate than those of the equal volume spheres, such as the homogeneous sphere and the coated sphere, with the errors of the scattering cross-section reduced by more than 10.7%. The calculated scattering phase function of the multi-component model is basically in good agreement with the experimental results. Compared to the normal growth phase, the lipid enrichment during the lipid-rich phase leads to a sharp increase in the scattering cross-section by three to four times, while the absorption cross-section remains stable. Remarkably, in the starch-rich phase, the abundant production of starch results in a reduction of two to three times in the absorption cross-section compared to the normal growth phase, while the scattering cross-section varies little. The results can provide basic data and theoretical support for the design and optimization of photobioreactors.

The Molecular Mechanism Underlying the Therapeutic Effect of Dihydromyricetin on Type 2 Diabetes Mellitus Based on Network Pharmacology, Molecular Docking, and Transcriptomics.

Type 2 diabetes mellitus (T2DM) is a chronic and complex disease, and traditional drugs have many side effects. The active compound dihydromyricetin (DHM), derived from natural plants, has been shown in our previous study to possess the potential for reducing blood glucose levels; however, its precise molecular mechanism remains unclear. In the present study, network pharmacology and transcriptomics were performed to screen the molecular targets and signaling pathways of DHM disturbed associated with T2DM, and the results were partially verified by molecular docking, RT-PCR, and Western blotting at in vivo levels. Firstly, the effect of DHM on blood glucose, lipid profile, and liver oxidative stress in db/db mice was explored and the results showed that DHM could reduce blood glucose and improve oxidative stress in the liver. Secondly, GO analysis based on network pharmacology and transcriptomics results showed that DHM mainly played a significant role in anti-inflammatory, antioxidant, and fatty acid metabolism in biological processes, on lipoprotein and respiratory chain on cell components, and on redox-related enzyme activity, iron ion binding, and glutathione transferase on molecular functional processes. KEGG system analysis results showed that the PI3K-Akt signaling pathway, IL17 signaling pathway, HIF signaling pathway, MAPK signaling pathway, AGE-RAGE signaling pathway in diabetic complications, and TNF signaling pathway were typical signaling pathways disturbed by DHM in T2DM. Thirdly, molecular docking results showed that VEGFA, SRC, HIF1A, ESR1, KDR, MMP9, PPARG, and MAPK14 are key target genes, five genes of which were verified by RT-PCR in a dose-dependent manner. Finally, Western blotting results revealed that DHM effectively upregulated the expression of AKT protein and downregulated the expression of MEK protein in the liver of db/db mice. Therefore, our study found that DHM played a therapeutic effect partially by activation of the PI3K/AKT/MAPK signaling pathway. This study establishes the foundation for DHM as a novel therapeutic agent for T2DM. Additionally, it presents a fresh approach to utilizing natural plant extracts for chemoprevention and treatment of T2DM.

Natural deep eutectic ready to use extract of astilbin: Super high in vitro bioaccessibility, α-amylase and α-glucosidase enzyme inhibition kinetics.

Astilbin, a natural flavonoid, possesses multiple functionalities, while the poor bioavailability seriously restricts its application in functional food and medicine. Therefore, in this study, a natural deep eutectic solvent (NaDES) with choline chloride: lactic acid (CHCL-LAC) is selected to deliver astilbin by evaluating the bioaccessibility and antioxidant capacity during in vitro gastrointestinal digestion, and the inhibitory effect with underlying mechanism of astilbin-CHCL-LAC against α-amylase/α-glucosidase were investigated. The CHCL-LAC showed significant high astilbin bioaccessibility (84.1% bioaccessible) and DPPH and ORAC antioxidant capacity with 75.7% and 57.7% respectively after 3 h in vitro digestion, which may be attributed by hydrogen bond based supramolecule formed between astilbin and CHCL-LAC. Moreover, significant inhibitions of astilbin-CHCL-LAC on α-amylase (IC50 of 0.67 g/L) and α-glucosidase (IC50 of 0.64 g/L) were observed in mixed competitive and non-competitive manners. The dominant binding force between enzymes and astilbin were the hydrogen and hydrophobic interaction. This is the first time that the underlying mechanisms for astilbin delivered by NaDESs were revealed, suggesting that CHCL-LAC-based NaDESs are promising ready-to-use vehicles of natural inhibitors for carbohydrate-hydrolyzing enzymes.

The Regulatory Effect of Phytochemicals on Chronic Diseases by Targeting Nrf2-ARE Signaling Pathway.

Redox balance is essential to maintain the body's normal metabolism. Once disrupted, it may lead to various chronic diseases, such as diabetes, neurodegenerative diseases, cardiovascular diseases, inflammatory diseases, cancer, aging, etc. Oxidative stress can cause or aggravate a series of pathological processes. Inhibition of oxidative stress and related pathological processes can help to ameliorate these chronic diseases, which have been found to be associated with Nrf2 activation. Nrf2 activation can not only regulate the expression of a series of antioxidant genes that reduce oxidative stress and its damage, but also directly regulate genes related to the above-mentioned pathological processes to counter the corresponding changes. Therefore, targeting Nrf2 has great potential for the prevention or treatment of chronic diseases, and many natural phytochemicals have been reported as Nrf2 activators although the defined mechanisms remain to be elucidated. This review article focuses on the possible mechanism of Nrf2 activation by natural phytochemicals in the prevention or treatment of chronic diseases and the regulation of oxidative stress. Moreover, the current clinical trials of phytochemical-originated drug discovery by targeting the Nrf2-ARE pathway were also summarized; the outcomes or the relationship between phytochemicals and chronic diseases prevention are finally analyzed to propose the future research strategies and prospective.

Multiomic analysis of dark tea extract on glycolipid metabolic disorders in db/db mice.

Glycolipid metabolic disorder is a serious threat to human health. Dark tea is a kind of traditional Chinese tea, which may regulate the glycolipid metabolic disorders. Dark tea extract (DTE) is the water extraction obtained from dark tea. Compared with traditional dark tea, DTE has the benefits of convenient consumption and greater potential for promoting health. However, the regulation of DTE on glycolipid metabolism and its molecular mechanism is rarely investigated. In our study, DTE was used as raw material to study the effect and molecular mechanism of its intervention on the glycolipid metabolic in db/db diabetic mice by using multiomics analysis and modern biological techniques. (1) DTE could significantly reduce fasting glucose in diabetic db/db mice, and the higher dose group has a better effect. Histopathological examination showed that DTE slightly improve the number of islets and decrease the number of islet ß cells in the pancreatic tissue in db/db mice. (2) RNA-Seq was used to analyze the gene expression in liver tissue. In terms of biological processes, DTE mainly affected the inflammation and fatty acid metabolism. In terms of cell components, the lipoprotein and respiratory chain are mainly affected. In the aspect of molecular function, DTE mainly affected the redox related enzyme activity, iron ion binding and glutathione transferase. Arachidonic acid metabolism pathway, glutathione metabolism and PPAR signaling pathway were enriched by DTE with the results of KEGG pathway enrichment. In addition, real-time PCR results confirmed that DTE could significantly activate key genes of PPAR signaling pathway like Fabp1, Cyp4a1, Ehhadh, Cyp4a32, Aqp7 and Me1. (3) 16s rDNA showed that DTE could significantly decrease the ratio of Firmicutes/Bacteroidetes and the abundance of Proteobacteria, and increased the relative abundance of Verrucomicrobia at the phylum level. At the genus level, the relative abundance of Akkermansia, Prevotellaceae, Bacteroides and Alloprevotella was significantly increased after DTE treatment. This study provides multiomics molecular evidence for the intervention effect of DTE on abnormal glucose and lipid metabolism and the application of precise nutritional diet intervention of dark tea extract.

Beneficial Effects of Theaflavins on Metabolic Syndrome: From Molecular Evidence to Gut Microbiome.

In recent years, many natural foods and herbs rich in phytochemicals have been proposed as health supplements for patients with metabolic syndrome (MetS). Theaflavins (TFs) are a polyphenol hydroxyl substance with the structure of diphenol ketone, and they have the potential to prevent and treat a wide range of MetS. However, the stability and bioavailability of TFs are poor. TFs have the marvelous ability to alleviate MetS through antiobesity and lipid-lowering (AMPK-FoxO3A-MnSOD, PPAR, AMPK, PI3K/Akt), hypoglycemic (IRS-1/Akt/GLUT4, Ca2+/CaMKK2-AMPK, SGLT1), and uric-acid-lowering (XO, GLUT9, OAT) effects, and the modulation of the gut microbiota (increasing beneficial gut microbiota such as Akkermansia and Prevotella). This paper summarizes and updates the bioavailability of TFs, and the available signaling pathways and molecular evidence on the functionalities of TFs against metabolic abnormalities in vitro and in vivo, representing a promising opportunity to prevent MetS in the future with the utilization of TFs.

Molecular Mechanism Underlying the Regulatory Effect of Vine Tea on Metabolic Syndrome by Targeting Redox Balance and Gut Microbiota.

Metabolic syndrome (MS) is a metabolic disorder that arises from the increasing prevalence of obesity. The pathophysiology seems to be largely attributable to the imbalance of lipid and glucose metabolism, redox signaling pathways, and gut microbiota. The increased syndromes, such as type 2 diabetes and cardiovascular disease demands natural therapeutic attention for those at high risk. Vine tea, as a traditional medicinal and edible resource rich in flavonoids, especially for dihydromyricetin (DHM), exhibits promising health benefits on the intervention of MS, but the specific molecular mechanism has not been systematically elucidated. The present article aims to summarize the regulatory effects and biological targets of vine tea or DHM on MS, and analyze the underlying potential molecular mechanisms in cells, animals, and humans, mainly by regulating the redox associated signaling pathways, such as Nrf2, NF-κB, PI3K/IRS2/AKT, AMPK-PGC1α-SIRT1, SIRT3 pathways, and the crosstalk among them, and by targeting several key biomarkers. Moreover, vine tea extract or DHM has a positive impact on the modulation of intestinal microecology by upregulating the ratio of Firmicutes/Bacteroidetes (F/B) and increasing the relative abundance of Akkermansia muciniphila. Therefore, this review updated the latest important theoretical basis and molecular evidence for the development and application of vine tea in dietary functional products or drugs against MS and also imputed the future perspectives to clarify the deep mechanism among vine tea or DHM, redox associated signaling pathways, and gut microbiota.

Instant Dark Tea Alleviates Hyperlipidaemia in High-Fat Diet-Fed Rat: From Molecular Evidence to Redox Balance and Beyond.

Instant dark tea (IDT) is a new product gaining increasing attention because it is convenient and can endow significant health benefit to consumers, which is partially attributed to its high concentration of functional ingredients. However, the molecular mechanism underlying its regulatory effect on hyperlipidaemia is rarely studied. In this study, we performed omics and molecular verification in high-fat diet (HFD)-fed rat, aiming to reveal the mechanism and provide molecular evidence. The results showed that the major bioactive components in IDT were 237.9 mg/g total polysaccharides, 336.6 mg/g total polyphenols, and 46.9 mg/g EGCG. Rats fed with IDT (0.27-0.54 g/kg for 12 weeks) significantly reduced the body weight and TC, TG, LDL-C, blood glucose, and MDA and induced the level of serum HDL-C and also the levels of liver SOD, CAT, GSH-Px, and Nrf2, compared to HFD group. For molecular mechanism study, HIDT feeding had significant impact on the gene expressions of biomarkers in lipogenesis (FABP, CD36, SCD1, Cyp4a1, and Kcnn2), lipid oxidation (PPARγ), and glucose glycolysis (Gck and ENO2) in liver tissue. Moreover, gut microbiome study found that rats fed with IDT dramatically modified the gut microbial species at the family level, such as suppressing the increase abundance of Proteobacteria and Firmicutes induced by HFD. HIDT significantly boosted the relative composition of beneficial bacterium Akkermansia and Rikenellaceae_RC9_gut_group and decreased the relative abundance of the harmful bacterium Ruminococcaceae_UCG-005 and Ruminiclostridium_9, compared to HFD (p < 0.01). Correlation analysis between microbiome and animal indicators found that seven genera including Akkermansia, Clostridiales, Lachnospiraceae, Lachnospiraceae_UCG-010, Ruminiclostridium_9, Ruminococaceae-UCG-005, and Ruminocuccus_1 were found as potential biomarkers that were strongly correlated with oxidative stress and metabolism genes. For instance, Ruminococcaceae_UCG-005 was significantly correlated with body weight, TG, HDL-C, Nfr2, FABP3, SCD1, Cyp4a1, and Kcnn2. Collectively, the above data obtained in this study had provided the primary molecular evidence for the molecular mechanism and brought in novel insights based on omics for the regulatory effect of IDT on hyperlipidaemia.

Phytochemical activators of Nrf2: a review of therapeutic strategies in diabetes.

Insulin resistance (IR) is fundamental to the development of type 2 diabetes (T2D), and altered mitochondrial function and abnormal lipid distribution are closely associated with IR or T2D. Excess oxidative stress-induced mitochondrial damage leads to an imbalance in redox homeostasis, which is considered the major contributor to the progression of diabetes. A key cellular defense mechanism, namely, the nuclear factor-E2 p45-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway, plays an essential protective role in combating excess oxidative stress. A series of phytochemicals are reported to improve IR and restore mitochondrial function against excess oxidative stress by activating the Nrf2-ARE signaling pathway to maintain cellular reactive oxygen species (ROS) homeostasis. The present review focuses on key knowledge gaps in the Nrf2-ARE system targeted by phytochemicals and its correlation to diabetes both in the in vitro and in vivo models and recent achievements in human clinical trials to evaluate its efficiency and safety. In addition, we provide an overview of recent research progress in nutrigenomics, precision nutrition and the interactions occurring in gut microbiota associated with the Nrf2-ARE signaling pathway and diabetes chemoprevention by phytochemicals and finally propose a future research strategy for regulating redox and microbiota balance via the Nrf2-ARE pathway. The present review aims to help us comprehensively understand the critical chemopreventive role of the Nrf2-ARE pathway targeted by phytochemicals in diabetes.

Discovery of New Triterpenoids Extracted from Camellia oleifera Seed Cake and the Molecular Mechanism Underlying Their Antitumor Activity.

Theasaponin derivatives, which are reported to exert antitumor activity, have been widely reported to exist in edible plants, including in the seed cake of Camellia oleifera (C.), which is extensively grown in south of China. The purpose of this study was to isolate new theasaponin derivatives from C. seed cake and explore their potential antitumor activity and their underlying molecular mechanism. In the present study, we first isolated and identified four theasaponin derivatives (compounds 1, 2, 3, and 4) from the total aglycone extract of the seed cake of Camellia oleifera by utilizing a combination of pre-acid-hydrolysis treatment and activity-guided isolation. Among them, compound 1 (C1) and compound 4 (C4) are newly discovered theasaponins that have not been reported before. The structures of these two new compounds were characterized based on comprehensive 1D and 2D NMR spectroscopy and high-resolution mass spectrometry, as well as data reported in the literature. Secondly, the cytotoxicity and antitumor property of the above four purified compounds were evaluated in selected typical tumor cell lines, Huh-7, HepG2, Hela, A549, and SGC7901, and the results showed that the ED50 value of C4 ranges from 1.5 to 11.3 µM, which is comparable to that of cisplatinum (CDDP) in these five cell lines, indicating that C4 has the most powerful antitumor activity among them. Finally, a preliminary mechanistic investigation was performed to uncover the molecular mechanism underlying the antitumor property of C4, and the results suggested that C4 may trigger apoptosis through the Bcl-2/Caspase-3 and JAK2/STAT3 pathways, and stimulate cell proliferation via the NF-κB/iNOS/COX-2 pathway. Moreover, it was surprising to find that C4 can inhibit the Nrf2/HO-1 pathway, which indicates that C4 has the potency to overcome the resistance to cancer drugs. Therefore, C1 and C4 are two newly identified theasaponin derivatives with antitumor activity from the seed cake of Camellia oleifera, and C4 is a promising antitumor candidate not only for its powerful antitumor activity, but also for its ability to function as an Nrf2 inhibitor to enhance the anticancer drug sensitivity.

Effect of Garlic Organic Sulfides on Gene Expression Profiling in HepG2 Cells and Its Biological Function Analysis by Ingenuity Pathway Analysis System and Bio-Plex-Based Assays.

Garlic organic sulfides are dietary bioactive components with multiple biofunctions to prevent chronic diseases/inflammation and promote human health. DADS (diallyl disulfide), DATS (diallyl trisulfide), and DTS (diallyl tetrasulfide) are typical organic sulfides with similar structures from garlic. However, the structure-activity relationship of garlic organic sulfides remained unknown. The aim of the present study was to investigate the effect of DADS, DATS, and DTS on the gene expression profiling of human hepatocellular carcinoma cells (HepG2) by application of microarray and specialized analysis software, GO, Bio-Plex-based cytokines assay and IPA and analyze their structure-activity relationship according to antioxidant, anti-inflammatory, and metabolic-related properties. According to the microarray data, with the increase of S atom in garlic organic sulfides, its biological activity was gradually enhanced. In the general catalog of GO, garlic organic sulfides mainly affect biological process, molecular function, and cellular component. RT-qPCR results indicated that the microarray data is trustworthy, and the structure-activity analysis data found that more sulfur atoms have more powerful properties; thus, microarray data of DTS was preceded to the subsequent IPA analysis. The results of IPA analysis showed that the top 5 signaling pathways and molecular functions were disturbed by DTS; the molecular functions with the highest scores affected by DTS are cancer, cell apoptosis, and cell proliferation, which imply that the occurrence or metabolism of these diseases is related to the differential expression of the above-mentioned related genes and the activation of signaling channels, and the core of the most significant molecular network is inflammation. Finally, the results found that the secretions of 6 cytokines in macrophages were significantly inhibited by DTS treatment. This is the first study that analyzed the structure-activity relationship of garlic organic sulfides, which will provide useful genetic information for its multi-biofunction and promote their clinical application in the near future.

Dihydromyricetin Acts as a Potential Redox Balance Mediator in Cancer Chemoprevention.

Dihydromyricetin (DHM) is a flavonoid extracted from the leaves and stems of the edible plant Ampelopsis grossedentata that has been used for Chinese Traditional Medicine. It has attracted considerable attention from consumers due to its beneficial properties including anticancer, antioxidative, and anti-inflammatory activities. Continuous oxidative stress caused by intracellular redox imbalance can lead to chronic inflammation, which is intimately associated with the initiation, promotion, and progression of cancer. DHM is considered a potential redox regulator for chronic disease prevention, and its biological activities are abundantly evaluated by using diverse cell and animal models. However, clinical investigations are still scanty. This review summarizes the current potential chemopreventive effects of DHM, including its properties such as anticancer, antioxidative, and anti-inflammatory activities, and further discusses the underlying molecular mechanisms of DHM in cancer chemoprevention by targeting redox balance and influencing the gut microbiota.

Microarray Based Functional Analysis of Myricetin and Proteomic Study on Its Anti-Inflammatory Property.

Myricetin has been reported as a promising chemopreventive compound with multiple biofunctions. To evaluate its influence on gene expressions in genome-wide set and further investigate its anti-inflammatory property, the present study performed Gene Ontology and Ingenuity Pathway Analysis (IPA) to describe the basic gene expression characteristics by myricetin treatment in HepG2 cells, confirmed its multi-biofunction by real-time fluorescent quantitative PCR (RT-qPCR), and further verified its anti-inflammatory property by Western blotting and bio-plex-based cytokines assay. The IPA data showed that 337 gene expressions (48% of the top molecules) are disturbed over 2-fold, and the most possible biofunctions of myricetin are the effect on "cardiovascular disease, metabolic disease, and lipid metabolism," via regulation of 28 molecules with statistic score of 46. RT-qPCR data confirmed the accuracy of microarray data, and cytokines assay results indicated that 6 of the total 27 inflammatory cytokine secretions were significantly inhibited by myricetin pretreatment, including TNF-α, IFN-γ, IL-1α, IL-1ß, IL-2, and IL-6. The present study is the first time to elucidate the multi-function of myricetin in genome-wide set by IPA analysis and verify its anti-inflammatory property by proteomics of cytokines assay. Therefore, these results enrich the comprehensive bioactivities of myricetin and reveal that myricetin has powerful anti-inflammatory property, which provides encouragement for in vivo studies to verify its possible health benefits.

Extraction, identification, and antioxidant property evaluation of limonin from pummelo seeds.

Limonin, the main bioactive phytochemical constituent of limonoids with multi-functions, is enriched in citrus fruits and often found at a high concentration in citrus seeds. The present study was attempted to introduce a new and efficient extraction method to isolate limonoids from pummelo seeds, and to evaluate the antioxidant property of the main constituent limonin in HepG2 cells. Three key single factors were identified for the extraction of limonoids from pummelo seeds using the Box-Behnken experiment design of response surface methodology (RSM), and the optimized extraction parameters were treatment with 89.68 mL of anhydrous acetone for 4.62 h at 78.94 °C, while the yield of limonoids was 11.52 mg/g. The structure of isolated main constituent of the limonoids was further identified as limonin by Fourier transform infrared (FT-IR) spectrometer and nuclear magnetic resonance (NMR) spectrum. Moreover, the molecular data in HepG2 cells revealed that limonin exerted its anti-oxidant property mainly by the activation of nuclear factor (erythroid-2)-like 2 (Nrf2)/kelch-like ECH-associated protein 1 (Keap1)- antioxidant response element (ARE) pathway in the form of transcriptional regulation of Nrf2 mRNA and posttranscriptional regulation of Nrf2/Keap1 system. These results demonstrate that pummelo seeds are an ideal source of limonoids, and limonin is proved to exert its anti-oxidant property by the activation of Nrf2/Keap1 pathway.