A novel online preparative high-performance liquid chromatography system with the multiple trap columns-valve switch technique for the rapid and efficient isolation of main flavonoids from Epimedium koreanum Nakai.

In this work, a new online preparative high-performance liquid chromatography was developed for the fast and efficient separation of complex chemical mixtures from natural products. This system integrates two chromatographic systems into an online automatic separation system using the technique of multiple trap columns with valve switching. The sample was first separated into 18 subfractions in the online preparative high-performance liquid chromatography, and the sample eluents were then diluted and captured online on 18 trap columns by the multiple trap columns technique, respectively. Each subfraction retained on the trap column was transferred online to the separation column for the second separation. Finally, the target compounds were purified by appropriate separation conditions and multiple heart-cutting strategies. Importantly, the system was successfully used to separate 18 high-purity flavonoids from the crude extract of Epimedium koreanum Nakai online in one step. The entire separation time was approximately 20 h, and the structures were characterized by the high-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometry and nuclear magnetic resonance. This online preparative high-performance liquid chromatography system represents an efficient and rapid separation system that has the potential for a wide array of applications in the separation of complex chemical components from natural products.

Rapid classification and identification of chemical constituents in Epimedium koreanum Nakai by UPLC-Q-TOF-MS combined with data post-processing techniques.

INTRODUCTION: Epimedium koreanum Nakai (EKN), is a well-known Chinese herbal medicine for the treatment of osteoporosis, immunosuppression, tumours and cardiovascular diseases. Comprehensive component identification is essential for elucidation of its pharmacological mechanism and quality control. However, its complex chemical composition has caused certain difficulties in the analysis of this traditional Chinese medicine (TCM). Therefore, there is an urgent need to establish a method for rapid classification and identification of EKN chemical components. OBJECTIVE: To establish a method for rapid classification and identification of the main components of flavonoids, organic acids and alkaloids in EKN. METHODS: The samples were analysed by ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) and data post-processing techniques. The UPLC system used a BEH C18 column to separate the total extract of EKN. The mobile phase consisted of 0.1% formic acid in water and acetonitrile, and the EKN extract was analysed by gradient elution at a flow rate of 0.4 mL/min. In both the positive and negative ion modes, the fragment information was obtained and compared with those of the characteristic fragmentations and neutral losses described in the literature to quickly identify the target compounds. RESULTS: Finally, we successfully screened out 51 chemical components, including 40 flavonoids, nine organic acids, and two alkaloids. CONCLUSION: The established method not only comprehensively analysed the chemical compositions of EKN, solved the difficult problems of analysis and identification of the complex chemical compositions of the TCM, but also further promoted the development of the application of chemical compositions of TCM.

Effectiveness of Prenyl Group on Flavonoids from Epimedium koreanum Nakai on Bacterial Neuraminidase Inhibition.

In this study, the inhibitory potential of bacterial neuraminidase (NA) was observed on the leaves of Epimedium koreanum Nakai, which is a popular ingredient in traditional herbal medicine. This study attempted to isolate the relevant, responsible metabolites and elucidate their inhibition mechanism. The methanol extraction process yielded eight flavonoids (1⁻8), of which compounds 7 and 8 were new compounds named koreanoside F and koreanoside G, respectively. All the compounds (1⁻8) showed a significant inhibition to bacterial NA with IC50 values of 0.17⁻106.3 µM. In particular, the prenyl group on the flavonoids played a critical role in bacterial NA inhibition. Epimedokoreanin B (compound 1, IC50 = 0.17 µM) with two prenyl groups on C8 and C5' of luteolin was 500 times more effective than luteolin (IC50 = 85.6 µM). A similar trend was observed on compound 2 (IC50 = 0.68 µM) versus dihydrokaempferol (IC50 = 500.4 µM) and compound 3 (IC50 = 12.6 µM) versus apigenin (IC50 = 107.5 µM). Kinetic parameters (Km, Vmax, and Kik/Kiv) evaluated that all the compounds apart from compound 5 showed noncompetitive inhibition. Compound 5 was proven to be a mixed type inhibitor. In an enzyme binding affinity experiment using fluorescence, affinity constants (KSV) were tightly related to inhibitory activities.

Potential of Icariin Metabolites from Epimedium koreanum Nakai as Antidiabetic Therapeutic Agents.

The therapeutic properties of Epimediumkoreanum are presumed to be due to the flavonoid component icariin, which has been reported to have broad pharmacological potential and has demonstrated anti-diabetic, anti-Alzheimer's disease, anti-tumor, and hepatoprotective activities. Considering these therapeutic properties of icariin, its deglycosylated icaritin and glycosylated flavonoids (icaeriside II, epimedin A, epimedin B, and epimedin C) were evaluated for their ability to inhibit protein tyrosine phosphatase 1B (PTP1B) and α-glucosidase. The results show that icaritin and icariside II exhibit potent inhibitory activities, with 50% inhibition concentration (IC50) values of 11.59 ± 1.39 µM and 9.94 ± 0.15 µM against PTP1B and 74.42 ± 0.01 and 106.59 ± 0.44 µM against α-glucosidase, respectively. With the exceptions of icaritin and icariside II, glycosylated flavonoids did not exhibit any inhibitory effects in the two assays. Enzyme kinetics analyses revealed that icaritin and icariside II demonstrated noncompetitive-type inhibition against PTP1B, with inhibition constant (Ki) values of 11.41 and 11.66 µM, respectively. Moreover, molecular docking analysis confirmed that icaritin and icariside II both occupy the same site as allosteric ligand. Thus, the molecular docking simulation results were in close agreement with the experimental data with respect to inhibition activity. In conclusion, deglycosylated metabolites of icariin from E. koreanum might offer therapeutic potential for the treatment of type 2 diabetes mellitus.

Transcriptome Analysis on the Quality of Epimedium koreanum in Different Soil Moisture Conditions at Harvesting Stage.

Epimedium koreanum is a traditional Chinese tonic herb. Its main medicinal components are secondary metabolites such as flavonoids and flavonol glycosides, but the biosynthetic mechanism is still unclear. Moisture conditions are a key environmental factor affecting E. koreanum medicinal components during harvesting. Different stages of E. koreanum under natural conditions after rainfall were selected to study changes in physiological properties, herb quality, and transcriptome. Malondialdehyde (MDA) content increased significantly in the D3 stage after rainfall, and protective enzyme levels also rose. Additionally, the flavonol glycoside content was relatively high. We sequenced the transcriptomes of D1, D3, and D9 (R) and identified differentially expressed genes (DEGs) related to flavonoid synthesis. This analysis allowed us to predict the roadmap and key genes involved in flavonoid biosynthesis for E. koreanum. These results suggest that the E. koreanum quality can be enhanced by natural drought conditions in the soil after precipitation during harvest. The harvesting period of E. koreanum is optimal when soil moisture naturally dries to a relative water content of 26% after precipitation. These conditions help E. koreanum tolerate a certain level of water scarcity, resulting in increased expression of flavonoid-related genes and ultimately enhancing the quality of the herb.

Comparative studies on antioxidant activities of extracts and fractions from the leaves and stem of Epimedium koreanum Nakai.

The antioxidant potency of various extracts and fractions from the leaves and stem of Epimedium koreanum Nakai was evaluated using three esteblished methods, specifically the 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radica-scavenging activity assay, the inhibitory effect on lipid peroxidation induced by Fe(2+)/ascorbate (MDA) assay and the ferric reducing power (FRP) assay. The amounts of total phenolics and total flavonoids in the extracts and fractions were determined by spectrophotometric methods and the content of icariin was determined by HPLC. The results showed that all the extracts and fractions exhibited antioxidant activities at different magnitudes of potency. The leaf extract and fractions demonstrated superior antioxidant activity in most of the assays. The decreasing order of antioxidant activities among the extracts/fractions assayed through the three methods were found to be n-BuOH fraction>ethyl acetate fraction>ethanol extract>petroleum ether fraction>water fraction. A positive correlation was found between the amounts of total phenolics, total flavonoids and icariin and DPPH radical scavenging activity (R(2) = 0.9935, 0.9944 and 0.9997, respectively) and inhibitory activity on lipid peroxidation (R(2) = 0.9987, 0.9830 and 0.9886, respectively). The results suggested that Icariin was one of the main constituents contribute to the antioxidant activity of Epimedium koreanum Nakai and the n-BuOH fractions of leaf extract might be valuable antioxidant natural sources.

Two new prenylated flavonol glycosides from Epimedium koreanum Nakai leaves and their anti-pulmonary fibrosis activities.

Two new prenylated flavonol glycosides, namely Desmethylicaritin-3-O-ß-D-glucopyranosyl-(1→3)-α-L(4″-O-acetyl) rhamnopyranosyl-7-O-ß-D(6''''-O-acetyl)-glucopyranoside (1) and 5,7,3',4'-tetrahydroxyl-8-(3,3-dimethylallyl)-flavonol-3-O-α-L-rhamnopyranoside (2), and one with no NMR spectral data reported (3) were isolated from Epimedium koreanum Nakai. Their structures were elucidated by 1D, 2D NMR and HRESIMS analysis. The identification of the sugar moieties was carried out by means of acid hydrolysis and HPLC analysis of their derivatives. The anti-pulmonary fibrosis activities result showed that compound 2 had significant inhibitory effects on A549 cell fibrosis, which was similar to that of the positive control drug, pirfenidone.

Diversity analysis of leaf endophytic fungi and rhizosphere soil fungi of Korean Epimedium at different growth stages.

BACKGROUND: Rhizosphere fungi and endophytic fungi play key roles in plant growth and development; however, their role in the growth of Epimedium koreanum Nakai at different stages remains unclear. Here, we used the Illumina MiSeq system, a high-throughput sequencing technology, to study the endophytic fungi and rhizosphere microbiome of Korean Epimedium. RESULTS: Epimedium koreanum Nakai rhizosphere soil and leaves had highly diverse fungal communities during the growth process. The relative abundance of soil fungi in the rhizosphere stage was higher than that of leaf endophytic fungi in the early growth stage, but the overall abundance was basically equal. Sebacina is a significantly divergent fungal genera, and Sebacina sp. are present among leaf fungi species in the rhizosphere soil of Epimedium koreanum Nakai. Sebacina sp. can move to each other in rhizosphere soil fungi and leaf endophytes. VIF (variance inflation factor) analysis showed that soluble salt, whole nitrogen, alkaline lysis nitrogen, whole phosphorus, total potassium, and fast-acting potassium are useful environmental factors for rhizosphere soil and leaf endophytic fungi: potassium, total nitrogen, whole phosphorus, and three environmental factors were significantly and positively associated with the relative abundance of Sebacina sp. CONCLUSIONS: (1) This study is the first to clarify the species diversity of fungi in Epimedium koreanum Nakai leaf and rhizosphere soil. (2) Different fungal communities of rhizosphere soil fungi and leaf endophytic fungi at different growth stages of Epimedium koreanum Nakai were examined. (3) Sebacina sp. can move to each other between rhizosphere soil fungi and leaf endophytic fungi. (4) Nitrogen, phosphorus and potassium elements in the environment have a significant positive effect on the relative abundance of Sebacina sp.

Antiviral activity of Epimedium koreanum Nakai water extract against influenza viruses.

Epimedium koreanum Nakai (EKN) is a popular plant in Korean and Chinese medicine for treating a variety of ailments. The aqueous extract of EKN has a significant inhibitory impact on influenza A virus (IAV) infection by directly blocking viral attachment and having a virucidal effect, according to this study. Using fluorescent microscopy and fluorescence-activated cell sorting (FACS) with a green fluorescent protein (GFP)-tagged Influenza A/PR/8/34 virus, we examined the effect of EKN on viral infection. By viral infection, EKN strongly suppresses GFP expression, and at a dosage of 100 µg/mL, EKN decreased GFP expression by up to 90% of the untreated infected control. Immunofluorescence and Western blot analyses against influenza viral proteins revealed that EKN decreased influenza viral protein expression in a dose-dependent manner. EKN inhibited the H1N1 influenza virus's hemagglutinin (HA) and neuraminidase (NA), preventing viral attachment to cells. Furthermore, EKN had a virucidal impact and inhibited the cytopathic effects of H1N1, H3N2 and influenza B virus infection. Finally, our findings show that EKN has the potential to be developed as a natural viral inhibitor against influenza virus infection.

Epimedium koreanum Nakai-Induced Liver Injury-A Mechanistic Study Using Untargeted Metabolomics.

Epimedii Folium is widely used worldwide as an herbal supplement, and the risk of its induced liver damage has emerged in recent years. Our preliminary study has found that, among several Epimedii Folium species specified in the Chinese Pharmacopoeia, Epimedium koreanum Nakai has a more severe propensity for hepatotoxicity. However, the mechanism of hepatotoxicity of Epimedium koreanum Nakai is still unclear. In this study, untargeted metabolomics was performed to analyze the serum and liver tissue to explore the mechanism of hepatotoxicity of Epimedium koreanum Nakai. The results of experiments in vivo showed that, after 28 days of exposure to Epimedium koreanum Nakai ethanol extract (EEE), the liver weight, levels of AST, ALP, TBIL, etc. in serum of rats in the EEE group were significantly increased, as well as severe cytoplasmic vacuolation appeared in the liver tissue, which suggested that EEE has significant hepatotoxicity. Subsequently, the results of metabolomics revealed significant changes in the metabolic profile in the liver and serum of rats after EEE exposure, in which metabolites in serum such as flavin mononucleotide, phenylacetylglycine, glutathione, l-tryptophan, and sphingomyelin were able to accurately identify liver injury caused by EEE and could be used as serum markers to reflect EEE-induced liver injury. The KEGG pathway enrichment analysis revealed that EEE caused extensive effects on rats' metabolic pathways. Some of the most affected pathways included glutathione metabolism, glutamate metabolism pathway, primary bile acid biosynthesis pathway, and sphingolipid metabolism pathway, which were all directed to the biological process of ferroptosis. Then, the main markers related to ferroptosis in the liver were examined, and the results demonstrated that the content of malondialdehyde was significantly increased, the activity of superoxide dismutase was significantly reduced, the ferroptosis inhibitory proteins GPX4 and System xc - were significantly downregulated, and the ferroptosis-promoting protein ACSL4 was significantly up-regulated. Judging from these results, we concluded that the mechanism of hepatotoxicity of Epimedium koreanum Nakai was probably related to the induction of ferroptosis in hepatocytes.

Flavonoids from the leaves of Epimedium Koreanum Nakai and their potential cytotoxic activities.

Phytochemical studies on the leaves of Epimedium koreanum Nakai have resulted in the discovery of two new flavonol glycosides, koreanoside F (1) and koreanoside G (2), along with six known flavonoids. Their structures were elucidated on the basis of HRESIMS, UV, IR, 1 D NMR and 2 D NMR data. Absolute configurations of 1 and 2 was further determined by 13C-NMR spectra with gate decoupling (GD). All of the compounds were evaluated for cytotoxic activities by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazoliumbromide (MTT) assay. The results indicated that compounds 3, 5, 6, 7 and 8 inhibited the proliferation of A549 and NCI-292 cells with IC50 values of 5.7-23.5 µM. Real-time monitoring in three kinds of lung cancer cells and a kind of human bronchial epithelial cells treated with compound 6 was also assessed.

A novel herbal formulation consisting of red ginseng extract and Epimedium koreanum Nakai-attenuated dextran sulfate sodium-induced colitis in mice.

BACKGROUND: Ulcerative colitis (UC) is a commonly encountered large intestine disease in the contemporary world that terminates into colorectal cancer; therefore, the timely treatment of UC is of major concern. Panax ginseng Meyer is an extensively consumed herbal commodity in South East Asian countries, especially Korea. It exhibits a wide range of biologically beneficial qualities for almost head-to-toe ailments in the body. Epimedium koreanum Nakai (EKN) is also a widely used traditional Korean herbal medicine used for treating infertility, rheumatism, and cardiovascular diseases. MATERIALS AND METHODS: Separately the anti-inflammatory activities of both red ginseng extracts (RGEs) and EKNs had been demonstrated in the past in various inflammatory models; however, we sought to unravel the anti-inflammatory activities of the combination of these two extracts in dextran sulfate sodium (DSS)-induced ulcerative colitis in mice model because the allopathic remedies for UC involve more side effects than benefits. RESULTS: Our results have shown that the combination of RGE + EKN synergistically alleviated the macroscopic lesions in DSS-induced colitic mice such as colon shortening, hematochezia, and weight loss. Moreover, it restored the histopathological lesions in mice and decreased the levels of pro-inflammatory mediators and cytokines through the repression of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and nucleotide-binding domain (NOD)-like receptor protein 3 (NLRP-3) expression. In vitro, this combination also reduced the magnitude of nitric acid (NO), pro-inflammatory mediators and cytokine through NF-κB and mitogen-activated protein kinase (MAPK) pathways in RAW 264.7 mouse macrophage cells. CONCLUSION: In the light of these findings, we can endorse this combination extract as a functional food for the prophylactic as well as therapeutic treatment of UC in humans together with allopathic remedies.

Chemical constituents from Epimedium koreanum Nakai and their chemotaxonomic significance.

In this study, 21 compounds were isolated from the aerial parts of Epimedium koreanum Nakai, including 6 phenols (1-5 and 10), 4 lignans (6-9) and 11 flavonoids (11-21). The chemical structures of those isolates were established after extensive one-and two-dimensional nuclear magnetic resonance and mass spectroscopy analyses. This is the first report of compounds 2, 3, 4 and 9 in E. koreanum and of compounds 5 and 7 in the family Berberidaceae. The chemotaxonomic significance of the isolated compounds is discussed.

Structural analysis and antioxidant activities of neutral polysaccharide isolated from Epimedium koreanum Nakai.

The structure and antioxidant activity of a neutral polysaccharide (named as EFPN-1) from Epimedium koreanum Nakai were studied. Based on the results from high performance gel permeation chromatography, methylation GC-MS and 1D/2D nuclear magnetic resonance spectroscopy, EFPN-1 was a branched polysaccharide with a molecular weight of 21.9 kD and containing over 60% of glucose. Its backbone was composed of (1 → 4)-ß-d-Glcp and (1 → 3)-ß-d-Galp, branched partially at O-6 of both residues, with a degree of branching of 40.4% and terminated mainly with Ara and Gal, as well as trace amount of Gal. Antioxidant tests showed that EFPN-1 exhibited a strong capacity on DPPH and hydroxyl radical scavenging, ferrous ion chelation and reducing antioxidant powder. EFPN-1 has potential to be developed as a natural antioxidant.

Effect of drying processes on prenylflavonoid content and antioxidant activity of Epimedium koreanum Nakai.

Epimedium koreanum Nakai is a famous Chinese herbal medicine for the treatment of impotence, osteoporosis, immune suppression and cardiovascular diseases. Drying is the most common and fundamental procedure in post-harvest processing of E. koreanum, which contributes to the variations of flavonoid content, especially prenylflavonoids, the bioactive components. In present study, effect of drying processes on flavonoid content and antioxidant activity were investigated. High performance liquid chromatography coupled with diode-array detection and electrospray ionization quadrupole time-of-flight tandem mass spectrometry methods were employed. Twenty seven compounds were identified and 11 of them, including eight prenylflavonoids and three other types of flavonoids, were further quantified. The antioxidant activity of E. koreanum was evaluated by the 2,2-diphenyl-1-picrylhydrazyl free radical scavenging method. The results showed that content of the eight prenylflavonoids exhibited significant variations after different drying processes, especially icariin and baohuoside I. The variation tendency of antioxidant activity was positively correlated with the content of total flavonoid, afzelin and icariin.

Epimedium koreanum Ameliorates Oxidative Stress-Mediated Liver Injury by Activating Nuclear Factor Erythroid 2-Related Factor 2.

Oxidative stress induced by reactive oxygen species is the main cause of various liver diseases. This study investigated the hepatoprotective effect of Epimedium koreanum Nakai water extract (EKE) against arachidonic acid (AA)[Formula: see text][Formula: see text][Formula: see text]iron-mediated cytotoxicity in HepG2 cells and carbon tetrachloride (CCl4-)-mediated acute liver injury in mice. Pretreatment with EKE (30 and 100[Formula: see text][Formula: see text]g/mL) significantly inhibited AA[Formula: see text][Formula: see text][Formula: see text]iron-mediated cytotoxicity in HepG2 cells by preventing changes in the expression of cleaved caspase-3 and poly(ADP-ribose) polymerase. EKE attenuated hydrogen peroxide production, glutathione depletion, and mitochondrial membrane dysfunction. EKE also increased the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2), transactivated anti-oxidant response element harboring luciferase activity, and induced the expression of anti-oxidant genes. Furthermore, the cytoprotective effect of EKE against AA[Formula: see text][Formula: see text][Formula: see text]iron was blocked in Nrf2 knockout cells. Ultra-performance liquid chromatography analysis showed that EKE contained icariin, icaritin, and quercetin; icaritin and quercetin were both found to protect HepG2 cells from AA[Formula: see text][Formula: see text][Formula: see text]iron via Nrf2 activation. In a CCl4-induced mouse model of liver injury, pretreatment with EKE (300[Formula: see text]mg/kg) for four consecutive days ameliorated CCl4-mediated increases in serum aspartate aminotransferase activity, histological activity index, hepatic parenchyma degeneration, and inflammatory cell infiltration. EKE also decreased the number of nitrotyrosine-, 4-hydroxynonenal-, cleaved caspase-3-, and cleaved poly(ADP-ribose) polymerase-positive cells in hepatic tissues. These results suggest EKE is a promising candidate for the prevention or treatment of oxidative stress-related liver diseases via Nrf2 activation.

A new flavonol glycoside from the aerial parts of Epimedium koreanum Nakai.

A new prenylated flavonol glycoside (1) was isolated from a 95% methanol extract of the dried and powdered aerial parts of Epimedium koreanum Nakai (Herba Epimedii), along with seven previously known flavonoids (2-8). The chemical structure of the new compound (1) was established to be 5-hydroxy-4'-methoxy-8-(2-hydroxy-3-methyl-3-butenyl)flavone 3-O-α-l-rhamnopyranosyl-7-O-ß-D-gluco pyranoside on the basis of spectroscopic methods. The antioxidant activities of these compounds were determined by the DPPH (2,2-diphenyl-1-picrylhydrazyl) free radical scavenging assay and kaempferitrin (8) showed a high reactivity with DPPH.

Polysaccharides from Epimedium koreanum Nakai with immunomodulatory activity and inhibitory effect on tumor growth in LLC-bearing mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Epimedium koreanum Nakai is documented as tonic herbal in China for over a thousand years and has the potential to enhance the body's immunity according to the theory of traditional Chinese medicine. Polysaccharides are one of the most important effective compounds in Epimedium koreanum Nakai. Accumulating evidence indicated polysaccharides derived from traditional Chinese medicine have potent immune-enhancing properties and relatively nontoxic effects in cancer treatment. However, information about immunological regulation in tumor of Epimedium koreanum Nakai polysaccharides is limited and the reports of purification, characterization of polysaccharides have remained less. The purpose of our study was to further investigate the active polysaccharides from Epimedium koreanum Nakai by evaluating the immune-regulation activities in tumor-bearing mice and provide reasonable explanation for traditional application. MATERIALS AND METHODS: We firstly purified Epimedium koreanum polysaccharide (EPS) from crude extracts and evaluated EPS in vitro using immunological experiments including maturation and Ag presentation function of DCs, CD4 T-cell differentiation and secretion of anti-cancer cytokines. In LLC-bearing mice model, we investigated its antitumor activities through evaluation of tumor cell proliferative activity, calculation of immune organ indexes and relative host immune system function tests. RESULTS: Results showed that EPS (180 × 104Da) was composed of mannose (Man), rhamnose (Rha), glucuronic acid (GlcUA), galactosamine (GalN), glucose (Glc), galactose (Gal), arabinose (Ara) and fructose (Fuc). Chemical composition assay indicated EPS was a fraction with 28.20% uronic acid content. FT-IR suggested the presence of pyraoid ring in EPS and SEM displayed smooth surface embedded by several pores. Moreover, Our study suggested EPS could remarkably stimulate macrophages to secrete substantial anti-cancer cytokines and promote maturation as well as Ag presentation function of DCs. Strikingly, CD4 T-cell differentiation and increased INF-γ production stimulated by EPS-activated macrophages were observed in the research. Furthermore, EPS exhibited prominent antitumor activities through regulating host immune system function in LLC-bearing mice. Taken together, experimental findings suggested EPS could be regarded as a potential immune-stimulating modifier for cancer therapy. CONCLUSION: Our studies demonstrated the polysaccharide (180 × 104Da) purified from Epimedium koreanum Nakai could promote maturation and Ag presentation function of DCs, increase the level of immunomodulatory cytokines and activate CD4 T-cell differentiation. Furthermore, it may inhibit the tumor growth in LLC-bearing mice through regulating host immune system function.

The potential of Epimedium koreanum Nakai for herb-drug interaction.

OBJECTIVES: This study aims to investigate potential herb-drug interactions (HDI) of Epimedium koreanum Nakai. METHODS: Human liver microsomes (HLMs) were used to determine the enzyme kinetics of the major human cytochrome P450s (CYPs). Inducible potential of E. koreanum on CYP1A2, 2B6, 2C19 and 3A4 activities of human primary hepatocytes was also examined. KEY FINDINGS: Ethanol extract of E. koreanum showed direct inhibitory potency for CYP1A2 (IC50  = 121.8 µg/ml, Ki  = 110.7 ± 36.8 µg/ml) and CYP2B6 (IC50  = 59.5 µg/ml, Ki  = 18.1 ± 2.9 µg/ml). For CYP2C9, 2C19, 2D6, 2E1 and 3A4, only negligible effect was observed. Time-dependent (irreversible) inhibition by E. koreanum was observed for CYP1A2 (KI  = 32.9 ± 18.4 µg/ml, kinact  = 0.031 ± 0.006 min-1 ). However, ethanol extract of E. koreanum (1.5-150 µg/ml) did not change the activity or mRNA expressions for CYP3A4, 1A2, 2C19 and 2B6. CONCLUSIONS: The ethanol extract of E. koreanum is not likely to cause HDI via inducing the major human CYPs. But the potential for interactions between E. koreanum extract and substrates of CYP1A2 or 2B6 cannot be overlooked.

Neuroprotective effects of total flavonoid fraction of the Epimedium koreanum Nakai extract on dopaminergic neurons: In vivo and in vitro.

Flavonoids, the active components of Epimedii Genus, have been demonstrated to protect against osteoporosis, cardiovascular diseases and rheumatoid arthritis. The present study aimed to investigate the neuroprotective effects of total flavonoid (TF) fraction of Epimedium koreanum Nakai on dopaminergic neurons in the cellular and mice models of Parkinson's disease (PD). TF pretreatment could ameliorate the decrease of striatal dopamine (DA) content and the loss of tyrosine hydroxylase (TH)-immunoreactive neurons in the substantia nigra pars compacta (SNpc) induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). TF treatment could reverse the changes of Bcl-2 and Bax protein expressions in the striatum of PD mice. 1-Methyl-4-phenylpyridinium ion (MPP+) significantly decreased the cell viability and mitochondrial membrane potential in MES23.5 cells. These effects could be reversed by TF treatment. In addition, MPP+-induced changes of Bcl-2 and Bax mRNA and protein expressions were also reversed by TF pretreatment. These data demonstrated that TF of E. koreanum Nakai could protect against MPTP-induced dopaminergic neuronal death in mice and MPP+-induced neurotoxicity in dopaminergic MES23.5 cells. Anti-apoptosis might be involved in this process.