• Journal of Microbiology and Biotechnology
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• v.27 no.7
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• pp.1257-1265
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• 2017

The present study was carried out to investigate the antioxidative and neuroprotective effects of sea buckthorn (Hippophae rhamnoides L.) leaves (SBL) harvested at different times. Reversed-phase high-performance liquid chromatography analysis revealed five major phenolic compounds: ellagic acid, gallic acid, isorhamnetin, kaempferol, and quercetin. SBL harvested in August had the highest total phenolic and flavonoid contents and antioxidant capacity. Treatment of neuronal PC-12 cells with the ethyl acetate fraction of SBL harvested in August increased their viability and membrane integrity and reduced intracellular oxidative stress in a dose-dependent manner. The relative populations of both early and late apoptotic PC-12 cells were decreased by treatment with the SBL ethyl acetate fraction, based on flow cytometry analysis using annexin V-FITC/PI staining. These findings suggest that SBL can serve as a good source of antioxidants and medicinal agents that attenuate oxidative stress.

• Natural Product Sciences
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• v.22 no.1
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• pp.64-69
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• 2016

A method for simultaneously identifying antioxidative compounds was developed using time-based LC-MS coupled with DPPH assay regardless of the time consuming process. The methanolic extract of Polygonum aviculare (Polygonaceae) showed significant DPPH radical scavenging activity. Time-based DPPH assay for simultaneous identification of active compounds from the extracts of P. aviculare was used. Major peaks of ethyl acetate fraction of P. aviculare showed high DPPH radical scavenging activity. A simple phenolic compound (1) and six flavonoids (2-7) were isolated from the ethyl acetate fraction of P. aviculare by silica gel and sephadex LH-20 column chromatography. The structures of seven compounds were determined to be protocatechuic acid (1), catechin (2), myricitrin (3), epicatechin-3-O-gallate (4), avicularin (5), quercitrin (6), and juglanin (7) based on the analysis of the $^1H$-NMR, $^{13}C$-NMR and ESI-MS data. All compounds exhibited significant antioxidant activity on DPPH assay and active compounds were well correlated with predicted one.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2022.09a
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• pp.106-106
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• 2022

The cone of Red Pine (Pinus densiflora), which has been used as a drug in traditional medicine. Its ethyl acetate fraction was reported to exert antioxidant, anti-melanogenesis, and anti-inflammation activites. Apoptosis of hepatic stellate cells (LX-2) is regarding as a potential strategy for alleviation of hepatic fibrosis. We conducted to investigated whether the treatment of cone has a potential to control of some factors related in apoptosis and autophagy in cell signaling pathways. We suggest that the cone induced apoptosis through confirming the expression levels of genes (cPARP, Bcl-XL, Bax, p53, and caspase-3) in LX-2 cells. Also, the cone may regulate autophagy (LC3, p62, Beclin-1, and ATG12). Remarkably, the treatment of cone may affect to formation of autophagosomes in the immunofluorescence image in live cells. These findings suggest that the ethyl acetate fraction from the cone of Red Pine (P. densiflora) may have potential as an alternative therapeutic agent for the alleviation and prevention of liver fibrosis.

• Journal of Life Science
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• v.23 no.8
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• pp.989-997
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• 2013

In this study, based on the antioxidative effects in organic solvent fractions obtained from the main methanolic extract of L. japonica, the protective cellular effects and gene expression patterns of ethyl acetate fractions on $H_2O_2$-induced Raw 264.7 cell death ($IC_{50}$) were analyzed. The antioxidant activity of the fractions measured using DPPH free radical scavenging activity increased in a dose-dependent manner, and the $ED_{50}$ exhibited the highest $39.56{\mu}g/ml$ in the ethyl acetate fraction. In addition, the ethyl acetate fractions' cell viability on $H_2O_2$-induced Raw 264.7 cell damage increased in a concentration-dependent manner, showed a visible cell survival rate of 82.49% at a concentration of $100{\mu}g/ml$. The gene expression patterns related to the ethyl acetate fractions' cytoprotective effect in $H_2O_2$-induced Raw 264.7 cell damage presented similar patterns to those of BHA. In comparative analysis for antioxidant activity-related genes affected by ethyl acetate fractions and BHA in $H_2O_2$-induced Raw 264.7 cells, both ethyl acetate fractions and BHA showed very similar gene expression patterns, but the gene expression level of the heme oxygenase 1 (Hmox1) gene making antioxidant enzymes in cells was four times higher in ethyl acetate fractions than BHA. In inflammation-related genes in $H_2O_2$induced Raw 264.7 cells, the T-box transcription factor (Tbx21) gene was expressed about two times more frequently in the ethyl acetate fraction treatment group, while it was expressed half as frequently in the BHA treatment group.

• Applied Chemistry for Engineering
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• v.24 no.2
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• pp.177-183
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• 2013

In this study, the antioxidative activities and component analysis of Broussonetia kazinoki SIEB (B. kazinoki). extracts were investigated. B. kazinoki extract showed the effective free radical (1,1-diphenyl-2-picrylhydrazyl, DPPH) scavenging activity ($FSC_{50}=8.53{\mu}g/mL$). Reactive oxygen species (ROS) scavenging activity ($OSC_{50}$) of the ethyl acetate fraction of B. kazinoki. extracts in the luminol-dependent $Fe^{3+}-EDTA/H_2O_2$ system was $1.69{\mu}g/mL$. The ethyl acetate fraction of B. kazinoki. extracts also exhibited more prominent cellular protective effects (${\tau}_{50}$, 183.3 min at $10{\mu}g/mL$) than that of typical antioxidant $\alpha$-tocopherol (${\tau}_{50}$ = 38.00 min) in the $^1O_2$-induced photohemolysis of human erythrocytes. Components of the ethyl acetate fraction obtained from B. kazinoki extracts were analyzed by TLC, HPLC chromatogram, LC/ESI-MS/MS and $^1H$-NMR. Consequently, Components, components were identified as the kazinol J of kazinol series and luteolin (2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-4-chromenone) of flavonoid series having antioxidant activities. These results indicate that extract/ fraction of B. kazinoki can be used as antioxidants in biological systems, particularly skins exposed to UV radiation by quenching and/or scavenging $^1O_2$ and other ROS, and protecting cellular membranes against ROS. Thus, the extract/fraction of B. kazinoki could be applicable to new cosmeceuticals.

• Journal of Korean Society of Forest Science
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• v.95 no.4
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• pp.429-434
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• 2006

The dried needles (1.5 kg) of Picea abies Karsten were ground, extracted with acetone-$H_2O$ (7:3, v/v), concentrated, and fractionated with a series of n-hexane, methylene chloride, ethyl acetate and water on a separatory funnel. Each fraction was freeze dried, then a portion of ethyl acetate soluble powder was chromatographed on a Sephadex LH-20 column using a series of aqueous methanol and ethanol-n-hexane mixture as eluents. The isolated compounds were identified by cellulose TLC, $^1H$-, $^{13}C$-NMR, FAB and EI-MS. (+)-catechin (compound I), (-)-epicatechin (compound II), kaempferol-3-O-${\beta}$-D-glucopyranoside (compound III), 4-hydroxyacetophenone (compound IV) were isolated from the ethyl acetate soluble fraction and (+)-catechin (compound I), protocatechuic acid (compound V) were isolated from the $H_2O$ soluble fraction of P. abies needle. The antioxidative activities of each fraction and the isolated compounds were tested by DPPH radical scavenging method, and EtOAc soluble fraction, (+)-catechin and (-)-epicatechin showed similar values to ${\alpha}$-tocopherol and BHT as controls.

• Natural Product Sciences
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• v.5 no.3
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• pp.113-120
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• 1999

Nitric oxide (NO) is a free radical synthesized from L-arginine by nitric oxide synthase (NOS). It is believed that NO is an important mediator in numerous physiological and inflammatory responses. Particularly, a large amount of NO released from the inducible nitric oxide synthase (iNOS) is mostly associated with inflammatory processes. Overproduction of NO in these processes including sepsis and autoimmune diseases can have deleterious consequences and pathophysiologic relevance. Therefore, for the discovery of new inhibitory agents against iNOS activity, we have evaluated about 100 kinds of natural products after partition into three layers (n-hexane, ethyl acetate and aqueous) from 100% methanol extracts to study inhibitory effects on iNOS activity induced by lipopolysaccharide (LPS) in RAW264.7 cells culture system. As a positive control, curcumin, which is known as an anti-tumor promoter, anti-inflammatory agent as an iNOS inhibitor, was used and showed the dose-dependent inhibitory effect $(IC_{50},\;2.5\;{\mu}g/ml)$. Among tested fractions, the n-hexane fraction of Cimicifuga heracleifolia $(IC_{50}:\;9.65\;{\mu}g/ml)$, Forsythiae fructus $(IC_{50}:\;6.36\;{\mu}g/ml)$, Saposhnikovia divaricata $(IC_{50}:\;5.92\;{\mu}g/ml)$, and the ethyl acetate fraction of Chrysanthemum sibiricum $(IC_{50}:\;2.56\;{\mu}g/ml)$, Gastrodia elata $(IC_{50}:\;3.46\;{\mu}g/ml)$, and the aqueous fraction of Dianthus chinensis $(IC_{50}:\;6.73\;{\mu}g/ml)$, Euonymus alatus $(IC_{50}:\;6.78\;{\mu}g/ml)$, Mechania urticifoloria $(IC_{50}:\;8.01\;{\mu}g/ml)$ showed strong inhibitory activity against LPS-stimulated iNOS. Especially, the ethyl acetate fraction of Chrysanthemum sibiricum $(IC_{50}:\;2.56\;{\mu}g/ml)$, which exhibited the strongest inhibition against iNOS, was fractionated with silica-gel column chromatography. These subfractions exhibited dose-dependent inhibition against iNOS activity in the range of $2.59-5.6\;{\mu}g/ml$ except for fraction No. 3, 4, 5, 6, 8, 9, and 16. Our study shows that Chrysanthemum sibiricum has the strongest inhibitory effect against iNOS activity and has similar effect to curcumin. Therefore, further studies for the identification of active principles from Chrysanthemum sibiricum and investigation for the mechanism of the inhibition of iNOS by active principles will be performed.

• Journal of Life Science
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• v.26 no.1
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• pp.117-122
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• 2016

In the current study, the ethanol extracts and their subsequent organic solvent fractions from lees of sweet potato soju were prepared and the prepared samples were designated as from KSD-E8-1 to KSD-E8-5. Their effects on cell viability and nitric oxide (NO) production in mouse macrophage RAW 264.7 cells were investigated. The results showed that the ethyl acetate fraction (KSD-E8-3) of lees extracts from sweet potato soju significantly decreased nitric oxide (NO) production in LPS-activated RAW 264.7 cells, whereas they did not affect cell viabilities. The fraction KSD-E8-3 reduced the expression of pro-inflammatory genes such as COX-2, iNOS and TNF-alpha and also decreased protein expression of iNOS in a dose dependent manner, which were detected with RT-PCR and Western blot analysis, respectively. In addition, we detected the expression of mitogen-activated protein kinases (MAPKs) such as p38, JNK, and ERK1/2 and their phosphorylated forms. The results indicated that the treatment of the fraction KSD-E8-3 did inhibit phosphorylation of p38, JNK, and ERK1/2 MAPKs, indicating that the fraction KSD-E8-3 regulates LPS-induced inflammatory response via suppressing MAPK signaling pathway. Overall, these results may contribute to understand the molecular mechanism of anti-inflammatory effects by the ethyl acetate fraction of lees extracts from sweet potato soju.

• Journal of the East Asian Society of Dietary Life
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• v.17 no.3
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• pp.393-401
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• 2007

This study was performed to investigate the antioxidant activity of Korean ginseng using an ORAC(Oxygen Radical Absorbance Capacity) assay. Four fractions each (80% ethanol, ethyl acetate, water saturated 1-butanol, and water) were obtained from different ginseng samples (White Ginseng: ; 6 yrs-., 5 yrs-., ; Cork Ginseng: ; 5 yrs-., 4 yrs-.). The saponin content of each fraction was quantified by LC/MS, and the antioxidant capacity of the ginseng was measured by the ORAC assay. The ORAC method, which was recently validated using automatic liquid handling systems, has been adapted for manual handling with the use of a conventional fluorescence microplate reader. Furthermore, the ORAC assay provides a direct measure of hydrophilic chain-breaking antioxidant capacity against peroxy radical, which is the exiting and emission of 2,2'-Azobis (2-methylpropionamidine)-dihychloride (AAPH). As a result of our experiments, ginsenosides Rg1 and Rb1 were the two major saponins found in the ginseng samples, and Rc, Rb2, Re, Rd, Rg3, and Rh1 were detected in a small quantities. For the antioxidant capacities of the fractions (80% ethanol, ethyl acetate, butanol, and water), we found that the organic solvent fraction had similar antioxidant capacities, and were higher than the capacity of the water fraction. When determining the similarities in each fraction, only the ethyl acetate fraction showed similarity compared to other fractions (p>0.05). The antioxidant capacity of ginseng may come from phenolic compounds and some nonpolar saponins. However, based on the results of this study, we hypothesize that some acidic polysaccharides and other biological components may contribute to its antioxidant capacity. Additional research is required to determine other possible biological response modifiers that contribute to the antioxidant capacity of ginseng.

• Journal of Physiology & Pathology in Korean Medicine
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• v.22 no.4
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• pp.903-906
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• 2008

Woodfordia fruticosa Kurz (Lythraceae) is used in the treatment of various ailments in traditional medicines. DPPH activity guided fractionation and purification process was used to identify the free radical-scavenging components from the flowers of this plant. The methanolic extract of the plant was first fractionated into four extracts; namely, n-hexane, chloroform, ethyl acetate and water fractions. Among them, the ethyl acetate fraction was found to be the most effective and was further subjected to activity guided-fractionation and isolation procedures. After successive column chromatography on silica gel and Sephadex LH-20, gallic acid, which is responsible for the radical scavenging activity, was isolated and its structure was elucidated by spectral methods ($^1H$ NMR, $^{13}C$ NMR) and by comparison with literature.