The major active components of Astragalus membranaceus (AM) are isoflavones, which exist in the form of various glycosides. Nuruk is a traditional fermentation starter in Korea, and is suitable for the biotransformation of isoflavone glycosides because it contains various microorganisms and enzymes. This study was performed to determine changes in the isoflavones and antioxidant properties of AM fermented (AF) with nuruk over 24 hours. AF was sampled after 0, 2, 4, 6, 12, 18, and 24 h of fermentation, and calycosin 7-glucoside, ononin, calycosin, and formononetin content, and the antioxidant properties of AF were analyzed. The total phenolic content increased with fermentation time, and the ABTS radical scavenging activity increased until 6 h of fermentation and then decreased. During fermentation, the isoflavone glycosides decreased significantly as fermentation time increased. The contents of calycosin and formononetin, which are aglycons of calycosin-7-glucoside and ononin, increased from 100.54 μg/g to 276.84 μg/g and from 56.29 μg/g to 123.04 μg/g, respectively, at 18 h of fermentation. Significant correlations were observed between fermentation time, isoflavone content, and antioxidant properties. The results of this study showed that fermentation with nuruk is suitable for the biotransformation of isoflavones in AM.

Aster koraiensis Nakai (A. koraiensis) which has been used as a food and medicinal plant in the past, is valuable as functional food material. Therefore, the aim of this study was to determine the antioxidant properties and major phenolics of A. koraiensis extracts with different ethanol concentrations (0, 50, 70, and 100% aqueous ethanol solution). When ethanol concentration in the extraction solvent was increased, extraction yield decreased; 34.2, 23.2, 21.0, and 5.5% in 0, 50, 70, and 100% ethanolic extracts, respectively. Total phenolics content and antioxidant activities of extracts were increased in an ethanol concentration-dependant manner. The major phenolics in the extracts were chlorogenic acid (21.264~58.666 mg/g), isochlorogenic acid A (10.432~145.353 mg/g), and isochlorogenic acid C(0.239~13.148 mg/g), and these phenolic contents were higher in 70 and 100% ethanolic extracts than other extracts. Significant correlations were observed between ethanol concentration of extraction solvent, antioxidant properties, and major phenolics. These results indicated that the optimal ethanol concentration for extraction was 70%.

Fresh Omija (Schisandra chinensis) has good marketability, but its quality is difficult to maintain during storage and distribution. Freezing and freeze-thawing treatments can be utilized for the quality maintenance and processing of cold press juice. In this study, the color, antioxidant properties, and the major components of soaked liquor from Omija with freeze-thawing treatment were analyzed during the extraction periods. Each of the frozen and freeze-thawed Omija samples was soaked in 35% ethanol, extracted for 15 days, and used for analysis. The frozen and freeze-thawed samples showed a tendency toward better color and higher antioxidant activity and major component levels than the controls, and freeze-thawing was the best. The results of this study showed that freeze-thawing treatment improved the color, antioxidant properties, and level of the major components of Omija soaked liquor, and freeze storage is suitable for making soaked liquor.

Background: The ginsenosides Rb1 (G-Rb1) and Rg1 (G-Rg1) are used as marker compounds, and are the principal bioactive compounds assessed in the quality control of white ginseng. This study was conducted to analyze white ginseng samples of different and to obtain useful data for the quality control of white ginseng. Methods and Results: The variation in the content of G-Rb1 and G-Rg1 was evaluated among 35 samples of 4-, 5-, and 6-year-old white ginseng. The content of both G-Rb1 and G-Rg1 did not significantly differ among ages, and the relative ratio of the maximum to the minimum content of these within ginseng of the same ages was more than two. However, the ratio of G-Rb1 to G-Rg1 content in the 5- and 6-year-old ginseng was significantly higher than that in the 4-year-old one. According to the ‘Ginseng industrial act’, the standard (w/w, %) minimum G-Rg1 and G-Rb1 content is 0.10% and 0.20% or more, respectively. Among the 35 samples examined, the content of G-Rg1 was found to be 0.124 - 0.399% with none being less than the standard level, while that of G-Rb1, was 0.147 - 0.595%, with 4 samples (11.4%) failing to meet the standard levels. The content of G-Rg1 and G-Rb1 did not show a constant relationship with the size of ginseng. Conclusions: In our study, the content of both G-Rg1 and G-Rb1 varied widely, and there was no significant difference among cultivation ages. The results of the present study might provide useful information for the quality control of raw ginseng and processed white ginseng using marker compound.

Background : It is known that Platycodon grandiflorum has anti-inflammatory activity and inhibits the production of nitric oxide (NO) in inflammatory macrophages. But the change of bioactivity of platycodon grandiflorum according to steaming is not well known. In this study, We investigated the effects of steaming on anti-inflammatory activity of 70% ethanol extracts of platycodon grandiflorum. Methods and Result : The cytotoxicity of RAW264.7 cells treated with platycodon grandiflorum extracts was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and the concentration of NO in culture supernatants were determined using nitric oxide (NO) assay. And western blotting was performed to quantify the expression of iNOS, a protein related to NO production. As a results, it was confirmed that no cytotoxicity was observed at 25, 50 and 100 ㎍/㎖ platycodon grandiflorum extracts in RAW264.7 cells. The production of NO and the expression of iNOS were induced by LPS and suppressed by all platycodon grandiflorum extracts in proportion to the number of steaming in RAW264.7 cells. Conclusion : These results suggest that a steaming process can increase anti-inflammatory activity of platycodon grandiflorum extracts.

Background : Spiraea prunifolia var. simpliciflora (Rosaceae) called “Brial wreath” is a deciduous latifoliate shrub that is widely distributed throughout in Northeast Asia. Phytochemical and biological investigation of S. prunifolia have led to the discovery of biologically active compounds. Pharmacological studies revealed that the extract of the root of S. prunifolia possess antioxidant, antipyretic and anti-inflammatory activities. Some chemical constituents such as sterols, phenolics, terpene and fatty acid, as well as ethanolic extracts from the roots of S. prunifolia, have previously been reported to modulate the deleterious effects of diabetes, to prevent high-fat diet-induced obesity, and to prevent cisplatin-induced nephrotoxicity. Our continuing research was carried out to search for other phytochemical constituents from the leavess of S. prunifolia. The chemical structures of compounds were determined by NMR and FAB/MS spectroscopic data interpretation. Methods and Results : Multiple-preparative liquid chromatography (MPLC) purifications were carried out on YMC LC-forte/R instrument (YMC Kyoto, Japan) equipped with YMC-Pack ODS-A columns (ODS gel : 5 ㎛, 10 ㎜ × 250 ㎜). High-performance liquid chromatography (HPLC) was performed on Agilent Technologies instrument (Aglient Tec., Santa clara, CA, USA) equipped with YMC–Pack Pro C18 columns (ODS gel : 5 ㎛, 4.6 ㎜ × 250 ㎜). Next, quantitative analysis was carried out on UPLC-QqQ/MS 3200 Q-TRAP instrument (AB SCIEX Toronto, Canada) using a ACQUITY UPLC (waters corp.) with an ACQUITY BEH C18 column (2.1 ㎜ × 100 ㎜, 1.7 ㎛). The metabolite samples was applied to preparative reversed-phase HPLC and UPLC using gradient method, solvent A [water + 0.1% formic acid (v/v)] and solvent B [acetonitrile + 0.1% formic acid (v/v)]. Conclusion : In this study, we isolated the major metabolites from the stem of Spiraea prunifolia var. simpliciflora by using MPLC and HPLC. UPLC-QqQ/MS was also used to quantify target compounds. Finally, we established methodology and performed the quantitative analysis on target compounds from the stem of Spiraea prunifolia var. simpliciflora.

Background : Lythrum salicaria L. (LS), a herb that is found all around the world, has long been used as medicinal plant to treat inflammation, external wound bleeding, and diarrhea, while its sprouts (young leaves) can be utilized as a food material. The antioxidant and hepato-protective activities of LS have been reported in several articles. This study was conducted to compare the efficacy and cell proliferation of LS leaves according to their growth period, and to obtain information on the optimal harvesting time of LS as a food resource. Methods and results : LS leaves were collected at ten-day intervals between April 27 and June 26, 2016 in Eumseong-gun, South Korea. The LS leaves were extracted with 50% ethanol at room temperature, and seven LS extracts (LSE) were obtained. A peroxynitrite (ONOO-) scavenging assay and a 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay were performed to compare the antioxidant effects of LSE, while a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was performed on the BV-2 cell lines to determine cell viability. The total phenol contents of LSE were quantified by using the calibration curve of tannic acid. From these assays, LSE harvested on April 27 showed the lowest value, while LSE harvested on June 6 showed the highest DPPH scavenging activity at 10 ㎍/㎖. There was no difference among the extracts in terms of their peroxynitrite scavenging activity. The extract prepared on April 27 showed the highest value in terms of BV2 cell viability, while that obtained on June 6 showed the lowest value. The value in terms of the total phenol content of the LSE harvested on June 6 was the highest, whereas that of the LSE harvested on April 27 was the lowest. Conclusion : When comparing the activity of LSE according to its harvesting time, the extract dated June 6 showed the highest effect in terms of its antioxidant activity and its total phenol content, whereas the extract dated April 27 showed the highest cell viability. As such, this study suggests that LS leaves harvested in the early season could be utilized as a food material even though they display low efficacy.

Background : The development of an antioxidant to prevent disease by ROS-induced oxidative stress is necessary. This study investigated the changes of antioxidant capacities of two medicinal crops extracts by lactic acid fermentation. Methods and Results : The changes of free-radical scavenging activity of medicinal crops extracts by lactic acid fermentation were evaluated by using DPPH free-radical scavenging assay and ABTS free-radical scavenging assay. The DPPH free-radical scavenging activity of extracts or lactic acid fermented extracts were estimated as followed. samples were thoroughly mixed with 1 ㎖ ethanol solution of 0.1 mM DPPH. After stand for 30 min in the dark, the absorbance was measured at 570 ㎚ by using a UV Spectrophotometer. ABTS scavenging activity of extracts or lactic acid fermented extracts were estimated as followed. The working solution was prepared by mixing 1 ㎖ of ABTS solution with 88 ㎖ of 50% ethanol. A total of 25 ㎕ of samples were mixed with 225 ㎕ of ABTS working solution and allowed to stand for 10 min. The absorbance was read at 732 ㎚ in a UV spectrophotometer. The data were showed that lactic acid fermented extracts were higher antioxidant ability than the extracts. Conclusion : This study was showed that the antioxidant capacities of two medicinal crops extracts were improved by lactic acid fermentation.

Background : While the anti-inflammatory effects of 20 (S)-ginsenoside Rg3 (Rg3) have been studied, it remains unclear how Rg3 regulates lipid metabolism in inflammatory macrophages. Thus, in this study, we characterized some eicosanoids related to the anti-inflammatory effects of 20 (S)-ginsenoside Rg3 in murine macrophages. Methods and Results : UPLC-MS/MS was used to profile various eicosanoids from RAW264.7 cells treated with lipopolysaccharide (LPS) and Rg3. The profiling data were statistically analyzed by principal component analysis, hierarchical clustering analysis and analysis of variance. The anti-inflammatory effect of Rg3 was validated by assessing the levels of nitric oxide, tumor necrosis factor-α, and interleukin-6 in the activated macrophages treated with Rg3. A total of 69 eicosanoids were analyzed in RAW264.7 cells. Principal component and hierarchical cluster analyses differentiated control cells from cells treated with LPS, Rg3, or LPS + Rg3 for 12 or 24 h. Furthermore, some differentially regulated compounds were found between macrophages treated with LPS for 24 h and those treated with LPS + Rg3 for 24 h. Conclusion : Rg3 alters eicosanoid metabolism in activated macrophages treated with LPS. Furthermore, we identified several eicosanoids correlated with the anti-inflammatory activity of Rg3.

Background : Methicillin-Resistant Staphylococcus aureus (MRSA) is a multidrug-resistant (MDR) strain. Especially, MRSA is developing resistance to available antibacterial agents and causing complications in the treatment of infections related to skin, soft tissue, respiratory, bone, joint, and endovascular disorders. Therefore, antibacterial agent combination therapy appears to be a useful option, particularly in developing countries where antibiotic availability is limited. (+)-Usnic acid (UA) is uniquely found in lichens, and is especially abundant in genera such as Usnea and Cladonia. UA has antimicrobial activity against human and plant pathogens. Therefore, UA may be a good antibacterial drug candidate for clinical development. Methods and Results : In search of a natural products capable of inhibiting this multidrug-resistant bacteria, we have investigated the antimicrobial activity of UA against MRSA. In this study, the effects of a combination of UA and permeable agents against MRSA were investigated. For the measurement of cell wall permeability, UA with concentration of Ethylenediaminetetraacetic acid (EDTA) was used. In the other hand, Sodium azide (NaN3) was used as inhibitors of ATPase. These results suggest that the antibacterial effect of UA was potentiated by membrane-binding agents and ABC transporter-inhibiting agents, implying that antibacterial activity is associated with damage of the cell wall and inhibition of ATPase function by UA. Conclusion : UA and in combination with EDTA and NaN3 could lead to the development of new combination antibiotics against MRSA infection. The results of this study appear to be promising, and they are expected to enhance the use of natural products as drugs.

Background : Licorice has been used as a source of medicine and a food material in East-Asia. Recently, demand for licorice increased in market due to a growing interest in health. Thus we conducted breeding research to solve the problems associated with domestically cultivated licorice such as low productivity and low glycyrrhizin content. Methods and Results : We crossed European licorice (G. glabra L.; female parent) and Chinese licorice (G. uralensis Fisch; male parent) in the greenhouse in May 2007. In September 2007, crossed and germinated seeds were retrieved and sown in the greenhouse. In June 2008, stolons were separated from the F1 licorice seedlings and cultivated, resulting in 32 clonal lines of interspecific hybrids. Among them we selected good lines and then conducted the replicated yield trials (RYT) in 2012-2013 and local adaptability test (LAT) in 2014-2015. The results, GLYES9 showed that was elect of stem, oblong of leaf shape, red-brown of root color. Glycyrrhizin conten of GLYES9 (3.0%) was higher than G. uralensis (1.9%) at four regions from 2014 to 2015. GLYES9 was less than 10% in the desease of brown spot (G. uralensis was more than 30%). The root yield of GLYES9 was 4.31 ton per hectare, which was increased 193% compared with a check variety of G. uralensis. Therfore, we named GLYES9 as new cultivar ‘Dagam’. Conclusion : Depending on the above results, we have developed a new licorice cultivar ‘Dagam’ by the medicinal crop breeding team of National Institute of Horticulture and Herbal Science, RDA, in 2015. It showed brown spot disease resistant, high-glycyrrhizn content and high-yielding than colleted Glycyrrhiza spp.

Background: This study was performed to evaluate the protective effect of Saururus chinensis ethanol extract (SCE) against styrene toxicity in mouse spermatocyte cells [GC-2spd (ts) cell line]. Methods and Results: Cytotoxicity in mouse spermatocyte cells was measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Generation of reactive oxygen species (ROS) was determined using 2’,7’-dichlorodihydrofluorescein diacetate (DCF-DA) assay. Semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and western blotting were performed to quantify the mRNA and protein expression levels, resepectiviely, of stress or apoptosis-related genes including p21, p53, heat shock protein 70 (Hsp70), heat shock protein 90 (Hsp90), Bax, Bcl-2, and caspase-3. The results of the MTT assay showed that 50 ㎍/㎖ SCE did not affect cell viability. ROS generation in mouse spermatocyte cells increased by treatment with 100 μM styrene, and decreased by co-treatment with SCE. SCE repressed the mRNA expression of stress-related genes, which increased by styrene treatment. In addition, SCE inhibited the apoptosis of mouse spermatocyte cells by ameliorating mRNA and protein levels of apoptotic genes that were altered by styrene treatment. Conclusions: These results suggest that SCE may alleviate styrene toxicity in mouse spermatocyte cells by reducing ROS stress and regulating genes related to styrene toxicity.

Background : Acanthopanax sessiliflorus (Rupr. et Maxim) Seem, belonging to the Araliaceae family, is widely distributed in Korea, China, and Japan. The plants belonging to Acanthopanax species are traditionally used in Korea as anti-rheumatoid arthritis, anti-inflammatory and anti-diabetic drugs and are recognized to have ginseng-like activities. A simple and sensitive high-performance liquid chromatographic (HPLC) method was developed and validated for independent analysis of major compounds and chlorogenic acid in A. sessiliflorus fruits. Chlorogenic acid was reported that prevent cancer and cardiovascular disease in vivo. Also, it has antioxidant effect in vitro test. In the previous experiment, chlorogenic acid were found in A. sessiliflorus fruits. This study was performed to identification of the major compounds and investigate the method validation for the determination of chlorogenic acid in A. sessiliflorus fruits. Methods and Results : Three major compounds were recorded on a Varian Unity Inova AS-400 FT-NMR spectrometer and analyzed by the new HPLC analysis method. HPLC analysis was carried out using an Waters e2695 and PDA detector. The new analyasis method was validated by the measurement of intra-day, inter-day precision, accuracy, limit of detection (LOD, S/N=3), and limit of quantification (LOQ, S/N=10) of chlorogenic acid. The results showed that the correlation coefficient (R2) for the calibration curves of chlorogenic acid was 0.997 in terms of linearity. The limit of detection (LOD) and limit of quantification (LOQ) were 0.565 ㎍/ml and 2.88 ㎍/ml, respectively. There was no interfering peak observed each other and HPLC system was suitable for analysis showing goodness of peak and high precision. Conclusion : This method is suitable to detect and quantify major compounds in A. sessiliflorus fruits. Furthermore, the result will be applied to establish chlorogenic acid as an standard compound for A. sessiliflorus fruits.

Background : Cynanchum wilfordii and Cynanchum auriculatum belong to the Asclepiadaceae family and appear morphologically similar. In order to discriminate them, it is needed to find the presence of sap and the leaf shapes: C. auriculatum has a blade ovate leaf comparing to C. wilfordii. However, in the herbal medicine market, they have been handled as cut and dried roots. Due to their similar morphology, it is limited to distinguish the roots of C. wilfordii and C. auriculatum. Recently in Korea, it has been a critical issue to misuse these two roots in the herbal market and food industry. Thus, it is required to establish a robust tool for the discrimination and quality control of them. Methods and Results : To separation and characterization of flavor compounds, C. wilfordii and C. auriculatum samples were analyzed by head space solid phase micro extraction (HSS) fiber coupled with gas chromatography-mass spectrometry using Rtx-5MS (30 m x 0.25 mm x 0.25 μm) column. As a result, We have identified compounds of a few hundred in aliphatic aldehydes and aliphatic alcohols, alkenes and acids, aromatic compounds, aromatic compounds containing nitrogen & sulfur, etc,. In particular, The aliphatic and aromatic compounds had been clearly separated on the second dimensional direction by using two-dimensional GC. Conclusion : The volatile flavor compounds of C. wilfordii and C. auriculatum could easily analyzed without pre-treatment with improved resolution and sensitivity using HSS-GCxGC-TOFMS. We have identified compounds of a few hundred in C. wilfordii and C. auriculatum sample. And It was more accurately qualitative confirmed with separation of GCxGC and TSD. We have confirmed the PCA and PLS-DA Plot that was classified depending on C. wilfordii and C. auriculatum through multivariate statistical analysis of the identified flavor compounds.

Background : Allergy is a common disease caused by type I hypersensitivity reaction of the immune system. Unfortunately, there is no proper treatment for allergy. Therefore, the discovery of therapeutic drugs for allergy is essential. In this study, the crude extracts of 56 plant parts were screened for anti-allergy effects in RBL-2H3 cells. Methods and Results : IgE-sensitized RBL-2H3 cells were individually treated with 56 extracts of medicinal herbs at the final concentration of 20 ㎍/㎖ and stimulated with the antigen DNP-BSA. β-Hexosaminidase release, interleukin-4 (IL-4) secretion, and cell viability in the sample treated cells were measured. Among the tested samples, extracts from the root of Polygonatum stenophyllum Maxim., aerial part of Acer triflorum Kom., and leaf of Pyrus pyrifolia var. culta (Makino) Nakai showed inhibitory effects on β-hexosaminidase release. The aerial part of Peucedanum japonicum Thunberg and seed of Panax ginseng C.A.Mey. showed suppressive activities on IL-4 secretion. All of the extracts were not cytotoxic at the tested concentration. Conclusion : From the result, six extracts including Polygonatum stenophyllum Maxim. (root) and Acer triflorum Kom. (aerial part) inhibited both β-hexosaminidase release and IL-4 secretion in IgE-sensitized RBL-2H3 cells. The use of these extracts for developing anti-allergy materials is suggested.

Background : Five medicinal herbs have been selected from the preliminary screening for in vitro anti-allergy activity (in RBL-2H3 cells). The present study is conducted to investigate the inhibitory effects of the medicinal herbs on allergic inflammation in other kind of cells. Methods and Results : Cells treated with five extracts prepared from Betula costata Trautv. (aerial part), Camellia sinensis L. (aerial part), Polygonatum stenophyllum Maxim. (root), Pyrus pyrifolia var. culta (Makino) Nakai (leaf), and Rehmannia glutinosa (Gaertn.) Libosch. ex Steud. (root) were measured for mRNA levels of TNF-α on HaCaT keratinocytes stimulated by TNF-α /INF-γ and for mRNA levels of IL-2 in Jurkat T cells mediated by PMA/A23187. Pre-treatment with the five extracts reduced the mRNA levels of TNF-α in HaCaT cells and mRNA levels of IL-2 in Jurkat T cells. In particular, the leaf extract of Pyrus pyrifolia var. culta (Makino) Nakai significantly and dose-dependently decreased the mRNA levels of TNF-α and IL-2. To determine the toxicity of the extracts from the selected medicinal herbs to HaCaT cells and Jurkat T cells, the viabilities of the cells treated with several concentrations of the five extracts were measured by MTT assay. Extracts of Polygonatum stenophyllum Maxim. (root), Pyrus pyrifolia var. culta (Makino) Nakai (leaf), and Rehmannia glutinosa (Gaertn.) Libosch. ex Steud. (root) (up to 250 ㎍/㎖) did not show cytotoxic effects on HaCaT cells and Jurkat T cells. On the other hand, 250 ㎍/㎖ of extracts of Betula costata Trautv. (aerial part) and Camellia sinensis L. (aerial part) reduced cell viability in both cells. Conclusion : These results demonstrate that the leaf extract of Pyrus pyrifolia var. culta (Makino) Nakai has an anti-inflammatory effect by inhibiting pro-inflammatory cytokine expression. Therefore, the leaf of Pyrus pyrifolia var. culta (Makino) Nakai can be a useful resource for the development of anti-allergy/anti-inflammatory materials.

Background : Ginseng, one of most famous traditional oriental medicines, has been known for a number of pharmacological properties including anti-tumor, anti-diabetic, anti-fatigue, anti-stress, anti-oxidative, and anti-aging effects. 20(S)-Protopanaxadiol (PPD), a intestinal metabolite of ginsenosides, is one of the active ingredients in ginseng. In this study, we investigated the synergistic anticancer effect of 20(S)-PPD and temozolomide (TMZ) and the mechanism of 20(S)-PPD on glioblastoma cells. Methods and Results : We examined cell viability and the morphological changes of C6 cells after treatment of 20(S)-PPD and TMZ. 20(S)-PPD showed a potent antiproliferative activity against C6 cells by triggering apoptosis. 20(S)-PPD-induced apoptosis was characterized by a dose-dependent mitochondrial damage. 20(S)-PPD and TMZ had a synergistic effect in increasing mitochondrial damage via caspase 3 activation. Conclusion : These results revealed an unexpected mechanism of 20(S)-PPD and TMZ, triggering a mitochondrial-mediated apoptosis in C6 cells. Our findings encourage further studies of 20(S)-PPD as a promising chemopreventive agent against glioblastoma.