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Simultaneous HPLC Determination of Marker Compounds for the Standardization of Hedyotis diffusa (백운풀의 지표성분 설정 및 품질표준화를 위한 정량 분석법)

  • Bang, Han-Yeol;Yang, Eun-Ju;Kim, Jeong-Ah;Song, Kyung-Sik
    • Journal of Life Science
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    • v.23 no.8
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    • pp.1025-1031
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    • 2013
  • From a 95% ethanolic extract of H. diffusa, four marker compounds (HD1~HD4) were isolated, which were relatively unique and exist in comparably high contents. The structures of marker compounds were identified as digitolutein (1), 2-hydroxy-3-methylanthraquinone (2), (E/Z)-6-O-p-coumaroyl scandoside methyl ester (4:1 mixture) (3), and (E/Z)-6-O-p-methoxycinnamoyl scandoside methyl ester (4:1 mixture) (4), respectively, on the basis of $^{13}C$ and $^1H$-NMR analyses. The calibration curves of marker compounds showed high linearity, as their correlation coefficient ($R^2$) were in the range of 0.9991~0.9999. In addition, the limit of detection (LOD) and the limit of quantification (LOQ) were $0.03{\sim}0.07{\mu}g/ml$ and $0.099{\sim}0.231{\mu}g/ml$, respectively. The intra-day/inter-day precision and accuracy were 0.23~2.00%/0.25~1.16% and 94.60~108.44%/94.73-110.23%, respectively. The optimal HPLC conditions for the simultaneous quantification of HD1~HD4 were as follows: stationary phase; Merck Chromolith RP-18e ($100{\times}4.6mm$, $5{\mu}m$), column temp.; room temperature, UV detection at 280 nm, flow rate; 2.0 ml/min, injection volume; $10{\mu}l$, mobile phase; start with the mixture of 80% solvent A ($H_2O$ containing 0.5% acetic acid) and 20% solvent B (methanol containing 0.5% acetic acid) and gradually decrease solvent A to 40% in 9 min., then retain this condition to 18 min. Under the HPLC condition, the four marker compounds 1~4 were successfully separated without any interference of other constituents. The results obtained in this study are expected to be helpful for the development of nutraceutics and natural medicines and for the quality control of this plant.

Studies on the Lipid Components of Panax ginseng (인삼(人蔘)의 지방질(脂肪質) 성분(成分)에 관한 연구)

  • Shin, Hyo-Sun;Lee, Min-Woong
    • Korean Journal of Food Science and Technology
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    • v.12 no.3
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    • pp.185-192
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    • 1980
  • To study lipid components of Panax ginseng produced in Korea, the lipids of fresh ginsengs were extracted with the mixture of chloroform-methanol (2:1, v/v) and those of dried ginsengs were extracted with diethyl ether respectively. The lipid components extracted were separated and quantitated by column, thin layer and gas-liquid chromatographies. The results were summarized as follows : 1. Fresh ginseng contained 0.62% total lipid of which 45.28% were neutral lipids, 18.12% glycolipids, and 36.60% phospholipids. But dried ginseng contained 0.89% total lipids of which 86.48% were neutral lipids, 9.20% glycolipids, and 4.32% phospholipids. 2. Triglycerides (37.6 to 42.5% of the total neutral lipids) and sterol esters (16.5 to 19.6%) in all the fresh and dried ginseng were the major components among the neutral lipids. Monoglycerides, diglycerides, free fatty acids and free sterols were minor components. 3. Digalactosyl diglycerides (23.5% of the total glycolipids) in the fresh ginseng and steryl liglycosides (28.9%) in the dried ginseng were predominant components among the glycopids, respectively, Esterified steryl glycosides and monogalactosyl diglycerides were also identified, and four unknown spots in the fresh ginseng and two unknown spots in the dried ginseng were present. 4. Phosphatidyl cholines (31.3 to 31.9% of the total phospholipids) and phosphatidyl glycerols (34.8 to 36.7%) in all the fresh and dried ginseng were the major components among the phospholipids. Phosphatidyl inositols and phosphatidyl ethanolamines were also identified. 5. The major fatty acids in the fresh and dried ginseng were linoleic $(62.29{\sim}64.32%)$, palmitic $(13.16{\sim}15.63%)$, oleic $(5.73{sim}7.23%)$ and linolenic $(5.73{sim}7.23%)$. The fatty acid compositions in neutral lipid fraction was similar to the pattern in those of the total lipids. But glycolipid and phospholipid fractions contained a lower percent of linoleic acid and a higher percent of palmitic acid than the neutral lipid fraction.

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Identification of Antioxidant Compound Derived from Methanolic Extract of Houttuynia Cordata (어성초 메탄올 추출물로부터 항산화 효능을 가진 활성물질의 확인)

  • Kim, Hyeji;Hwang, Heesung;Park, Sumin;Kang, Sungwook;Kim, Hyejeong;Hong, Sugyeong;Kim, Moon-Moo;Oh, Yunghee
    • Journal of Life Science
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    • v.27 no.7
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    • pp.796-804
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    • 2017
  • This study was carried out to evaluate the antioxidant effect of methanolic extract of Houttuynia cordata (HCME) and to identify a compound having antioxidant effect. The ethyl acetate fraction of HCME showed the highest antioxidant effect in organic solvent fractions. The fraction was then separated into 12 fractions by open column chromatography. Among these fractions, the fraction 10 (Fr. 10) with the highest antioxidant activity was isolated, and its antioxidant effect was evaluated by DPPH radical scavenging activity, reducing power, TBARS, cell viability, DNA oxidation and DCF fluorescence. The Fr. 10 at a $64{\mu}g/ml$ showed 60% of inhibitory effect similar to that of vitamin C at $10{\mu}g/ml$, compared with blank group. The Fr. 10 at $64{\mu}g/ml$ showed 264% of reducing power, compared with blank group. TBARS assay showed that the Fr. 10 at $64{\mu}g/ml$ had 35.5% of inhibitory effect similar to that of vitamin E at $1,000{\mu}g/ml$, compared with blank group. The Fr. 10 above $32{\mu}g/ml$ displayed cytotoxicity. However, it was observed that the Fr. 10, above $1{\mu}g/ml$ reduced DNA damage. DCF fluorescence assay showed that the Fr. 10 inhibited oxidative stress by $H_2O_2$ in a dose dependent manner. The compound of Fr. 10 was identified to be rutin whose molecular weight is 610 by the IR and LC-MS analyses. Therefore, these results suggest that the rutin of Fr. 10 could use as a natural antioxidant for development of cosmetics and functional foods.

Isolation and Characterization of a Marine Bacterium, Pseudomonas sp. YJ-1 with Anti-Methicillin Resistant Staphylococcus aureus Activity (항 Methicillin Resistant Staphylococcus aureus 활성을 가지는 해양미생물 Pseudomonas sp. YJ-1의 분리와 특성)

  • Woo, Ye-Ju;Jeong, Seong-Yun
    • Korean Journal of Environmental Biology
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    • v.35 no.4
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    • pp.694-705
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    • 2017
  • The aim of this study was to isolate and identify marine bacterium with anti-methicillin-resistant Staphylococcus aureus (MRSA) activity, and to purify the anti-MRSA compound, as well as to determine its activity and synergistic effects. Among the marine bacteria isolated in this study, the YJ-1 isolate had the strongest anti-MRSA activity. The YJ-1 isolate was identified on the basis of its biochemical characteristics and an analysis of 16S rRNA gene sequences. The YJ-1 isolate showed over 99.2% homology with Pseudomonas stutzeri, and was designated as a Pseudomonas sp. YJ-1. The optimal culture conditions were $25^{\circ}C$ and initial pH 7.0. For the purification of the anti-MRSA compounds, the YJ-1 was cultured in Pa PES-II medium, and the culture filtrates were extracted by ethyl acetate, hexane, and 80% MeOH. The 80% MeOH fraction was separated by a $C_{18}$ ODS column, silica gel chromatography and a reverse phase HPLC, to yield three anti-MRSA agents, the MR1, MR2, and MR3 compounds. When the MR1 compound of $250{\mu}g\;mL^{-1}$ concentration was applied to the MRSA cells, over 95% of bacterial cells was killed within 48 hr. Compared with vancomycin and ampicillin, the MR1 compound showed significant anti-MRSA activity. In addition, the anti-MRSA activity was increased by dose and time dependent manners. Furthermore, the combination of an MR1 compound with vancomycin produced a more rapid decrease in the MRSA cells than did the MR1 compound alone. Taken together, our results suggest that the Pseudomonas sp. YJ-1 and its anti-MRSA compounds could be employed as a natural antibacterial agent in MRSA infections.

Radiosynthesis of $[^{11}C]6-OH-BTA-1$ in Different Media and Confirmation of Reaction By-products. ($[^{11}C]6-OH-BTA-1$ 조제 시 생성되는 부산물 규명과 반응용매에 따른 표지 효율 비교)

  • Lee, Hak-Jeong;Jeong, Jae-Min;Lee, Yun-Sang;Kim, Hyung-Woo;Lee, Eun-Kyoung;Lee, Dong-Soo;Chung, June-Key;Lee, Myung-Chul
    • Nuclear Medicine and Molecular Imaging
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    • v.41 no.3
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    • pp.241-246
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    • 2007
  • Purpose: $[^{11}C]6-OH-BTA-1$ ([N-methyl-$^{11}C$]2-(4'-methylaminophenyl)-6-hydroxybenzothiazole, 1), a -amyloid imaging agent for the diagnosis of Alzheimer's disease in PET, can be labeled with higher yield by a simple loop method. During the synthesis of $[^{11}C]1$, we found the formation of by-products in various solvents, e.g., methylethylketone (MEK), cyclohexanone (CHO), diethylketone (DEK), and dimethylformamide (DMF). Materials and Methods: In Automated radiosynthesis module, 1 mg of 4-aminophenyl-6-hydroxybenzothiazole (4) in 100 l of each solvent was reacted with $[^{11}C]methyl$ triflate in HPLC loop at room temperature (RT). The reaction mixture was separated by semi-preparative HPLC. Aliquots eluted at 14.4, 16.3 and 17.6 min were collected and analyzed by analytical HPLC and LC/MS spectrometer. Results: The labeling efficiencies of $[^{11}C]1$ were $86.0{\pm}5.5%$, $59.7{\pm}2.4%$, $29.9{\pm}1.8%$, and $7.6{\pm}0.5%$ in MEK, CHO, DEK and DMF, respectively. The LC/MS spectra of three products eluted at 14.4, 16.3 and 17.6 mins showed m/z peaks at 257.3 (M+1), 257.3 (M+1) and 271.3 (M+1), respectively, indicating their structures as 1, 2-(4'-aminophenyl)-6-methoxybenzothiazole (2) and by-product (3), respectively. Ratios of labeling efficiencies for the three products $([^{11}C]1:[^{11}C]2:[^{11}C]3)$ were $86.0{\pm}5.5%:5.0{\pm}3.4%:1.5{\pm}1.3%$ in MEK, $59.7{\pm}2.4%:4.7{\pm}3.2%:1.3{\pm}0.5%$ in CHO, $9.9{\pm}1.8%:2.0{\pm}0.7%:0.3{\pm}0.1%$ in DEK and $7.6{\pm}0.5%:0.0%:0.0%$ in DMF, respectively. Conclusion: The labeling efficiency of $[^{11}C]1$ was the highest when MEK was used as a reaction solvent. As results of mass spectrometry, 1 and 2 were conformed. 3 was presumed.