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Definition of the peptide mimotope of cellular receptor for hepatitis C virus E2 protein using random peptide library (Random peptide library를 이용한 C형 간염바이러스 E2 단백질 세포막 수용체의 peptide mimotope 규명)

  • Lee, In-Hee;Paik, Jae-Eun;Seol, Sang-Yong;Seog, Dae-Hyun;Park, Sae-Gwang;Choi, In-Hak
    • IMMUNE NETWORK
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    • v.1 no.1
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    • pp.77-86
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    • 2001
  • Background: Hepatitis C virus(HCV), a family of Flaviviridae, has a host cell-derived envelope containing a positive-stranded RNA genome, and has been known as the maj or etiological agent for chronic hepatitis, hepatic cirrhosis, and hepatocellular carcinoma. There remains a need to dissect a molecular mechanism of pathogenesis for the development of therapeutic and effective preventive measure for HCV. Identification of cellular receptor is of central importance not only to understand the viral pathogenesis, but also to exploit strategies for prevention of HCV. This study was aimed at identifying peptide mimotopes inhibiting the binding of E2 protein of HCV to MOLT-4 cell. Methods: In this study, phage peptide library displaying a random peptides consisting of 7 or 12 random peptides was employed in order to pan against E2 protein. Free HCV particles were separated from the immune complex forms by immunoprecipitation using anti-human IgG antibody, and used for HCV-capture ELISA. To identify the peptides inhibiting E2-binding to MOLT-4 cells, E2 protein was subj ect to bind to MOLT-4 cells under the competition with phage peptides. Results: Several phage peptides were selected for their specific binding to E2 protein, which showed the conserved sequence of SHFWRAP from 3 different peptide sequences. They were also able to recognize the HCV particles in the sera of HCV patients captured by monoclonal antibody against E2 protein. Two of them, showing peptide sequence of HLGPWMSHWFQR and WAPPLERSSLFY respectively, were revealed to inhibit the binding of E2 protein to MOLT-4 cell efficiently in dose dependent mode. However, few membrane-associated receptor candidates were seen using Fasta3 programe for homology search with these peptides. Conclusion: Phage peptides containing HLGPWMSHWFQR and WAPPLERSSLFY respectively, showed the inhibition of E2-binding to MOLT-4 cells. However, they did not reveal any homologues to cellular receptors from GenBank database. In further study, cellular receptor could be identified through the screening of cDNA library from MOLT-4 or hepatocytes using antibodies against these peptide mimotopes.

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A study on the ulcerous inhibitory effects of Taeumchowetang (태음조위탕(太陰調胃湯)의 유탕억제효능(遺瘍抑制效能)에 관(關)한 연구(硏究))

  • Park, Dong-Un;Kim, Dal-Rae
    • Journal of Sasang Constitutional Medicine
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    • v.9 no.2
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    • pp.227-243
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    • 1997
  • The experimental study used in this paper was done to examine the clinical effects of Taeumchowetang for the gastric ulcer and the function of gastrointestinal tract using rats and mice which were administered orally the water extraction from Taeumchowetang. Then, the counter-action of the water extraction on the isolated ileum and gastric fundus, the inhibitory effects of pylorus-ligated ulcer and indomethacin-induced ulcer, the associations with gastric juice secretion, total acidity, pepsin output, the transportability in the small and large intestine, were studied with administering acetylcholine chloride and barium chloride. In addition, it was investigated whether the central nervous system related to pain control and sleeping time was influenced by Taeumchowetang. The following results have been obtained; 1. As resulting from injection of acetylcholine chloride and barium chloride into the isolated ileum of rats and mice, Taeumchowetang led to have an inhibitory effect on the muscle contraction of the ileum. Then, acetylcholine chloride was measured as lower effect than barium chloride 2. For the inhibitory effect on contraction for the gastric fundus strip in rats by either acetylcholine chloride or barium chloride, the one showed low inhibitory effect, on the other hand the other showed density-dependent effect. 3. The water extraction was given on the pylorus ligated ulcer with using two different administration groups of 1,300mg/kg and 2,600mg/kg, each result was measured as 22.9% and 36.5% for an ulcerous inhibitory effect (p<0.05). 4. According to the two administration groups, the preventive effect was tabulated 18.1% and 29.3% for indomethacin-induced ulcer (p<0.05, P<0.01). 5. For associations with gastric juice secretion, total acidity, pepsin output in the administration group 2,600mg/kg, Taeumchowetang was recognized as having an inhibitory effects related to suppressive actions involving gastric juice secretion(p<0.05), and free acidity(p<0.01), but there was no significant association with total acidity and pepsin output. 6. To know the transportability in the intestine, BaSO4 solution was used. The transportability of the small intestine in the administration group 2,600mg/kg was 22.2% which was statistically significant compared with the large intestine's transportability(P<0.01). 7. In the administration group 1,300mg/kg and 2,600mg/kg, analgesic effect with against acetic acid was measured as being 16.8% and 24.4% which was shown as statistically siginificant(p<0.01). 8. No statistically significant association between Taeumchowetang and sleeping time was found in both 1,300mg/kg and 2,600mg/kg by administering phenobarbital-Na. According to the results of this study, Taeumchowetang has agreed with the effects of literature review. Further more in this study, Taeumchowetang also has preventive effects on pylorus-ligated ulcer. Hence, Teaumchowetang can be significant effect such as both anti ulcer agent and increasing gastric activity for the patients who suffer from gastric ulcer.

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Protective effect of euonymus alatus extract on experimental liver injury in mice (Euonymus alatus 추출물의 실험적 간 손상 억제)

  • Shin, Sook-Jeong;Lee, Byung-Yong;Shin, Dong-Keun;Lee, Jeong-Ho
    • IMMUNE NETWORK
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    • v.1 no.3
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    • pp.213-220
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    • 2001
  • Background: A previous study has shown that Euonymus alatus (EA) has an antidotic activities against inflammation, suggesting possibility that EA can exert this beneficial effects to liver injury by an initial protection against drug-induced hepatocyte demage. The present study was undertaken to evaluate the protective effect of EA-extract on experimentally induced hepatitis in ICR mice and to investigate some mechanisms responsible for its action. Methods: Water EA extract was used in this experiments. The mice received i.p. a dose of 700 mg/kg galactosamine (GalN) together with $5{\mu}g/kg$ of endotoxin (LPS), or received i.v. 12 mg/kg of concanavalin A (Con A). EA (4 mg/mouse) was administrated on day -2, -1 and 0 before induction of liver injury. Liver injury was assessed by measurement of serum alanin amino-transferase (SGPT) levels on 9 hr after GaIN.LPS, or 8 hr after con A administration. Results: Treatment with either GaIN or LPS alone did not cause hepatitis. However, simultaneous administration of GalN and LPS to mice resulted in LPS-dose dependent fulminant hepatitis. GaLN/LPS-induced liver injury was reduced when mice were given EA for 3 days before induction. This preventive effect of Ea was more prominent when EA was given by intraperitoneal route rather then by oral route. Pretreatment of EA or dexamethasone inhibited significantly $TNF{\alpha}$ production in GalL/LPS-injured mice. However, EA-treatment did not influence $TNF{\alpha}$-induced hepatitis in GalN-sensitized mice, suggesting that $TNF{\alpha}$ is likely to act as one of final mediators of endotoxin action and the protective effect of EA might be manifested chiefly by inhibition of endotoxin-induced $TNF{\alpha}$ production, not by blocking the $TNF{\alpha}$-action. Injection of Con A into mice evoked remarkable liver injury in a dose dependent fashion. This liver damage was reduced by EA-pretreatment. Dexamethasone significantly reduced both GalL/LPS-induced and Con A-induced liver damages, showing synergism with EA. However, indomethacin reduced only GalN/ LPS-induced hepatitis, not for Con A-induced hepatitis. Conclusion: These results led to the conclusion that EA may be able to contribute at least in part to prevent the drug-induced hepatotoxicity, and that its anti-hepatitis effects might be manifested directly by modulation of endogenous mediators, such as leukotriese D4, $TNF{\alpha}$ and free radical, and indirectly by regulation of immune mediated responses. Also these results suggested that EA could be developed as a potential antidotic agent.

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Effect of 4-hexylresorcinol on Blood Coagulation and Healing of Injured Vessel in a Rat Model

  • Park, Yong-Tae;Park, Si-Yeok;Kim, Min-Keun;Kim, Seong-Gon;Park, Young-Wook;Kwon, Kwang-Jun
    • Maxillofacial Plastic and Reconstructive Surgery
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    • v.35 no.5
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    • pp.284-293
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    • 2013
  • Purpose: For reconstruction of craniomaxillofacial defects caused by tumor, trauma, infection etc, free flap transplantation with microvascular surgery is a very useful method. Thrombus formation at the anastomosis site is the major cause of graft failure. 4-Hexylresorcinol (4-HR) is generally known as an antiseptic and antiparasitic agent. This study was conducted in order to evaluate the effect of 4-HR on blood coagulation in vitro. In addition, we investigated thrombus formation and endothelial repair of an injured vessel in an animal model. Methods: In the in vitro experiment, we compared blood coagulation time between the 4-HR treated group and normal blood. Thirty rats were used for in vivo animal experiments. After exposure of the right femoral vein, a micro vessel clamp was placed and the femoral vein was intentionally cut. Microvascular anastomosis was performed on all rats using 10-0 nylon under microscopy. The animals were divided into two groups. In the experimental group (n=15), 4-HR (250 mg/kg) mixed with olive oil (10 mL/kg) was administered per os daily. Animals in the control group (n=15) were given olive oil only. The animals were sacrificed at three days, seven days, and fourteen days after surgery and rat femoral vein samples were taken. Vascular patency and thrombus formation were investigated just before sacrifice. Histologic analysis was performed under a microscope. Results: Results of an in vitro blood coagulation test showed that coagulation time was delayed in the 4-HR treated group. The results obtained from an in vivo 4-HR administered rat model showed that the patency of all experimental groups was better at thirty minutes, seven days, and fourteen days after microvascular anastomosis than that of the control group at seven and fourteen days after anastomosis, and the amount of thrombus in the experimental groups was much less than that of the control group. Endothelial repair was observed in the histologic analysis. Conclusion: Findings of this study demonstrated that blood coagulation was delayed in the vitro 4-HR treated group. In addition, good vascular patency, anti-thrombotic effect, and repair of venous endothelial cells were observed in the vivo 4-HR administered rat group.

Protective Effect of Nitric Oxide against Oxidative Stress under UV-B Radiation in Maize Leaves (UV-B 조사시 옥수수 잎의 산화적 스트레스에 대한 Nitric Oxide의 보호효과)

  • Kim, Tae-Yun;Jo, Myung-Hwan;Hong, Jung-Hee
    • Journal of Environmental Science International
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    • v.19 no.12
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    • pp.1323-1334
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    • 2010
  • The effect of nitric oxide (NO) on antioxidant system and protective mechanism against oxidative stress under UV-B radiation was investigated in leaves of maize (Zea mays L.) seedlings during 3 days growth period. UV-B irradiation caused a decrease of leaf biomass including leaf length, width and weight during growth. Application of NO donor, sodium nitroprusside (SNP), significantly alleviated UV-B stress induced growth suppression. NO donor permitted the survival of more green leaf tissue preventing chlorophyll content reduction and of higher quantum yield for photosystem II than in non-treated controls under UV-B stress, suggesting that NO has protective effect on chloroplast membrane in maize leaves. Flavonoids and anthocyanin, UV-B absorbing compounds, were significantly accumulated in the maize leaves upon UV-B exposure. Moreover, the increase of these compounds was intensified in the NO treated seedlings. UV-B treatment resulted in lipid peroxidation and induced accumulation of hydrogen peroxide ($H_2O_2$) in maize leaves, while NO donor prevented UV-B induced increase in the contents of malondialdehyde (MDA) and $H_2O_2$. These results demonstrate that NO serves as antioxidant agent able to scavenge $H_2O_2$ to protect plant cells from oxidative damage. The activities of two antioxidant enzymes that scavenge reactive oxygen species, catalase (CAT) and ascorbate peroxidase (APX) in maize leaves in the presence of NO donor under UV-B stress were higher than those under UV-B stress alone. Application of 2-(4-carboxyphenyl)-4, 4, 5, 5-tetramethylimidazoline-1-oxyl-3- oxide (PTIO), a specific NO scavenger, to the maize leaves arrested NO donor mediated protective effect on leaf growth, photosynthetic pigment and free radical scavenging activity. However, PTIO had little effect on maize leaves under UV-B stress compared with that of UV-B stress alone. $N^{\omega}$-nitro-L-arginine (LNNA), an inhibitor of nitric oxide synthase (NOS), significantly increased $H_2O_2$ and MDA accumulation and decreased antioxidant enzyme activities in maize leaves under UV-B stress. This demonstrates that NOS inhibitor LNNA has opposite effects on oxidative resistance. From these results it is suggested that NO might act as a signal in activating active oxygen scavenging system that protects plants from oxidative stress induced by UV-B radiation and thus confer UV-B tolerance.

Changes in the Dyeing Property of Tencel Blended Fabrics as Susceptible Functional Fiber through Chitosan Finishing (키토산 가공에 의한 감성기능소재로서의 텐셀 혼방직물의 염색성 변화)

  • Park, Youn-Hee;Kang, In-Sook;Bae, Hyun-Sook
    • Journal of the Korean Society of Clothing and Textiles
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    • v.31 no.12
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    • pp.1672-1681
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    • 2007
  • Cationization is effective to complement the defects of Tencel blended fabrics by introducing new functions. For this purpose, we used chitosan, which is congenial to the human body, free of pollution, and easily reacted. Then, we compared it to the Tencel single fabrics. To perform such effective cationization, the fabrics were treated with chitosan after NaOH pretreatment and enzyme treatment thereof. After that, the fabrics were treated with a crosslinking agent and a softner. The dyeing property of the cationized Tencel blended fabrics and reactive dye, which is a type of anionic dye, show a high concentration in neutral salt and excellent repulsive power between the fabrics and the decreased dyes. The dyeing property of the chitosan treated fabrics represented better performances than that of untreated fabric in the lower concentration of neutral salt. Meanwhile, when it was dyed with certain acid dyes, the dyeing property of the chitosan treated fabrics showed better results due to the reaction of an amine group, which was introduced by chitosan treatment. Thus, the verification of the cationization of the Tencel blended fabrics was performed. The washing fastness of the Tencel blended fabrics showed a little bit better than that of the Tencel single fabric, and it represented a better performance in the dye with a reactive dye than that of an acid dye.

Effects of hyperbaric oxygen and α-tocopherol on full-thickness skin grafts in rats (쥐(rats)의 full-thickness skin grafts에 대한 hyperbaric oxygen과 α-tocopherol의 효과)

  • Kim, Jong-shu;Kim, Chung-hul;Kim, Gon-sup;Hah, Dae-sik;Park, Sun-gun;Kim, Yang-mi
    • Korean Journal of Veterinary Research
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    • v.40 no.3
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    • pp.497-504
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    • 2000
  • To document that effects of hyperbaric oxygen(HBO) and ${\alpha}-tocopherol$ on full-thickness skin grafts in rat, we performed full-thickness skin grafts bilaterally on each rats. The HBO-treated rats were received HBO twice daily for 90 minutes at 2 ATA. Surgical control rats were not treated with HBO. ${\alpha}-tocopherol$ treated rats were received the agent via oral gastric tube daily for 3 days preoperative and a fourth dose 1 to 2 hours postoperative. HBO plus ${\alpha}-tocopherol$ treated rats were received HBO and ${\alpha}-tocopherol$ as mentioned above. Biopsy specimens were taken from each rat at the time of grafting and on days 2, 4, 7, 10, 14, 21, and 28, then were processed for tissue-concentration of total glutathione(GSHt), oxidized/reduced glutathione level, and thiobarbituric acid-reactive substance(TBARS) levels. The percentage of viable graft on day 10 ranged from 67 to 93%, and was not significantly different among the each other groups. The percentage of viable graft were, however, higher in HBO plus ${\alpha}-tocopherol$ treated rats(78.6%) than in HBO alone treated rats(59.1%), ${\alpha}-tocopherol$ alone treated rats(66.7%) and surgical control rats(58.2%). TBARS concentration had a significant increase from preoperative concentration at day 2, and peak concentration at day 4(p<0.01). Concentration then decreased to preoperative concentration at day 28. GSHt concentration of free skin graft had a similar patteren of change in four groups and decreased significantly from preoperative concentration at day 2, returning to preoperative concentration by day 7(surgical control, HBO-treated, and ${\alpha}-tocopherol-treated$, alone) and 28(HBO plus ${\alpha}-tocopherol-treated$). Percentage of the concentration of reduced glutathione decreased in surgical control, HBO-treated and, ${\alpha}-tocopherol-treated$(p<0.05), and HBO plus ${\alpha}-tocopherol-treared$(p<0.01) on day 7 after surgery, whereas the concentration of oxidized increased significantly in HBO-treated(p<0.05), ${\alpha}-tocopherol-treated$(p<0.05), and HBO plus ${\alpha}-tocopherol-treated$(p<0.01).

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Simultaneous Analysis of Bangpungtongseong-san and Its Antioxidant Effect (방풍통성산의 동시분석 및 항산화 효능 연구)

  • Seo, Chang-Seob;Kim, Ohn Soon;Shin, Hyeun-Kyoo
    • Herbal Formula Science
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    • v.21 no.2
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    • pp.133-143
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    • 2013
  • Objectives : We carry out the simultaneous quantification for quality control of four components in Bangpungtongseong-san (BPTSS) sample. In addition, we assessed the antioxidant effects of BPTSS sample. Methods : The used column for separation and analysis of four compounds was Luna C18 column and column oven temperature was maintained at $40^{\circ}C$. The mobile phase for simultaneous determination consisted of two solvent systems, 1.0% acetic acid in water and 1.0% acetic acid in acetonitrile. High performance liquid chromatography-photodiode array (HPLC-PDA) method for analysis was performed at a flow rate of 1.0 mL/min with PDA detection at 254 and 280 nm. The injection volume was 10 ${\mu}L$. The antioxidant activities of BPTSS were evaluated by measuring free radical scavenging activities on 2,2'-Azinobis-3-ethyl-benzothiazoline-6-sulfonic acid (ABTS) and 1-1-diphenyl-2-picrylhydrazyl (DPPH). The inhibitory effects on low-density lipoprotein (LDL) oxidation were evaluated by the formation of thiobarbituric acid relative substances (TBARS) and relative electrophoretic mobility (REM). Results : Calibration curves were acquired with $r^2{\geq}0.9999$. The values of limit of detection (LOD) and quantification (LOQ) were 0.06-0.29 ${\mu}g/mL$ and 0.20-0.98 ${\mu}g/mL$, respectively. The amounts of geniposide, liquiritin, baicalin, and glycyrrhizin in BPTSS were 5.06, 7.33, 27.56, and 7.81 mg/g, respectively. The BPTSS showed the radical scavenging activity in a dose-dependent manner. The concentration required for 50% reduction (RC50) against ABTS and DPPH radicals were 72.51 ${\mu}g/mL$ and 128.49 ${\mu}g/mL$. Furthermore, GMGHT reduced the oxidation properties of LDL induced by CuSO4. Conclusions : The established HPLC-PDA method will be helpful to improve quality control of BPTSS. In addition, BPTSS has potentials as therapeutic agent on anti-atherosclerosis.

Determination of Acethylsalicylic acid (Aspirin) and Salicylic acid in Eel (Anguilla japonica) Plasma by High-performance Liquid Chromatography (HPLC에 의한 뱀장어 (Anguilla japonica) 혈장중의 acethylsalicylic acid (aspirin)과 salicylic acid의 측정)

  • Kim, Dong-Wan;Koo, Jae-Geun;Park, Sung-Woo
    • Journal of fish pathology
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    • v.20 no.2
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    • pp.139-145
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    • 2007
  • To decrease stress in eel (Anguilla japonica) during its culture or transportation, aspirin (ASA) known as analgesic, antiinflammatory and antithrombic agent was administrated by dipping or oral routes. Concentrations of aspirin (ASA) and salicylic acid (SA) in eel plasma were simultaneously measured by a high performance liquid chromatography (HPLC). The plasma was acidified with 0.2 M HCl and 0.2 M orthophosphoric acid, and mixed with acetonitrile. ASA and SA extracted with acetonitrile were analyzed by the HPLC equipped with reversed phase Novapak C18 column (4 ㎛ silica, 150×4 mm) and UV detector(237 nm). The mobile phase consisted of 740 ㎖ water, 900 ㎕ orthophosphoric acid (85%) and 180 ㎖ acetonitrile. The retention times of ASA, SA and 2-methylbenzoic acid(MBA) were 4.8 min, 8.4 min and 11.5 min, respectively. The limit of quantification was 0.01 ㎍/㎖ for SA and 0.05 ㎍/㎖ for ASA. The mean recovery from eel plasma was 70.8~99.6% for ASA and 95.2~100.3% for SA. This HPLC method was applied to analyze ASA and SA of eel plasma after either dipping in a concentration of 20 ppm or feeding the feed supplemented with 50 ㎎/kg BW. Only SA was detected in eel plasma after the administration of ASA by dipping or oral routes because the drug was quickly decomposed into SA in eel plasma. The amount of SA in eel plasma reached the highest value at 3hr in dipping and 7 days in oral administration. When the ASA-administrated eel were kept in ASA free aquaria, 0.02-0.03 ㎍/㎖ of SA were detected 48 hr after the administration in both routes.

Isolation of Anthocyanin from Black Rice (Heugjinjubyeo) and Screening of its Antioxidant Activities (흑미(현진주벼)유래 안토시아닌의 항산화능 탐색)

  • Park, Young-Sam;Kim, Sun-Joong;Chang, Hyo-Ihl
    • Microbiology and Biotechnology Letters
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    • v.36 no.1
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    • pp.55-60
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    • 2008
  • Colored rices are a hulled grains having red or purple pigments in bran. Especially black rice (Heugjinjubyeo) is considered to be a healthy food in Asia. Black rice is of great interesting because of the possible biological activity with their anthocyanins. Anthocyanins are water-soluble plant pigments and representatives of flavonoids. The anthocyanins in black rice include cyanidin 3-O-glucoside, peonidin 3-O-glucoside, malvidin 3-O-glucoside, pelagonidin 3-O-glucoside and delphinidin 3-O-glucoside. In this study, anthocyanins in a black rice were analyzed quantitatively and qualitatively with HPLC and UV-Vis spectrophotometer. The anthocyanins contained approximately 95% of cyanidin-3-O-glucoside and 5% of peonidin-3-O-glucoside. Antioxidant activities of the anthocyanin extract were investigated by using various in vitro methods. The 100g/ml concentration of the anthocyanin extracted exhibited 88.83% inhibition on the peroxidation of linoleic acid, 55.20% DPPH free radical scavenging activity, 54.96% superoxide anion radical scavenging activity, and 72.67% hydrogen peroxide scavenging activity. And it also showed high ferrous ion reducing capability. These results suggest that the anthocyanin extracted from black rice may be utilized as a possible antioxdiant agent against ROS.