• Title/Summary/Keyword: DNA comet assay

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Evaluation of Environmental Mutagens-Complex Mixture in Diesel Exhaust Respirable Particulate Matter

  • Kim, Soung-Ho;Ryu, Byung-Tak;Jang, Hyoung-Seok;Kim, Yun-Hee;Lee, Do-Han;Han, Kyu-Tae;Oh, Seung-Min;Chung, Kyu-Hyuck
    • Proceedings of the Korea Society of Environmental Toocicology Conference
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    • 2003.05a
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    • pp.194-194
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    • 2003
  • The International Agency for Research on Cancer (IARC, 1989) has classified whole diesel exhaust as probably carcinogenic to humans. Diesel exhaust particulate matter (DPM) adsorbs different chemical substances including PAHs and nitroarenes. DPM is emphasized because it is a major component of diesel exhaust, it is suspected of contributing to a health hazard. Diesel exhaust is a complex mixture of carbon particles and associated organics and inorganics, and it is not known what fraction or combination of fractions cause the health effects [cancer effects, noncancer effects (respiratory tract irritation/inflammation and changes in lung function)] that have been observed with exposure to diesel exhaust. In order to identify which chemical classes are responsible for the majority of the observed biological activities, we performed a particular biological/chemical analysis. Respirable particulate matter (PM2.5: <2.5mm) was collected from diesel engine exhaust using a high-volume sampler equipped with a cascade impactor. Particulate oganic matter was extracted by the dichloromethane/sonication method and the crude extract was fractionated according to EPA recommended procedure into seven fractions by acid-base partitioning and silica gel column chromatography. We examined genotoxic potentials of diesel exhaust particulate matter using novel genotoxicity tests, which are rapid, simple and sensitive methods for assessing DNA-damage at the DNA and chromosomal level (comet assay, in vitro MN test and Ames test). Higher genotoxic potency was observed in non polar fractions and several PAHs were detected by GC-MS, such as 1,2,5,6 dibenzanthracene, chrysene, 1,2-benzanthracene, phenanthrene and fluoranthene.

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Protective effect of methanol extract from citrus press cakes prepared by far-infrared radiation drying on $H_2O_2$-mediated oxidative damage in Vero cells

  • Wijesinghe, W.A.J.P.;Senevirathne, Mahinda;Oh, Myung-Cheol;Jeon, You-Jin
    • Nutrition Research and Practice
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    • v.5 no.5
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    • pp.389-395
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    • 2011
  • In the present study, a suitable drying method was developed for citrus press cakes (CPCs), which are produced as a by-product in citrus juice plants, and the protective effect of methanol extract of CPCs prepared by far-infrared radiation (FIR) drying against $H_2O_2$-induced DNA damage was evaluated versus that of freeze-dried CPCs. Methanol extract of FIR-dried CPCs exhibited comparatively good ROS scavenging activity versus the freeze-dried CPCs at the concentration of 100 ${\mu}g$/mL. The extract strongly enhanced the cell viability against $H_2O_2$-induced oxidative damage in Vero cells. Lipid peroxidation inhibitory activity of the extract from FIR-dried CPCs was comparable to that of the extract from freeze-dried CPCs. This sample also exhibited good protective effects against $H_2O_2$-mediated cell apoptosis as demonstrated by decreased apoptotic body formation in the nuclear staining with Hoechst 33342. In the comet assay, the CPC extracts exhibited strong inhibitory effects against $H_2O_2$-mediated DNA damage in a dose-dependent manner. Thus, this study demonstrated that FIR drying effectively preserves CPC as a functionally important natural antioxidant source and the FIR drying can be adapted for drying CPCs and is more economical for massive production than freeze drying.

Antioxidant Effects of Spinach (Spinacia oleracea L.) Supplementation in Hyperlipidemic Rats

  • Ko, Sang-Heui;Park, Jae-Hee;Kim, So-Yun;Lee, Seon Woo;Chun, Soon-Sil;Park, Eunju
    • Preventive Nutrition and Food Science
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    • v.19 no.1
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    • pp.19-26
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    • 2014
  • Increased consumption of fresh vegetables that are high in polyphenols has been associated with a reduced risk of oxidative stress-induced disease. The present study aimed to evaluate the antioxidant effects of spinach in vitro and in vivo in hyperlipidemic rats. For measurement of in vitro antioxidant activity, spinach was subjected to hot water extraction (WE) or ethanol extraction (EE) and examined for total polyphenol content (TPC), oxygen radical absorbance capacity (ORAC), cellular antioxidant activity (CAA), and antigenotoxic activity. The in vivo antioxidant activity of spinach was assessed using blood and liver lipid profiles and antioxidant status in rats fed a high fat-cholesterol diet (HFCD) for 6 weeks. The TPC of WE and EE were shown as $1.5{\pm}0.0$ and $0.5{\pm}0.0mg$ GAE/g, respectively. Increasing the concentration of the extracts resulted in increased ORAC value, CAA, and antigenotoxic activity for all extracts tested. HFCD-fed rats displayed hyperlipidemia and increased oxidative stress, as indicated by a significant rise in blood and liver lipid profiles, an increase in plasma conjugated diene concentration, an increase in liver thiobarbituric acid reactive substances (TBARS) level, and a significant decrease in manganese superoxide dismutase (Mn-SOD) activity compared with rats fed normal diet. However, administration of 5% spinach showed a beneficial effect in HFCD rats, as indicated by decreased liver TBARS level and DNA damage in leukocyte and increased plasma conjugated dienes and Mn-SOD activity. Thus, the antioxidant activity of spinach may be an effective way to ameliorate high fat and cholesterol diet-induced oxidative stress.

Lymphocyte DNA damage and plasma antioxidant status in Korean subclinical hypertensive patients by glutathione S-transferase polymorphism

  • Han, Jeong-Hwa;Lee, Hye-Jin;Choi, Hee Jeong;Yun, Kyung Eun;Kang, Myung-Hee
    • Nutrition Research and Practice
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    • v.11 no.3
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    • pp.214-222
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    • 2017
  • BACKGROUND/OBJECTIVES: Glutathione S-transferase (GST) forms a multigene family of phase II detoxification enzymes which are involved in the detoxification of xenobiotics by conjugating substances with glutathione. The aim of this study is to assess the antioxidative status and the degree of DNA damage in the subclinical hypertensive patients in Korea using glutathione S-transferase polymorphisms. SUBJECTS/METHODS: We examined whether DNA damage and antioxidative status show a difference between GSTM1 or GSTT1 genotype in 227 newly diagnosed, untreated (systolic blood pressure $(BP){\geq}130mmHg$ or diastolic $BP{\geq}85mmHg$) subclinical hypertensive patients and 130 normotensive subjects (systolic BP < 120 mmHg and diastolic BP < 80 mmHg). From the blood of the subjects, the degree of the DNA damage in lymphocyte, the activities of erythrocyte superoxide dismutase, the catalase, and the glutathione peroxidase, the level of glutathione, plasma total radical-trapping antioxidant potential (TRAP), anti-oxidative vitamins, as well as plasma lipid profiles and conjugated diene (CD) were analyzed. RESULTS: Of the 227 subjects studied, 68.3% were GSTM1 null genotype and 66.5% were GSTT1 null genotype. GSTM1 null genotype had an increased risk of hypertension (OR: 2.104, CI: 1.38-3.35), but no significant association in GSTT1 null genotype (OR 0.982, CI: 0.62-1.55). No difference in erythrocyte activities of superoxide dismutase, catalase, or glutathione peroxidase, and plasma TRAP, CD, lipid profiles, and GSH levels were observed between GSTM1 or GSTT1 genotype. Plasma levels of ${\alpha}-tocopherol$ increased significantly in GSTT1 wild genotype (P < 0.05); however, plasma level of ${\beta}-carotene$ increased significantly in GSTT1 null genotype (P < 0.01). DNA damage assessed by the Comet assay was significantly higher in GSTM1 null genotype than wild genotype (P < 0.05). CONCLUSIONS: These results confirm the association between GSTM1 null genotype and risk of hypertension as they suggest that GSTM1 null genotype leads to an increased oxidative stress compared with wild genotype.

Effects of Chungkukjang Added with Onion on Lipid and Antioxidant Metabolisms in Rats Fed High Fat-Cholesterol Diet (양파 첨가 청국장이 고콜레스테롤혈증 유발 흰쥐의 지질 및 항산화 대사에 미치는 영향)

  • Park, Jae-Hee;Kim, Jung-Mi;Park, Eun-Ju;Lee, Kyung-Hea
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.37 no.10
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    • pp.1244-1250
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    • 2008
  • This study was carried out investigate the effects of chungkukjang added with onion (OC) and chungkukjang (C) on the lipid and antioxidant metabolisms of hypercholesterolemic rats. Thirty-two male Sprague-Dawley rats were divided into four groups after 1-week adaptation period and were fed a normal diet, high fat-cholesterol diet (HC), HC-OC or HC-C for 8 weeks, respectively. The supplementation of HC-OC and HC-C groups significantly reduced hepatic total cholesterol and AST activity. HC-C group increased high density lipoprotein, while decreasing low density lipoprotein and AI compared with HC-OC group. Conjugated dienes of HC-C group was significantly lower than those of HC-OC group. $H_2O_2$ induced DNA damage reduced significantly in HC-OC and HC-C groups compared with high fat-cholesterol diet group. $H_2O_2$ induced DNA damage exhibited significant positive correlations with hepatic total cholesterol, AST and CD. These results suggested that supplementation of chungkukjang or chungkukjang added with onion might be helpful in preventing lipid oxidation and leukocytic DNA damage. However, the health beneficial effect has not improved by the addtion of onion in chungkukjang.

No Relationship between the Amount of DNA Damage and the Level of hMLH1 and RASSF1A Gene Expression in Bladder Cancer Cells Treated with Cisplatin and Gemcitabine

  • de Camargo, Elaine Aparecida;da Silva, Glenda Nicioli;Gobette, Camila Pereira;de Castro Marcondes, Joao Paulo;Salvadori, Daisy Maria Favero
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.10
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    • pp.5941-5948
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    • 2013
  • Tumor response to antineoplastic drugs is not always predictable. This is also true for bladder carcinoma, a highly recurrent neoplasia. Currently, the combination of cisplatin and gemcitabine is well accepted as a standard protocol for treating bladder carcinoma. However, in some cases, this treatment protocol causes harmful side effects. Therefore, we investigated the roles of the genes TP53, RASSF1A (a tumor suppressor gene) and hMLH1 (a gene involved in the mismatch repair pathway) in cell susceptibility to cisplatin/gemcitabine treatment. Two bladder transitional carcinoma cell (TCC) lines, RT4 (wild-type TP53) and 5637 (mutated TP53), were used in this study. First, we evaluated whether the genotoxic potential of cisplatin/gemcitabine was dependent on TP53 status. Then, we evaluated whether the two antineoplastic drugs modulated RASSF1A and hMLH1 expression in the two cell lines. Increased DNA damage was observed in both cell lines after treatment with cisplatin or gemcitabine and with the two drugs simultaneously, as depicted by the comet assay. A lack of RASSF1A expression and hypermethylation of its promoter were observed before and after treatment in both cell lines. On the other hand, hMLH1 downregulation, unrelated to methylation status, was observed in RT4 cells after treatment with cisplatin or with cisplatin and gemcitabine simultaneously (wild-type TP53); in 5637 cells, hMLH1 was upregulated only after treatment with gemcitabine. In conclusion, the three treatment protocols were genotoxic, independent of TP53 status. However, cisplatin was the most effective, causing the highest level of DNA damage in both wild-type and mutated TP53 cells. Gemcitabine was the least genotoxic agent in both cell lines. Furthermore, no relationship was observed between the amount of DNA damage and the level of hMLH1 and RASSF1A expression. Therefore, other alternative pathways might be involved in cisplatin and gemcitabine genotoxicity in these two bladder cancer cell lines.

Antioxidant Activity and Radioprotection of Two Flavonoids from Propolis (프로폴리스에서 분리한 플라보노이드 화합물의 항산화 활성 및 방사선 방어효과)

  • Jeong, Ill-Yun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.34 no.2
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    • pp.162-166
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    • 2005
  • Two flavonoids, 7-O-methyl-3',4'-didehydroxy quercetin (MDQ) and quercetin, isolated from Chinese propolis, which is the generic name for the resinous substance collected by honeybees from various plant sources, were tested for their antioxidant activity and protective effect against radiation-induced DNA damage in mouse lymphocytes. In antioxidant test, both compounds provided a dose-dependent scavenging effect on DPPH radical and a dose-dependent inhibitory effect on lipid peroxidation in mouse liver. Quercetin showed stronger scavenging and inhibitory effect than MDQ, and it also provided strong inhibition on superoxide anion radical generated in xanthine-xanthine oxidase system, but there was no inhibitory ability for MDQ. In comet assay using single cell gel electrophoresis, MDQ and quercetin showed a protective effect against DNA damage caused by gamma irradiation. They reduced DNA damage to 54% (p<0.01) and 53% (p<0.01) at 25 $\mu$mol, respectively. These results suggest that free radical scavenging seems to be associated with their catechol form on the B ring, and radioprotection appears to be a likely mechanism of antioxidant activity by these flavonoids.

Effects of Gamma Irradiation on Color Changes and Antioxidative Activities of Caesalpinia sappan L. (감마선 조사에 의한 소목추출물의 색상변화 및 항산화 활성 평가)

  • Kwon, Hyung-Jun;Jung, U-Hee;Park, Hae-Ran;Shin, Dong-Hwa;Jo, Sung-Kee
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.36 no.8
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    • pp.1055-1061
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    • 2007
  • Caesalpinia sappan L. is an indeciduous tree distributed in China, India, Miyanmar and Vietnam. Its heartwood has long been used in oriental folk medicines to treat diseases. In this study, antioxidative activities of Caesalpinia sappan L. and the effect of gamma irradiation on its chemical and biological properties were investigated. The ethyl acetate fraction (EtOAc fr.) of Caesalpinia sappan L. was irradiated with 100 kGy of gamma ray. The dark red color of EtOAc fr. was significantly (p<0.05) removed by irradiation (Hunter L and b values increased and a value decreased). The total phenolic content of EtOAc fr. was 865 mg/g and it was increased to 1195 mg/g by gamma irradiation. DPPH radical and superoxide anion radical scavenging activities and lipid peroxidation inhibitory activity of EtOAc fr. were very high and its activities were also increased by gamma irradiation. EtOAc fr. also inhibited the irradiation-induced DNA damage of lymphocyte as determined by comet assay. In conclusion, EtOAc fr. of Caesalpinia sappan L. extract showed high antioxidative activities in vitro. Furthermore, gamma irradiation on EtOAc fr. ameliorated the color and antioxidative properties. Therefore, it can be suggested that Caesalpinia sappan L. may be a good material for antioxidant function and gamma irradiation may be applied for the improvement of chemical and biological properties of Caesalpinia sappan L.

Antioxidative Activity of Extracts from Wisteria floribunda Flowers (등나무 꽃 추출물의 항산화 활성)

  • Oh, Won-Gyeong;Jang, In-Cheol;Jeon, Gyeong-Im;Park, Eun-Ju;Park, Hae-Ryong;Lee, Seung-Cheol
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.37 no.6
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    • pp.677-683
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    • 2008
  • The antioxidant activities of Wisteria floribunda flowers (WFF) were evaluated. The samples were prepared by extracting separately two different colored flowers (purple and white) with four different solvents (methanol, ethanol, acetone, and water). The antioxidant properties were evaluated by determining total phenolic contents (TPC), radical scavenging activity (RSA), and reducing power (RP). Water extract from purple WFF and ethanol extract of white WFF showed the highest total phenol contents (491 and 787 ${\mu}M$ gallic acid equivalents), respectively. Water extracts of purple and white WFF also showed higher RSA. In the case of RP, ethanol extract of purple WFF, methanol and water extracts of white WFF showed relatively higher values. The 200 ${\mu}M$ $H_2O_2$ induced oxidative DNA damage in human leukocytes was significantly inhibited with WFF extracts excluding ethanol and acetone extracts of purple flowers. These results suggest that W. floribunda flowers have significant antioxidative activity and protective effect against oxidative DNA damage.

Changes in the constituents and UV-photoprotective activity of Astragalus membranaceus caused by roasting (황기의 볶음 조건에 따른 성분 및 자외선 광보호 활성 변화)

  • Park, Jeong-Yong;Lee, Ji Yeon;Kim, Hyung Don;Jang, Gwi Yeong;Seo, Kyung Hye
    • Journal of Nutrition and Health
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    • v.52 no.5
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    • pp.413-421
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    • 2019
  • Purpose: Astragalus membranaceus (AM) is an important traditional medicinal herb. Pharmacological research has indicated that AM has various physiological activities such as antioxidant, anti-inflammatory, immunoregulatory, anticancer, hypolipidemic, antihyperglycemic, and hepatoprotective activities. The bioactive substances responsible for the physiological activities in AM, including many antioxidant substances, change during the roasting process. This study investigated and compared the changes in the antioxidant constituents of AM caused by roasting. Methods: DPPH (1,1-diphenyl-2-picryl hydrazyl) and $ABTS^+$ (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt) radical scavenging activities and their total phenolic content (TPC) were measured. High-performance liquid chromatography (HPLC) analysis was performed to confirm any changes in the isoflavonoids of roasted AM (R-AM),. The cell viability of UVB-induced HDF (Human dermal fibroblast) cells treated with AM and R-AM extracts was investigated. The comet assay was used to examine the inhibitory effects of R-AM extracts on DNA damage caused by oxidative stress. Results: The DPPH and $ABTS^+$ radical scavenging activities were $564.6{\pm}20.9$ and $108.2{\pm}3.1$ ($IC_{50}$ value) respectively, from the 2R-AM. The total phenol content was $47.80{\pm}1.40mg$ GAE/g from the 1R-AM. The values of calycosin and formononetin, which are the known isoflavonoid constituents of AM, were $778.58{\pm}2.72$ and $726.80{\pm}3.45{\mu}g/g$ respectively, from the 2R-AM. Treatment of the HDF cells with R-AM ($50{\sim}200{\mu}g/mL$) did not affect the cell viability. Furthermore, the R-AM extracts effectively protected against UVB-induced DNA damage. Conclusion: The findings of this study indicate that R-AM increases its isoflavonoid constituents and protects against UVB-induced DNA damage in HDF cells.