• Journal of Ginseng Research
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• v.13 no.1
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• pp.42-48
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• 1989

The binding of bay region diol-epoxides of polycyclic aromatic hydrocarbons (PAHs) to target tissue DNA is thought to be essential for the initiation of cancer by these compounds. In this study we investigated the effect of polyacetylenes such as panaxynol and panaxydol on the formation of benzo(a)pyreno (BP)-metabolite-DNA adduct in the liver of ICR mice. Treatment of mice by i.p. administration of polyacetylenes produced a marked reduction in BP metabolite binding to DNA in vitro. Following i.v. administration of (3H)BP(300, ${\mu}$Ci/21 nmoles/0.1 nt DMSO) to mice, radioactivity was detected in the DNA of the liver in vivo. The result of tentative identification of the 4 peaks between the two standard markers for high pressure liquid chromatography showed that the peaks. I, II, III, and IV were BP-phenol oxide-DNA adduct (or BP-diol-epoxide-dCyt. adduct), (-) BP$.$diolepoxide I:dGuO adduct, (+) BP-diol-epoxide I: dGuo adduct, and BP-diol-epoxide II:dGuO adduct, respectively. The minor adduct, (-) BP-diol epoxide I: dGuo was reduced to 6971 of the amount of the control, while the major adduct, (+) BP-diolepoxide I: dGuO(peak II) which was produced from (-) BP-7, 8-diol was reduced to 78% of that of the control. The amount of the minor adduct, BP-diol-epoxide II:dGuo adduct(peak IV) which formed from (+) BP-7, 8-diol was 58% of the control. These results show that the panaxydol is more related to inhibition of the formation of the minor ad- ducts than of the major adducts, which were generally produced from ($\pm$) BP-7, 8-dihydro-dials.

• BMB Reports
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• v.36 no.1
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• pp.1-11
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• 2003

The causative role of polycyclic aromatic hydrocarbons (PAH) in human carcinogenesis is undisputed. Measurements of PAH-DNA adduct levels in easily accessible white blood cells therefore represent useful early endpoints in exposure intervention of chemoprevention studies. The successful applicability of DNA adducts as early endpoints depends on several criteria:i.adduct levels in easily accessible surrogate tissues should reflect adduct levels in target-tissues, ii. toxicokinetics and the temporal relevance should be properly defined.iii. sources of inter- and intra-individual variability must be known and controllable, and finally iv. adduct analyses must have advantages as compared to other markers of PAH-exposure. In general, higher DNA adduct levels or a higher proportion of subjects with detectable DNA adduct levels were found in exposed individuals as compared with non-exposed subjects, but saturation may occur at high exposures. Furthermore, DNA adduct levels varied according to changes in exposure, for example smoking cessation resulted in lower DNA adduct levels and adduct levels paralleled seasonal variations of air-pollution. Intra-individual variation during continuous exposure was low over a short period of time (weeks), but varied significantly when longer time periods (months) were investigated. Inter-individual variation is currently only partly explained by genetic polymorphisms in genes involved in PAH-metabolism and deserves further investigation. DNA adduct measurement may have three advantages over traditional exposure assessment: i. they can smooth the extreme variability in exposure which is typical for environmental toxicants and may integrate exposure over a longer period of time. Therefore, DNA adduct assessment may reduce the monitoring effort. ii. Biological monitoring of DNA adducts accounts for all exposure routes. iii. DNA adducts may account for inter-individual differences in uptake, elimination, distribution, metabolism and repair amongst exposed individuals. In conclusion, there is now a sufficiently large scientific basis to justify the application of DNA adduct measurement as biomarkers in exposure assessment and intervention studies. Their use in risk-assessment, however, requires further investigation.

• Journal of Environmental Health Sciences
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• v.27 no.3
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• pp.63-70
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• 2001

To identify and evaluate the benzidine-DNA adducts in liver and bladder, we exposed the 80 ppm benzidine to 40 sprague-dawley rats by drinking water for 4 weeks(6.2 mg/kg body wt./day). Only one benzidine-DNA adduct was found at the same site of thin layer chromatogram of $^{32}$ P-postlabeling method in the liver and bladder of exposed rats. So we know the DNA adduct formed at liver and bladder were similar to each other, which was N-(deoxyguanosin-8-yl)-N'-acetylbenzidine. Relative adduct labeling(RAL) of DNA adduct was similar to each other for 1 and 2 weeks, but that in liver was significantly higher than in bladder for 3 and 4 weeks. RAL$\times$10$^{9}$ of DNA adduct were 84.45$\pm$11.31 and 152.8$\pm$5.53 in liver, and were 24.76$\pm$7.06 and 38.00$\pm$10.57 in bladder for 3 and 4 weeks, respectively. Regression equation between liver and bladder was Y=-3.801+2.507 X(r=0.6036, p<0.01). In conclusion, benzidine-DNA adduct formed in liver was significantly higher than that in bladder, with the similar compound structure in sparague-dewley rates treated benzidine in drinking water.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.193-193
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• 1994

발암물질의 조기검색법 개발 및 chemoprevention연구의 일환으로 발암물질과 DNA 및 단백질의 공유결합체인 발암물질-DNA 및 -단백질 adduct를 연구하였다. 발암물질(예, 밴조피렌)-단백질 adduct에 관한 연구에서는 시료(단백질)에 soluble protease를 이용하는 간편하고 손쉬운 ELISA(Enzyme Linked Immunosorbent Assay)분석법을 확립했다. 발암물질(예,벤조피렌,아플라톡신 B1) -DNA 및 -단백질 adduct를 이용한 발암성 조기검색법의 개발을 Ames test 및 염색체이상시험과 비교 연구한 결과 본 연구에서 새로이 개발한 DNA 및 Protein-adduct형성 시험법은 저농도에서 고농도에 이르기까지 뚜렷한 용량-반응 관계를 나타냈으며 Ames test 및 Chromosomal test에서 일어날 수 있는 false positive나 false negative의 결과를 나타낼 우려가 없었다. 벤조피렌-DNA adduct를 이용한 chemoprevention 연구에서는 항산화제로 알려진 비타민 E,C 및 $\beta$-carotene을 시험한 결과 용량의존적으로 벤조피렌-DNA adduct 형성을 억제하였다.

• Korean Journal of Food Science and Technology
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• v.24 no.4
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• pp.367-370
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• 1992

Aflatoxins are highly carcinogenic agents consistantly found as contaminants in human food supplies in many areas of the world and epidemiologically linked to incidence of human liver cancer. To examine the carcinogenic action of aflatoxin $B_1$, aflatoxin $B_1-DNA$ adducts were chemically synhtesized with the reaction of 20 mg calf thymus DNA, and 8 mg standard aflatoxin $B_1$. Since DNA molecule was too large for analysis, it was fragmented by acid hydrolysis and heat. The fragmented aflatoxin $B_1-DNA$ adducts were selectively concentrated by immunoaffinity column procedure and confirmed by HPLC method. The main component was aflatoxin $B_1-guanine$ adduct, which was quantatively measured as 5.2 mg aflatoxin $B_1$.

• Proceedings of the Korean Environmental Health Society Conference
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• 2002.04a
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• pp.49-51
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• 2002

To identify and evaluate the dichlorobenzidine(DCB)-DNA adducts in liver cell and bladder epithelial cells by $^{32}$ P-postlabeling and GC/MS-SIM, we orally exposed the dichlorobenzidine (20mg/kh body wt.,/day)to male sprague-dawley rats for 14 days. Two kinds of DCB-DNA adduct were found at the same site of thin layer chromatogram of $^{32}$ P-postlabeling method in liver cells and bladder epithelial cells. In liver cells, relative adduct labeling(RAL) $\times$ 10$^{12}$ of DCB-DNA adduct A1 were 34.1$\pm$3.71 and 69.9$\pm$5.02, that of adduct A2 were 74.1$\pm$10.1 and 105.1$\pm$10.1 on 10 and 14 days after treatment, respectively. And in bladder epithelia cells, RAL $\times$ 10$^{12}$ of DCB-DNA adduct A1 were 5.92$\pm$1.60 and 15.9$\pm$1.31, that of adduct A2 were 9,81$\pm$2.81 and 22.8$\pm$1.79 on 10 and 14 days after treatment, respectively. DCB metabolites formed DNA adducts were monoacetyl-dichlorobenzidine(acDCB) and diacety1-dichlorobenzidine(di-acDCB), which was identify by gas chromatography/mass spectrometry-scan ionization mode(GC/MS-SIM), along with hydrolysis, extraction and TFA(trifluoroacetyl anhyride) derivatization with DCB-DNA adducts isolated from live cells and bladder epithelial cells. The base peak of acDCB were 252 and 294 m/z, and that of di-acDCB were 252, 294 and 336 m/z. In conclusion, the exposed DCB formed two kinds of DCB-DNA adduct, the proximate materials of that were acDCB and di-acDCB in liver and bladder epithlial cells. And the above GC/MS-SIM method was found the DCB-DNA adducts could be monitoring by gas chromatography.

• Journal of Environmental Health Sciences
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• v.40 no.1
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• pp.55-62
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• 2014

Objectives: $^{32}P$-postlabeling assay is the most sensitive method of detecting DNA adduct formation. However, it is limited by a low sample throughput and use of radioisotopes (RI). In this study, we modified it to minimize these limitations and applied it to Z. platypus exposed to Benzo(a)pyrene (BaP) in order to investigate DNA adduct formation (effect biomarker for pollutants) in Z. platypus for assessing risk of waterborne BaP exposure. Methods: DNA hydrolysis was performed only with Micrococcal nuclease (MNase), RI reduction test was performed and the overlapping steps between thin layer chromatography (TLC) and radioisotope high-performance liquid chromatography (RI-HPLC) were omitted. The application of a modified method to Z. platypus exposed to BaP was performed. Results: The results revealed that the amount of RIs used can be reduced roughly 10-fold. Because the analysis time was shortened by 8.5 hours, the sample throughput per hour was increased compared with the previous method. The results of applying modified $^{32}P$-postlabeling assay to Z. platypus, DNA adduct formation in Z. platypus showed dose-dependency with the BaP concentration. Only BPDE-dGMP was detected as a DNA adduct. Conclusion: These results demonstrate that the modified $^{32}P$-postlabeling assay is a suitable method for detecting DNA adduct formation in Z. platypus exposed to waterborne BaP and will be useful in risk assessment of carcinogenic effect in aquatic environment due to BaP.

• Journal of Environmental Health Sciences
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• v.24 no.3
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• pp.92-98
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• 1998

작업장에서 근로자들이 엔진오일 등 각종 오일에 피부가 폭로되었을 때 이것을 쉽게 세척하기 위하여 일반적으로 솔벤트를 사용한다. 그러나 솔벤트를 사용하면 피부를 건조하게 만들 뿐만 아니라 오일에 함유되어 있는 각종 성분들을 피부내에 흡수되는 것을 촉진시킬 수 있어서 이에대한 대처방법이 요구된다. 특히 폐가솔닐엔진오일데는 방향족탄화수소(PAHs)와 같은 물질이 함유되어 있어 체내에 흡수되면 발암물질로 대사되어 표적장기(피부와 폐조직)에서 DNA adducts를 높은 수준으로 형성한다고 알려져 있다. 본 연구에서는 식물기름에서 구할 수 있는 d-라이모닌(Limonene)를 세척제로 사용하여 폐가솔린엔진오일의 폭로로 인하여 형성되는 DNA adducts를 $^{32}P-postlabeling방법으로 분석함으로써 d-라이모닌의 세척효과를 평가하고자 하였다. HDC(ICR) Br 자성마우스의 견갑골 부위에 있는 털을 제거하고 그 부위에 폐가솔린엔진오일을 폭로시키고 1시간과 8시간이 지난 다음에 d-라이모닌으로 각각 세척을 하였다. 마지막 폭로를 마치고 24시간이 지난 다음에 실험동물을 희생시켜 표적장기(폭로된 피부와 폐)에서 시료를 채취하였다. 먼저 시료에서 DNA를 분리하여 가수분해한 다음에 $^{32}P-postlabeling하여 DNA adducts를 분리하였다. 폐가솔린엔진오일만 폭로시킨 그룹의 피부와 폐조직에 형성된 DNA adducts가 각각 30.3$\pm$3.7과 15.7$\pm$2.4로서 대조군(2.5$\pm$1.0과 1.4$\pm$0.4)에 비하여 통계적으로 유의하게 높았고 (p<0.01), 또한 폐조직에서 보다 피부조직에서 통계적으로 유의하게 높았다(p<0.01). 폐가솔린엔진오일을 폭로시킨 후에 d-라이모닌으로 세척한 그룹에서는 피부와 폐조직에 형성된 DNA adducts가 통계적으로 유의하게 감소하였는데(p<0.01), 8시간 보다는 1시간이 지난 다음에 세척한 그룹에서 DNA adducts의 감소현상이 더 크게 나타났다. 결론적으로 피부에 폭로된 폐가솔린 엔진오일을 d-라이모닌으로 세척하면 폐가솔닐엔진오일내에 함유된 발암물질이 체내흡수되는 것이 억제되고, 피부와 폐조직 모두에서 DNA adducts의 형성을 감소시킬 수 있으며, 폐오일이 폭로된 후 빨리 세척하는 것이 더 효과적임을 증명하였다.

• Journal of Korean Society of Occupational and Environmental Hygiene
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• v.3 no.1
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• pp.14-21
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• 1993

N-Heterocyclic aromatics (NHA) are widely occurring environmental pollutants formed during the pyrolysis of nitrogen-containing organic chemicals. NAH are found in significant amounts in tobacco condensates, synthetic fuels, polluted river sediment, and effluents from the heating of coal. Following topical application 7H-dibenzo[c, g]carbazole (DBC) induces cancer in liver as well as skin, indicating that dermal exposure can lead to systemic effect. DBC and dibenz[a,j]acridine (DBA) are examples of NHA. The potency of many carcinogenic compounds is related, at least in part, to the efficiency of their biological activation. We undertook studies to determine which initial metabolites lead to the formation of high levels of carcinogen-DNA adducts in vivo. DBC and DBA's, DBA, trans-DBA-1,2-dihydrodiol (DBA-1,2-DHD), trans-DBA-3,4-dihydrodiol (DBA-3,4-DHD), and trans-DBA-5,6-dihydrodiol (DBA-5,6-DHD), were applied to the skin of mice. There were six adducts that were related to DBC application. These addusts were seen in the target organ, liver at high levels, but at very low levels in non-target organs, skin, lung and kidney. In skin, DBA produced two distinct adducts. The same two adducts were seen when DBA-3,4-DHD was applied. In addition the total adduct level elicited by DBA-3,4-DHD higher than that of parent compound. Two adducts were seen when DBA-5,6-DHD was applied, but these were very different from adducts seen with DBA. These results suggested that activation of DBA to DNA-binding compounds in skin includes initial formation of DBA-3,4-DHD.

• Archives of Pharmacal Research
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• v.25 no.1
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• pp.11-24
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• 2002

The antitumor antibiotic (+)-CC-1065 can alkylate N3 of guanine in certain sequences. A previous high-field $^1H$ NMR study on the$(+)-CC-1065d[GCGCAATTG*CGC]_2$ adduct ($^*$ indicates the drug alkylation site) showed that drag modification on N3 of guanine results in protonation of the cross-strand cytosine [Park, H-J.; Hurley, L. H. J. Am. Chem. Soc.1997, 119,629]. In this contribution we describe a further analysis of the NMR data sets together with restrained molecular dynamics. This study provides not only a solution structure of the (+)-CC-1065(N3- guanine) DNA duplex adduct but also new insight into the molecular basis for the sequence- specific interaction between (+)-CC-1065 and N3-guanine in the DNA duplex. On the basis of NOESY data, we propose that the narrow minor groove at the 7T8T step and conformational kinks at the junctions of 16C17A and 18A19T are both related to DNA bending in the drugDNA adduct. Analysis of the one-dimensional $^1H$ NMR (in $H_2O$) data and rMD trajectories strongly suggests that hydrogen bonding linkages between the 8-OH group of the (+)-CC-1065 A-sub-unit and the 9G10C phosphate via a water molecule are present. All the phenomena observed here in the (+)-CC-1065(N3-guanine) adduct at 5'$-AATTG^*$are reminiscent of those obtained from the studies on the (+)-CC-1065(N3-adenine) adduct at $5'-AGTTA^*$, suggesting that (+)-CC-1065 takes advantage of the conformational flexibility of the 5'-TPu step to entrap the bent structure required for the covalent bonding reaction. This study reveals a common molecular basis for (+)-CC-1065 alkylation at both $5'-TTG^*$ and $5'-TTA^*$, which involves a trapping out of sequence-dependent DNA conformational flexibility as well as sequence-dependent general acid and general base catalysis by duplex DNA.