•  
  •  
 

Abstract

This study was conducted to observe the profile of DNA Adduct (8-OHdG) formation as DNA damage indicators, by using calf thymus DNA incubated with toxic and carcinogenic compounds. The compounds which could trigger free radicals in this research were PAH(Benzo[a]Pyrene), TiO2, and CuCl2. Calf thymus DNA was incubated with Benzo[a]Pyrene and CuCl2 compounds under pH and temperature variations. The incubation of calf thymus DNA with TiO2-UV radiation (254 nm) was used to induce the formation of reactive oxygen species (ROS) in the process of oxidative DNA damage. From this research, all of compounds have potency to trigger the formation of DNA Adduct (8-OHdG). The ratio of absorbance to assess the purity of DNA at 260 nm and 280 nm (λ260/ λ280 ) was measured at ~1.9. The shifted peaks at λmax were indicating changes on structures of DNA as a result of calf thymus DNA incubation with B[a]P and CuCl2. The highest level of 8-OHdG results in calf thymus DNA incubation with B[a]P and CuCl2 under pH 8.5 and incubation temperature at 60°C, was about 120.856 μg/L. Calf thymus DNA incubation with TiO2-UV radiation (254 nm) under pH 8.5 resulting 8-OHdG level at 57.025 μg/L.

Bahasa Abstract

Untuk melihat profil pembentukan DNA Adduct (8-OHdG) sebagai indikator kerusakan DNA, dilakukan studi in vitro pembentukan DNA Adduct (8-OHdG) dengan calf thymus DNA dan beberapa senyawa yang memiliki toksisitas tinggi sebagai pemicu radikal bebas dan dapat menyebabkan kerusakan DNA secara oksidatif yang memicu peristiwa karsinogenesis, seperti PAH (Benzo[a]piren), TiO2 dan CuCl2. Pada percobaan ini, calf thymus DNA diinkubasi dengan senyawa Benzo[a]piren dan CuCl2 di bawah kondisi variasi pH serta suhu. Dilakukan pula inkubasi calf thymus DNA dan TiO2 dengan bantuan radiasi sinar UV (254 nm) yang dapat menginduksi terbentuknya spesies oksigen reaktif (ROS) sehingga menyebabkan kerusakan oksidatif DNA. Dari hasil penelitian ini, ketiga senyawa tersebut memiliki potensi dalam pembentukan DNA Adduct (8-OHdG). Rasio kemurnian calf thymus DNA pada λ260/ λ280 yang digunakan dalam penelitian ini adalah ~1.9. Terjadi pergeseran puncak panjang gelombang maksimum dari hasil inkubasi calf thymus DNA dan senyawa B[a]P serta CuCl2 yang menandakan bahwa terjadi perubahan struktur DNA. Konsentrasi 8-OHdG tertinggi dihasilkan pada inkubasi calf thymus DNA dengan senyawa B[a]P dan CuCl2 pada pH 8.5 dan suhu 60°C, yaitu 120.856 μg/L. Inkubasi calf thymus DNA dengan TiO2 dan sinar UV (25y4 nm) pada pH 8.5 dapat menghasilkan 8-OHdG sebesar 57.025 μg/L.

References

Askoxylakis, V., Thieke, C., Pleger, Sven T., et.al. (2010). Long-term survival of cancer patients compared to heart failure and stroke: A systematic review. BMC Cancer, 10:105

Beach, A.C., Gupta, R.C. (1992). Human Biomonitoring and the 32P-Postlabeling assay. Carcinogenesis, 13(7), 1053-1074.

Bryla, P., and Weyand, Eric H. (1991). Role of activated oygen species in Benzo[a]Pyrene : DNA –Adduct Formation in vitro. Free Radical Biology & Medicine, 11, 17-24.

Budiawan and Eder, Erwin. (2000). Detection of 1,N2-Propanodeoxyguanosine Adduct in DNA of Fischer 344 Rats by An Adapted 32P-Post-Labeling Technique After Per Os Application of Crotonaldehyde. Carcinogenesis, 21(6), 1191-1196.

E. Eder and Budiawan. (2001). Cancer risk assessment for the environmental mutagen and carcinogen crotonaldehyde on the basis of TD50 and comparison with 1,N2-propanodeoxyguanosine adduct levels, Cancer Epidem. Biom. Prev. 10, 883-888.

Guindon, K. (2008). Investigation of The Role of Oxidative DNA Damage in Aflatoxin B1-Induced Pulmonary Carcinogenesis. Departemen Farmakologi dan Toksikologi: Universitas Queen.

Hanahan, D., and Weinberg, R.A. (2000). The hallmarks of cancer. Cell 100, 57–70.

Huang, Ning-ping., Xu, Min-hua., Yuan, Chun-wei., Yu, Rui-rong. (1997). The study of the photokilling effect and mechanism of ultrafine TiO2 particles on U937 cells. China Journal of Photochemistry and Photobiology A: Chemistry, 108, 229-233.

Gupta. R. C. (1985). Enhanced sensitivity of 12P-postlabeling analysis of aromatic carcinogen: DNA adducts. Cancer Res. 45, 5656-5662.

Halliwell, B., M. Whiteman. 2004. Measuring reactive species and oxidative damage in vivo and in cell culture :how should you di it and what do the results mean?. British Journal of Pharmacology, 142, 231-255.

Hofer, T. 2001. Method Develpoment for Analysis of 8-oxodG as a Biomarker for Oxidative Stress. Departemen Biosains: Institut Karolinska.

Hiraku, Y. dan Kawanishi, S. 1996. Oxidative DNA Damage and Apoptosis Induced by Benzene Metabolites. Cancer Research, 56, 5172-5178.

IARC. 1989. Occupational Exposures in Petroleum Refining; Crude Oil and Major Petroluem Fuels. IARC Monographs on the Evaluation of Carcinogenic Risk of Chemicals to Humans, vol. 45. Lyon, France:International Agency for Research on Cancer. 322 pp.

IARC. 1989. Titanium Dioxide. IARC Summary & Evaluation, Volume 47.

IPCS WHO (International Programme on Chemical Safet – World Health Organization). 1998. EHC 202 : Selected Non- Heterocyclic Polycyclic Aromatic Hydrocarbons, World Health Organization, Geneva.

Inoue, O., et al. 2000. Urinary Phenylmercapturic Acid as a Marker of Occupational Exposure to Benzene. Industrial Health, 38, 195-204.

Kasai, H., Kawai, K. dan Li, Y. 2008. Analysis of 8-OH-dG and 8-OH-Gua as Biomarkers of Oxidative Stress. Genes and Environment. Vol 30, 33-40.

Kasai H. 1997. Analysis of a form of oxidative DNA damage, 8-hydroxy-2deoxyguanosine, as a marker of cellular oxidative stress during carcinogenesis. Mutation Research ;387:147–163.

Khan, S.A., Carmichael, P.L., Robinson, S.D.Taylor., Habib, N., Thomas H.C. 2003. DNA Adducts, detected by 32P postlabelling, in human cholangiocarcinoma. Gut 2003; 52:586-591.

Kim EJ, Choi SD, Chang YS. 2011. Levels and patterns of polycyclic aromatic hydrocarbons (PAHs) in soils after forest fires in South Korea. Environ Sci PollutRes Int. Nov;18(9):1508-17.

Kim, S., dkk. 2006. Using urinary biomarkers to elucidate dose-related patterns of human benzene metabolism. Carcinogenesis, Vol.27, No. 4, 772-781.

Kim, Sun Young., Taguchi, Takao., Nishioka, Motomu., Taya, Masahito. 2004. Quantitative Assessment of DNA Damage Accompanied with No Substantial Loss in Its Molecular Weight During Exposure to Oxidative Stress. Biochemical Engineering Journal 22 : 81–87.

Klauning, J.E., Z. Ang et al. 2011. Oxidative Stress and Oxidative Damage in Chemical Carcinogenesis. Toxicology and Applied Pharmacology., 24: 86-99.

Liu, Z., Lu, Y., R, Barry., L, Mark., Wei, Huachen. 1998. Benzo[a]Pyrene Enhances the Formation of 8-Hydroxy-2’-deoyguanosine by Ultraviolet A Radiation in Calf thymus DNA and Human Epidermoid Carcinoma Cells. Biochemistry, 37, 10307-10312.

Lodovici, Maura and Bigagli, Elisabetta. 2009. Biomarkkers of Induced Active and Passive Smoking Damage. Int. J. Environ. Res. Public Health, 6 874-888.

Lorentzen, Ronald J.,Caspary, William J., Lesko, Stephen A. 1975. Autooxidation of 6-hydroxybenzo[a]pyrene and 6-oxobenzo[a]pyrene radical, reactive metabolites of benzo[a]pyrene. Biochemistry, 14(18), pp 3970-3977.

Minotti, G., Aust, S.D. 1989. The role of iron in oygen radical mediated lipid peroidation. Chem. Biol. 244::6049-6055.

Mizoue, T., Kasai, H., Kubo, T., dan Tokunaga, S. 2006. Leanness, Smoking, and Enhanced Oxidative DNA Damage. Cancer Epidemiol, Biomarkers Prev., 582-585.

Phillips, D.H. and Castegnaro, M. 1999. Standardization and Validation of DNA Adduct Postlabelling Methods : Report of Interlaboratory Trials and Production of Recommended Protocols. Mutagenesis, vol.14 no.3 pp.301-315.

Ramkumae, Kunga Mohan., Manjula, Chinnasamy., Kumar, Georgepeter Gnana., Kanjwal, Muzafar A. 2012. Oxidative stress-mediated cytotoxicity and apoptosis induction by TiO2 nanofibers in HeLa cells. European Journal of Pharmaceutics and Biopharmaceutics 81 (2012) 324–333.

Teare, J.M., R., Islam., Flanagan, R., Gallagher, S., Davies, M.G., and Grabau, C. 1997. Measurement of Nucleic Acid Concentrations Using the DyNa Quant TM and the GeneQuantTM. Biotechniques 22:1170-1174.

Tharnpoophasiam, P., Kongtip, P., Wongwit, W., Fungladda, W., dan Kitayaporn, D. 2004. Simultaneous Determination of Trans, Trans-Muconic Acid and S-Phenylmercapturic Acid by High Pressure Liquid Chromatography and Its Application. South East Asian J. Trop. Med. Public Health, Vol. 35, No.3.

Valavanidis, A., Vlachogianni, T., dan Fiotakis, C. 2009. 8-hydroxy-2_-deoxyguanosine (8-OHdG): A Critical Biomarker of Oxidative Stress and Carcinogenesis. Journal of Environmental Science and Health Part C, 27:120–139.

Valko, Marian., Leibfritz, Dieter., Moncol, Jan., Cronin, Mark.T.D., Mazur, Milan., Telser, Joshua. 2007. Free radicals and antioidants in normal physiological functions and human disease. The International Journal of Biochemistry & Cell Biology 39 44-84.

Vergnoux A, Malleret L, Asia L, Doumenq P, Theraulaz F. 2011. Impact of forest fires on PAH level and distribution in soils. Environ Res, Feb 111(2):193-8.

Weisburger J.H., and G.M.Williams. 1984. Bioassay of carcinogens : in vitro and in vivo test, in : Chemical Carcinogens, 2nd edition, Vol.2, C.E Searle (ed), ACS Monograph 182, 1323-1373, American Chemical Society, Washington D.C.

World Health Organization. 2014. News Realeses : Seven million premature deaths annually linked to air pollution. WHO, Geneva.

Zhang,Li, et al. 2013. Interception of Benzo (a) pyrene- 7,8-dione by UDP Glucuronosyltransferase (UGTs) in Human Lung Cells.

Zhang, Zhihua and Miriam C. Poirier. 1997. Cisplatin-DNA Adduct Determination in the Hepatic Albumin Gene as Compared to Whole Genomic DNA. 10,971-977

Zhang, Yu Jing, et al. 1999. Development of Monoclonal Antibody Rcogniing Beno(c)phenanthrenediol Epoxide-DNA Adducts: Application to Immunohistochemical Detection of DNA Damage. 10,948-952

Download

Share

COinS
 
 

To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.