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Aijalon Shantavia Bettis
Monitoring DNA Interactions with Small Ring Polycyclic Aromatic Hydrocarbons, their Alkylated Analogs Aijalon Shantavia Bettis
Mentor: Harshica Fernando Department of Chemistry
Introduction: Polycyclic aromatic hydrocarbons (PAHs) are a group of more than 100 chemicals, also called polynuclear aromatic hydrocarbons. PAHs are environmental pollutants and can easily be added to food, water, and air. They are commonly found in coal, hydrocarbon fuel, and crude oil. When oil, gas, wood, and various other items are burned, PAHs can be produced and become airborne. In addition, PAHs can be found in cigarette smoke as well as meat cook at high temperatures. You can be exposed to PAHs by 1) ingesting or swallowing food that contains them, 2) inhaling or breathing fumes or smoke, or 3) touching food that contains PAHs. Exposure to some PAHs can cause less serious issues such as irritation of the eyes and or breathing passages as well as more serious issues, such as cancer. Environmental Protection Agency has identified sixteen hazardous parents PAHs based on their potential to human and ecological health effects, and many studies are carried out with some of these parent PAHs. However, new evidence suggests that alkylated PAHs are more potent and toxic than the parent PAHs. In oil, there exist a high amount of alkylated analogs of the PAHs than the parent hydrocarbons. In the past, studies have focused on higher ring PAH's (5-6) as they were found to be carcinogenic. Recently more attention is given to small ring PAH's as they are found to be toxic and present in large amounts. Monitoring interactions of the small ring PAHs and their alkylated analogs with DNA is important, as human health effects from exposure to lower ring PAHs are not clearly defined. In this work, DNA and PAH interactions are monitored using a UV-Visible and a fluorescence approach. Materials and Methods: Calf thymus-DNA was purchased from Worthington Biomedical Corp, NJ. 9, 10 bis-phenylethynylanthracene was obtained from Sigma Chemical Company. All other chemicals and solvents were of analytical grade. DNA solution was prepared in Tris-HCl buffer at pH = 7.4. The purity of the DNA was measured by observing the absorbance ratio at 260 and 280. A ratio of 1.80 confirmed purity of the DNA. The stock solution of the PAH's was prepared in ethanol. The interaction of DNA with the fluorophore 9, 10 bisphenylethnylanthracene was monitored using both spectroscopic methods. The DNA concentration were calculated based on !!"# =6600&$% '( $% . A Shimatzu UV-Vis spectrometer was used to scan the spectra from 200 to 600 nm and the interactions observed with the fluorescence aspect was monitored using images being taken with an image app, and ImageJ, a program used for analyzing fluorescence intensity, in turn analyzed these images.
Results and Discussion:
In the images, DNA interaction is being compared with the blank experiment with the well containing the added DNA on the bottom left at 4 seconds of exposure. Both wells with added aliquots show an interaction that is represented with the darker green color at the rim of the wells.
Conclusion: Our results show that 9, 10 bisphenylethnylanthracene exhibit interaction with DNA. The nature of the interaction still needs to be identified. Future work will include kinetics, continued use of
ImageJ, and compare the intensity using a fluorimeter, and have more alkylated PAH's to compare the binding kinetics and the nature of binding.
References
1. Polycyclic Aromatic Hydrocarbons (PAHs): Your Environment, Your Health | National Library of Medicine. (n.d.). Retrieved from https://toxtown.nlm.nih.gov/chemicals-andcontaminants/polycyclic-aromatic-hydrocarbons-pahs#what-happens-when-i-am-exposed-topolycyclic-aromatic-hydrocarbons-(pahs) 2. Polycyclic Aromatic Hydrocarbons (PAHs). (n.d.). Retrieved from http://www.epa.gov/ 3. Shi, J.-H., Lou, Y.-Y., Zhou, K.-L., & Pan, D.-Q. Exploration of intermolecular interaction of calf thymus DNA with sulfosulfuron using multi-spectroscopic and molecular docking techniques. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 204, 209–216, 2018. doi: 10.1016/j.saa.2018.06.054 4. Fernando, H., Ju, H., Kakumanu, R., Bhopale, K.K., Croisant, S., Elferink, C., Kaphalia, B.S., and Ansari, G.A.S. Distribution of petrogenic polycyclic aromatic hydrocarbons (PAHs) in seafood following Deepwater Horizon oil spill. Mar. Pollut. Bull. 145, 200-207, 2019. 5. Fernando, H., Halpert, J.R. and Davydov, D.R., Two substrate binding sites for pyrene derivatives demonstrated by FRET and absorbance spectroscopy. Biochemistry 45:4199-4209, 2006. 6. Fernando, H., Huang, C.R., Shu, L., Milliman, A. and LeBreton, P.R., Influence of Na+ on DNA reactions with aromatic epoxides and diol epoxides: Evidence that DNA catalyzes the formation of benzo[a]pyrene and benz[a]anthracene adducts at intercalation sites, J Chem Res In Toxicol 9:1391-1402, 1996.
