Investigation of the effect of Aflatoxin B1 and Aflatoxin G1 on DNA Hybridization by Using Electrochemical DNA Biosensor

Yıl 2019, Cilt: 40 Sayı: 2, 332 - 339, 30.06.2019

Gültekin Gökçe Songül Gökçe Şenay Çetinus

https://doi.org/10.17776/csj.544897

Öz

Biomolecular
detection has become a current issue together with the increase in
electrochemical sensor studies. The effect of Aflatoxin B1 and Aflatoxin G1 on
DNA hybridization was analyzed with electrochemically by using pencil graphite
electrode. For this purposes; the changes in guanine currents were monitored
before and after hybridization occurred between DNA complementary base
sequences using differential pulse voltammetry (DPV). In the first step of this
work the capture probe was immobilized by wet adsorption onto the surface of
PGE optimizing concentration and immobilization time and then hybridization
event was performed between capture and its target probe. In the second step
the hybridization experiments were repeated using inosine modified capture
probe instead of capture probe which used in the first step. As differently
from the first step the inosine modified probe was interacted with 2 µg/mL and
4 µg/mL of AFB1 for 10 min before allowing hybridization and the same protocol
was repeated for AFG1. The decrease in guanine current after hybridization was
the reference for the genotoxic effects of aflatoxins. The decrease in guanine
oxidations in the cases of 2 µg/mL and 4 µg/mL AFG1 and 2 µg/mL and 4 µg/mL
AFB1 concentrations were found to be 18% and 26 % for AFG1; and 50% and 61% for
AFB1, respectively. AFB1 showed the maximum genotoxic effect to DNA
hybridization.

Anahtar Kelimeler

Aflatoxin B1, Aflatoxin G1, Pencil Graphite Electrode, Hybridization, Differential Pulse Voltammetry

Aflatoksin B1 ve Aflatoksin G1'in DNA Hibridizasyonuna Etkisinin Elektrokimyasal DNA Biyosensörü ile İncelenmesi

Öz

Biyomoleküler analizlerde yeni teknolojilerin gelişmesi, elektrokimyasal
sensör alanındaki çalışmaların da katılmasıyla güncel bir konu olmuştur. Bu
çalışmada Aflatoksin B1'in (AFB1) ve Aflatoksin G1'in (AFG1) DNA hibridizasyonu
üzerindeki etkisi, kalem grafit elektrot (PGE) kullanılarak elektrokimyasal yöntemlerle
analiz edildi. Bu amaçla; guanin oksidasyon akımlarındaki değişiklikler,
diferansiyel puls voltametrisi (DPV) ile birbirlerini eşleniği olan DNA baz
sekansları arasında hibridizasyon oluşmadan önce ve sonra izlendi. Çalışmanın
ilk aşamasında, yakalama probu ıslak adsorpsiyonla PGE yüzeyine immobilize
edilirken prob derişimi ve adsorpsiyon süresi optimize edildi ve sonra, hedef
prop ile hibritleşmeye tabi tutulurken hedef proba ait konsantrasyon ve
hibridizasyon süresi optimize edildi. İkinci aşamada, hibridizasyon deneyleri,
birinci aşamada kullanılan yakalama probu yerine inosin modifiye edilmiş
yakalama probu kullanılarak tekrarlandı. Ancak, ilk aşamadan farklı bir şekilde
inosin modifiye edilmiş prob, hibridizasyona tabi tutulmadan önce 10 dakika süre
ile 2 µg / mL ve 4 µg / mL AFB1 ile etkileştirildi ve aynı protokol, aynı
koşullarda AFG1 için de tekrarlandı. Hibridizasyondan sonra guanin oksidasyon
akımındaki azalma, aflatoksinlerin DNA sekanslarının hibridizasyonuna etkisinin
bir sonucu olup, AFB1 ve AFGl'in DNA üzerine genotoksik etkisinin bir
göstergesidir. Guanin oksidasyon sinyalindeki azalmalar 2 µg/mL ve 4 µg/mL AFG1
için % 18 ve % 26; 2 µg/mL ve 4 µg/mL AFB1 için % 50 ve % 61 olarak bulundu.
Sonuç olarak AFB1'in DNA üzerindeki genotoksik etkisi AFG1'e göre çok daha
fazla olarak tespit edildi.

Anahtar Kelimeler

Aflatoksin B1, Aflatoksin G1, Kalem Grafit Elektrot, Hibridizasyon, Diferansiyel Puls Voltametrisi

Kaynakça

• [1] Kerman, K., Meric, B., Ozkan, D., Kara., P., Erdem, A. and Ozsoz, M., Electrochemical DNA biosensor for the determination of Benzo[a]pyrene–DNA adducts, Analytica Chimica Acta, 450 (2001) 45-52.
• [2] Ozsoz M., Erdem A., Kerman K., Ozkan D., Tugrul B., Topcuoglu N., Ekren H. and Taylan M., Electrochemical genosensor based on colloidal gold nanoparticles for the detection of factor V Leiden mutation using disposable pencil graphite electrodes, Anal Chem., 75 (2003) 2181-2187.
• [3] Palecek E., Fojta, M., Tomschik M., and Wang J., Electrochemical biosensors for DNA damage, Biosensors and Bioelectronics, 13 (1998) 621-628.
• [4] Del Giallo M.L., Ariksoysal D.O., Marrazza G., Mascini M. and Ozsoz M., Disposable Electrochemical Enzyme-Amplified Genosensor for Salmonella Bacteria Detection, Analytical Letters, 38 (2005) 2509–2523.
• [5] Lin Y., Lu F. and Wang J., Disposable Carbon Nanotube Modified Screen-Printed Biosensor for Amperometric Detection of Organophosphorus Pesticides and Nerve Agents, Electroanalysis, 16(1-2) (2004) 145-149.
• [6] Sing Virendra V., Recent Advances in Electrochemical Sensors for Detecting Weapons of Mass Destruction. A Review, ELECTROANALYSIS, 28 (5) (2016) 920-935.
• [7] Chen, J. H., Zhang, J., Huang, L. Y., Lin, X. H. and Chen, G. N., Hybridization biosensor using 2-nitroacridone as electrochemical indicator for detection of short DNA species of Chronic Myelogenous Leukemia, Biosensors & Bioelectronics, 24 (2008) 349-355.
• [8] Marrazza, G., Chianella, I. and Mascini, M., Disposable DNA electrochemical sensor for hybridization detection, Biosensors & Bioelectronics, 14 (1999) 43-51.
• [9] Brabec V. and Koudelka J., Oxidation of DNA at carbon electrodes the effect of the quality of the DNA sample, J. Electroanal. Chem., 116 (1980) 793-799.
• [10] Erdem A, Meriç B., Kerman K., Dalbastı T. and Özsöz M., Detection of interaction between metal complex indicator and DNA by using electrochemical biosensor, Electroanal., 11 (1999) 1372-1377.
• [11] Jelen, F., Erdem A. and Palecek, E., Cyclic voltammetry of echinomycin and its interaction with double-stranded and single-stranded DNA adsorbed at the electrode, Bioelectrochemistry,55 (2002) 165-173.
• [12] Özkan D., Karadeniz H., Erdem A., Mascini M. and Özsöz M., Electrochemical genosensor for Mitomycin C- DNA interaction based on guanine signal, J. Pharmaceutical and Biomedical Analysis, 35 (2004) 905-912.
• [13] Wang A.H.J., Interactions between anti tumor drugs and DNA, Nucleic Acids and Molecular Biology, 1 (1987) 52-58.
• [14] Pohland, A.E., Mycotoxins in review, Food Additives Contaminants, 10, (1993) 17-28.
• [15] Gündüz, S Kimyacılar için istatistik, Gazi Kitabevi, Ankara (1998) 89.
• [16] Hazır, Z. and Çoksöyler, N., Farklı bölgelerde ve farklı yöntemlerle elde edilen kırmızı biberlerde aflatoksin düzeyleri, Gıda Mühendisliği Kongre ve Sergisi (1998) 479-483.
• 17] Trucksess, M.W. and Scott, P.M., Mycotoxins in botanicals and dried fruits: a review. Food Additives and Contaminants. 25 (2008) 181–192.
• [18] Williams, J.H., Phillips, T.D., Jolly, P.E., Stiles, J.K., Jolly, C.M., Aggerwal, D. Human aflatoxicosis in developing countries: a review of toxicology, exposure, potential health consequences, and interventions, The American Journal Clinical Nutrition. 80 (2004) 1106-1122.
• [19] Fernández-Cruz, M.L., Mansilla, M.L. and Tadeo, J.L., Mycotoxins in fruits and their processed products: Analysis, occurrence and health implications, Journal of Advanced Research, 1 (2010) 113–122.
• [20] Zinedine, A., González-Osnaya, L., Soriano, J.M., Moltó, J.C., Idrissi and L., Mañes, J. Presence of aflatoxin M1 in pasteurized milk from Morocco. International journal of food microbiology, 114 (2007) 25–29.
• [21] Wang, J., Kawde, A. N. , Erdem A. and Salazar, M., Magnetic bead-based label-free electrochemical detection of DNA hybridization, Analyst, 126 (2001) 2020-2025.
• [22] Kuralay, F., Erdem, A., Abacı, S. Ozyoruk, H. and Yıldız, A., Poly (vinylferrocenium) coated disposable pencil graphite electrode for DNA hybridization, Electrochem. Commun., 11(6) (2009) 1242-1246.
• [23] Smrdel K.S., Seret M., Duh D., Knap N. and Avsic-Zupanc T., Anaplasma phagocytophilum in ticks in Slovenia, Parasitesand Vectors, 3 (2010) 210-215.
• [24] Wang J., Cai X., Rivas G. and Shiraishi H., Stripping potentiometric transduction of DNA hybridization processes, Anal. Chim. Act., 326 (1996) 141-145.
• [25] Wang, J.S. and Groopman, J.D., DNA damage by mycotoxins, Mutation Research, 424 (1999) 167–181.