Eskişehir Teknik Üniversitesi Bilim ve Teknoloji Dergisi - C Yaşam Bilimleri Ve Biyoteknoloji 2667-4203 Eskisehir Technical University 10.18036/estubtdc.1609497 Structural Biology Yapısal Biyoloji FENTANYL GENOTOXICITY EVALUATION VIA COMET ASSAY FENTANYL GENOTOXICITY EVALUATION VIA COMET ASSAY https://orcid.org/0000-0001-9389-4450 Başoğlan Artagan Öge MUĞLA SITKI KOÇMAN ÜNİVERSİTESİ, MUĞLA SAĞLIK HİZMETLERİ MESLEK YÜKSEKOKULU https://orcid.org/0009-0007-0316-6769 Köklü Bahar ESKİŞEHİR TEKNİK ÜNİVERSİTESİ 07 25 2025 14 2 59 65 12 29 2024 02 15 2025 Copyright © 2010, Eskişehir Technical University Journal of Science and Technology C - Life Sciences and Biotechnology 2010 Eskişehir Technical University Journal of Science and Technology C - Life Sciences and Biotechnology

DNA is our main genetic structure that can be affected by various molecules we come into contact with. Therefore, there is a high probability of DNA damage. Genotoxicity tests involve systematic methods designed to assess the safety of drugs and chemicals. Comet assay evaluates the possible genotoxicity resulting from DNA single-strand breaks.Fentanyl is a powerful painkiller from the group called opioid analgesics administered intravenously. In this research, we evaluated the possible DNA damage in human lymphocytes after fentanyl exposure via single-cell gel electrophoresis (Comet Assay). DNA damage in healthy human peripheral lymphocytes treated with fentanyl was investigated via comet assay. Lymphocytes were treated with 5, 10, 20, and 40 µg/mL doses of fentanyl for 1 hour. After the incubation period, the cells' DNA tail length, tail intensity, and tail moment values were evaluated by comparing them with the spontaneous control and positive control data. EtBr stained slides were visualized under the fluorescent microscope. Fentanyl induced the comet parameters such as tail length, tail intensity, and tail moment but dose-dependent increase was not obtained after fentanyl administration. Fentanyl showed the highest tail length and tail moment value at the dose of 10 µg/mL. The highest tail intensity value was obtained at the dose of 40 µg/mL fentanyl administration.This study aimed to reveal previously undiscovered genotoxicity of fentanyl on healthy human lymphocytes in vitro via comet assay. Fentanyl exposure induces DNA damage in healthy human lymphocytes, as shown via comet parameters (tail length, tail intensity, and tail moment). Genotoxic effect does not display a consistent dose-dependent increase.

DNA is our main genetic structure that can be affected by various molecules we come into contact with. Therefore, there is a high probability of DNA damage. Genotoxicity tests involve systematic methods designed to assess the safety of drugs and chemicals. Comet assay evaluates the possible genotoxicity resulting from DNA single-strand breaks.Fentanyl is a powerful painkiller from the group called opioid analgesics administered intravenously. In this research, we evaluated the possible DNA damage in human lymphocytes after fentanyl exposure via single-cell gel electrophoresis (Comet Assay). DNA damage in healthy human peripheral lymphocytes treated with fentanyl was investigated via comet assay. Lymphocytes were treated with 5, 10, 20, and 40 µg/mL doses of fentanyl for 1 hour. After the incubation period, the cells' DNA tail length, tail intensity, and tail moment values were evaluated by comparing them with the spontaneous control and positive control data. EtBr stained slides were visualized under the fluorescent microscope. Fentanyl induced the comet parameters such as tail length, tail intensity, and tail moment but dose-dependent increase was not obtained after fentanyl administration. Fentanyl showed the highest tail length and tail moment value at the dose of 10 µg/mL. The highest tail intensity value was obtained at the dose of 40 µg/mL fentanyl administration.This study aimed to reveal previously undiscovered genotoxicity of fentanyl on healthy human lymphocytes in vitro via comet assay. Fentanyl exposure induces DNA damage in healthy human lymphocytes, as shown via comet parameters (tail length, tail intensity, and tail moment). Genotoxic effect does not display a consistent dose-dependent increase.

 Comet assay Single-cell gel electrophoresis DNA damage Lymphocyte culture Genotoxicity Fentanyl Komet Testi Tek Hücre Jel Elektroforezi DNA Hasarı Lenfosit Kültürü Genotoksisite Fentanil [1] Waskell L. A study of the mutagenicity of anesthetics and their metabolites. Mutation Research 1978; 57(2): 141–153. [2] Kant I, van Rijssen-Moll M, Borm PJ. Simulation of nitrous oxide concentrations in operating and recovery rooms. Annals of Occupational Hygiene 1990; 34(6): 575–583. [3] Stanley TH, Philbin DM, Coggins CH. Fentanyl-oxygen anaesthesia for coronary artery surgery: cardiovascular and antidiuretic hormone responses. Canadian Anaesthetists' Society Journal 1979; 26(3): 168–172. [4] Stanley TH. The fentanyl story. The Journal of Pain 2014; 15(12): 1215–1226. [5] Gasperini S, et al. The genotoxicity of acrylfentanyl, ocfentanyl and furanylfentanyl raises the concern of long-term consequences. International Journal of Molecular Sciences 2022; 23(22): 14406. [6] Collins AR. The comet assay for DNA damage and repair. Mol Biotechnol. 2004;26(3):249-261. [7] Singh NP, McCoy MT, Tice RR, Schneider EL. A simple technique for quantitation of low levels of DNA damage in individual cells. Experimental Cell Research 1988; 175(1): 184–191. [8] Olive PL, Banáth JP, Durand RE. Heterogeneity in radiation-induced DNA damage and repair in tumor and normal cells measured using the "comet" assay. Radiation Research 1990; 122(1): 86–94. [9] Yeager MP, et al. Intravenous fentanyl increases natural killer cell cytotoxicity and circulating CD16(+) lymphocytes in humans. Anesthesia and Analgesia 2002; 94(1): 113–119. [10] Sardaş S, et al. DNA damage evaluated by the alkaline comet assay in lymphocytes of humans anaesthetized with isoflurane. Mutation Research 1998; 418(1): 1–6. [11] Brambilla G, Martelli A. Update on genotoxicity and carcinogenicity testing of 472 marketed pharmaceuticals. Mutation Research 2009; 681(2-3): 209–229. [12] Kadioglu E, et al. Determination of DNA damage by alkaline halo and comet assay in patients under sevoflurane anesthesia. Toxicology and Industrial Health 2009; 25(3): 205–212. [13] Kılıc M, Tuylu BA. An in vitro investigation of genotoxic effects of dexketoprofen trometamol on healthy human lymphocytes. Drug and Chemical Toxicology 2020; 43(2): 174–181. [14] OECD. Test No. 489: In vivo mammalian alkaline comet assay. OECD Guidelines for the Testing of Chemicals 2016; Section 4. [15] Kassie F, Parzefall W, Knasmüller S. Single cell gel electrophoresis assay: a new technique for human biomonitoring studies. Mutation Research 2000; 463(1): 13–31. [16] Albertini RJ, et al. IPCS guidelines for the monitoring of genotoxic effects of carcinogens in humans. Mutation Research 2000; 463(2): 111–172. [17] Santovito A, Cervella P, Delpero M. Increased frequency of chromosomal aberrations and sister chromatid exchanges in peripheral lymphocytes of radiology technicians chronically exposed to low levels of ionizing radiations. Environmental Toxicology and Pharmacology 2014; 37(1): 396–403. [18] Avuloglu Yilmaz Y, et al. Evaluation of cytogenetic and DNA damage induced by the antidepressant drug-active ingredients, trazodone and milnacipran, in vitro. Drug and Chemical Toxicology 2017; 40(1): 57–66. [19] Güzel Bayülken D, et al. Assessment of in vitro cytotoxic and genotoxic activities of some trimethoprim conjugates. Cytotechnology 2018; 70(3): 1051–1059. [20] Güzel Bayülken D, Ayaz Tüylü B, Sinan H, Sivas H. Investigation of genotoxic effects of paraben in cultured human lymphocytes. Drug and Chemical Toxicology 2019; 42(4): 349–356.