MALAHİT YEŞİLİNİN GÖKKUŞAĞI ALABALIĞINDA SİTOKROM P450 VE ANTİOKSİDAN ENZİM AKTİVİTELERİ ÜZERİNE İN VİVO MODÜLATÖR ETKİSİ

Öz

Malahit yeşili, tekstil ve kağıt endüstrisinde boya olarak ve balık çiftliklerinde ektoparazitlere, bakteriyel ve fungal enfeksiyonlara karşı kullanılan mutajenik ve karsinojenik bir kimyasaldır. Sucul organizmalar malahit yeşiline maruz kalma riski altındadır. Bu çalışmada amacımız gökkuşağı alabalığında (Oncorhynchus mykiss) iki farklı konsantrasyonda malahit yeşilinin sitokrom P450 ve antioksidan sistem üzerine etkisini belirlemektir. Bu amaçla, 24 balık üç gruba rastgele ayrılmıştır: kontrol, 0,1 mg/L malahit yeşili ile muamele edilen grup ve 0,5 mg/L malahit yeşili ile muamele edilen grup. Oldukça yüksek 7-etoksiresorufin O-deetilaz (EROD) aktivitesi balıklarda 0,5 mg/L malahit yeşili ile muamele edilen grupta ölçülmüştür. 7-pentoksiresorufin O-depentilaz (PROD) aktivitesi artan malahit yeşili dozu ile artmıştır. Katalaz (CAT) ve glutatyon redüktaz aktiviteleri 0,1 mg/L malahit yeşili ile muamele edilen grupta kontrol grubundan yüksektir. Eritromisin N-demetilaz (ERND) ve glutatyon S-transferaz (GST) aktiviteleri her iki muamele grubunda malahit yeşilinden önemli ölçüde etkilenmemiştir. Bu çalışmanın sonuçları malahit yeşilinin gökkuşağı alabalığında EROD, PROD, CAT ve GR aktiviteleri üzerinde modülatör bir etkiye sahip olduğunu açıkça göstermektedir.

Anahtar Kelimeler

• Antioksidan enzim aktiviteleri
• sitokrom P450
• gökkuşağı alabalığı (Oncorhynchus mykiss)
• ksenobiyotik metabolize eden enzimler
• malahit yeşili

In VIVO MODULATORY EFFECT of MALACHITE GREEN on CYTOCHROME P450 AND ANTIOXIDANT ENZYME ACTIVITIES in RAINBOW TROUT LIVER

Öz

Malachite green is a mutagenic and carcinogenic chemical used as a dye in textile and paper industry and in fish farms against ectoparasites, bacterial and fungal infections. Aquatic organisms are at risk of malachite green exposure. In this study, our aim was to determine the effect of two different doses of malachite green on cytochrome P450 system and antioxidant system in rainbow trout (Oncorhynchus mykiss). For this purpose, 24 fish were randomly divided into three groups: the control, the 0.1 mg/L malachite green treatment group and the 0.5 mg/L malachite green treatment group. Highly elevated 7-ethoxyresorufin-O-deethylase activities (EROD) were measured in fish treated with 0.5 mg/L malachite green. 7-pentoxyresorufin O-depentylase (PROD) activities increased with increasing doses of malachite green. Catalase (CAT) and glutathione reductase (GR) activities in the 0.1 mg/L malachite green administration group were higher than those in the control group. Erythromycin N-demethylase (ERND) and glutathione S-transferase (GST) activities were not significantly affected by malachite green treatment in either treatment group. The results of this study clearly demonstrate that malachite green has a modulatory effect on EROD, PROD, CAT, and GR activities in rainbow trout.

Anahtar Kelimeler

• Antioxidant enzyme activities
• Cytochrome P450
• Malachite green
• Rainbow trout (Oncorhynchus mykiss)
• Xenobiotic metabolizing enzymes

Kaynakça

1. Aebi, H. (1984). Catalase in vitro. Methods in Enzymology, 105, 121-136.
2. Arinç, E., & Şen, A. (1993). Characterization of cytochrome P450 dependent mixed-function oxidase system of gilthead seabream (Sparus aurata; Sparidae) liver. Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 104(1), 133-139.
3. Bozcaarmutlu, A., & Arinç, E. (2008). Purification of CYP2B-like protein from feral leaping mullet (Liza saliens) liver microsomes and its biocatalytic, molecular, and immunological characterization. Journal of Biochemical and Molecular Toxicology, 22(4), 284-298.
4. Bozcaarmutlu, A., Sapmaz, C., Kaleli, G., Turna, S., & Yenisoy-Karakaş, S. (2015). Combined use of PAH levels and EROD activities in the determination of PAH pollution in flathead mullet (Mugil cephalus) caught from the West Black Sea coast of Turkey. Environmental Science and Pollution Research, 22(4), 2515-2525.
5. Bozcaarmutlu, A., Sapmaz, C., Kaleli-Can, G., Turna, S., Aygun, Z., & Arinç, E. (2020). Monitoring of pollution in the western Black Sea coast of Turkey by striped red mullet (Mullus surmuletus). Environmental Monitoring and Assessment, 192(9), 586.
6. Burke, M. D., & Mayer, R. T. (1974). Ethoxyresorufin: direct fluorimetric assay of a microsomal O-dealkylation which is preferentially inducible by 3-methylcholanthrene. Drug Metabolism and Disposition, 2(6), 583-588. Carlberg, I., & Mannervik, B. (1985). Glutathione reductase. Methods in Enzymology, 113, 484-490.
7. Cochin, J., & Axelrod, J. (1959). Biochemical and pharmacological changes in the rat following chronic administration of morphine, nalorphine, and normorphine. Journal of Pharmacology and Experimental Therapeutics, 125(2), 105-110.
8. Culp, S. J., Blankenship, L. R., Kusewitt, D. F., Doerge, D. R., Mulligan, L. T., & Beland, F. A. (1999). Toxicity and metabolism of malachite green and leucomalachite green during short-term feeding to Fischer 344 rats and B6C3F1 mice. Chemico-Biological Interactions, 122(3), 153-170.

9. Das, J. K., Sarkar, S., Hossain, S. U., Chakraborty, P., Das, R. K., & Bhattacharya, S. (2013). Diphenylmethyl selenocyanate attenuates malachite green-induced oxidative injury through antioxidation & inhibition of DNA damage in mice. Indian Journal of Medical Research, 137(6), 1163-1173.
10. Doerge, D. R., Churchwell, M. I., Gehring, T. A., Pu, Y. M., & Plakas, S. M. (1998). Analysis of malachite green and metabolites in fish using liquid chromatography atmospheric pressure chemical ionization mass spectrometry. Rapid Communications in Mass Spectrometry, 12(21), 1625-1634.
11. Dutta, S., Adhikary, S., Bhattacharya, S., Roy, D., Chatterjee, S., Chakraborty, A., Banerjee, D., Ganguly, A., Nanda, S., & Rajak, P. (2024). Contamination of textile dyes in aquatic environment: Adverse impacts on aquatic ecosystem and human health, and its management using bioremediation. Journal of Environmental Management, 353, 120103.
12. Esteves, F., Rueff, J., & Kranendonk, M. (2021). The central role of cytochrome P450 in xenobiotic metabolism—A brief review on a fascinating enzyme family. Journal of Xenobiotics, 11(3), 94-114.
13. Fakhri, Y., Mahmoudizeh, A., Hemmati, F., Adiban, M., Esfandiari, Z., & Khaneghah, A. M. (2025). The concentration of malachite green in fish: a systematic review, meta-analysis, and probabilistic risk assessment. International Journal of Environmental Health Research, 2025 (Early Access).
14. Gharavi-Nakhjavani, M. S., Niazi, A., Hosseini, H., Aminzare, M., Dizaji, R., Tajdar-Oranj, B., & Alizadeh, A. M. (2023). Malachite green and leucomalachite green in fish: a global systematic review and meta-analysis. Environmental Science and Pollution Research, 30(17), 48911-48927.
15. Goksøyr, A. (1995). Use of cytochrome P450 1A (CYP1A) in fish as a biomarker of aquatic pollution. Archives of Toxicology, Supplement, 17, 80-95.
16. Hakami, A. A. H., Ahmed, M. A., Khan, M. A., AlOthman, Z. A., Rafatullah, M., Islam, M. A., & Siddiqui, M. R. (2021). Quantitative analysis of malachite green in environmental samples using liquid chromatography-mass spectrometry. Water, 13(20), 2864.
17. Hassan, N. A. M., Elgendy, F. S., & Mansy, A. E. (2014). Can malachite green concentrations in edible fish cause genetic and hepatic toxic effects? An experimental study on adult albino rats. Ain Shams Journal of Forensic Medicine and Clinical Toxicology, 22, 37-49.
18. Habig, W.H., Pabst, M.J., Jakoby, W.B. (1974). Glutathione S-transferases. The first enzymatic step in mercapturic acid formation. Journal of Biological Chemistry, 249(22), 7130-7139.
19. Lemos, E. S., Fiorentini, E. F., Bonilla-Petriciolet, A., & Escudero, L. B. (2023). Malachite green removal by grape stalks biosorption from natural waters and effluents. Adsorption Science & Technology, 2023, 6695937.
20. Li, G., Shen, D., Liang, X. F., He, Y., & He, S. (2013). Effects of malachite green on the mRNA expression of detoxification-related genes in Nile tilapia (Oreochromis niloticus) and other major Chinese freshwater fishes. Environmental Toxicology, 28(3), 137-145.
21. Lowry, O. H., Rosebrough, N. J., Farr, A. L., & Randall, R. J. (1951). Protein measurement with the Folin phenol reagent. Journal of Biological Chemistry, 193(1), 265-275.
22. Lubet, R. A., Mayer, R. T., Cameron, J. W., Nims, R. W., Burke, M. D., Wolff, T., & Guengerich, F. P. (1985).
23. Dealkylation of pentoxyresorufin: A rapid and sensitive assay for measuring induction of cytochrome(s) P-450 by phenobarbital and other xenobiotics in the rat. Archives of Biochemistry and Biophysics, 238(1), 43-48.
24. Khan, M. R., Wabaidur, S. M., Busquets, R., Khan, M. A., Siddiqui, M. R., & Azam, M. (2019). Identification of malachite green in industrial wastewater using lignocellulose biomass composite bio-sorbent and UPLC-MS/MS: A green environmental approach. Process Safety and Environmental Protection, 126, 160-166.
25. Kovacic, P., & Somanathan, R. (2014). Toxicity of imine-iminium dyes and pigments: Electron transfer, radicals, oxidative stress, and other physiological effects. Journal of Applied Toxicology, 34(8), 825-834.
26. Nash, T. (1953). The colorimetric estimation of formaldehyde by means of the Hantzsch reaction. Biochemical Journal, 55(3), 416-421.
27. Nemoto, N., & Sakurai, J. (1995). Glucocorticoid and sex hormones as activating or modulating factors for expression of Cyp2b-9 and Cyp2b-10 in the mouse liver and hepatocytes. Archives of Biochemistry and Biophysics, 319(1), 286-292.
28. Nebbia, C., Girolami, F., Carletti, M., Gasco, L., Zoccarato, I., & Giuliano Albo, A. (2017). In vitro interactions of malachite green and leucomalachite green with hepatic drug-metabolizing enzyme systems in the rainbow trout (Oncorhynchus mykiss). Toxicology Letters, 280, 41-47.
29. Panandiker, A., Fernandes, C., & Rao, K. V. (1992). The cytotoxic properties of malachite green are associated with the increased demethylase, aryl hydrocarbon hydroxylase, and lipid peroxidation in primary cultures of Syrian hamster embryo cells. Cancer Letters, 67, 93-101.
30. Rabuffetti, G., Brola, J., Pérez, D., Somoza, G. M., & Carriquiriborde, P. (2024). EROD activity in the native fish Cnesterodon decemmaculatus as a biomarker for assessing aquatic pollution by AhR agonist chemicals within the Rio de la Plata Basin. Science of the Total Environment, 947, 174721.
31. Sadauskas-Henrique, H., Braz-Mota, S., Campos, D. F., Barroso, H. D., Kochhann, D., Val, A. L., & de Almeida-Val, V. M. F. (2024). Oil spill in an Amazon blackwater environment: Biochemical and physiological responses of local fish species. Regional Studies in Marine Science, 61, 102865.
32. Sinha, R., Jindal, R., & Faggio, C. (2021). Protective effect of Emblica officinalis in Cyprinus carpio against hepatotoxicity induced by malachite green: Ultrastructural and molecular analysis. Applied Sciences, 11(8), 3507. Srivastav, A. (2016). Malachite green (Triarylmethane dye) and Pyceze (Bronopol) induced histopathological and biochemical changes in the liver of a catfish Heteropneustes fossilis (Bloch.). Indian Journal of Fisheries, 63(1), 135-139.
33. Tew, K. D., & Ronai, Z. (1999). GST function in drug and stress response. Drug Resistance Updates, 2(3), 143-147. Xu, C., Li, C. Y., & Kong, A. N. (2005). Induction of phase I, II and III drug metabolism/transport by xenobiotics. Archives of Pharmacal Research, 28(3), 249-268.
34. Veith, A., & Moorthy, B. (2018). Role of cytochrome P450s in the generation and metabolism of reactive oxygen species. Current Opinion in Toxicology, 7, 44-51.
35. Vilchis-Landeros, M. M., Vázquez-Meza, H., Vázquez-Carrada, M., Uribe-Ramírez, D., & Matuz-Mares, D. (2024). Antioxidant enzymes and their potential use in breast cancer treatment. International Journal of Molecular Sciences, 25(11), 5675.
36. Yonar, M. E., & Yonar, S. M. (2010). Changes in selected immunological parameters and antioxidant status of rainbow trout exposed to malachite green (Oncorhynchus mykiss, Walbaum, 1792). Pesticide Biochemistry and Physiology, 97, 19-23.
37. Zu-Yi, C., Bing-lin, H., Yu, B., Yuan, Z., & Xue-song, F. (2024). Malachite green and related substances in environmental samples: Updates on pretreatment and analysis methods. Journal of Environmental Chemical Engineering, 12(5), 113812.