INVESTIGATIONS OF BIOLOGICAL EFFECTS OF GRAPHENE OXIDE NANOSTRUCTURES ON BROWN TROUT (SALMO TRUTTA)

Gytis Martusevičius; Gintarė Sauliutė; Janina Pažusienė; Živilė Jurgelėnė; Sergej Šemčuk; Milda Stankevičiūtė

Open Readings 2021 • P8-22

INVESTIGATIONS OF BIOLOGICAL EFFECTS OF GRAPHENE OXIDE NANOSTRUCTURES ON BROWN TROUT (SALMO TRUTTA)

Gytis Martusevičius^{1, 2}, Gintarė Sauliutė¹, Janina Pažusienė¹, Živilė Jurgelėnė¹, Sergej Šemčuk³, Milda Stankevičiūtė¹

¹ Nature Research Centre, Akademijos St. 2, LT-08412 Vilnius, Lithuania

² Vilnius University, Life Sciences Center, Department of Biochemistry and Molecular Biology, Saulėtekio av. 7, 10223 Vilnius, Lithuania

³ SRI Center for Physical Sciences and Technology, Savanorių av. 231, LT-02300 Vilnius, Lithuania

[email protected]

Graphene oxide (GO) nanoparticles are useful and promising material for graphene-based applications in electronic, optics, chemistry, energy storage, and biology. Recently developed methods for preparation of graphene oxide derivatives open the new attractive application areas in green technologies including energy storage and utilizing nuclear wastes [1]. Extensive application of GO increases its probability to enter the environment. GO belong to the class of carbon nanomaterials, which have the ability to cross cellular barriers, interact with many of cellular components, including the plasma membrane, cytoplasmic organelles and nucleus [2]. For these reasons GO may cause toxic effects on aquatic invertebrates and fish (in vitro and in vivo) [3].

The aim of this study was to determine genotoxicity responses (nuclear abnormalities assay) and metallothioneins (MT) level in Salmo trutta larvae exposed to graphene oxide nanostructures (GON), metal mixture (MIX), and GON+MIX. Toxicity of metal mixture (MIX) at 40-fold (40↑) increased maximum-permissible-concentrations (MPC) set for EU inland waters to S. trutta larvae were evaluated using the whole-mixture approach. Induction of micronuclei (MN), nuclear buds (NB), nuclear buds on filament (NBf), bi-nucleated erythrocytes with nucleoplasmic bridges (BNb) and blebbed nuclei (BL) cells were assessed as genotoxicity endpoints. The total genotoxicity (ΣGentox) level was assessed as the sum of frequencies of the analysed genotoxicity (MN+NB+NBf+BNb+BL) endpoints. Metallothioneins induction was measured by colorimetric reaction using Elman reagent.

Results showed that after exposure to GON40↑, the amount of MT in S. trutta larvae increased, as well after the exposure to GON40↑+MIX40↑, but was not statistically significant. Exposure to MIX40↑ resulted in 100% mortality of fish larvae. The genotoxicity assay showed that the highest frequencies of analysed endpoints were found in GON40↑+MIX40↑ exposed larvae. S. trutta larvae exposure to GON40↑+MIX40↑ resulted in a significant elevation of MN, NB and ΣGentox frequencies compare to control and GON40↑ exposure groups. GON40↑ induced significant MN and ΣGentox formation compare to control group; and MN, NB and ΣGentox levels compared to GON40↑+MIX40↑ group. This research highlights the genotoxic and cytotoxic potential of GON in early stages of S. trutta. The results of the MT analysis may indicate the potential of GON to bind heavy metals in S. trutta larvae.

Acknowledgments: This work was funded by the Research Council of Lithuania, Project No. S-MIP-20-22.

[1] A. T. Dideikin, A. Y. Vul, Graphene Oxide and Derivatives: The Place in Graphene Family, Frontiers in Physics 6, 149 (2019).

[2] A. K. Dasmahapatra, T. P. S. Dasari, P. B. Tchounwou, Graphene-Based Nanomaterials Toxicity in Fish. Rev Environ Contam Toxicol 247, 1-58 (2019).

[3] N. Malhotra, O. B. Villaflores, G. Audira, P. Siregar, J-S. Lee, T-R. Ger, C-D Hsiao, Toxicity studies on graphene-based nanomaterials in aquatic organisms: current understanding, Molecules 25 (16), 3618 (2020).