Potential toxicity of graphene (oxide) quantum dots via directly covering the active site of anterior gradient homolog 2 protein.

Graphene quantum dots (GQDs) have attracted significant attention in biomedicine, while extensive investigations have revealed a reverse regarding the potential biotoxicity of GQDs. In order to supplementing the understanding of the toxicity profile of GQDs, this study employs a molecular dynamics (MD) simulation approach to systematically investigate the potential toxicity of both GQDs and Graphene Oxide Quantum Dots (GOQDs) on the Anterior Gradient Homolog 2 (AGR2) protein, a key protein capable of protecting the intestine. We construct two typical simulation systems, in which an AGR2 protein is encircled by either GQDs or GOQDs. The MD results demonstrate that both GQDs and GOQDs can directly make contact with and even cover the active site (specifically, the Cys81 amino acid) of the AGR2 protein. This suggests that GQDs and GOQDs have the capability to inhibit or interfere with the normal biological interaction of the AGR2 active site with its target protein. Thus, GQDs and GOQDs exhibit potential detrimental effects on the AGR2 protein. Detailed analyses reveal that GQDs adhere to the Cys81 residue due to van der Waals (vdW) interaction forces, whereas GOQDs attach to the Cys81 residue through a combination of vdW (primary) and Coulomb (secondary) interactions. Furthermore, GQDs aggregation typically adsorb onto the AGR2 active site, while GOQDs adsorb to the active site of AGR2 one by one. Consequently, these findings shed new light on the potential adverse impact of GQDs and GOQDs on the AGR2 protein via directly covering the active site of AGR2, providing valuable molecular insights for the toxicity profile of GQD nanomaterials.

Graphene oxide quantum dots (GOQDs) induce behavioral disorders via the disturbance of kynurenine pathway in zebrafish larvae.

The emergence of graphene quantum dots (GQDs) expands the use of graphene derivatives in nanomedicine for its direct therapeutic applications in treating neurodegeneration, inflammation, metabolic dysfunction, and among others. Nevertheless, the biosafety assessment of GQDs remains deficient mostly because of the diverse surface characteristics of the nanoparticles. Our prior work demonstrated that GQDs can induce strong thigmotactic effects in zebrafish larvae over a wide range of concentrations, yet the underlying metabolic mechanisms remain largely unknown. In this study, we conducted a further exploration about graphene oxide quantum dots (GOQDs) for its potential neurotoxic effect on the behaviors of zebrafish larvae by combining neurotransmitter-targeted metabolomics with locomotion analysis. After continuous exposure to a concentration gradient of GOQDs (12.5 - 25 - 50 - 100 - 200 µg/mL) for 7 days, the thigmotactic activities of zebrafish larvae were observed across all exposure concentrations relative to the control group, while the basal locomotor activities, including distance moved and average velocity, were significantly changed by low concentrations of GOQDs. Targeted metabolomics was performed using zebrafish larvae at 7 days post-fertilization (dpf) that were exposed to 12.5 and 200 µg/mL, both of which were found to perturb the kynurenine pathway by regulating the levels of kynurenine, 3-hydroxyanthranilic acid (3-HAA), and quinolinic acid (QA). Furthermore, the thigmotaxis of larval fish induced by GOQDs during exposure could be counteracted by supplementing Ro-61-8048, an agonist acting on kynurenine 3-monooxygenase (KMO). In conclusion, our study establishes the involvement of the kynurenine pathway in GOQDs-induced thigmotaxis, which is independent of the transcriptional modulation of glutamate receptor families.

Computational Assessment of the Biocompatibility of Two-Dimensional g-C3N3 Toward Lipid Membranes.

One of the most recent additions to the family of two-dimensional (2D) materials, graphitic C3N3 (g-C3N3), has been considered a viable contender for biomedical applications, although its potential toxicity remains elusive. We perform all-atom molecular dynamics simulations to decipher the interactions between model lipid membranes and g-C3N3 as a first step toward exploring the cytotoxicity induced at the nanoscale. We show that g-C3N3 can easily insert into the cellular membranes following a multistage mechanism consisting of simultaneous desolvation of the 2D material along with enrichment of nanomaterial-lipid interactions. Free energy calculations indicate that g-C3N3 is more stable in a membrane-bound state compared to an aqueous solution; however, the insertion of the material does not disturb the structural integrity of lipid membranes. After being inserted into a membrane, g-C3N3 is unlikely to be released into the cellular environment and is incapable of extracting lipid molecules from the membrane. The nature of interaction between the 2D material and membranes is found to be independent of the nanomaterial size. Also, the performance of g-C3N3 toward biomolecular delivery is shown to be significantly improved compared to the state-of-the-art 2D materials graphene and hexagonal boron nitride (h-BN). It is revealed that, the affinity of g-C3N3 toward lipid membranes is weaker compared to the nanotoxic graphene and h-BN, while being marginally higher than h2D-C2N, which in turn, increases the biocompatibility of the material, thereby brightening its future as a noncytotoxic material for forthcoming biomedical applications.

Magnetic graphene oxide nanocomposites induce cytotoxicity in ADSCs via GPX4 regulating ferroptosis.

Magnetic graphene oxide nanocomposites (MGO NPs) have been widely studied in biomedical applications. However, their cytotoxicity and underlying mechanisms remain unclear. In this study, the biosafety of MGO NPs was investigated, and the mechanism involved in ferroptosis was further explored. MGO can produce cytotoxicity in ADSCs, which is dependent on their concentration. Ferroptosis was involved in MGO NP-induced ADSC survival inhibition by increasing total ROS and lipid ROS accumulation as well as regulating the expression levels of ferroptosis-related genes and proteins. GPX4 played a critical role in the MGO NP-induced ADSC ferroptosis process, and overexpressing GPX4 suppressed ferroptosis to increase cell survival. This study provides a theoretical basis for the biosafety management of MGO NPs used in the field of biomedical treatment.

EPS-corona formation on graphene family nanomaterials (GO, rGO and graphene) and its role in mitigating their toxic effects in the marine alga Chlorella sp.

GFNs have widespread applications but can harm marine systems due to excessive use and improper disposal. Algae-secreted EPS can mitigate nanomaterial harm, but their impact on GFN toxicity is understudied. Hence, in the present study, we investigated the toxicity of three GFNs, graphene oxide (GO), reduced graphene oxide (rGO), and graphene, in pristine and EPS-adsorbed forms in the marine alga Chlorella sp. At an environmentally relevant concentration of 1 mgL-1, all three GFNs induced considerable oxidative stress and impeded growth and photosynthetic activity of the algae. The order of the toxic potential followed GO > rGO > graphene. The various facets of adsorption of EPS (1:1 mixture of loosely bound, and tightly bound EPS) on GFNs were investigated through microscopy, surface chemical analyses, fluorescence quenching studies, and isotherm and kinetics studies. Amongst the pristine GFNs treated with algal cells, GO was found to exert the maximum negative effects on algal growth. Upon adsorption of EPS over the GFNs, a significant decline in growth inhibition was observed compared to the respective pristine forms which strongly correlated with reduced oxidative stress and enhanced photosynthetic parameters in the cells. The formation of a layer of eco-corona after interaction of GFNs with EPS possibly caused a barrier effect which in turn diminished their toxic potential. The findings from the present investigation offer valuable insights into the environmental toxicity of GFNs and show that the eco-corona formation may lessen the risk posed by these materials in the marine environment.

Graphene-Based Engineered Living Materials.

With the rise of engineered living materials (ELMs) as innovative, sustainable and smart systems for diverse engineering and biological applications, global interest in advancing ELMs is on the rise. Graphene-based nanostructures can serve as effective tools to fabricate ELMs. By using graphene-based materials as building units and microorganisms as the designers of the end materials, next-generation ELMs can be engineered with the structural properties of graphene-based materials and the inherent properties of the microorganisms. However, some challenges need to be addressed to fully take advantage of graphene-based nanostructures for the design of next-generation ELMs. This work covers the latest advances in the fabrication and application of graphene-based ELMs. Fabrication strategies of graphene-based ELMs are first categorized, followed by a systematic investigation of the advantages and disadvantages within each category. Next, the potential applications of graphene-based ELMs are covered. Moreover, the challenges associated with fabrication of next-generation graphene-based ELMs are identified and discussed. Based on a comprehensive overview of the literature, the primary challenge limiting the integration of graphene-based nanostructures in ELMs is nanotoxicity arising from synthetic and structural parameters. Finally, we present possible design principles to potentially address these challenges.

Photostability and phototoxicity of graphene quantum dots interacting with red blood cells.

Here we discuss fluorescent properties of graphene quantum dots (GQDs) interacting with the membranes of red blood cells. We report the results of spectroscopic, microscopic, and photon-counting measurements of the GQDs in different surroundings for uncovering specific features of the GQD fluorescence, and describe two observed phenomena important for implementation of the GQDs as fluorescent labels and agents for drug delivery. Firstly, the GQDs can suffer from photodegradation but also can be stabilized in the presence of antioxidants (reduced glutathione, N-acetylcysteine, or 1,4-hydroquinone). Secondly, GQDs can accumulate in red blood cell membranes without compromising the viability of the cells but also can induce hemolysis in the presence of visible light. We discuss mechanisms and regimes of the photodegradation, stabilization, interaction of the GQDs with red blood cell membranes, and hemolysis. Notably, photohemolysis for the case is dependent on the light dose and GQD concentration but not caused by the production of reactive oxygen species.

Abnormality of mussel in the early developmental stages induced by graphene and triphenyl phosphate: In silico toxicogenomic data-mining, in vivo, and toxicity pathway-oriented approach.

Increasing number of complex mixtures of organic pollutants in coastal area (especially for nanomaterials and micro/nanoplastics associated chemicals) threaten aquatic ecosystems and their joint hazards are complex and demanding tasks. Mussels are the most sensitive marine faunal groups in the world, and their early developmental stages (embryo and larvae) are particularly susceptible to environmental contaminants, which can distinguish the probable mechanisms of mixture-induced growth toxicity. In this study, the potential critical target and biological processes affected by graphene and triphenyl phosphate (TPP) were developed by mining public toxicogenomic data. And their combined toxic effects were verified by toxicological assay at early developmental stages in filter-feeding mussels (embryo and larvae). It showed that interactions among graphene/TPP with 111 genes (ABCB1, TP53, SOD, CAT, HSP, etc.) affected phenotypes along conceptual framework linking these chemicals to developmental abnormality endpoints. The PPAR signaling pathway, monocarboxylic acid metabolic process, regulation of lipid metabolic process, response to oxidative stress, and gonad development were noted as the key molecular pathways that contributed to the developmental abnormality. Enriched phenotype analysis revealed biological processes (cell proliferation, cell apoptosis, inflammatory response, response to oxidative stress, and lipid metabolism) affected by the investigated mixture. Combined, our results supported that adverse effects induced by contaminants/ mixture could not only be mediated by single receptor signaling or be predicted by the simple additive effect of contaminants. The results offer a framework for better comprehending the developmental toxicity of environmental contaminants in mussels and other invertebrate species, which have considerable potential for hazard assessment of coastal mixture.

Three-dimensional graphene aerogel mitigated the toxic impact of chloramphenicol wastewater on microorganisms in an EGSB reactor.

Anaerobic treatment of chloramphenicol wastewater holds significant promise due to its potential for bioenergy generation. However, the high concentration of organic matter and residual toxic substances in the wastewater severely inhibit the activity of microorganisms. In this study, a three-dimensional graphene aerogel (GA), as a conductive material with high specific surface area (114.942 m2 g-1) and pore volume (0.352 cm3 g-1), was synthesized and its role in the efficiency and related mechanism for EGSB reactor to treat chloramphenicol wastewater was verified. The results indicated that synergy effects of GA for Chemical Oxygen Demand (COD) removal (increased by 8.17 %), chloramphenicol (CAP) removal (increased by 4.43 %) and methane production (increased by 70.29 %). Furthermore, GA increased the average particle size of anaerobic granular sludge (AGS) and promoted AGS to secrete more redox active substances. Microbial community analysis revealed that GA increased the relative abundance of functional bacteria and archaea, specifically Syntrophomonas, Geobacter, Methanothrix, and Methanolinea. These microbial species can participate in direct interspecific electron transfer (DIET). This research serves as a theoretical foundation for the application of GA in mitigating the toxic impact of refractory organic substances, such as antibiotics, on microorganisms during anaerobic treatment processes.

Applications of graphene-based nanomaterials in drug design: The good, the bad and the ugly.

INTRODUCTION: Graphene-based materials (GBMs) have unique physicochemical properties that make them extremely attractive as platforms for the design of new drugs. Indeed, their bidimensional (2D) morphology, high surface area, mechanical and optical properties, associated to different possibilities for functionalization of their surface, provides opportunities for their use as nanomedicines for drug delivery and/or phototherapies. AREAS COVERED: This opinion paper provides an overview of the current status of GBMs in drug design, with a focus on their therapeutic applications, potential environmental and health risks, and some controversial results. The authors discuss the chemical modifications of GBMs for the treatment of various diseases. The potential toxicity associated with some GBMs is also presented, along with a safe-by-design approach to minimize the risks. Finally, the authors address some issues associated to the use of GBMs in the biomedical field, such as contradictory antibacterial effects, fluorescence quenching and imprecise chemical functionalization. EXPERT OPINION: GBMs are a promising and exciting area of research in drug delivery. It is however important that responsible and safe use of these materials is ensured to fully exploit their advantages and overcome their drawbacks.

In vitro assessment of skin irritation and corrosion properties of graphene-related materials on a 3D epidermis.

The increasing use of graphene-related materials (GRMs) in many technological applications, ranging from electronics to biomedicine, needs a careful evaluation of their impact on human health. Skin contact can be considered one of the most relevant exposure routes to GRMs. Hence, this study is focused on two main adverse outcomes at the skin level, irritation and corrosion, assessed following two specific Test Guidelines (TGs) defined by the Organization for Economic Co-operation and Development (OECD) (439 and 431, respectively) that use an in vitro 3D reconstructed human epidermis (RhE) model. After the evaluation of their suitability to test a large panel of powdered GRMs, it was found that the latter were not irritants or corrosive. Only GRMs prepared with irritant surfactants, not sufficiently removed, reduced RhE viability at levels lower than those predicting skin irritation (≤50%, after 42 min exposure followed by 42 h recovery), but not at levels lower than those predicting corrosion (<50%, after 3 min exposure or <15% after 1 h exposure). As an additional readout, a hierarchical clustering analysis on a panel of inflammatory mediators (interleukins: IL-1α, IL-1ß, IL-6, and IL-18; tumor necrosis factor-α and prostaglandin E2) released by RhE exposed to these materials supported the lack of irritant and pro-inflammatory properties. Overall, these results demonstrate that both TGs are useful in assessing GRMs for their irritant or corrosion potential, and that the tested materials did not cause these adverse effects at the skin level. Only GRMs prepared using toxic surfactants, not adequately removed, turned out to be skin irritants.

Environmental biotechnology and the involving biological process using graphene-based biocompatible material.

Biotechnology is a promising approach to environmental remediation but requires improvement in efficiency and convenience. The improvement of biotechnology has been illustrated with the help of biocompatible materials as biocarrier for environmental remediations. Recently, graphene-based materials (GBMs) have become promising materials in environmental biotechnology. To better illustrate the principle and mechanisms of GBM application in biotechnology, the comprehension of the biological response of microorganisms and enzymes when facing the GBMs is needed. The review illustrated distinct GBM-microbe/enzyme composites by providing the GBM-microbe/enzyme interaction and the determining factors. There are diverse GBM modifications for distinct biotechnology applications. Each of these methods and applications depends on the physicochemical properties of GBMs. The applications of these composites were mainly categorized as pollutant adsorption, anaerobic digestion, microbial fuel cells, and organics degradation. Where information was available, the strategies and mechanisms of GBMs in improving application efficacies were also demonstrated. In addition, the biological response, from microbial community changes, extracellular polymeric substances changes to biological pathway alteration, may become important in the application of these composites. Furthermore, we also discuss challenges facing the environmental application of GBMs, considering their fate and toxicity in the ecosystem, and offer potential solutions. This research significantly enhances our comprehension of the fundamental principles, underlying mechanisms, and biological pathways for the in-situ utilization of GBMs.

Graphene oxide had adverse effects on sperm motility and morphology through oxidative stress.

Graphene oxide (GO) is a new type of graphene material, but its effects on the male reproductive system are unclear. Here, we investigated the effects of GO on human sperm in vitro. Sperms were incubated with various doses of GO (0, 10, 20, or 40 µg/mL) for different times (1, 3, or 6 h) at 37 °C, followed by analyses of the sperm motility, viability, abnormalities, and DNA fragmentations. GO exposure significantly decreased sperm motility and viability, increased sperm abnormalities, and DNA fragmentation. Moreover, GO exposure resulted in a significant reduction of sperm mitochondrial membrane potential (MMP), which was confirmed by the ultrastructural changes of chromatin and mitochondria caused by GO. These data revealed the adverse effects of GO on sperm. Further research showed that GO exposure led to a significant increase in malondialdehyde (MDA) and reactive oxygen species (ROS) in sperm cells and a significant decrease in total superoxide dismutase (T-SOD) and glutathione peroxidase (GSH-Px). In addition, western blot analysis showed that the levels of Nrf-2 and HO-1 protein expression in GO-treated sperm cells were significantly increased compared to the control. These results indicated that GO had adverse effects on human sperm through oxidative stress, which was associated with Nrf-2/HO-1 signaling pathway.

Toxicological assessment of pristine and degraded forms of graphene functionalized with MnOx nanoparticles using human in vitro models representing different exposure routes.

The development of novel advanced nanomaterials (NMs) with outstanding characteristics for their use in distinct applications needs to be accompanied by the generation of knowledge on their potential toxicological impact, in particular, that derived from different occupational risk exposure routes, such as inhalation, ingestion, and skin contact. The harmful effects of novel graphene-metal oxide composites on human health are not well understood, many toxicological properties have not been investigated yet. The present study has evaluated several toxicological effects associated with graphene decorated with manganese oxide nanoparticles (GNA15), in a comparative assessment with those induced by simple graphene (G2), on human models representing inhalation (A549 cell line), ingestion (HT29 cell line) and dermal routes (3D reconstructed skin). Pristine and degraded forms of these NMs were included in the study, showing to have different physicochemical and toxicological properties. The degraded version of GNA15 (GNA15d) and G2 (G2d) exhibited clear structural differences with their pristine counterparts, as well as a higher release of metal ions. The viability of respiratory and gastrointestinal models was reduced in a dose-dependent manner in the presence of both GNA15 and G2 pristine and degraded forms. Besides this, all NMs induced the production of reactive oxygen species (ROS) in both models. However, the degraded forms showed to induce a higher cytotoxicity effect. In addition, we found that none of the materials produced irritant effects on 3D reconstructed skin when present in aqueous suspensions. These results provide novel insights into the potentially harmful effects of novel multicomponent NMs in a comprehensive manner. Furthermore, the integrity of the NMs can play a role in their toxicity, which can vary depending on their composition and the exposure route.

In vitro safety assessment of reduced graphene oxide in human monocytes and T cells.

Considering the increase in the use of graphene derivatives in different fields, the environmental and human exposure to these materials is likely, and the potential consequences are not fully elucidated. This study is focused on the human immune system, as this plays a key role in the organism's homeostasis. In this sense, the cytotoxicity response of reduced graphene oxide (rGO) was investigated in monocytes (THP-1) and human T cells (Jurkat). A mean effective concentration (EC50-24 h) of 121.45 ± 11.39 µg/mL and 207.51 ± 21.67 µg/mL for cytotoxicity was obtained in THP-1 and Jurkat cells, respectively. rGO decreased THP-1 monocytes differentiation at the highest concentration after 48 h of exposure. Regarding the inflammatory response at genetic level, rGO upregulated IL-6 in THP-1 and all cytokines tested in Jurkat cells after 4 h of exposure. At 24 h, IL-6 upregulation was maintained, and a significant decrease of TNF-α gene expression was observed in THP-1 cells. Moreover, TNF-α, and INF-γ upregulation were maintained in Jurkat cells. With respect to the apoptosis/necrosis, gene expression was not altered in THP-1 cells, but a down regulation of BAX and BCL-2 was observed in Jurkat cells after 4 h of exposure. These genes showed values closer to negative control after 24 h. Finally, rGO did not trigger a significant release of any cytokine at any exposure time assayed. In conclusion, our data contributes to the risk assessment of this material and suggest that rGO has an impact on the immune system whose final consequences should be further investigated.

Graphene oxide worsens copper-mediated embryo-larval toxicity in the pacific oyster while reduced graphene oxide mitigates the effects.

Due to their properties, graphene-based nanomaterials (GBMs) are triggering a great interest leading to an increase of their global production and use in new applications. As a consequence, their release into the environment is expected to increase in the next years. When considering the current knowledge in the evaluation of GBMs ecotoxic potential, studies aiming to evaluate the hazard associated to these nanomaterials towards marine species and particularly considering potential interactions with other environmental pollutants such as metals are scarce. Here we evaluated the embryotoxic potential of GBMs, which include graphene oxide (GO) and its reduced form (rGO), both individually and in combination with copper (Cu) as a referent toxicant, towards early life stages of the Pacific oyster through the use of a standardized method (NF ISO 17244). We found that following exposure to Cu, dose-dependent decrease in the proportion of normal larvae was recorded with an Effective Concentration leading to the occurrence of 50% of abnormal larvae (EC50) of 13.85 ± 1.21 µg/L. Interestingly, the presence of GO at a non-toxic dose of 0.1 mg/L decreased the Cu EC50 to 12.04 ± 0.85 µg/L while it increased to 15.91 ± 1.57 µg/L in presence of rGO. Based on the measurement of copper adsorption, the obtained results suggest that GO enhances Cu bioavailability, potentially modifying its toxic pathways, while rGO mitigates Cu toxicity by decreasing its bioavailability. This research underscores the need to characterize the risk associated to GBMs interactions with other aquatic contaminants and supports the adoption of a safer-by-design strategy using rGO in marine environments. This would contribute to minimize the potential adverse effects on aquatic species and to reduce the risk for economic activities associated to coastal environments.

Environmental and health impacts of functional graphenic materials and their ultrasonically altered products.

Graphenic materials have excited the scientific community due to their exciting mechanical, thermal, and optoelectronic properties for a potential range of applications. Graphene and graphene derivatives have demonstrated application in areas stretching from composites to medicine; however, the environmental and health impacts of these materials have not been sufficiently characterized. Graphene oxide (GO) is one of the most widely used graphenic derivatives due to a relatively easy and scalable synthesis, and the ability to tailor the oxygen containing functional groups through further chemical modification. In this paper, ecological and health impacts of fresh and ultrasonically altered functional graphenic materials (FGMs) were investigated. Model organisms, specifically Escherichia coli, Bacillus subtilis, and Caenorhabditis elegans, were used to assess the consequences of environmental exposure to fresh and ultrasonically altered FGMs. FGMs were selected to evaluate the environmental effects of aggregation state, degree of oxidation, charge, and ultrasonication. The major findings indicate that bacterial cell viability, nematode fertility, and nematode movement were largely unaffected, suggesting that a wide variety of FGMs may not pose significant health and environmental risks.

Comparative ecotoxicity of graphene, functionalized multi-walled CNTs, and their mixture in freshwater microalgae, Scenedesmus obliquus: analyzing the role of oxidative stress.

Due to their remarkable properties, the applications of carbon-based nanomaterials (CNMs) such as graphene and functionalized multi-walled carbon nanotubes (f-MWCNTs) are increasing. These CNMs can enter the freshwater environment via numerous routes, potentially exposing various organisms. The current study assesses the effects of graphene, f-MWCNTs, and their binary mixture on the freshwater algal species Scenedesmus obliquus. The concentration for the individual materials was kept at 1 mg L-1, while graphene and f-MWCNTs were taken at 0.5 mg L-1 each for the combination. Both the CNMs caused a decrease in cell viability, esterase activity, and photosynthetic efficiency in the cells. The cytotoxic effects were accompanied by increased hydroxyl and superoxide radical generation, lipid peroxidation, antioxidant enzyme activity (catalase and superoxide dismutase), and mitochondrial membrane potential. Graphene was more toxic compared to f-MWCNTs. The binary mixture of the pollutants demonstrated a synergistic enhancement of the toxic potential. Oxidative stress generation played a critical role in toxicity responses, as noted by a strong correlation between the physiological parameters and the biomarkers of oxidative stress. The outcomes from this study emphasize the significance of considering the combined effects of various CNMs as part of a thorough evaluation of ecotoxicity in freshwater organisms.

The impact of various carbon nanomaterials on the morphological, behavioural, and biochemical parameters of rainbow trout in the early life stages.

With the increasing production and the number of potential applications of carbon nanomaterials, mainly from the graphene family, their release into the natural environment, especially to aquatic ecosystems, is inevitable. The aim of the study was to determine the effects of various carbon nanomaterials (graphene nanoflakes (GNF), graphene oxide (GO), reduced graphene oxide (RGO) and silicon carbide nanofibers (NFSiC) in the concentration of 4 mg L-1 on the early life stages of the rainbow trout Oncorhynchus mykiss. The survival rates of O. mykiss were not affected after 36 days of exposure to studied materials, except for RGO, which caused significant mortality of both embryos and larvae compared to the control conditions. Larvae exposed to GO and NFSiC were characterized by a smaller standard body length at hatch, whereas at the end of the experiment, the growth of fish exposed to all materials was accelerated, especially in GO and RGO treatment, in which higher body weight and length were accompanied by lower volume of the yolk sac. Neither the markers of the oxidative damage nor the antioxidant enzymes activities were significantly affected in embryos, newly hatched larvae and larvae after 26-day exposure to studied carbon nanomaterials. Also, no neurotoxic effect expressed by the activity of the whole-body acetylcholinesterase was observed. Nevertheless, the significant increase in the velocity and the overall activity of larvae exposed to GNF (not investigated after exposure to other materials) must be highlighted. The most pronounced effect of RGO might be connected with its large particle size, sharp edges, and the presence of TiO2 nanoparticles. The results indicate for the first time that various carbon nanomaterials potentially released into aquatic ecosystems may have serious developmental implications for the early life stages of salmonid fish.

Surface Area of Graphene Governs Its Neurotoxicity.

Due to their unique physicochemical properties, graphene and its derivatives are widely exploited for biomedical applications. It has been shown that graphene may exert different degrees of toxicity in in vivo or in vitro models when administered via different routes and penetrated through physiological barriers, subsequently being distributed within tissues or located within cells. In this study, in vitro neurotoxicity of graphene with different surface areas (150 and 750 m2/g) was examined on dopaminergic neuron model cells. SH-SY5Y cells were treated with graphene possessing two different surface areas (150 and 750 m2/g) in different concentrations between 400 and 3.125 µg/mL, and the cytotoxic and genotoxic effects were investigated. Both sizes of graphene have shown increased cell viability in decreasing concentrations. Cell damage increased with higher surface area. Lactate dehydrogenase (LDH) results have concluded that the viability loss of the cells is not through membrane damage. Neither of the two graphene types showed damage through lipid peroxidation (MDA) oxidative stress pathway. Glutathione (GSH) values increased within the first 24 and 48 h for both types of graphene. This increase suggests that graphene has an antioxidant effect on the SH-SY5Y model neurons. Comet analysis shows that graphene does not show genotoxicity on either surface area. Although there are many studies on graphene and its derivatives on their use with different cells in the literature, there are conflicting results in these studies, and most of the literature is focused on graphene oxide. Among these studies, no study examining the effect of graphene surface areas on the cell was found. Our study contributes to the literature in terms of examining the cytotoxic and genotoxic behavior of graphene with different surface areas.