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Graphene, graphene oxide, and reduced graphene oxide have been widely considered as promising candidates for industrial and biomedical applications due to their exceptionally high mechanical stiffness and strength, excellent electrical conductivity, high optical transparency, and good biocompatibility. In this article, we reviewed several techniques that are available for the synthesis of graphene-based nanomaterials, and discussed the biocompatibility and toxicity of such nanomaterials upon exposure to mammalian cells under in vitro and in vivo conditions. Various synthesis strategies have been developed for their fabrication, generating graphene nanomaterials with different chemical and physical properties. As such, their interactions with cells and organs are altered accordingly. Conflicting results relating biocompatibility and cytotoxicity induced by graphene nanomaterials have been reported in the literature. In particular, graphene nanomaterials that are used for in vitro cell culture and in vivo animal models may contain toxic chemical residuals, thereby interfering graphene-cell interactions and complicating interpretation of experimental results. Synthesized techniques, such as liquid phase exfoliation and wet chemical oxidation, often required toxic organic solvents, surfactants, strong acids, and oxidants for exfoliating graphite flakes. Those organic molecules and inorganic impurities that are retained in final graphene products can interact with biological cells and tissues, inducing toxicity or causing cell death eventually. The residual contaminants can cause a higher risk of graphene-induced toxicity in biological cells. This adverse effect may be partly responsible for the discrepancies between various studies in the literature.

Graphene and its oxide are nanomaterials considered currently to be very promising because of their great potential applications in various industries. The exceptional physiochemical properties of graphene, particularly thermal conductivity, electron mobility, high surface area, and mechanical strength, promise development of novel or enhanced technologies in industries. The diverse applications of graphene and graphene oxide (GO) include energy storage, sensors, generators, light processing, electronics, and targeted drug delivery. However, the extensive use and exposure to graphene and GO might pose a great threat to living organisms and ultimately to human health. The toxicity data of graphene and GO is still insufficient to point out its side effects to different living organisms. Their accumulation in the aquatic environment might create complex problems in aquatic food chains and aquatic habitats leading to debilitating health effects in humans. The potential toxic effects of graphene and GO are not fully understood. However, they have been reported to cause agglomeration, long-term persistence, and toxic effects penetrating cell membrane and interacting with cellular components. In this review paper, we have primarily focused on the toxic effects of graphene and GO caused on aquatic invertebrates and fish (cell line and organisms). Here, we aim to point out the current understanding and knowledge gaps of graphene and GO toxicity.

Graphene is a two-dimensional atomic crystal, and since its development it has been applied in many novel ways in both research and industry. Graphene possesses unique properties, and it has been used in many applications including sensors, batteries, fuel cells, supercapacitors, transistors, components of high-strength machinery, and display screens in mobile devices. In the past decade, the biomedical applications of graphene have attracted much interest. Graphene has been reported to have antibacterial, antiplatelet, and anticancer activities. Several salient features of graphene make it a potential candidate for biological and biomedical applications. The synthesis, toxicity, biocompatibility, and biomedical applications of graphene are fundamental issues that require thorough investigation in any kind of applications related to human welfare. Therefore, this review addresses the various methods available for the synthesis of graphene, with special reference to biological synthesis, and highlights the biological applications of graphene with a focus on cancer therapy, drug delivery, bio-imaging, and tissue engineering, together with a brief discussion of the challenges and future perspectives of graphene. We hope to provide a comprehensive review of the latest progress in research on graphene, from synthesis to applications.

Due to their unique physicochemical properties, graphene-family nanomaterials (GFNs) are widely used in many fields, especially in biomedical applications. Currently, many studies have investigated the biocompatibility and toxicity of GFNs in vivo and in intro. Generally, GFNs may exert different degrees of toxicity in animals or cell models by following with different administration routes and penetrating through physiological barriers, subsequently being distributed in tissues or located in cells, eventually being excreted out of the bodies. This review collects studies on the toxic effects of GFNs in several organs and cell models. We also point out that various factors determine the toxicity of GFNs including the lateral size, surface structure, functionalization, charge, impurities, aggregations, and corona effect ect. In addition, several typical mechanisms underlying GFN toxicity have been revealed, for instance, physical destruction, oxidative stress, DNA damage, inflammatory response, apoptosis, autophagy, and necrosis. In these mechanisms, (toll-like receptors-) TLR-, transforming growth factor β- (TGF-β-) and tumor necrosis factor-alpha (TNF-α) dependent-pathways are involved in the signalling pathway network, and oxidative stress plays a crucial role in these pathways. In this review, we summarize the available information on regulating factors and the mechanisms of GFNs toxicity, and propose some challenges and suggestions for further investigations of GFNs, with the aim of completing the toxicology mechanisms, and providing suggestions to improve the biological safety of GFNs and facilitate their wide application.

Metal-free, low-cost, organic photocatalytic graphitic carbon nitride (g-C3N4) has become a promising and impressive material in numerous scientific fields due to its unique physical and chemical properties. As a semiconductor with a suitable band gap of ~2.7 eV, g-C3N4 is an active photocatalytic material even after irradiation with visible light. However, information regarding the toxicity of g-C3N4 is not extensively documented and there is not a comprehensive understanding of its potential adverse effects on human health or the environment. In this context, the term “toxicity” can be perceived in both a positive and a negative light, depending on whether it serves as a benefit or poses a potential risk. This review shows the applications of g-C3N4 in sensorics, electrochemistry, photocatalysis, and biomedical approaches while pointing out the potential risks of its toxicity, especially in human and environmental health. Finally, the future perspective of g-C3N4 research is addressed, highlighting the need for a comprehensive understanding of the toxicity of this material to provide safe and effective applications in various fields.

Background Carbon nanotubes, graphene, graphite nanoplatelets and carbon black are seemingly chemically identical carbon-based nano-materials with broad technological applications. Carbon nanotubes and carbon black possess different inhalation toxicities, whereas little is known about graphene and graphite nanoplatelets. Methods In order to compare the inhalation toxicity of the mentioned carbon-based nanomaterials, male Wistar rats were exposed head-nose to atmospheres of the respective materials for 6 hours per day on 5 consecutive days. Target concentrations were 0.1, 0.5, or 2.5 mg/m 3 for multi-wall carbon nanotubes and 0.5, 2.5, or 10 mg/m 3 for graphene, graphite nanoplatelets and low-surface carbon black. Toxicity was determined after end of exposure and after three-week recovery using broncho-alveolar lavage fluid and microscopic examinations of the entire respiratory tract. Results No adverse effects were observed after inhalation exposure to 10 mg/m 3 graphite nanoplatelets or relatively low specific surface area carbon black. Increases of lavage markers indicative for inflammatory processes started at exposure concentration of 0.5 mg/m 3 for multi-wall carbon nanotubes and 10 mg/m 3 for graphene. Consistent with the changes in lavage fluid, microgranulomas were observed at 2.5 mg/m 3 multi-wall carbon nanotubes and 10 mg/m 3 graphene. In order to evaluate volumetric loading of the lung as the key parameter driving the toxicity, deposited particle volume was calculated, taking into account different methods to determine the agglomerate density. However, the calculated volumetric load did not correlate to the toxicity, nor did the particle surface burden of the lung. Conclusions The inhalation toxicity of the investigated carbon-based materials is likely to be a complex interaction of several parameters. Until the properties which govern the toxicity are identified, testing by short-term inhalation is the best option to identify hazardous properties in order to avoid unsafe applications or select safer alternatives for a given application.

Background Graphene and graphene derivative nanoplatelets represent a new generation of nanomaterials with unique physico-chemical properties and high potential for use in composite materials and biomedical devices. To date little is known about the impact graphene nanomaterials may have on human health in the case of accidental or intentional exposure. The objective of this study was to assess the cytotoxic potential of graphene nanoplatelets with different surface chemistry towards a human hepatoma cell line, Hep G2, and identify the underlying toxicity targets. Methods Graphene oxide (GO) and carboxyl graphene (CXYG) nanoplatelet suspensions were obtained in water and culture medium. Size frequency distribution of the suspensions was determined by means of dynamic light scattering. Height, lateral dimension and shape of the nanoplatelets were determined using atomic force and electron microscopy. Cytotoxicity of GO and CXYG nanoplatelets was assessed in Hep G2 cells using a battery of assays covering different modes of action including alterations of metabolic activity, plasma membrane integrity and lysosomal function. Induction of oxidative stress was assessed by measuring intracellular reactive oxygen species levels. Interaction with the plasma membrane, internalization and intracellular fate of GO and CXYG nanoplatelets was studied by scanning and transmission electron microscopy. Results Supplementing culture medium with serum was essential to obtain stable GO and CXYG suspensions. Both graphene derivatives had high affinity for the plasma membrane and caused structural damage of the latter at concentrations as low as 4 μg/ml. The nanoplatelets penetrated through the membrane into the cytosol, where they were concentrated and enclosed in vesicles. GO and CXYG accumulation in the cytosol was accompanied by an increase in intracellular reactive oxygen species (ROS) levels, alterations in cellular ultrastructure and changes in metabolic activity. Conclusions GO and CXYG nanoplatelets caused dose- and time-dependent cytotoxicity in Hep G2 cells with plasma membrane damage and induction of oxidative stress being important modes of toxicity. Both graphene derivatives were internalized by Hep G2, a non-phagocytotic cell line. Moreover, they exerted no toxicity when applied at very low concentrations (< 4 μg/ml). GO and CXYG nanoplatelets may therefore represent an attractive material for biomedical applications.

Graphene oxide (GO) has promising biomedical applications, but its potential toxicity to the female reproductive system is underexplored. This study investigates the short-term effects of a single dose of GO nanosheets on human ovarian granulosa cells, focusing on mitochondrial damage. First, cell viability was detected by CCK-8 and apoptosis was detected by flow cytometry to assess the cytotoxicity of GO on KGN. Second, reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and mitochondrial morphology were observed by confocal microscopy, mitochondrial and sub-mitochondrial structure by transmission electron microscopy (TEM), quantitative analysis of ATP and mitochondrial complex I enzyme activity by luminosity value and autophagy by flow cytometry to assess the mitochondrial toxicity of GO on KGN cells. The 72h half-maximum effective concentration (EC50) value of GO was determined to be 29.73 μg/mL. GO induced cell death in a dose-dependent manner, with significant effects on cell viability even at low doses (1 μg/mL). Exposure to low GO concentrations resulted in abnormal mitochondrial morphology and function, including mitochondrial breakage, membrane damage, reduced mitochondrial cristae, enhanced autophagy, decreased ATP production, decreased MMP, and decreased enzymatic activity of mitochondrial complex I. Mitochondrial function returned to normal levels on day 7 after KGN cells left the GO-exposed environment. This study demonstrates that short-term exposure to low-dose GO causes mitochondrial damage in human ovarian granulosa cells, highlighting the need for further research on the safety of GO, particularly regarding its potential effects on reproductive health. However, GO-induced transient mitochondrial damage is highly likely to negatively affect ovarian reserve function, which needs to be further verified in animal models.

This review (with 239 refs.) summarizes the progress that has been made in applications of graphene-based nanomaterials (such as plain graphene, graphene oxides, doped graphene oxides, graphene quantums dots) in biosensing, imaging, drug delivery and diagnosis. Following an introduction into the field, a first large section covers the toxicity of graphene and its derivatives (with subsections on bacterial toxicity and tissue toxicity). The use of graphene-based nanomaterials in sensors is reviewed next, with subsections on electrochemical, FET-based, fluorescent, chemiluminescent and colorimetric sensors and probes. The large field of imaging is treated next, with subchapters on optical, PET-based, and magnetic resonance based methods. A concluding section summarizes the current status, addresses current challenges, and gives an outlook on potential future trends. Graphical Abstract Schematic presentation of the potential applications of graphene-based materials in life science and biomedicine, emphatically reflected in some vital areas such as DNA analysis, biological monitoring, drug delivery, in vitro labelling, in vivo imaging, tumor target, etc.

The extensive application of graphene nanosheets (GNSs) has raised concerns over risks to sensitive species in the aquatic environment. The humic acid (HA) corona is traditionally considered to reduce GNSs toxicity. Here, we evaluate the effect of sorbed HA (GNSs-HA) on the toxicity of GNSs to Gram positive Bacillus tropicus . Contrary to previous data, GNSs-HA exhibits greater toxicity compared to GNSs. Multi-omics combined with sensitive bioassays and electrochemical methods reveals GNSs disrupt oxidative phosphorylation by causing physical membrane damage. This leads to the accumulation of intracellular reactive oxygen species and inhibition of ATP production, subsequently suppressing synthetic and metabolic processes and ultimately causing bacterial death. Conversely, GNSs-HA directly extracts electrons from bacteria and oxidized biomolecules due to HA-improved electron transfer. This finding suggests that the HA corona does not always mitigate the toxicity of nanoparticles, thereby introducing uncertainty over the interaction between environmental corona and nanoparticles during ecological risk evaluation. Wide use of graphene nanosheets has raised concerns about the potential for environmental toxicity. Here, the authors explore the use of humic acid, traditionally considered to reduce graphene nanosheet toxicity, and show the coating has increased toxicity to model bacteria.