Elsevier

FlatChem

Volume 26, March 2021, 100231
FlatChem

Oxidation degree or sheet size: What really matters for the photothermal effect and ecotoxicity of graphene oxide?

https://doi.org/10.1016/j.flatc.2021.100231Get rights and content

Highlights

  • A straight correlation exists between photothermal effect and ecotoxicity of GO.

  • The photothermal effect is stronger in more oxidized GO.

  • Aqueous suspensions of carboxylated GO samples can reach 80 °C under NIR.

  • GO sheets larger than 200 nm delay hatching of zebrafish embryos.

  • GO ecotoxicity is more physical than chemical.

Abstract

A set of thirteen GO samples obtained following different methodologies and described by eight structural parameters – hydrodynamic diameter (658 nm–142 nm), size polydispersity index, optical absorption coefficient, lambda maximum, D/G peak intensity ratio, crystallite size, distance between defects, and pH – was submitted to NIR 808 nm irradiation (experiment 1) and fish embryo toxicity (FET) test performed with zebrafish embryos (experiment 2). Principal component analysis (PCA) applied to the obtained data shows that the optical absorption coefficient and lambda maximum, which are strongly dependent on the GO oxidation degree and presence of carboxylic acid groups, are the most relevant features for the photothermal effect. Conversely, size is the predominant parameter for the ecotoxicity. Only samples exhibiting the smallest sheet size (below 200 nm) do not interfere either on the hatching time or the rate of alive hatched individuals. GO samples composed of larger sheets, regardless their oxidation and carboxylation degrees, impart negative effects to the embryo’s development stages, including hatching delay, death of embryos and abnormalities on hatched individuals. It is therefore concluded that the photothermal effect is more dependent on the oxidation degree, whereas ecotoxicity is more dependent on the size of GO sheets.

Introduction

In recent years, graphene oxide (GO), the oxidized version of graphene, has become an independent field of research owing to its unique structural features, including the presence of several types of oxygenated functional groups that ensures water dispersibility and long-term colloidal stability, besides scalable synthesis at much lower cost than other two-dimensional nanomaterials [1], [2], [3]. Indeed, the presence of oxygenated groups has made it possible to show photoluminescence [4] anchoring harbor for the attachment of specific drugs and photosensitizers [5] photothermal [6] and electrochemical activities [7] all of them easily tunable by means of controlled chemical processes. Finally, functionalized GO has proven to be potentially promising for biomedical applications, including cell imaging [8] photothermal and photodynamic therapies [9] gene therapy [10] antibacteria substrates [11] scaffolds for mesenchymal cells [12] and many others [13], [14], [15], [16].

Despite the multiple purposes and technological potentialities, the large-scale production of GO samples with controlled size and molecular structure is still highly challenging. For example, the actual commercial availability of high-purity, single- and few-layer graphene and GO has been highly criticized [17]. Moreover, the extensive usage of GO in different applications will be inevitably accompanied by the production of a large amount of waste. Thus, toxicity of the GO waste needs to be properly addressed in order to mitigate its eventual occupational and environmental impacts.

The photothermal effect consists on the conversion of luminous energy into heat [18]. Depending upon the absorbed wavelength, for example near infrared (NIR), it can be harnessed in photothermal therapies of different types of malign tumors [19]. Since this wavelength is sufficient to penetrate into tissues, whereas water and hemoglobin are transparent to it, the therapy can be carried out while displaying minimum side effects. The photothermal effect can be also extended to kill antibiotic-resistant bacteria, as the generated heat causes membranes rupture, protein denaturation and irreversible bacterial destruction [20]. The effects of GO structure (reduction degree or functionalization) and size on its photothermal effect, photothermal therapy, protein adsorption, drug loading, UV absorption and natural toxicity to bacteria have been extensively investigated [21], [22], [23], [24].

It has been argued that GO absorbs more effectively NIR wavelengths whenever the lateral size of its individual sheets is below sub-micrometer ranges, forming the so-called nano-GO. In order to reach such small sizes by means of top-down approaches, graphitic oxide and/or GO suspensions are submitted to extensive ultrasonication [25]. This process can also be conducted in the presence of chemical agents, for example chloroacetic acid/sodium hydroxide [8] hypochlorite [26] Fenton reagent [27] or even assisted by microwave irradiation [28]. Nonetheless, the sheet size and molecular structure of the resulting nano-GO is dependent on several reaction pathways induced by the ultrasound cavitation [25]. Furthermore, sonotrodes may undergo extensive erosion when employed for several hours, which affects the power delivered to the precursor suspension in the course of the ultrasonication process. Consequently, the preparation of nano-GO samples with reproducible molecular structures is still quite difficult. In addition, the photothermal effect appears to be dependent not only on the sheet size but also on the molecular structure, for example, the type and distribution of oxygenated groups at basal plane and edge of the sheets [29].

The cyto- and ecotoxicity of GO and nano-GO probed by means of in-vitro/in-vivo tests are focus of intensive research nowadays. Whereas several papers in the literature have shown that, at low doses, GO and nano-GO exhibit relatively low toxicity to cells and tissues [30] a similar number of publications have conversely shown that these carbon-based nanomaterials represent a serious threat to living aquatic resources [31], [32], [33]. Indeed, several structural factors, including lateral size, surface structure, functionalization, charge, impurities, aggregation, and corona effect, may determine the toxicity of these nanomaterials, making it the analysis even more difficult [30].

In this regard, the present contribution aims at answering whether size or molecular structure, herein represented by the oxidation degree, is the most relevant feature for the photothermal effect and ecotoxicity of GO. As the large-scale production of GO and its subsequent use in photothermal conversion should increase its occurrence in the aquatic environment, both properties should be, somehow, highly correlated. For that purpose, a set of thirteen GO samples obtained following different methodologies and described by eight structural parameters was submitted to NIR 808 nm irradiation (experiment 1) and fish embryo toxicity (FET) test performed with zebrafish embryos (experiment 2). The structure of all samples was evaluated by attenuated total reflectance Fourier transform infrared (ATR-FTIR), UV–Vis absorption and micro Raman spectroscopies, and CHN elemental analysis. The sizes of GO sheets were determined by dynamic light scattering (DLS) and confirmed further by atomic force microscopy (AFM). Since many structural variables have been gathered, the data were processed by principal component analysis (PCA), which permitted us to evaluate the effective contribution of each structural variable to the targeted properties, namely, photothermal effect and ecotoxicity.

Section snippets

Sample preparation

Graphite powder (particle size <0.45 μm), potassium permanganate 98%, sulfuric acid 98%, nitric acid 67%, sodium nitrate, hydrochloric acid 37%, phosphorous pentoxide 98%, ammonium persulfate 98%, and hydrogen peroxide 30% were all purchased from Sigma-Aldrich (USA) and used as received. Monochloroacetic acid and sodium hydroxide were purchased from Sigma-Vetec (Brazil) and used without any further purification. Double distilled and ultrapure water (resistivity: 18 MΩ cm) were used throughout

Structural features of GrO and GO samples

In order to answer the main question of this study, whether size or molecular structure is the most relevant feature for the photothermal effect and ecotoxicity, GO samples had to be prepared in different sizes and molecular structures. The way we found to do that was to start with GrO produced under different oxidation procedures. This approach was quite satisfactory, since it allowed us the production of thirteen samples that could be described by eight different structural parameters –

Conclusions

The present study has demonstrated that the two most important structural features of GO, namely oxidation degree and size of sheets, work in opposite directions for the photothermal effect and ecotoxicity. By one hand, the oxidation degree and the presence of carboxylic acid groups are more important than the size of sheets for GO suspensions achieving the highest temperature under NIR irradiation. This is because the presence of oxygenated groups creates mid-gap states in the GO electronic

CRediT authorship contribution statement

Caio C. C. Moreira: Conceptualization, Data curation, Formal analysis, Investigation, Methodology. Ítalo A. Costa: Investigation, Methodology, Formal analysis. Diego S. Moura: Conceptualization, Methodology, Validation, Formal analysis, Writing – original draft, Writing – review and editing. Cesar K. Grisolia: Conceptualization, Methodology, Validation, Formal analysis, Writing – original draft, Writing – review and editing. Carlos A. E. M. Leite: Investigation, Methodology, Formal analysis.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

The financial support of Brazilian funding agencies Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação de Apoio à Pesquisa do Distrito Federal (FAPDF) (process n. 0193.001630/2017), and Instituto Nacional de Ciência e Tecnologia em Bioanalítica (INCTBio) (process no. 5736672/2008-3) are greatly acknowledged. This study was also financed by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES) – Finance Code 001.

References (54)

  • K.E. Pelka et al.

    Size does matter – determination of the critical molecular size for the uptake of chemicals across the chorion of zebrafish (Danio rerio) embryos

    Aquat. Toxicol.

    (2017)
  • D.R. Dreyer et al.

    The chemistry of graphene oxide

    Chem. Soc. Rev.

    (2010)
  • O.C. Compton et al.

    Graphene oxide, highly reduced graphene oxide, and graphene: versatile building blocks for carbon-based materials

    Small

    (2010)
  • D.R. Dreyer et al.

    Harnessing the chemistry of graphene oxide

    Chem. Soc. Rev.

    (2014)
  • Z. Luo et al.

    Photoluminescence and band gap modulation in graphene oxide

    Appl. Phys. Lett.

    (2009)
  • B. Tian et al.

    Photothermally enhanced photodynamic therapy delivered by nano-graphene oxide

    ACS Nano

    (2011)
  • M.C. Wu et al.

    Graphene-based photothermal agent for rapid and effective killing of bacteria

    ACS Nano

    (2013)
  • M. Pumera

    Graphene-based nanomaterials and their electrochemistry

    Chem. Soc. Rev.

    (2010)
  • X. Sun et al.

    Nano-graphene oxide for cellular imaging and drug delivery

    Nano Res.

    (2008)
  • L.Z. Feng et al.

    Graphene based gene transfection

    Nanoscale

    (2011)
  • O. Akhavan et al.

    Toxicity of graphene and graphene oxide nanowalls against bacteria

    ACS Nano

    (2010)
  • O.N. Ruiz et al.

    Graphene oxide: a nonspecific enhancer of cellular growth

    ACS Nano

    (2011)
  • K. Yang et al.

    Graphene in mice: ultrahigh in vivo tumor uptake and efficient photothermal therapy

    Nano Lett.

    (2010)
  • L.M. Zhang et al.

    Functional graphene oxide as a nanocarrier for controlled loading and targeted delivery of mixed anticancer drugs

    Small

    (2010)
  • V.K. Rana et al.

    synthesis and drug-delivery behavior of chitosan-functionalized graphene oxide hybrid nanosheets

    Macromol. Mater. Eng.

    (2011)
  • J. Lee et al.

    Laser desorption/ionization mass spectrometric assay for phospholipase activity based on graphene oxide/carbon nanotube double-layer films

    J. Am. Chem. Soc.

    (2010)
  • A.P. Kauling et al.

    The worldwide graphene flake production

    Adv. Mater.

    (2018)
  • Cited by (2)

    • Applicability of OECD TG 201, 202, 203 for the aquatic toxicity testing and assessment of 2D Graphene material nanoforms to meet regulatory needs

      2023, NanoImpact
      Citation Excerpt :

      Toxicological effects have also been evidenced to vary as a function of these distinct properties or their alterations (Liu et al., 2012; Sydlik et al., 2015; Jiang et al., 2021; Lu et al., 2017; Chen et al., 2021). For example lateral sheet size seemed to influence adverse effects towards zebrafish embryos in a comparative study, with larger sheets (>200–600 nm) showing increased toxicity through physical interactions and smaller sheets (<200 nm) not showing any effects (Moreira et al., 2021). In Table 8 all relevant material properties, both inherent and system-dependent, are presented building on work by Wick et al. (2014), considering (i) information requirements to fulfil reporting obligations by REACH (Annex VI) (EC, 2006), (ii) particle properties that are relevant for ecotoxicological hazard assessment in the aquatic environment, and (iii) those defined by the graphene classification framework Graphene Flagship and IEC/ISO.

    1

    ORCID id: 0000-0001-7716-0318.

    View full text