Sonochemically Covalent Functionalized Graphene Oxide Towards Photoluminescence and Nanocytotoxicity Activities

  • Gopal Avashthi
  • Shrikant S. Maktedar
  • Man SinghEmail author
Part of the Carbon Nanostructures book series (CARBON)


The greener mechanistic cavitation method has been applied for synthesis of graphene oxide (GrO) based functionalized materials. The GrO functionalization with various amine substituted heterocyclic moieties (ASHM) have an emerging technology towards biomedical processing of graphene. Hence, an ultrasound energy has been applied for GrO functionalization with 2-Amino-1,3,4-thidiazole (ATDZ) to synthesize Covalent functionalized product f-(ATDZ)GrO. Structural investigations have confirmed the covalent functionalization (CF) of GrO to synthesize f-(ATDZ)GrO. The structure of f-(ATDZ)GrO has confirmed with Fourier-transform infrared spectroscopy (FTIR), ultraviolet–visible spectroscopy (UV), RAMAN, X-ray diffraction (XRD), thermogravimetric analysis (TGA)/differential thermal analysis (DTA)/Differential thermal Gravimetry (DTG), Dynamic Light Scattering (DLS), high-resolution transmission electron microscopy (HRTEM), selected area electron diffraction (SAED), atomic force microscopy (AFM), scanning electron microscopy (SEM). The structural insights provide a mechanistic understanding of functional expression, through the contribution of atomic domains (CAD). TGA of f-(ATDZ)GrO validates total percentage weight loss of 95.5% at 198.17 °C. Thermal stability of f-(ATDZ)GrO as temperature aspects also certified an exothermic curve obtained with DTA. The calculated PL band gap of 3.87 eV in noncompatible f-(ATDZ)GrO is indicating towards biosensing applications. In extension of functionalization series of GrO with heterocyclic derivative, the cytotoxicity of f-(ATDZ)GrO has evaluated with Sulforhodamine B (SRB) assay to living cells, HaCaT and Vero cell lines. The average estimated cell viabilities have observed ~91.575% with HaCaT cell lines over a wide concentration range of 10–80 μg mL−1. The high cytocompatibility of f-(ATDZ)GrO has further extent with Vero cell lines of ~36.825% biocompatibility. However, the morphological effect on HaCaT cell line and some extinct significant with Vero have evidently confirmed that higher cytocompatibility of f-(ATDZ)GrO can be explore for the cytocompatibility as Nanotoxicity aspects. Therefore, f-(ATDZ)GrO appeared as an advanced material which can be further used for development of various biomedical applications.


Graphene oxide Covalent functionalization 2-Amino-1,3,4-thidiazole HaCaT and vero Cytocompatibility Photoluminescence activity 



Authors are thankful to Central University of Gujarat, India for support. Dr. Vasant Sathe, UGC-DAE CSR Indore, India is acknowledged for providing Raman facility. Dr. Jyoti A. Kode, ACTREC, Tata Memorial Centre, Mumbai, India is acknowledged for providing in vitro cytotoxicity screening facility.


  1. 1.
    Geim, A.K., Novoselov, K.S.: The rise of graphene. Nat. Mate. 6, 183–191 (2007)CrossRefGoogle Scholar
  2. 2.
    Georgakilas, V., Otyepka, M., Bourlinos, A.B., Chandra, V., Kim, N., Kemp, K.C.P., Zboril, R., Kim, K.S.: Functionalization of graphene: covalent and non-covalent approaches, derivatives and applications. Chem. Rev. 112, 6156–6214 (2012)CrossRefGoogle Scholar
  3. 3.
    Dai, L.: Functionalization of graphene for efficient energy conversion and storage. Acc. Chem. Res. 1, 31–42 (2013)CrossRefGoogle Scholar
  4. 4.
    Park, J., Yan, M.: Covalent functionalization of graphene with reactive intermediates. Acc. Chem. Res. 46, 181–189 (2013)CrossRefGoogle Scholar
  5. 5.
    Chng, E.L.K., Pumera, M.: The toxicity of graphene oxides: dependence on the oxidative methods used. Chem. Eur. J. 19, 8227–8235 (2013)CrossRefGoogle Scholar
  6. 6.
    Chng, E.L.K., Sofer, Z., Pumera, M.: Cytotoxicity profile of highly hydrogenated graphene. Chem. Eur. J. 20, 6366–6373 (2014)CrossRefGoogle Scholar
  7. 7.
    Pinto, A.M., Gonçalves, C., Sousa, D.M., Ferreira, A.R., Moreira, J.A., Gonçalves, I.C., Magalhaes, F.D.: Smaller particle size and higher oxidation improves biocompatibility of graphene-based materials. Carbon 99, 318–329 (2016)CrossRefGoogle Scholar
  8. 8.
    Kumar, A.M., Suresh, B., Ramakrishna, S., Kim, K.S.: Biocompatible responsive polypyrrole/GO nanocomposite coatings for biomedical applications. RSC Adv. 5, 99866–99874 (2015)CrossRefGoogle Scholar
  9. 9.
    Hasanzadeh, M., Mokhtari, F., Shadjou, N., Eftekhari, A., Mokhtarzadeh, A., Jouyban-Gharamaleki, V., Soltanali, M.: Poly arginine-graphene quantum dots as a biocompatible and non-toxic nanocomposite: layer-by-layer electrochemical preparation, characterization and non-invasive malondialdehyde sensory application in exhaled breath condensate. J. Mater. Sci. Eng. C 75, 247–258 (2017)CrossRefGoogle Scholar
  10. 10.
    Barua, S., Chattopadhyay, P., Phukan, M.M., Konwar, B.K., Islam, J., Karak, N.: Biocompatible hyperbranched epoxy/silver–reduced graphene oxide–curcumin nanocomposite as an advanced antimicrobial material. RSC Adv. 4, 47797–47805 (2014)CrossRefGoogle Scholar
  11. 11.
    Barahuiea, F., Saifullaha, B., Dorniania, D., Fakurazid, S., Karthivashand, G., Husseina, M.Z., Elfghi, F.M.: Graphene oxide as a nanocarrier for controlled release and targeted delivery of an anticancer active agent, chlorogenic acid. J. Mater. Sci. Eng. C 74, 177–185 (2017)CrossRefGoogle Scholar
  12. 12.
    Zhang, H., Grüner, G., Zhao, Y.: Recent advancements of graphene in biomedicine. J. Mater. Chem. B 1, 2542 (2013)CrossRefGoogle Scholar
  13. 13.
    Shi, S., Chen, F., Ehlerding, E.B., Cai, W.: Surface engineering of graphene-based nanomaterials for biomedical applications. Bioconjugate Chem. 25, 1609–1619 (2014)CrossRefGoogle Scholar
  14. 14.
    Pattnaik, S., Swain, K., Lin, Z.: Graphene and graphene-based nanocomposites: biomedical applications and biosafety. J. Mater. Chem. B 4, 7813–7831 (2016)CrossRefGoogle Scholar
  15. 15.
    Yousefi, M., Dadashpour, M., Hejazi, M., Hasanzadeh, M., Behnam, B., de la Guardia, M., Shadjou, N., Mokhtarzadeh, A.: Anti-bacterial activity of graphene oxide as a new weapon nanomaterial to combat multidrug-resistance bacteria. J. Mater. Sci. Eng. C 74, 568–581 (2017)CrossRefGoogle Scholar
  16. 16.
    Barua, S., Thakur, S., Aidew, L., Buragohain, A.K., Chattopadhyay, P., Karak, N.: One step preparation of a biocompatible, antimicrobial reduced graphene oxide–silver nanohybrid as a topical antimicrobial agent. RSC Adv. 4, 9777–9783 (2014)CrossRefGoogle Scholar
  17. 17.
    Kostarelos, K., Novoselov, K.S.: Materials science. exploring the interface of graphene and biology. Science 344, 261–263 (2014)CrossRefGoogle Scholar
  18. 18.
    An, J., Gou, Y., Yang, C., Hu, F., Wang, C.: Synthesis of a biocompatible gelatin functionalized graphene nanosheets and its application for drug delivery. J. Mater. Sci. Eng. C 33, 2827–2837 (2013)CrossRefGoogle Scholar
  19. 19.
    Mitra, T., Manna, P.J., Raja, S.T.K., Gnanamani, A., Kundu, P.P.: Curcumin loaded nano graphene oxide reinforced fish scale collagen—a 3D scaffold biomaterial for wound healing applications. RSC Adv. 5, 98653–98665 (2015)CrossRefGoogle Scholar
  20. 20.
    Zhou, L., Wang, W., Tang, J., Zhou, J.-H., Jiang, H.-J., Shen, J.: Graphene oxide noncovalent photosensitizer and its anticancer activity in vitro. Chem. Eur. J. 17, 12084–12091 (2011)CrossRefGoogle Scholar
  21. 21.
    Yang, Y., Zhang, Y.-M., Chen, Y., Zhao, D., Chen, J.-T., Liu, Y.: Construction of a graphene oxide based noncovalent multiple nanosupramolecular assembly as a scaffold for drug delivery. Chem. Eur. J. 18, 4208–4215 (2012)CrossRefGoogle Scholar
  22. 22.
    Gies, V., Zou, S.: Systematic toxicity investigation of graphene oxide: evaluation of assay selection, cell type, exposure period and flake size. Toxicol. Res. 7, 93–101 (2018)CrossRefGoogle Scholar
  23. 23.
    Pelin, M., Fusco, L., León, V., Martín, C., Criado, A., Sosa, S., Vázquez, E., Tubaro, A., Prato, M.: Differential cytotoxic effects of graphene and graphene oxide on skin Keratinocytes. Sci. Rep. 7, 40572-12 (2016)Google Scholar
  24. 24.
    Liao, K.-H., Lin, Y.-S., Macosko, C.W., Haynes, C.L.: Cytotoxicity of graphene oxide and graphene in human erythrocytes and skin fibroblasts. ACS Appl. Mater. Interfaces. 3, 2607–2615 (2011)CrossRefGoogle Scholar
  25. 25.
    Seabra, A.B., Paula, A.J., de Lima, R., Alves, O.L., Duran, N.: Nanotoxicity of graphene and graphene oxide. Chem. Res. Toxicol. 27, 159–168 (2014)CrossRefGoogle Scholar
  26. 26.
    Guo, X., Mei, N.: Assessment of the toxic potential of graphene family nanomaterials. J. Food Drug Anal. 22, 105–115 (2014)CrossRefGoogle Scholar
  27. 27.
    Kumar, S., Modak, D.M., Paik, P.: Graphene oxide for biomedical applications. J. Nanomed. Res. 5(6), 00136 (2017)Google Scholar
  28. 28.
    Dubey, P., Dr. Gopinath, P.: Functionalized graphene oxide based nanocarrier for tumor-targeted combination therapy to elicit enhanced cytotoxicity against breast cancer cells in vitro. Chem. Select. 1, 4845–4855 (2016)CrossRefGoogle Scholar
  29. 29.
    Li, Y., Feng, L., Shi, X., Wang, X., Yang, Y., Yang, K., Liu, T., Yang, G., Liu, Z.: Surface coating-dependent cytotoxicity and degradation of graphene derivatives: towards the design of non-toxic, degradable nano-graphene. Small 10, 1544–1554 (2014)CrossRefGoogle Scholar
  30. 30.
    Yang, K., Li, Y., Tan, X., Peng, R., Liu, Z.: Behavior and toxicity of graphene and its functionalized derivatives in biological systems. Small 9, 1492–1503 (2013)CrossRefGoogle Scholar
  31. 31.
    Peña-Bahamondea, J., Miguela, V.S., Nguyenb, H.N., Ozisikc, R., Rodrigues, D.F.: Functionalization of reduced graphene oxide with polysulfone brushes enhance antibacterial properties and reduce human cytotoxicity. Carbon 111, 258–268 (2017)CrossRefGoogle Scholar
  32. 32.
    Dong, H., Li, Y., Yu, J., Song, Y., Cai, X., Liu, J., Zhang, J., Ewing, R.C., Shi, D.: A versatile multicomponent assembly via β-cyclodextrin Host-guest chemistry on graphene for biomedical applications. Small 9, 446–456 (2013)CrossRefGoogle Scholar
  33. 33.
    Chena, J., Shia, X., Rena, L., Wanga, Y.: Graphene oxide/PVA inorganic/organic interpenetrating hydrogels with excellent mechanical properties and biocompatibility. Carbon 111, 18–27 (2017)CrossRefGoogle Scholar
  34. 34.
    Liu, Y., Zhang, Y., Zhang, T., Jiang, Y., Liu, X.: Synthesis, characterization and cytotoxicity of phosphorylcholine oligomer grafted graphene oxide. Carbon 71, 166–175 (2014)CrossRefGoogle Scholar
  35. 35.
    Bao, H., Pan, Y., Ping, Y., Sahoo, N.G., Wu, T., Li, L., Li, J., Gan, L.H.: Chitosan-functionalized graphene oxide as a nanocarrier for drug and gene delivery. Small 7, 1569–1578 (2011)CrossRefGoogle Scholar
  36. 36.
    Sayyar, S., Murray, E., Thompson, B.C., Gambhir, S., Officer, D.L., Wallace, G.G.: Covalently linked biocompatible graphene/polycaprolactone composites for tissue engineering. Carbon 52, 296–304 (2013)CrossRefGoogle Scholar
  37. 37.
    Yan, R., Wu, H., Zheng, Q., Wang, J., Huang, J., Ding, K., Guo, Q., Wang, J.: Graphene quantum dots cut from graphene flakes: high electrocatalytic activity for oxygen reduction and low cytotoxicity. RSC Adv. 4, 23097–23106 (2014)CrossRefGoogle Scholar
  38. 38.
    Maktedar, S.S., Mehetre, S.S., Singh, M., Kale, R.K.: Ultrasound irradiation: a robust approach for direct functionalization of graphene oxide with thermal and antimicrobial aspects. Ultrason. Sonochem. 21, 1407–1416 (2014)CrossRefGoogle Scholar
  39. 39.
    Maktedar, S.S., Avashthi, G., Singh, M.: Understanding the significance of O-doped graphene towards biomedical applications. RSC Adv. 6, 114264–114275 (2016)CrossRefGoogle Scholar
  40. 40.
    Mehetre, S.S., Maktedar, S.S., Singh, M.: Understanding the mechanism of surface modification through enhanced thermal and electrochemical stabilities of N-doped graphene oxide. Appl. Surf. Sci. 366, 514–522 (2016)CrossRefGoogle Scholar
  41. 41.
    Wang, T., Zhu, S., Jiang, X.: Toxicity mechanism of graphene oxide and nitrogen- doped graphene quantum dots in RBCs revealed by surface-enhanced infrared absorption spectroscopy. Toxicol. Res. 4, 885–894 (2015)CrossRefGoogle Scholar
  42. 42.
    Hirsch, A., Englert, J.M., Hauke, F.: Wet chemical functionalization of graphene. Acc. Chem. Res. 46, 87–96 (2013)CrossRefGoogle Scholar
  43. 43.
    James, E.J., Hersam, M.C.: Atomic covalent functionalization of graphene. Acc. Chem. Res. 46, 77–86 (2013)CrossRefGoogle Scholar
  44. 44.
    Hangxun, X., Kenneth, S.S.: Sonochemical preparation of functionalized graphenes. J. Am. Chem. Soc. 133, 9148–9151 (2011)CrossRefGoogle Scholar
  45. 45.
    Marcano, D.C., Kosynkin, D.V., Berlin, J.M., Sinitskii, A., Sun, Z., Slesarev, A., Alemany, L.B., Lu, W., Tour, J.M.: Improved synthesis of graphene oxide. ACS Nano 4, 4806–4814 (2010)CrossRefGoogle Scholar
  46. 46.
    Clauss, A., Plass, R., Boehm, H.P., Hofmann, U.: Untersuchungen zur structur des Graphitoxyds. Anorg. Allergy. Chem. 291, 205–220 (1957)CrossRefGoogle Scholar
  47. 47.
    Scholz, W., Boehm, H.P.: Untersuchungen am Graphitoxid. VI. Betrachtungen zur Struktur des Graphitoxids. Anorg. Allg. Chem. 369, 327–340 (1969)CrossRefGoogle Scholar
  48. 48.
    Kenry, T., Lim, C.T.: Biocompatibility and nanotoxicity of layered two-dimensional nanomaterials. Chem. Nano. Mat. 3, 5–16 (2017)Google Scholar
  49. 49.
    Majeeda, W., Bourdoa, S., Petiboneb, D.M., Sainia, V., Vanga, K.B., Nimaa, Z.A., Darriguesa, E., Ghosha, A., Watanabea, F., Cascianoa, D., Alid, S.F., Birisa, A.S.: The role of surface chemistry in the cytotoxicity profile of graphene. J. Appl. Toxicol. 37, 462–470 (2017)CrossRefGoogle Scholar
  50. 50.
    Crisana, L., Crisanb, B., Soritauc, O., Baciutb, M., Radu, Birisd A., Baciuta, G., Lucaciu, O.: In vitro study of biocompatibility of a graphene composite with gold nanoparticles and hydroxyapatite on human osteoblasts. J. Appl. Toxicol. 35, 1200–1210 (2015)CrossRefGoogle Scholar
  51. 51.
    Bitounis, D., Ali-Boucetta, H., Hong, B.H., Min, D.-H., Kostarelos, K.: Prospects and challenges of graphene in biomedical applications. Adv. Mater. 25, 2258–2268 (2013)CrossRefGoogle Scholar
  52. 52.
    Farshid, B., Lalwani, G., Sitharaman, B.: In vitro cytocompatibility of one-dimensional and two-dimensional nanostructure-reinforced biodegradable polymeric nanocomposites. J. Biomed. Mater. Res. A 103, 2309–2321 (2015)CrossRefGoogle Scholar
  53. 53.
    Maktedar, S.S., Mehetre, S.S., Avashthi, G., Singh, M.: In situ sonochemical reduction and direct functionalization of graphene oxide: a robust approach with thermal and biomedical applications. Ultrason. Sonochem. 34, 67–77 (2017)CrossRefGoogle Scholar
  54. 54.
    Maktedar, S.S., Avashthi, G., Singh, M.: Ultrasound assisted simultaneous reduction and direct functionalization of graphene oxide with thermal and cytotoxicity profile. Ultrason. Sonochem. 34, 856–864 (2017)CrossRefGoogle Scholar
  55. 55.
    Maktedar, S.S., Malik, P., Avashthi, G., Singh, M.: Dispersion enhancing effect of sonochemically functionalized graphene oxide for catalysing antioxidant efficacy of curcumin. Ultrason. Sonochem. 39, 208–217 (2017)CrossRefGoogle Scholar

Copyright information

© Springer Nature Singapore Pte Ltd. 2019

Authors and Affiliations

  • Gopal Avashthi
    • 1
  • Shrikant S. Maktedar
    • 1
  • Man Singh
    • 1
    Email author
  1. 1.School of Chemical SciencesCentral University of GujaratGandhinagarIndia

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