Relevance of Physicochemical Characterization of Nanomaterials for Understanding Nano-cellular Interactions

  • Henriqueta LouroEmail author
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 1048)


The manufactured nanomaterials (NMs) have specific physicochemical properties that confer unique mechanical, optical, electrical and magnetic characteristics that are beneficial for biomedical and industrial applications. However, recent studies have suggested that such specific physicochemical properties of the NMs may define nano-bio interactions thereby determining their toxic potential.

One of the major concerns about NMs is the potential to induce cancer, suggested by some experimental studies, as seen for titanium dioxide nanomaterials or carbon nanotubes. To analyze in a short term the carcinogenic properties of a compound, genotoxicity assays in mammalian cell lines or animal models are frequently used. However, the investigation of the genotoxic properties of NMs has been inconclusive, up to date, since divergent results have been reported throughout the literature. While trying to understand how the NMs’ characteristics may encompass increased toxicological effects that harbor uncertainties for public health, the use of correlation analysis highlights some physicochemical properties that influence the genotoxic potential of these NM.

In this chapter, it is hypothesized that the different genotoxicity observed in closely related NMs may be due to subtle differences in their physicochemical characteristics. The present work provides an overview of the studies exploring the correlation between physicochemical properties of nanomaterials and their genotoxic effects in human cells, with focus on the toxicity of two groups of NMs, titanium dioxide nanomaterials and multiwalled-carbon nanotubes. It is suggested that, for tackling NMs’ uncertainties, the in-depth investigation of the nano-bio interactions must be foreseen, where in vitro research must be integrated with in vivo and biomonitoring approaches, to cope with the complex dynamic behaviour of nanoscale materials.


Nanomaterials Public health Genotoxicity Cytotoxicity Titanium dioxide Multiwalled carbon nanotubes Physicochemical properties 



The author wishes to thank Dr. Maria João Silva and all the team from the Genetic Toxicology Laboratory (INSA, Portugal). HL acknowledges the support of the partners of the Nanogenotox Joint Action (Health Programme under Grant Agreement no. 2009 21), NANOREG Project (A common European approach to the regulatory testing of nanomaterials, Grant Agreement 310584) and Dr. José Catita (Paralab, Portugal) for the help with the DLS analysis.


  1. 1.
    IARC – International Agency for Research on Cancer (2010) Volume 93: carbon black, titanium dioxide and talc. In: World Health Organization (ed) IARC monographs on the evaluation of carcinogenic risks to humans. World Health Organization, LyonGoogle Scholar
  2. 2.
    NIOSH - National Institute for Occupational Safety and Health. Department Health and Human Services. Centers for Disease Control and Prevention. 2011. Current intelligence bulletin 63: occupational exposure to titanium dioxide. Available at:
  3. 3.
    Sycheva LP, Zhurkov VS, Iurchenko VV et al (2011) Investigation of genotoxic and cytotoxic effects of micro- and nanosized titanium dioxide in six organs of mice in vivo. Mutat Res 726:8–14CrossRefPubMedGoogle Scholar
  4. 4.
    SCCS - Scientific Committee on Consumer Safety (2013) Opinion on titanium dioxide (nano form). COLIPA n° S75Google Scholar
  5. 5.
    Grosse Y, Loomis D, Guyton KZ et al (2014) Carcinogenicity of fluoro-edenite, silicon carbide fibres and whiskers, and carbon nanotubes. Lancet Oncol 15:1427–1428CrossRefPubMedGoogle Scholar
  6. 6.
    Cveticanin J, Joksic G, Leskovac A et al (2010) Using carbon nanotubes to induce micronuclei and double strand breaks of the DNA in human cells. Nanotechnology 21:015102CrossRefPubMedGoogle Scholar
  7. 7.
    Szendi K, Varga C (2008) Lack of genotoxicity of carbon nanotubes in a pilot study. Anticancer Res 28:349–352PubMedGoogle Scholar
  8. 8.
    Donaldson K, Murphy FA, Duffin R et al (2010) Asbestos, carbon nanotubes and the pleural mesothelium: a review of the hypothesis regarding the role of long fibre retention in the parietal pleura, inflammation and mesothelioma. Part Fibre Toxicol 7:5CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Rittinghausen S, Hackbarth A, Creutzenberg O et al (2014) The carcinogenic effect of various multi-walled carbon nanotubes (MWCNTs) after intraperitoneal injection in rats. Part Fibre Toxicol 11:59CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Pelaz B, Charron G, Pfeiffer C et al (2012) Interfacing engineered nanoparticles with biological systems: anticipating adverse nano-bio interactions. Small 9(9–10):1573–1584PubMedGoogle Scholar
  11. 11.
    Tavares AM, Louro H, Antunes S et al (2014) Genotoxicity evaluation of nanosized titanium dioxide, synthetic amorphous silica and multi-walled carbon nanotubes in human lymphocytes. Toxicol In Vitro 28(1):60–69CrossRefPubMedGoogle Scholar
  12. 12.
    Lynch I, Weiss C, Valsami-Jones E (2014) A strategy for grouping of nanomaterials based on key physico-chemical descriptors as a basis for safer-by-design NMs. Nano Today 9(3):266–270CrossRefGoogle Scholar
  13. 13.
    Fadeel B, Fornara A, Toprak MS et al (2015) Keeping it real: the importance of material characterization in nanotoxicology. Biochem Biophys Res Commun 468(3):498–503CrossRefPubMedGoogle Scholar
  14. 14.
    Maynard AD (2016) Navigating the risk landscape. Nat Nanotechnol 11(3):211–212CrossRefPubMedGoogle Scholar
  15. 15.
    Rasmussen K, De Temmerman PI, Verleysen E et al (2014) Titanium dioxide, NM-100, NM-101, NM-102, NM-103, NM-104, NM-105: characterisation and physicoChemical properties. NM-series of Representative Manufactured Nanomaterials. JRC, Joint Research Centre. Available at:
  16. 16.
    Rasmussen K, Mast J, De Temmerman P-J et al (2014). Multi-walled carbon nanotubes, NM-400, N,M-401, NM-402, NM-403: characterisation and physico-chemical properties. JRC Repository: NM-series of Representative Manufactured Nanomaterials European Commission- Joint Research Centre, Institute for Health and Consumer Protection. Available at:
  17. 17.
    Oberdorster G (2010) Safety assessment for nanotechnology and nanomedicine: concepts of nanotoxicology. J Intern Med 267(1):89–105CrossRefPubMedGoogle Scholar
  18. 18.
    Chen R, Riviere JE (2017) Biological surface adsorption index of nanomaterials: modelling surface interactions of nanomaterials with biomolecules. Adv Exp Med Biol 947:207–253CrossRefPubMedGoogle Scholar
  19. 19.
    Monopoli MP, Aberg C, Salvati A et al (2012) Biomolecular coronas provide the biological identity of nanosized materials. Nat Nanotechnol 7(12):779–786CrossRefPubMedGoogle Scholar
  20. 20.
    Hussain SM, Warheit DB, Ng SP et al (2015) At the crossroads of nanotoxicology in vitro: past achievements and current challenges. Toxicol Sci 147:5–16CrossRefPubMedGoogle Scholar
  21. 21.
    Atluri R, Jensen KA (2017) Engineered nanomaterials: their physicochemical characteristics and how to measure them. Adv Exp Med Biol 947:3–23CrossRefPubMedGoogle Scholar
  22. 22.
    Ren G, Hu D, Cheng EW et al (2009) Characterisation of copper oxide nanoparticles for antimicrobial applications. Int J Antimicrob Agents 33(6):587–590CrossRefPubMedGoogle Scholar
  23. 23.
    Seoane JR, Llovet X (2012) Handbook of instrumental techniques for materials, chemical and biosciences research. Centres Científics i Tecnològics, Barcelona, Spain, BarcelonaGoogle Scholar
  24. 24.
    Jensen KA, Kembouche Y, Christiansen E et al (2011) The generic NANOGENOTOX dispersion protocol – Standard Operation Procedure (SOP). Available at:
  25. 25.
    OECD – Organisation for Economic Co-operation and Development (2010) OECD guideline for the testing of chemicals – in vitro mammalian cell micronucleus test, Test Guideline 487. OECD, ParisGoogle Scholar
  26. 26.
    Magdolenova Z, Lorenzo Y, Collins A et al (2012) Can standard genotoxicity tests be applied to nanoparticles? J Toxicol Environ Health A 75:800–806CrossRefPubMedGoogle Scholar
  27. 27.
    Gonzalez L, Sanderson BJ, Kirsch-Volders M (2011) Adaptations of the in vitro MN assay for the genotoxicity assessment of nanomaterials. Mutagenesis 26(1):185–191CrossRefPubMedGoogle Scholar
  28. 28.
    Louro H, Pinhão M, Santos J et al (2016) Evaluation of the cytotoxic and genotoxic effects of benchmark multi-walled carbon nanotubes in relation to their physicochemical properties. Toxicol Lett 262:123–134CrossRefPubMedGoogle Scholar
  29. 29.
    JRC – Joint Research Center (2011) Impact of engineered nanomaterials on health: considerations for benefit-risk assessment. EASAC Policy Report - JRC Reference Report. IspraGoogle Scholar
  30. 30.
    Osman IF, Baumgartner A, Cemeli E et al (2010) Genotoxicity and cytotoxicity of zinc oxide and titanium dioxide in HEp-2 cells. Nanomedicine 5:1193–1203CrossRefPubMedGoogle Scholar
  31. 31.
    Shukla RK, Sharma V, Pandey AK et al (2011) ROS-mediated genotoxicity induced by titanium dioxide nanoparticles in human epidermal cells. Toxicol In Vitro 25:231–241CrossRefPubMedGoogle Scholar
  32. 32.
    Uboldi C, Urban P, Gilliland D et al (2016) Role of the crystalline form of titanium dioxide nanoparticles: rutile, and not anatase, induces toxic effects in Balb/3T3 mouse fibroblasts. Toxicol In Vitro 31:137–145CrossRefPubMedGoogle Scholar
  33. 33.
    Di Bucchianico S, Cappellini F, Le Bihanic F et al (2017) Genotoxicity of TiO2 nanoparticles assessed by mini-gel comet assay and micronucleus scoring with flow cytometry. Mutagenesis 32(1):127–137CrossRefPubMedGoogle Scholar
  34. 34.
    Magdolenova Z, Collins A, Kumar A et al (2014) Mechanisms of genotoxicity. A review of in vitro and in vivo studies with engineered nanoparticles. Nanotoxicology 8(3):233–278CrossRefPubMedGoogle Scholar
  35. 35.
    Magdolenova Z, Bilanicova D, Pojana G et al (2012) Impact of agglomeration and different dispersions of titanium dioxide nanoparticles on the human related in vitro cytotoxicity and genotoxicity. J Environ Monit 14(2):455–464CrossRefPubMedGoogle Scholar
  36. 36.
    Nagai H, Okazaki Y, Chew SH et al (2011) Diameter and rigidity of multiwalled carbon nanotubes are critical factors in mesothelial injury and carcinogenesis. Proc Natl Acad Sci U S A 108:E1330–E1338CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Takagi A, Hirose A, Nishimura T et al (2008) Induction of mesothelioma in p53+/− mouse by intraperitoneal application of multi-wall carbon nanotube. J Toxicol Sci 33:105–116CrossRefPubMedGoogle Scholar
  38. 38.
    Takagi A, Hirose A, Futakuchi M et al (2012) Dose-dependent mesothelioma induction by intraperitoneal administration of multi-wall carbon nanotubes in p53 heterozygous mice. Cancer Sci 103:1440–1444CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Sakamoto Y, Nakae D, Fukumori N et al (2009) Induction of mesothelioma by a single intrascrotal administration of multi-wall carbon nanotube in intact male Fischer 344 rats. J Toxicol Sci 34:65–76CrossRefPubMedGoogle Scholar
  40. 40.
    Jackson P, Kling K, Jensen KA et al (2015) Characterization of genotoxic response to 15 multiwalled carbon nanotubes with variable physicochemical properties including surface functionalizations in the FE1-Muta(TM) mouse lung epithelial cell line. Environ Mol Mutagen 56(2):183–203CrossRefPubMedGoogle Scholar
  41. 41.
    Lindberg HK, Falck GCM, Suhonen S et al (2009) Genotoxicity of nanomaterials: DNA damage and micronuclei induced by carbon nanotubes and graphite nanofibres in human bronchial epithelial cells in vitro. Toxicol Lett 186(3):166–173CrossRefPubMedGoogle Scholar
  42. 42.
    Atsuta J, Sterbinsky SA, Plitt J et al (1997) Phenotyping and cytokine regulation of the BEAS-2B human bronchial epithelial cell: demonstration of inducible expression of the adhesion molecules VCAM-1 and ICAM-1. Am J Respir Cell Mol Biol 17(5):571–582CrossRefPubMedGoogle Scholar
  43. 43.
    Foster KA, Oster C, Mayer M et al (1998) Characterization of the A549 cell line as a type II pulmonary epithelial cell model for drug metabolism. Exp Cell Res 243(2):359–366CrossRefPubMedGoogle Scholar
  44. 44.
    Haniu H, Saito N, Matsuda Y, Tsukahara T et al (2013) Culture medium type affects endocytosis of multi-walled carbon nanotubes in BEAS-2B cells and subsequent biological response. Toxicol In Vitro 27(6):1679–1685CrossRefPubMedGoogle Scholar
  45. 45.
    Asakura M, Sasaki T, Sugiyama T et al (2010) Genotoxicity and cytotoxicity of multi-wall carbon nanotubes in cultured Chinese hamster lung cells in comparison with chrysotile fibers. J Occup Health 52:155–166CrossRefPubMedGoogle Scholar
  46. 46.
    Migliore L, Saracino D, Bonelli A et al (2010) Carbon nanotubes induce oxidative DNA damage in RAW 264.7 cells. Environ Mol Mutagen 51:294–303PubMedGoogle Scholar
  47. 47.
    Catalan J, Siivola KM, Nymark P et al (2016) In vitro and in vivo genotoxic effects of straight versus tangled multi-walled carbon nanotubes. Nanotoxicology 10:794–806CrossRefPubMedGoogle Scholar
  48. 48.
    Stearns RC, Paulauskis JD, Godleski JJ (2001) Endocytosis of ultrafine particles by A549 cells. Am J Respir Cell Mol Biol 24(2):108–115CrossRefPubMedGoogle Scholar
  49. 49.
    Tabet L, Bussy C, Amara N et al (2009) Adverse effects of industrial multiwalled carbon nanotubes on human pulmonary cells. J Toxicol Environ Health A 72(2):60–73CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Maruyama K, Haniu H, Saito N et al (2015) Endocytosis of multiwalled carbon nanotubes in bronchial epithelial and mesothelial cells. Biomed Res Int 2015:793186. CrossRefPubMedPubMedCentralGoogle Scholar
  51. 51.
    Louro H, Silva MJ (2011) Cost/benefit of mutation induction under PARP1 deficiency: from genomic instability to therapy. In: Urbano KV (ed) Advances in genetics research. Nova Science Publishers, New York, pp 109–134Google Scholar
  52. 52.
    Louro H, Silva MJ (2010) In vivo mutagenic effects of alkylating agents eliciting different DNA-adducts. In: Emerson A, Cunha R (eds) DNA adducts: formation, detection and mutagenesis. Nova Science Publishers, New York, pp 39–60Google Scholar
  53. 53.
    ICH- International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (2012) S2(R1) genotoxicity testing and data interpretation for pharmaceuticals intended for human use. Food and Drug Administration, HHS. (Ed)Google Scholar
  54. 54.
    OECD – Organisation for Economic Co-operation and Development (2011) OECD guidelines for the testing of chemicals: transgenic rodent somatic and germ cell gene mutation assays, Test Guideline 488. OECD, ParisGoogle Scholar
  55. 55.
    Louro H (2013) Nanomateriais manufaturados: avaliação de segurança através da caracterização dos seus efeitos genéticos. Ph.D. Dissertation, Nova University of LisbonGoogle Scholar
  56. 56.
    Louro H, Pinto M, Vital N et al (2014) The LacZ plasmid-based transgenic mouse model: an integrative approach to study the genotoxicity of nanomaterials. In: Sierra LM, Gaivão I (eds) Genotoxicity and DNA repair – a practical approach. Methods in pharmacology and toxicology. Humana Press, Springer, New York, pp 451–477Google Scholar
  57. 57.
    Louro H, Tavares A, Vital N et al (2014) Integrated approach to the in vivo genotoxic effects of a titanium dioxide nanomaterial using LacZ plasmid-based transgenic mice. Environ Mol Mutagen 55(6):500–509CrossRefPubMedGoogle Scholar
  58. 58.
    Tice RR, Agurell E, Anderson D et al (2000) Single cell gel/comet assay: Guidelines for in vitro and in vivo genetic toxicology testing. Environ Mol Mutagen 35(3):206–221CrossRefPubMedGoogle Scholar
  59. 59.
    Bruno ME, Tasat DR, Ramos E et al (2013) Impact through time of different sized titanium dioxide particles on biochemical and histopathological parameters. J Biomed Mater Res A 102(5):1439–1448CrossRefPubMedGoogle Scholar
  60. 60.
    Olmedo DG, Tasat DR, Evelson P et al (2008) Biological response of tissues with macrophagic activity to titanium dioxide. J Biomed Mater Res A 84(4):1087–1093CrossRefPubMedGoogle Scholar
  61. 61.
    Sadauskas E, Wallin H, Stoltenberg M et al (2007) Kupffer cells are central in the removal of nanoparticles from the organism. Part Fibre Toxicol 4:10CrossRefPubMedPubMedCentralGoogle Scholar
  62. 62.
    Oh N, Park JH (2014) Endocytosis and exocytosis of nanoparticles in mammalian cells. Int J Nanomedicine 9(Suppl 1):51–63PubMedPubMedCentralGoogle Scholar
  63. 63.
    Bhattacharya K, Davoren M, Boertz J et al (2009) Titanium dioxide nanoparticles induce oxidative stress and DNA-adduct formation but not DNA-breakage in human lung cells. Part Fibre Toxicol 2009:6–17Google Scholar
  64. 64.
    Singh N, Manshian B, Jenkins GJ et al (2009) NanoGenotoxicology: the DNA damaging potential of engineered nanomaterials. Biomaterials 30(23–24):3891–3914CrossRefPubMedGoogle Scholar
  65. 65.
    Naya M, Kobayashi N, Ema M, Kasamoto S, Fukumuro M, Takami S, Nakajima M, Hayashi M, Nakanishi J (2012) In vivo genotoxicity study of titanium dioxide nanoparticles using comet assay following intratracheal instillation in rats. Regul Toxicol Pharmacol 62:1–6CrossRefPubMedGoogle Scholar
  66. 66.
    Landsiedel R, Ma-Hock L, Van Ravenzwaay B et al (2010) Gene toxicity studies on titanium dioxide and zinc oxide nanomaterials used for UV-protection in cosmetic formulations. Nanotoxicology 4:364–381CrossRefPubMedGoogle Scholar
  67. 67.
    Suzuki T, Miura N, Hojo R et al (2016) Genotoxicity assessment of intravenously injected titanium dioxide nanoparticles in gpt delta transgenic mice. Mutat Res 802:30–37CrossRefGoogle Scholar
  68. 68.
    Li Y, Yan J, Ding W et al (2017) Genotoxicity and gene expression analyses of liver and lung tissues of mice treated with titanium dioxide nanoparticles. Mutagenesis 32:33–46CrossRefPubMedGoogle Scholar
  69. 69.
    Chen Z, Wang Y, Ba T et al (2014) Genotoxic evaluation of titanium dioxide nanoparticles in vivo and in vitro. Toxicol Lett 226:314–319CrossRefPubMedGoogle Scholar
  70. 70.
    Joris F, Manshian BB, Peynshaert K et al (2013) Assessing nanoparticle toxicity in cell-based assays: influence of cell culture parameters and optimized models for bridging the in vitro-in vivo gap. Chem Soc Rev 42(21):8339–8359CrossRefPubMedGoogle Scholar
  71. 71.
    De Boo J, Hendriksen C (2005) Reduction strategies in animal research: a review of scientific approaches at the intra-experimental, supra-experimental and extra-experimental levels. Altern Lab Anim 33(4):369–377PubMedGoogle Scholar
  72. 72.
    Bello D, Hart AJ, Ahn K et al (2008) Particle exposure levels during CVD growth and subsequent handling of vertically-aligned carbon nanotube films. Carbon 46(6):974–977CrossRefGoogle Scholar
  73. 73.
    Bello D, Wardle B, Yamamoto N et al (2009) Exposure to nanoscale particles and fibers during machining of hybrid advanced composites containing carbon nanotubes. J Nanopart Res 11(1):231–249CrossRefGoogle Scholar
  74. 74.
    Han JH, Lee EJ, Lee JH et al (2008) Monitoring multiwalled carbon nanotube exposure in carbon nanotube research facility. Inhal Toxicol 20(8):741–749CrossRefPubMedGoogle Scholar
  75. 75.
    Lee JH, Lee S-B, Bae GN et al (2010) Exposure assessment of carbon nanotube manufacturing workplaces. Inhal Toxicol, 2010 22(5):369–381CrossRefGoogle Scholar
  76. 76.
    Tsai SJ, Hofmann M, Hallock M et al (2009) Characterization and evaluation of nanoparticle release during the synthesis of single-walled and multiwalled carbon nanotubes by chemical vapor deposition. Environ Sci Technol 43(15):6017–6023CrossRefPubMedGoogle Scholar
  77. 77.
    Pelclova D, Barosova H, Kukutschova J et al (2015) Raman microspectroscopy of exhaled breath condensate and urine in workers exposed to fine and nano TiO2 particles: a cross-sectional study. J Breath Res 9(3):036008CrossRefPubMedGoogle Scholar
  78. 78.
    Pelclova D, Zdimal V, Kacer P et al (2016) Oxidative stress markers are elevated in exhaled breath condensate of workers exposed to nanoparticles during iron oxide pigment production. J Breath Res 10(1):016004CrossRefPubMedGoogle Scholar
  79. 79.
    Gonzalez L, Kirsch-Volders M (2016) Biomonitoring of genotoxic effects for human exposure to nanomaterials: the challenge ahead. Mutat Res 768:14–26CrossRefGoogle Scholar
  80. 80.
    Jimenez AS, van Tongeren M (2017) Assessment of human exposure to ENMs. Adv Exp Med Biol 947:27–40CrossRefPubMedGoogle Scholar
  81. 81.
    Bonassi S, Znaor A, Ceppi M et al (2007) An increased micronucleus frequency in peripheral blood lymphocytes predicts the risk of cancer in humans. Carcinogenesis 28(3):625–631CrossRefPubMedGoogle Scholar
  82. 82.
    Holland N, Bolognesi C, Kirsch-Volders M et al (2008) The micronucleus assay in human buccal cells as a tool for biomonitoring DNA damage: the HUMN project perspective on current status and knowledge gaps. Mutat Res 659(1–2):93–108CrossRefPubMedGoogle Scholar
  83. 83.
    Tay CY, Fang W, Setyawati MI et al (2014) Nano-hydroxyapatite and nano-titanium dioxide exhibit different subcellular distribution and apoptotic profile in human oral epithelium. ACS Appl Mater Interfaces 6(9):6248–6256CrossRefPubMedGoogle Scholar
  84. 84.
    Sayes CM, Smith PA, Ivanov I (2013) A framework for grouping nanoparticles based on their measurable characteristics. Int J Nanomedicine 8:45–56CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© The Author(s) 2018

Authors and Affiliations

  1. 1.Department of Human GeneticsNational Institute of Health Dr. Ricardo Jorge (INSA)LisbonPortugal
  2. 2.Toxicogenomics and Human Health (ToxOmics), Nova Medical School/Faculdade de Ciências MédicasUniversidade Nova de LisboaLisbonPortugal

Personalised recommendations