Skip to main content

Screening of Fullerene Toxicity by Hemolysis Assay

  • Protocol
  • First Online:
Book cover Nanotoxicity

Part of the book series: Methods in Molecular Biology ((MIMB,volume 926))

Abstract

Fullerene is a compound formed during carbon burst that has been produced synthetically starting from the 1990s. The spherical shape and the characteristic carbon bonds of this allotrope (C60) have made it a suitable molecule for many applications. During the last decade, the low aqueous solubility of this molecule has been improved by chemical functionalization allowing the use of fullerene derivatives in biological fluids. The characterization of the toxicity potential of fullerenes is therefore of growing interest for any biomedical application. Intravenous injection is one of the possible routes of their administrations and therefore red blood cells are among the first targets of fullerene cytotoxicity. Human red blood cells are easily available and separated from plasma. Membrane disruption by toxic compounds is easily detected in red blood cells as release of hemoglobin in the cell medium, which can be assayed spectrophotometrically at λ = 415 nm. Due to the high molar extinction coefficient of hemoglobin, the assay can be performed on a small amount of both red blood cells and the test compounds, which might be available only in small quantities. So, the hemolysis assay is a simple screening test, whose results can guide further investigations on cytotoxicity in more complex experimental models.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 159.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Prato M (1997) [60]Fullerene chemistry for materials science applications. J Mater Chem 7:1097–1109

    Article  CAS  Google Scholar 

  2. Beuerle F, Hirsch A (2009) Synthesis and orthogonal functionalization of [60]fullerene e, e, e-trisadducts with two spherically defined addend zones. Chemistry 15:7434–7446

    Article  PubMed  CAS  Google Scholar 

  3. Bianco A, Da Ros T, Prato M, Toniolo C (2001) Fullerene-based amino acids and peptides. J Pept Sci 4:208–219

    Article  Google Scholar 

  4. Jagadeesan D, Eswaramoorthy M (2010) Functionalized carbon nanomaterials derived from carbohydrates. Chem Asian J 5:232–243

    Article  PubMed  CAS  Google Scholar 

  5. Nakamura E, Isobe H (2003) Functionalized fullerenes in water. The first 10 years of their chemistry, biology, and nanoscience. Acc Chem Res 36:807–815

    Article  PubMed  CAS  Google Scholar 

  6. Witte P, Beuerle F, Hartnagel U, Lebovitz R, Savouchkina A, Sali S, Guldi D, Chronakis N, Hirsch A (2007) Water solubility, antioxidant activity and cytochrome C binding of four families of exohedral adducts of C60 and C70. Org Biomol Chem 5:3599–3613

    Article  PubMed  CAS  Google Scholar 

  7. Gharbi N, Pressac M, Hadchouel M, Szwarc H, Wilson SR, Moussa F (2005) [60]Fullerene is a powerful antioxidant in vivo with no acute or subacute toxicity. Nano Lett 5:2578–2585

    Article  PubMed  CAS  Google Scholar 

  8. Sun T, Xu Z (2006) Radical scavenging activities of alpha-alanine C60 adduct. Bioorg Med Chem Lett 16:3731–3734

    Article  PubMed  CAS  Google Scholar 

  9. Enes RF, Tomé AC, Cavaleiro JA, Amorati R, Fumo MG, Pedulli GF, Valgimigli L (2006) Synthesis and antioxidant activity of [60]fullerene-BHT conjugates. Chemistry 12:4646–4653

    Article  PubMed  CAS  Google Scholar 

  10. Hu Z, Guan W, Wang W, Huang L, Xing H, Zhu Z (2007) Synthesis of β-alanine C60 derivative and its protective effect on hydrogen peroxide-induced apoptosis in rat pheochromocytoma cells. Cell Biol Int 31:798–804

    Article  PubMed  CAS  Google Scholar 

  11. Yang J, Alemany LB, Driver J, Hartgerink JD, Barron AR (2007) Fullerene-derivatized amino acids: Synthesis, characterization, antioxidant properties, and solid-phase peptide synthesis. Chem Eur J 13:2530–2545

    Article  PubMed  CAS  Google Scholar 

  12. Dugan L, Gabrielsen J, Yu S, Lin T, Choi D (1996) Buckminsterfullerenol free radical scavengers reduce excitotoxic and apoptotic death of cultured cortical neurons. Neurobiol Dis 3:129–135

    Article  PubMed  CAS  Google Scholar 

  13. Dugan LL, Turetsky DM, Du C, Lobner D, Wheeler M, Almli CR, Shen CKF, Luh TY, Choi DW, Lin TS (1997) Carboxyfullerenes as neuroprotective agents. Proc Natl Acad Sci USA 94:9434–9439

    Article  PubMed  CAS  Google Scholar 

  14. Jin H, Chen WQ, Tang XW, Chiang LY, Yang CY, Schloss JV, Wu JY (2000) Polyhydroxylated C(60), fullerenols, as glutamate receptor antagonists and neuroprotective agents. J Neurosci Res 62:600–607

    Article  PubMed  CAS  Google Scholar 

  15. Yang X, Chen Z, Meng X, Li B, Tan X (2007) Inhibition of DNA restrictive endonucleases and Taq DNA polymerase by trimalonic acid C60. Chin Sci Bull 52:1802–1806

    Article  CAS  Google Scholar 

  16. Iwata N, Mukai T, Yamakoshi Y, Hara S, Yanase T, Shoji M, Endo T, Miyata N (1998) Effects of C60, a fullerene, on the activities of glutathione S-transferase and glutathione-related enzymes in rodent and human livers. Fullerenes Nanotubes Carbon Nanostruct 6:213–226

    Article  CAS  Google Scholar 

  17. Lai YL, Chiang LY (1997) Water-soluble fullerene derivatives attenuate exsanguination-induced bronchoconstriction of guinea-pigs. J Auton Pharmacol 17:229–235

    Article  PubMed  CAS  Google Scholar 

  18. Nielsen GD, Roursgaard M, Jensen KA, Poulsen SS, Larsen ST (2008) In vivo biology and toxicology of fullerenes and their derivatives. Basic Clin Pharmacol Toxicol 103:197–208

    Article  PubMed  CAS  Google Scholar 

  19. Yang X, Meng X, Li B, Chen Z, Zhao D, Tan X, Yu Q (2008) Inhibition of in vitro amplification of targeted DNA fragment and activity of exonuclease I by a fullerene-oligonucleotide conjugate. Biologicals 36:223–226

    Article  PubMed  CAS  Google Scholar 

  20. Wang Z, Zhao J, Li F, Gao D, Xing B (2009) Adsorption and inhibition of acetylcholinesterase by different nanoparticles. Chemosphere 77:67–73

    Article  PubMed  CAS  Google Scholar 

  21. Innocenti A, Durdagi S, Doostdar N, Strom TA, Barron AR, Supuran CT (2010) Nanoscale enzyme inhibitors: fullerenes inhibit carbonic anhydrase by occluding the active site entrance. Bioorg Med Chem 18:2822–2828

    Article  PubMed  CAS  Google Scholar 

  22. Bosi S, Da Ros T, Castellano S, Banfi E, Prato M (2000) Antimycobacterial activity of ionic fullerene derivatives. Bioorg Med Chem Lett 10:1043–1045

    Article  PubMed  CAS  Google Scholar 

  23. Klumpp C, Lacerda L, Chaloin O, Da Ros T, Kostarelos K, Prato M, Bianco A (2007) Multifunctionalised cationic fullerene adducts for gene transfer: design, synthesis and DNA complexation. Chem Commun (Camb) 36:3762–3764

    Article  Google Scholar 

  24. Bosi S, Da Ros T, Spallato G, Prato M (2003) Fullerene derivatives: an attractive tool for ­biological applications. Eur J Med Chem 38:913–923

    Article  PubMed  CAS  Google Scholar 

  25. Johnston HJ, Hutchison GR, Frans M, Christensen FM, Aschberger K, Stone V (2010) The biological mechanisms and physicochemical characteristics responsible for driving fullerene toxicity. Toxicol Sci 114:162–182

    Article  PubMed  CAS  Google Scholar 

  26. Trpkovic A, Todorovic-Markovic B, Kleut D, Misirkic M, Janjetovic K, Vucicevic L, Pantovic A, Jovanovic S, Dramicanin M, Markovic Z, Trajkovic V (2010) Oxidative stress-mediated hemolytic activity of solvent exchange-prepared fullerene (C60) nanoparticles. Nanotechnology 21:375102

    Article  PubMed  Google Scholar 

  27. Bosi S, Feruglio L, Da Ros T, Spalluto G, Gregoretti B, Terdoslavich M, Decorti G, Passamonti S, Moro S, Prato M (2004) Haemolytic effects of water-soluble fullerene derivatives. J Med Chem 47:6711–6715

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Sabina Passamonti .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2012 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Tramer, F., Da Ros, T., Passamonti, S. (2012). Screening of Fullerene Toxicity by Hemolysis Assay. In: Reineke, J. (eds) Nanotoxicity. Methods in Molecular Biology, vol 926. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-002-1_15

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-002-1_15

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-001-4

  • Online ISBN: 978-1-62703-002-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics