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Photoinduced protein modifications by methylene blue and naproxen

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Abstract

HPLC and emission spectroscopy were used to investigate UVA photosensitization of methylene blue (MB) or naproxen (NAP) towards bovine serum albumin (BSA). In addition, time resolved singlet oxygen measurements were carried out. The most stable drug : protein adducts stoichiometry of MB-BSA (1 : 1) and NAP–BSA (9 : 1) were verified by means of binding constant determination. UVA photosensitization of MB or NAP on BSA was studied by monitoring tryptophan (Trp) residue integrity. The sensitized photodegradation of the BSA resulted in different degrees of Trp damage. Thus, protein damage was determined by quantitative measurements of the different Trp (photo)-products. Indeed, many of these Trp derivatives are diagnostic for the photosensitization mechanism and some of them, for the first time in this work, were obtained by UVA photosensitization in proteins. The analysis of quantum yields of the photoproduct distribution allowed to weigh up the type I/II contribution to the UVA photosensitization mechanism. As expected, additional experiments in deuterated solvent resulted in an increase of the photodegradation quantum yields for those species where a singlet oxygen mechanism was involved. The UVA mediated generation of these Trp derivatives is consistent with the occurrence of singlet oxygen formation (almost dominant in MB), and photoionization (significant in NAP) within the protein matrix. Additional experiments at lower NAP concentration, as well as with human serum albumin (which differs for Trp content and, partially, localization), support further the molecular mechanism of photosensitization proposed. The results obtained in the case of this more complex system are in agreement with the free Trp model, even if, in almost all cases, the Trp photoproduct formation quantum yields are lower, due to the higher number of sensitization targets in the proteins.

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Abbreviations

NAP:

naproxen

MB:

methylene blue

Trp:

tryptophan

BSA:

bovine serum albumin

HSA:

human serum albumin

References

  1. B. A. Kerwin, R. L. Remmele Jr., Protect from light: photodegradation and protein biologics, J. Pharm. Sci., 2007, 96, 1468–1479.

    Article  CAS  PubMed  Google Scholar 

  2. A. Kawada, K. Hatanaka, H. Gomi and I. Matsuo, In vitro phototoxicity of new quinolones: production of active oxygen species and photosensitized lipid peroxidation, Photodermatol., Photoimmunol. Photomed., 1999, 15, 226–230.

    Article  CAS  Google Scholar 

  3. M. Mauget-Fa, M. Quaranta, N. Francoz and D. BenEzra, Incidental retinal phototoxicity associated with ingestion of photosensitizing drugs, Graefe’s Arch. Clin. Exp. Ophthalmol., 2001, 239, 501–508.

    Article  Google Scholar 

  4. G. T. Wondrak, M. K. Jacobson and E. L. Jacobson, Endogenous UVA photosensitizers: mediators of skin photodamage and novel targets for skin photoprotection, Photochem. Photobiol. Sci., 2006, 5, 215–237.

    Article  CAS  PubMed  Google Scholar 

  5. M. A. Miranda, J. V. Castell, D. Hernandez, M. J. Gomez-Lechon, F. Bosca, I. M. Morera and Z. Sarabia, Drug-photosensitized protein modification: identification of the reactive sites and elucidation of the reaction mechanisms with tiaprofenic acid/albumin as model system, Chem. Res. Toxicol., 1998, 11, 172–177.

    Article  CAS  PubMed  Google Scholar 

  6. A. Hideki, I. Kenji, H. Junichi and I. Hisami, Methylene blue photosensitization: mechanisms of virus inactivation and its clinical application, in Recent Research Developments in Photochemistry and Photobiology, ed. S. G. Pandalai, Transworld Research Network, Trivandrum, India, 2001, vol. 5, pp. 163–173. ISBN 81-7895-004-9.

    Google Scholar 

  7. Y. Usui, M. Tsukada and H. Nakamura, Kinetic studies of photosensitized oxygenation by singlet oxygen in aqueous micellar solutions, Bull. Chem. Soc. Jpn., 1978, 51, 379–384.

    Article  CAS  Google Scholar 

  8. W. Vonach and N. Getoff, Formation of eaq− from excited thionine and methylene blue in aqueous solutions, J. Photochem., 1983, 23, 233–239.

    Article  CAS  Google Scholar 

  9. L. Zhi-Ren, A. M. Wilkie, M. J. Clemens and C. W. J. Smith, Detection of double-stranded RNA–protein interactions by methylene blue-mediated photo-crosslinking, RNA, 1996, 2, 611–621.

    Google Scholar 

  10. E. Silva, C. De Landea, A. M. Edwards and E. Lissi, Lysozyme photo-oxidation by singlet oxygen: properties of the partially inactivated enzyme, J. Photochem. Photobiol., B, 2000, 55, 196–200.

    Article  CAS  Google Scholar 

  11. A. Michaeli and J. Feitelson, Reactivity of singlet oxygen toward proteins: the effect of structure in basic pancreatic trypsin inhibitor and in ribonuclease A, Photochem. Photobiol., 1997, 65, 309–315.

    Article  CAS  PubMed  Google Scholar 

  12. S. G. Bertolotti, N. A. Garcia, G. A. Argüello, Effect of the peptide bond on the singlet molecular oxygen-mediated photo-oxidation of tyrosine and tryptophan dipeptides, J. Photochem. Photobiol., B, 1991, 10, 57–70.

    Article  CAS  Google Scholar 

  13. N. R. Parker, J. F. Jamie, M. J. Davies and R. J. W. Truscott, Protein-bound kynurenine is a photosensitizer of oxidative damage, Free Radical Biol. Med., 2004, 37, 1479–1489.

    Article  CAS  Google Scholar 

  14. U. Khan and T. Wilson, Reactive oxygen species as cellular messengers, Chem. Biol., 1995, 2, 437–445.

    Article  CAS  PubMed  Google Scholar 

  15. R. M. Danziger, K. H. Bar-Eli and K. Weiss, The laser photolysis of methylene blue, J. Phys. Chem., 1967, 71, 2633–2640.

    Article  CAS  Google Scholar 

  16. J. G. Lombardino, Nonsteroidal Antiinflammatory Drugs, Wiley, New York, 1985.

    Google Scholar 

  17. G. Condorelli, L. L. Costanzo, G. De Guidi, S. Giuffrida, P. Miano, S. Sortino and A. Velardita, Photosensitization induced by non-steroidal antiinflammatory drugs: an overview of molecular mechanisms in biological systems, EPA Newsletter, 1996, 58, 60–77.

    CAS  Google Scholar 

  18. F. Boscá and M. A. Miranda, Photosensitizing drugs containing the benzophenone chromophore, J. Photochem. Photobiol., B, 1998, 43, 120–123.

    Article  Google Scholar 

  19. M. A. Miranda, J. V. Castell and M. J. Gòmez-Lechòn, Phototoxicity of Drugs. In Vitro Methods in Pharmaceutical Research, Academic Press, San Diego, 1997.

    Google Scholar 

  20. C. J. Le Coz, A. Bottlaender, J.-N. Scrivener, F. Santinelli, B. J. Cribier, E. Heid and E. M. Grosshans, Photocontact dermatitis from ketoprofen and tiaprofenic acid: cross-reactivity study in 12 consecutive patients, Contact Dermatitis, 1998, 38, 245–252.

    Article  CAS  PubMed  Google Scholar 

  21. J. V. Castell, M. J. Gomez-Lechon, C. Grassa, L. A. Martinez, M. A. Miranda and P. Tarrega, Involvement of drug-derived peroxides in the phototoxicity of naproxen and tiaprofenic acid, Photochem. Photobiol., 1993, 57, 486–490.

    Article  CAS  PubMed  Google Scholar 

  22. M. A. Miranda, I. Morera, F. Vargas, M. J. Gomez-Lechon and J. V. Castell, In vitro assessment of the phototoxicity of anti-inflammatory 2-arylpropionic acids, Toxicol. in Vitro, 1991, 5, 451–455.

    Article  CAS  PubMed  Google Scholar 

  23. L. L. Costanzo, G. De Guidi, G. Condorelli, A. Cambria, M. Famà, Molecular mechanism of Naproxen photosensitization in red blood cells, J. Photochem. Photobiol., B, 1989, 3, 223–235.

    Article  CAS  Google Scholar 

  24. F. Boscà, M. L. Marin and M. A. Miranda, Photoreactivity of the nonsteroidal anti-inflammatory 2-arylpropionic acids with photosensitizing side effects, Photochem. Photobiol., 2001, 74, 637–655.

    Article  PubMed  Google Scholar 

  25. A. Catalfo, G. Bracchitta, G. De Guidi, Role of aromatic amino acid tryptophan UVA-photoproducts in the determination of drug photosensitization mechanism: a comparison between methylene blue and naproxen, Photochem. Photobiol. Sci., 2009, 8, 1467–1475.

    Article  CAS  PubMed  Google Scholar 

  26. G. De Guidi, R. Chillemi, L. L. Costanzo, S. Giuffrida and G. Condorelli, Molecular mechanism of drug photosensitization. 4. Photohemolysis sensitized by carprofen, J. Photochem. Photobiol., B, 1993, 17, 239–246.

    Article  Google Scholar 

  27. T. Banerjee, S. K. Singh and N. Kishore, Binding of naproxen and amitriptyline to bovine serum albumin: biophysical aspects, J. Phys. Chem. B, 2006, 110, 24147–24156.

    Article  CAS  PubMed  Google Scholar 

  28. M. Maruthamuthu and G. Selvakumar, Selective quenching of tryptophanyl fluorescence in bovine serum albumin by the iodide ion, J. Chem. Soc., 1995, 107, 79–86.

    CAS  Google Scholar 

  29. T. Peters Jr., Serum albumin, Adv. Protein Chem., 1985, 37, 161–245.

    Article  CAS  PubMed  Google Scholar 

  30. R. G. Eckenhoff and J. S. Johansson, Molecular interactions between inhaled anesthetics and proteins, Pharmacol. Rev., 1997, 49, 343–367.

    CAS  PubMed  Google Scholar 

  31. Y. J. Hu, Y. Liu, X. S. Shen, X. Y. Fang and S. S. J. Qu, Studies on the interaction between 1-hexylcarbamoyl-5-fluorouracil and bovine serum albumin, J. Mol. Struct., 2005, 738, 143–147.

    Article  CAS  Google Scholar 

  32. Y. Ping-gui, L. Jun-feng, S. Zhi-cai, Y. Qing-sen, Study on the interaction between methylene blue and bovine serum albumin by fluorescence spectroscopy, Guangpuxue Yu Guangpu Fenxi, 2001, 21, 826–828.

    Google Scholar 

  33. D. C. Carter and X. M. He, Structure of human serum albumin, Science, 1990, 249, 302–303.

    Article  CAS  PubMed  Google Scholar 

  34. P. Miskovsky, D. Jancura, S. Sanchez-Cortes, E. Kocisova and L. Chinsky, Antiretrovirally active drug hypericin binds the IIA subdomain of human serum albumin: resonance Raman and surface-enhanced Raman spectroscopy study, J. Am. Chem. Soc., 1998, 120, 6374–6379.

    Article  CAS  Google Scholar 

  35. Y.-J. Hu, Y. Liu, R.-M. Zhao, J.-X. Dong, S.-S. Qu, Spectroscopic studies on the interaction between methylene blue and bovine serum albumin, J. Photochem. Photobiol., A, 2006, 179, 324–329.

    Article  CAS  Google Scholar 

  36. Y.-J. Hu, W. Li, Y. Liu, J.-X. Dong, S.-S. Qu, Short communication Fluorometric investigation of the interaction between methylene blue and human serum albumin, J. Pharm. Biomed. Anal., 2005, 39, 740–745.

    Article  CAS  PubMed  Google Scholar 

  37. S. Wei, L. Zhao, X. Cheng, J.-M. Lin, Determination of naproxen with flow injection chemiluminescence of Ru(bpy)32+–PbO2 system and its application for the binding study of naproxen to protein, Anal. Chim. Acta, 2005, 545, 65–73.

    Article  CAS  Google Scholar 

  38. M. M. Rahman, M. H. Rahman and N. N. Rahaman, Competitive binding of Ibuprofen and Naproxen to serum bovine albumin: modified form of drug-drig displacement interaction at the binding site, Pak. J. Pharm. Sci., 2005, 18, 43–47.

    CAS  PubMed  Google Scholar 

  39. C. B. Murphy, Y. Zhang, T. Troxler, V. Ferry, J. J. Martin and W. E. Jones, Probing förster and dexter energy-transfer mechanisms in fluorescent conjugated polymer chemosensors, J. Phys. Chem. B, 2004, 108, 1537–1543.

    Article  CAS  Google Scholar 

  40. Y. P. Tsentalovich, O. A. Snytnikova, M. D. E. Forbes, E. I. Chernyak and S. V. Morozov, Photochemical and thermal reactivity of kynurenine, Exp. Eye Res., 2006, 83, 1439–1445.

    Article  CAS  PubMed  Google Scholar 

  41. K. Schafer, D. Goddinger and H. Hocker, Photodegradation of tryptophan in wool, J. Soc. Dyers Colour., 1997, 113, 350–355.

    Article  CAS  Google Scholar 

  42. D. Creed, The photophysics and photochemistry of the near-uv absorbing amino acids–I. Tryptophan and its simple derivatives, Photochem. Photobiol., 1984, 39, 537–562.

    Article  CAS  Google Scholar 

  43. J. M. Dyer, S. D. Bringans and W. G. Bryson, Characterisation of photo-oxidation products within photoyellowed wool proteins: tryptophan and tyrosine derived chromophores, Photochem. Photobiol. Sci., 2006, 5, 698–706.

    Article  CAS  PubMed  Google Scholar 

  44. M. Wood and R. J. W. Truscott, Ultraviolet filter coupounds in human lenses: 3-hydroxykynurenine glycoside formation, Virus Res., 1994, 34, 1369–1374.

    CAS  Google Scholar 

  45. I. Balzer and R. Hardeland, Action of kynuramine in a dinoflagellate: stimulation of bioluminescence in gonyaulax polyedra, Comp. Biochem. Physiol., C, 1989, 94, 129–132.

    Article  Google Scholar 

  46. W. Wang, B. Qiu, X. Xu, L. Zhang and G. Chen, Separation and determination of l-tryptophan and its metabolites by capillary micellar electrokinetic chromatography with amperometric detection, Electrophoresis, 2004, 25, 903–910.

    Article  CAS  Google Scholar 

  47. P. Walrant and R. Santus, N-Formylkynurenine, a tryptophan photooxidation product, as a photodynamic sensitizer, Photochem. Photobiol., 1974, 19, 411–417.

    Article  CAS  PubMed  Google Scholar 

  48. A. Pirie, Formation of N-formylkynurenine in proteins from lens and other sources by exposure to sunlight, Biochem. J., 1971, 125, 203–208.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. H. Salminen and M. Heinonen, Plant phenolics affect oxidation of tryptophan, J. Agric. Food Chem., 2008, 56, 7472–7481.

    Article  CAS  PubMed  Google Scholar 

  50. P. R. Ogilby and C. S. Foote, Chemistry of singlet oxygen. Effect of solvent, solvent isotopic substitution, and temperature on the lifetime of singlet molecular oxygen (1Δg), J. Am. Chem. Soc., 1983, 105, 3423–3430.

    Article  CAS  Google Scholar 

  51. E. Skovsen, J. W. Snyder, J. D. C. Lambert and P. R. Ogilby, Lifetime and diffusion of singlet oxygen in a cell, J. Phys. Chem. B, 2005, 109, 8570–8573.

    Article  CAS  PubMed  Google Scholar 

  52. J. Davila and A. Harriman, Photoreactions of macrocyclic dyes bound to human serum albumin, Photochem. Photobiol., 1990, 51, 9–19.

    Article  CAS  PubMed  Google Scholar 

  53. E. Reddi, C. R. Lambert, G. Jori and M. A. Rodgers, Photokinetic and photophysical measurements of the sensitized photooxidation of the tritptophyl residue in N-acetyl tryphophanasmide and in human serum albumine, Photochem. Photobiol., 1987, 45, 345–351.

    Article  CAS  PubMed  Google Scholar 

  54. A. Michaeli and J. Feitelson, Reactivity of singlet oxygen toward amino acids and peptides, Photochem. Photobiol., 1994, 59, 284–289.

    Article  CAS  PubMed  Google Scholar 

  55. W. M. Sharman, J. E. van Lier and C. M. Allen, Targeted photodynamic therapy via receptor mediated delivery systems, Adv. Drug Delivery Rev., 2004, 56, 53–76.

    Article  CAS  Google Scholar 

  56. U. Kragh-Hansen, Structure and ligand binding properties of human serum albumin, Dan. Med. Bull., 1990, 37, 57–84.

    CAS  PubMed  Google Scholar 

  57. A. Siemiarczuk, C. E. Petersen, C.-E. Ha, J. Yang and N. V. Bhagavan, Analysis of tryptophan fluorescence lifetimes in a series of human serum albumin mutants with substitutions in subdomain 2A, Cell Biochem. Biophys., 2004, 40, 115–122.

    Article  CAS  PubMed  Google Scholar 

  58. E. Alarcon, A. M. Edwards, A. Aspee, F. E. Moran, C. D. Borsarelli, E. A. Lissi, D. Gonzalez-Nilo, H. Poblete and J. C. Scaiano, Photophysics and photochemistry of dyes bound to human serum albumin are determined by the dye localization, Photochem. Photobiol. Sci., 2010, 9, 93–102.

    Article  CAS  PubMed  Google Scholar 

  59. V. Lhiaubet-Vallet, Z. Sarabia, F. Boscá and M. A. Miranda, Human serum albumin-mediated stereodifferentiation in the triplet state behavior of (S)- and (R)-Carprofen, J. Am. Chem. Soc., 2004, 126, 9538–9539.

    Article  CAS  PubMed  Google Scholar 

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Authors and Affiliations

  1. Dipartimento di Scienze Chimiche, Università di Catania, Viale Andrea Doria 6, 95125, Catania, Italy

    Giuseppina Bracchitta, Alfio Catalfo & Guido De Guidi

  2. Inca - Consorzio Interuniversitario Chimica per l’Ambiente, Via delle Industrie 21/8, 30175, Marghera, VE, Italy

    Guido De Guidi

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  1. Giuseppina Bracchitta
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  2. Alfio Catalfo
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  3. Guido De Guidi
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Correspondence to Guido De Guidi.

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Bracchitta, G., Catalfo, A. & De Guidi, G. Photoinduced protein modifications by methylene blue and naproxen. Photochem Photobiol Sci 11, 1886–1896 (2012). https://doi.org/10.1039/c2pp25067e

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