Analytical and Bioanalytical Chemistry

, Volume 386, Issue 2, pp 244–248 | Cite as

Fluorometry of hydrogen peroxide using oxidative decomposition of folic acid

  • Kazutaka Hirakawa
Original Paper


Hydrogen peroxide (H2O2) is one of the most important reactive oxygen species. In the present study, a fluorometry method for detecting H2O2 utilizing folic acid was evaluated. Folic acid was decomposed by H2O2 in the presence of Cu(II) into pterine-6-carboxylic acid, leading to strong fluorescence enhancement. In the absence of the metal ion, superoxide and H2O2 could not decompose folic acid. Also, H2O2 plus sodium hypochlorite (a source of singlet oxygen) could not induce fluorescence enhancement. These results demonstrate that H2O2 can be selectively detected using folic acid plus Cu(II). The limit of detection (LOD; at S/N=3) for H2O2 is 0.5 μM. This method based on the fluorescence enhancement of folic acid was applied in order to determine small amounts of H2O2 generated through the autooxidation of semicarbazide (generation rate: ∼0.01 μM min−1), a carcinogenic compound.


Hydrogen peroxide Reactive oxygen species Fluorometry Folic acid Copper(II) ion 



The author wishes to thank Professor Motoshi Nakamura (Faculty of Engineering, Shizuoka University) for his helpful discussion. This work was supported by a Grant-in-Aid for Scientific Research on Priority Areas (417) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) of the Japanese Government.


  1. 1.
    Soh N, Ariyoshi T, Fukaminato T, Nakano K, Irie M, Imato T (2006) Bioorg Med Chem Lett 16:2943–2946CrossRefGoogle Scholar
  2. 2.
    Soh N, Sakawaki O, Makihara K, Odo Y, Fukaminato T, Kawai T, Irie M, Imato T (2005) Bioorg Med Chem 13:1131–1139CrossRefGoogle Scholar
  3. 3.
    Soh N (2006) Anal Bioanal Chem (accepted)Google Scholar
  4. 4.
    Lei W, Duerkop A, Lin Z, Wu M, Wolfbeis OS (2003) Microchim Acta 143:269–274Google Scholar
  5. 5.
    Wolfbeis OS, Dürkop A, Wu M, Lin Z (2004) Angew Chem Int Ed 41:4495–4498CrossRefGoogle Scholar
  6. 6.
    Kozhevnikov VN, Mandl C, Miltschitzky S, Duerkop A, Wolfbeis OS, Koenig B (2005) Inorg Chim Acta 358:2445–2448CrossRefGoogle Scholar
  7. 7.
    Setsukinai K, Urano Y, Kakinuma K, Majima HJ, Nagano T (2003) J Biol Chem 278:3170–3175CrossRefGoogle Scholar
  8. 8.
    Miura T, Urano Y, Tanaka K, Nagano T, Ohkubo K, Fukuzumi S (2003) J Am Chem Soc 125:8666–8671CrossRefGoogle Scholar
  9. 9.
    Cadet J, Douki T, Gasparutto D, Ravanat JL (2003) Mutat Res 531:5–23Google Scholar
  10. 10.
    Martinez GR, Loureiro AP, Marques SA, Miyamoto S, Yamaguchi LF, Onuki J, Almeida EA, Garcia CC, Barbosa LF, Medeiros MH, Di Mascio P (2003) Mutat Res 544:115–127CrossRefGoogle Scholar
  11. 11.
    Kawanishi S, Hiraku Y, Oikawa S (2001) Mutat Res 488:65–76CrossRefGoogle Scholar
  12. 12.
    Hirakawa K, Mori M, Yoshida M, Oikawa S, Kawanishi S (2004) Free Radical Res 38:439–447Google Scholar
  13. 13.
    Hirakawa K, Oikawa S, Hiraku Y, Hirosawa I, Kawanishi S (2002) Chem Res Toxicol 15:76–82CrossRefGoogle Scholar
  14. 14.
    Akhtar MJ, Khan MA, Ahmad I (1999) J Pharm Biomed Anal 19:269–275CrossRefGoogle Scholar
  15. 15.
    Akhtar MJ, Khan MA, Ahmad I (2000) J Pharm Biomed Anal 23:1039–1044CrossRefGoogle Scholar
  16. 16.
    Akhtar MJ, Khan MA, Ahmad I (2003) J Pharm Biomed Anal 31:579–588CrossRefGoogle Scholar
  17. 17.
    Hirakawa K, Suzuki H, Oikawa S, Kawanishi S (2003) Arch Biochem Biophys 410:261–268CrossRefGoogle Scholar
  18. 18.
    Thomas AH, Lorente C, Capparelli AL, Pokhrel MR, Braun AM, Oliveros E (2002) Photochem Photobiol Sci 1:421–426CrossRefGoogle Scholar
  19. 19.
    Akhtar MJ, Khan MA, Ahmad I (1997) J Pharm Biomed Anal 16:95–99CrossRefGoogle Scholar
  20. 20.
    Taher MM, Lakshmaiah N (1987) J Inorg Biochem 31:133–141CrossRefGoogle Scholar
  21. 21.
    Taher MM, Lakshmaiah N (1987) Arch Biochem Biophys 257:100–106CrossRefGoogle Scholar
  22. 22.
    Land EJ, Ebert M (1967) Trans Faraday Soc 63:1181–1190CrossRefGoogle Scholar
  23. 23.
    Liao JC, Roider J, Jay DG (1994) Proc Natl Acad Sci USA 91:2659–2663CrossRefGoogle Scholar
  24. 24.
    Hirakawa K, Midorikawa K, Oikawa S, Kawanishi S (2003) Mutat Res 536:91–101Google Scholar

Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  1. 1.Department of Basic Engineering (Chemistry), Faculty of EngineeringShizuoka UniversityHamamatsuJapan

Personalised recommendations