Graphite Furnace Atomic Absorption Spectrometric Evaluation of Iron Excretion in Mouse Urine Caused by Whole-Body Gamma Irradiation

  • Makoto Yoshiyama
  • Yasuaki OkamotoEmail author
  • Shunsuke Izumi
  • Daisuke Iizuka


A procedure for the determination of iron in mice urine using graphite furnace atomic absorption spectrometry was developed. The mice urinary samples contain many organic compounds in the matrix, whose concentrations are approximately 20%, and the value is 30-fold higher compared to those found in human urine. Moreover, only 0.2 mL or less of urine was obtained as a sample volume per urination event. It was difficult to decompose the organic materials in the samples by wet digestion using mineral acids and oxidising agents, because of the tiny volumes. In this experiment, raw urinary samples were placed directly into the graphite tube furnace for analysis. The organic contents were simply ashed during the preheating stages. To facilitate ashing in the furnace, air was invaded from the surroundings by interrupting the stream of argon gas. Atomic absorption was measured at 248.3270 nm (wavelength for atomic absorption), with the background monitored at 247.0658 nm (wavelength for background correction). The optimised instrument operating conditions precluded the use of chemical modification technique. The analytical procedures used are quite simple, i.e. an aliquot of raw urine sample was injected directly into the graphite tube furnace and was followed by a suitable heating programme with no chemical modifier. Therefore, this method is useful for scientists who are not familiar with delicate chemical experiments. The proposed analytical method was applied as a kind of biomarker by determining iron concentrations in urinary samples of mice, which were irradiated with 4 Gy of gamma irradiation to their whole body. The time dependence of the iron concentration was determined, and the iron concentrations increased within 1 day of irradiation exposure, then decreased to ordinal values after several days.


Graphite furnace atomic absorption spectrometry Iron determination Mouse urine sample Iron metabolism Whole-body irradiation exposure Radiation injury 


Compliance with Ethical Standards

All experiments using mice were approved by the Animal Experimentation Committee of Hiroshima University.

Conflict of Interest

The authors declare that they have no conflict of interest.

Supplementary material

12011_2018_1589_MOESM1_ESM.pdf (69 kb)
ESM 1 (PDF 69 kb)
12011_2018_1589_MOESM2_ESM.pdf (113 kb)
ESM 2 (PDF 112 kb)


  1. 1.
    De Souza CC, Fabrino JHF, Beinner MA, Neto WB, Cangussu SD, Tafuri WL, Da Silva JBB (2013) Development and validation of methods for the determination of copper and iron in serum of dogs with canine visceral leishmaniasis using multivariate optimization and GFAAS. Anal Methods 5:3129–3135. CrossRefGoogle Scholar
  2. 2.
    Zhang XH, Lou ZC, Wang AL, Hu XD, Zhang HQ (2013) Development of serum iron as a biological dosimeter in mice. Radiat Res 179:684–689.
  3. 3.
    Correia PRM, Oliveira E, Oliveira PV (2002) Minimalism approach for determination of Cu, Fe and Zn in serum by simultaneous electrothermal atomic absorption spectrometry. Anal Chim Acta 458:321–329. CrossRefGoogle Scholar
  4. 4.
    Donnici CL, Souza CC, Beinner MA, Da Silva JBB (2016) Fast determination of iron and zinc in hair and human serum samples after alkaline solubilization by GFAAS. J Braz Chem Soc 27:119–126. CrossRefGoogle Scholar
  5. 5.
    Forte G, Bocca B, Senofonte O, Petrucci F, Brusa L, Stanzione P, Zannino S, Violante N, Alimonti A, Sancesario G (2004) Trace and major elements in whole blood, serum, cerebrospinal fluid and urine of patients with Parkinson’s disease. J Neural Transm 111:1031–1040. CrossRefPubMedGoogle Scholar
  6. 6.
    Tahán JE, Granadillo VA, Romero RA (1994) Electrothermal atomic absorption spectrometric determination of Al, Cu, Fe, Pb, V and Zn in clinical samples and in certified environmental reference materials. Anal Chim Acta 295:187–197. CrossRefGoogle Scholar
  7. 7.
    Choengchan N, Mantim T, Inpota P, Nacapricha D, Wilairat P, Jittangprasert P, Waiyawat W, Fucharoen S, Sirankpracha P, Morales NP (2015) Tandem measurements of iron and creatinine by cross injection analysis with application to urine from thalassemic patients. Talanta 133:52–58. CrossRefPubMedGoogle Scholar
  8. 8.
    Blażewicz A, Klatka M, Astel A, Partyka M, Kocjan R (2013) Differences in trace metal concentrations (Co, Cu, Fe, Mn, Zn, Cd, and Ni) in whole blood, plasma, and urine of Obese and nonobese children. Biol Trace Elem Res. 155:190–200. CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Zettner A, Mansbach L (1965) Application of atomic absorption spectrometry in the determination of iron in urine. Am J Clin Pathol 44:517–519. CrossRefPubMedGoogle Scholar
  10. 10.
    He G, Chen X, Zhang G, Lin H, Li R, Wu X (2014) Detection of urine C2C and trace element level in patients with knee osteoarthritis. Cell Biochem Biophys 70:457–479. CrossRefGoogle Scholar
  11. 11.
    Chen H, Tan C, Lin Z, Wu T (2014) The diagnostics of diabetes mellitus based on ensemble modeling and hair/urine element level analysis. Comput Biol Med 50:70–75. CrossRefPubMedGoogle Scholar
  12. 12.
    Brodzka R, Trzcinka-Ochocka M, Janasik B (2013) Multi-element analysis of urine using dynamic reaction cell inductively coupled plasma mass spectrometry (ICP-DRC-MS)—a practical application. Int J Occup Med Environ Health 26:302–312. CrossRefPubMedGoogle Scholar
  13. 13.
    Crowder JM, Bates N, Roberts J, Torres AS, Bonitatibus PJ Jr (2016) Determination of tantalum from tantalum oxide nanoparticle X-ray/CT contrast agents in rat tissues and bodily fluids by ICP-OES. J Anal At Spectrom 31:1311–1317. CrossRefGoogle Scholar
  14. 14.
    Rakoczy R, Kopeć A, Piatkowska E, Smoleń S, Skoczylas L, Leszczyńska T, Sady W (2016) The iodine content in urine, faeces and selected organs of rats fed lettuce biofortified with iodine through foliar application. Biol Trace Elem Res 174:347–355. CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Pronk C, Oldenziel H, Lequin HC (1974) A method for determination of serum iron, total iron binding capacity and iron in urine by atomic absorption spectrometry with manganese as internal standard. Clin Chim Acta 50:35–41. CrossRefPubMedGoogle Scholar
  16. 16.
    Saracoglu S, Soylak M, Peker DSK, Elci L, Dos Santos WNL, Lemos VA, Ferreira SLC (2006) A preconcentration procedure using coprecipitation for determination of lead and iron in several samples using flame atomic absorption spectrometry. Anal Chim Acta 575:133–137. CrossRefPubMedGoogle Scholar
  17. 17.
    Kandhro GA, Kazi TG, Biag JA, Sirajuddin, Afridi HI, Shah AQ, Sheikh HR, Kolachi NF, Wadhwa SK (2010) Zinc and iron determination in serum and urine samples of thyroid patients using cloud point extraction. J AOAC Int 93:1589–1594Google Scholar
  18. 18.
    Singh SP, Rahman MF, Murty USN, Mahboob M, Grover P (2013) Comparative study of genotoxicity and tissue distribution of nano and micron sized iron oxide in rats after acute oral treatment. Toxicol Appl Phamacol 266:56–66 Accessed 28 Nov 2018
  19. 19.
    Gómez T, Bequer L, Mollineda A, Molina JL, Álvarez A, Lavastida M, Clapés S (2017) Concentration of zinc, copper, iron, calcium, and magnesium in the serum, tissues, and urine of streptozotocin-induced mild diabetic rat model. Biol Trace Elem Res 179:237–246. CrossRefPubMedGoogle Scholar
  20. 20.
    Chen H, Tan C, Lin Z, Wu T, Diao Y (2013) A feasibility study of diagnosing cardiovascular diseases based on blood/urine element analysis and consensus models. Comput Biol Med 43:865–869. CrossRefPubMedGoogle Scholar
  21. 21.
    Fieten H, Hugen S, Van den Ingh TSGAM, Hendriks WH, Vernooij JCM, Bode P, Watson AL, Leegwater PAJ, Rothuizen J (2013) Urinary excretion of copper, zinc and iron with and without D-penicillamine administration in relation to hepatic copper concentration in dogs. Veterinary J 197:468–473. CrossRefGoogle Scholar
  22. 22.
    Musumeci M, Maccari S, Massimi A, Stati T, Sestili P, Corritore E, Pastorelli A, Stacchini P, Marano G, Catalano L (2014) Iron excretion in iron dextran-overloaded mice. Blood Transfus 12:485–490. CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Ma D, Okamoto Y, Kumamaru T, Iwamoto E (1999) Determination of gallium by graphite furnace atomic absorption spectrometry with combined use of a tungsten-coated L’vov platform tube and a chemical modification technique. Anal Chim Acta 390:201–206. CrossRefGoogle Scholar
  24. 24.
    Ma D, Okamoto Y, Kumamaru T, Iwamoto E (1999) Determination of indium in aluminum alloy by electrothermal atomic absorption spectrometry with a tungsten-coated L’vov platform tube. Anal Sci 15:193–195. CrossRefGoogle Scholar
  25. 25.
    Glenn MT, Savory J, Fein SA, Reeves RD, Molnar CJ, Winefordner JD (1973) Graphite rod atomizer in atomic absorption spectrometry for direct determination of iron in serum. Anal Chem 45:203–205. CrossRefPubMedGoogle Scholar
  26. 26.
    Sturgeon RE, Berman SS, Desaulniers A, Russell DS (1979) Determination of iron, manganese, and zinc in seawater by graphite furnace atomic absorption spectrometry. Anal Chem 51:2364–2369. CrossRefGoogle Scholar
  27. 27.
    Liang L, D’Haese PC, Lamberts LV, De Broe ME (1989) Direct determination of iron in urine and serum using graphite furnace atomic absorption spectrometry. Analyst 114:143–147. CrossRefPubMedGoogle Scholar
  28. 28.
    Iwamoto E, Shimazu H, Yokota K, Kumamaru T (1992) Determination of tin by electrothermal atomic absorption spectrometry with a tungsten-coated tube. J Anal At Spectrom 7:421. CrossRefGoogle Scholar
  29. 29.
    Slavin W (1984) Graphite furnace AAS: a source book, part No. 0993–8139. Perkin-Elmer, Norwalk, p 18Google Scholar
  30. 30.
    de Queiroz JV, Vieira JCS, Bataglioli IC, Bittarello AC, Braga CP, de Oliveira G, Padilha CCF, Padilha PM (2018) Total mercury determination in muscle and liver tissue samples from Brazilian amazon fish using slurry sampling. Biol Trace Elem Res 184:517–522. CrossRefPubMedGoogle Scholar
  31. 31.
    Gladney ES, O’Malley BT, Roelandts I, Gills TE (1987) Compilation of elemental concentration data for nbs clinical, biological, geological, and environmental standard reference materials. National Bureau of Standards, U. S. Department of Commerce, GaithersburgCrossRefGoogle Scholar
  32. 32.
    Iizuka D, Yoshioka S, Kawai H, Okazaki E, Kiriyama K, Izumi S, Nishimura M, Shimada Y, Kamiya K, Suzuki F (2016) Hepcidin-2 in mouse urine as a candidate radiation-responsive molecule. J Radiat Res 57:142–149. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Graduate School of ScienceHiroshima UniversityHigashihiroshimaJapan
  2. 2.Department of Radiation Effects ResearchNational Institute for Quantum and Radiological Science and TechnologyChibaJapan

Personalised recommendations