the atomic energy organization of iran (aeoi) a-10!1!43-7c4aa3a

7/31/2019 The Atomic Energy Organization of Iran (AEOI) a-10!1!43-7c4aa3a

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J. of Nuclear Sci. and Tech.

5279-721389 No. 52, pp. 72-79, 2010

Sci. and Tech. note

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Discernment of Irradiated Chicken Meat by Determination of O-Tyrosine

Using High Performance Liquid Chromatography and Fluorescence Detection

F. Aflaki*, A. Roozbahani, M. Salahinejad

Nuclear Science Research School, Nuclear Science and Technology Research Institute, AEOI, P.O.Box: 11365-3486, Tehran – Iran

Abstract: O-Tyrosine is proposed as a marker for identification of irradiated protein-rich foods. In this

study, HPLC/ Fluorescence method that allows accurate quantification of 0.1ng of o-tyrosine has been

used. The method involves freeze-drying of sample, acid hydrolysis and fractionation by HPLC. Byusing Spherisorb ODS2 column, the base-line separation of o-tyrosine from impurities was performed.The yield of o-tyrosine in the irradiated chicken meat was proportional to the irradiation dose. Since thevariable levels of o-tyrosine were found in unirradiated chicken meat (0.15-0.42 µg/g per wet weight),this method is able to identify the irradiated chicken meat at 4kGy or higher. Because the dose responsecurve can be extended over 50kGy, the method is suitable for detecting the overdosed samples.

Keywords: O-Tyrosine, Irradiated Food, Detection, High Performance Liquid Chromatography

:[email protected] *email 20/2/89: پ 8/5/88:



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Detection: Fluorescence: Excitation 275nm, Emission 305

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1- CEN: Comite Europeen de Normalisation

2- ESR: Electron Spin Resonance

3- TL: Thermoluminescence

4- HC: Hydrocarbons Hydrocarbons

5- CB: Cyclobutanone

6- DNA Comet Assay

7- DEFT/APC: Direct Epifluorescent Filter

Technique/Aerobic Plate Count

8- FDA: Food and Drug Administration

9- Fenton- Type Reaction

10- HPLC: High Performance Liquid Chromatography

11- Freeze- Dryer

12- Theoretical Plate

13- Base Line

14- Gradient Elution

References:

1. IAEA-TECDOC-587, “Analytical detection

methods for irradiated foods, A review of

current literature,” Joint FAO/IAEA Division

of Nuclear Techniques in Food and Agriculture,

Austria, March (1991).

2. C.H. McMurray, “Detection methods for

irradiated foods: Current status,” Cambridge,

UK, Royal Society of Chemistry (1996).

3. H. Delincee, “Analytical methods to identify

irradiated food-a review,” Radiat. Phy. Chem,

63, 455-458 (2002).

4. The European Parliament and of the Council,

“Food and food ingredients treated with

ionizing radiation,” Off.J. L006, 16-23 (1999).

5. FDA, “Irradiation in the production, processing

and handling,” Fed. Reg. 62, 64107-64112

(1997).

6. R.A. Molins, “Food irradiation: Principles and

applications,” Johan Wiley & Sons, New York

(2001).

7. M. Miyahara, H. Ito, T. Nagasawa, T.

Kamimura, A. Saito, “Determination of o-

tyrosine production in aqueous solutions of phenylalanine irradiated with gamma ray, using

HPLC with automated pre-column

derivatization and LASER fluorometric

detection,” J. Health. Sci. 46, 192-199 (2000).

8. L.R. Karam, M.G. Simic, “Detecting irradiated

foods: Use of hydroxyl radical biomarkers,”

Anal. Chem. 60, 1117A-1120A (1988).

9. H. Zegota, K. Kolodziejczyk, M. Krol, B. Krol,

“O-tyrosine hydroxylation by OHº

Radicals. 2,

3-Dopa and 2, 5-Dopa formation in γ-irradiated

aqueous solution,” Rad. Phys. Chem. 72. 25-33

(2005).

10. R.J. Hart, J.A. White, W.J. Ried, “Technical

note: Occurrence of o-tyrosine in non-

irradiated foods,” International J. food sci.

Tech. 23, 643-647 (2207).

11. L.R. Karam, M.G. Simic, “Formation of o-

Tyrosine by radiation and organic solvents in

chicken tissue,” J. Biol. Chem. 265, 11581-11585 (1990).



521389

12. M. Grootveld, R. Jain, “Methods for the

detection of irradiated food stuffs: Aromatic

hydroxylation and degradation of poly

unsaturated fatty acids,” Radiat. Phys. Chem.

34, 925-931 (1989).

13. J. Hopkins, L.J. Davis, “Detection of irradiationof meat by o-tyrosine determination,” J.

Chromatogr, 426, 155-161 (1988).

14. M. Miyahara, H. Ito, A. Saito, T. Nagasawa, M.

Kariya, “Detection of irradiation of meats by

HPLC determination fir tyrosine using novel

LASER Fluorometric detection with automatic

pre-column reaction,” J. Health Sci. 46, 304-

309 (2000).

15. M. Miyahara, T. Nagasawa, T. Kamimura, H.

Ito, M. Toyoda, “Identification of irradiation of boned chicken by determination of o- tyrosine

and electron spin resonance spectrometry,” J.

Health, sci. 48, 79-82 (2002).

16. W. Meier, R. Burgin, D. Frohlich, “Analysis of

o-tyrosine as a method for the identification of

irradiated chicken meat,” Beta-Gamma, 1, 34-

38 (1988).

17. C. Willemot, R. Couture, J. Makhlouf, F.

Cheour, “Generation of oxyradicals in

biosystems,” Can. Inst. Food Sci. Technol. J.22, 350-355 (1989).

18. N. Chuaqui-Offermanns, T. Mcdougall, “An

HPLC method to determination o-tyrosine in

chicken meat,” J. Agric. Food Chem. 39, 300-

302 (1991).

19. F.I. Ibe, R. Grinter, R. Massey, R. Homer,

“Detection of o-tyrosine in irradiated chiken byrevese-phase HPLC and fluorescence

detection,” Food additive & contamin. Part A,8

(6), 787-792 (1991).

20. L.L. Zwart, J.H.N. Commandeur, N.P.E.

Vermeulen, “Biomarkers of free radical

damage, application in experimental animals

and in humans,” Free Radic. Biol. 26, 202-206

(1999).

21. F. Aflaki, M. Salahinejad, A. Roozbehani,

“Formation of tyrosine isomers in aquous phenylalanine solutions by gamma irradiation,”

47, 23-31 (2009).

22. J.N. Miller, J.C. Miller, “Statistical and chemo

metrics for analytical chemistry,” Fifth Ed.

Pearson Education (2007).

the atomic energy organization of iran (aeoi) a-10!1!43-7c4aa3a

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