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Int J Sports Med 2009; 30(2): 80-86
DOI: 10.1055/s-0028-1103283
Physiology & Biochemistry

© Georg Thieme Verlag KG Stuttgart · New York

Identification of Darbepoetin Alfa in Human Plasma by Liquid Chromatography Coupled to Mass Spectrometry for Doping Control

F. Guan 1 , C. E. Uboh 2 , L. R. Soma 1 , E. Birksz 1 , J. Chen 1
  • 1School of Veterinary Medicine, University of Pennsylvania, Kennett Square, Pennsylvania, United States
  • 2PA Equine Toxicology and Research Center, Department of Chemistry, West Chester University, West Chester, Pennsylvania, United States
Further Information

Publication History

accepted after revision July 1, 2008

Publication Date:
28 January 2009 (online)

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Abstract

Darbepoetin alfa (DPO) or Novel Erythropoiesis Erythropoiesis Stimulating Protein (NESP), an analog of recombinant human erythropoietin (rhEPO), is abused as a blood doping agent along with the latter in human sports. This paper describes a new method for unequivocal identification of DPO in human plasma. The analyte was extracted from plasma by immunoaffinity separation with anti-rhEPO antibodies, digested by trypsin followed by PNGase F, and analyzed by liquid chromatography coupled to tandem mass spectrometry. The deglycosylated tryptic peptide, T9, was employed in DPO identification using liquid chromatographic retention time and major product ions of the T9 peptide. The limit of detection of this method for DPO was 0.1 ng/mL in plasma, and that of identification was 0.2 ng/mL. This method is definitive and devoid of false positive results, providing “mass fingerprints” for identification of DPO in human plasma samples. Although this method is not applicable to identification of rhEPO in human plasma because it cannot differentiate rhEPO from endogenous EPO, it is the first successful attempt towards establishing a reliable and selective method for definitive identification of exogenously administered EPOs in doping control analyses.

Key words

erythropoietin - novel erythropoiesis stimulating protein - Aranesp

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Correspondence

Dr. F. GuanPhD 

School of Veterinary Medicine

University of Pennsylvania

220 East Rosedale Avenue

West Chester, PA 19382

United States

Phone: +610\436\35 01

Fax: +610\436\35 04

Email: guanf@vet.upenn.edu

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