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Keywords:

  • doping;
  • erythropoietin;
  • epoetin kappa;
  • isoelectric focusing;
  • SDS-PAGE;
  • mass spectrometry;
  • biosimilar

Since the expiration of patent protection, a number of new recombinant erythropoietin (rEPO) biosimilars have appeared on the worldwide market. In 2010, epoetin kappa, which is biosimilar to epoetin alfa, was clinically approved in Japan. Currently, both isoelectric focusing polyacrylamide gel electrophoresis (IEF-PAGE) and sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) are approved by the World Anti-Doping Agency (WADA) for detection of rEPO doping. Because it was unclear whether epoetin kappa could be detected by WADA-accredited detection methods, intravenous administration studies of epoetin kappa, epoetin alfa, and epoetin beta were performed to test the applicability of these methods. The isoform bands of epoetin kappa expanded more widely towards the basic area and the profile appeared to be composed of at least eight bands, which were clearly different from those of other epoetins. The results showed that epoetin kappa also contains isoforms of higher molecular masses than those of originator epoetins on SDS-PAGE; the mass distribution was confirmed by electrospray ionization time-of-flight mass spectrometry. We clearly detected epoetin kappa after its administration up to 10 h by IEF-PAGE and 24 h by SDS-PAGE; the detection window of the SDS-PAGE is longer than that of the IEF-PAGE. SDS-PAGE compensates for the disadvantages of IEF-PAGE in detecting urinary epoetin kappa. We also concluded that athletes abusing rEPO might move to intravenous injections for shorter clearance times instead of subcutaneous injections. In conclusion, out-of-competition tests need to be applied more frequently to improve the effectiveness of the rEPO detection. Copyright © 2011 John Wiley & Sons, Ltd.