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Drug Test Anal. 2017 Sep;9(9):1363-1371. doi: 10.1002/dta.2205. Epub 2017 May 31.

Doping control study of AICAR in post-race urine and plasma samples from horses.

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Racing Laboratory, The Hong Kong Jockey Club, Sha Tin Racecourse, Sha Tin, N.T, Hong Kong, China.
Laboratoire des Courses Hippiques, 15 rue de Paradis, 91370, Verrieres le Buisson, France.
Australian Racing Forensic Laboratory, Racing NSW, Sydney, NSW, 2000, Australia.
Canadian Pari-Mutuel Agency, 1130 Morrison Dr. Suite 101, Ottawa, Ontario, K2H 9N6, Canada.
Equine Drug Evaluation Centre, Canadian Pari-Mutuel Agency, 115 Sunnyridge, RR#1, Jerseyville, Ontario, L0R 1R0, Canada.
Department of Statistics, The Chinese University of Hong Kong, Sha Tin, N.T, Hong Kong, China.


Acadesine, 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside, commonly known as AICAR, is a naturally occurring adenosine monophosphate-activated protein kinase (AMPK) activator in many mammals, including humans and horses. AICAR has attracted considerable attention recently in the field of doping control because of a study showing the enhancement of endurance performance in unexercised or untrained mice, resulting in the term 'exercise pill'. Its use has been classified as gene doping by the World Anti-Doping Agency (WADA), and since it is endogenous, it may only be possible to control deliberate administration of AICAR to racehorses after establishment of an appropriate threshold. Herein we report our studies of AICAR in post-race equine urine and plasma samples including statistical studies of AICAR concentrations determined from 1,470 urine samples collected from thoroughbreds and standardbreds and analyzed in Australia, France, and Hong Kong. Quantification methods in equine urine and plasma using liquid chromatography-mass spectrometry were developed by the laboratories in each country. An exchange of spiked urine and plasma samples between the three countries was conducted, confirming no significant differences in the methods. However, the concentration of AICAR in plasma was found to increase upon haemolysis of whole blood samples, impeding the establishment of a suitable threshold in equine plasma. A possible urine screening cut-off at 600 ng/mL for the control of AICAR in racehorses could be considered for adoption. Application of the proposed screening cut-off to urine samples collected after intravenous administration of a small dose (2 g) of AICAR to a mare yielded a short detection time of approximately 4.5 h.


AICAR; doping control; equine urine; liquid chromatography-mass spectrometry; urine

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