Investigations into the analysis of intact drug conjugates in animal sport doping control – Development and assessment of a rapid and economical approach for screening greyhound urine

Abstract

Animal sport doping control laboratories are constantly reviewing ways in which they can improve their service offering whilst ensuring that they remain economically viable. This paper describes the development and assessment of a rapid and economical method for the detection of intact glucuronide conjugates of three anabolic steroids and their metabolites along with three corticosteroids in canine urine. The analysis of intact drug conjugates for animal sport doping control is generally not performed routinely as it presents a number of analytical challenges, not least of which is the lack of availability of appropriate reference standards. Here, we report the development of a UHPLC–MS/MS method using APCI in the negative ion mode for the detection of intact phase II conjugates, including the importance of in vitro incubations in order to provide appropriate reference materials. Cross‐validation of the developed method demonstrated that the detection capability of the intact phase II conjugates of stanozolol, boldenone, nandrolone, and their metabolites along with the corticosteroids dexamethasone and methylprednisolone was equivalent to that achieved in routine race‐day screens. The new process has been in operation for approximately 2 years and has been used to analyze in excess of 13500 canine urine samples, resulting in a number of positive screening findings. To the best of our knowledge, this is the first reported use of a routine screen for intact drug conjugates within animal sport doping control.

Development of a Standardized Microflow LC Gradient to Enable Sensitive and Long-Term Detection of Synthetic Anabolic-Androgenic Steroids for High-Throughput Doping Controls

“…Interestingly, stanozolol could be identified in the urine extract as previously reported, which supports that stanozolol can be directly metabolized into phase II N- and/or O-glucuronides conjugates by the equine enzymatic machinery. These phase II metabolites have been observed in humans and dogs and in vitro using horse S9 liver fractions …”

“…Interestingly, stanozolol could be identified in the urine extract as previously reported, 24 which supports that stanozolol can be directly metabolized into phase II N-and/or Oglucuronides conjugates by the equine enzymatic machinery. These phase II metabolites have been observed in humans 29 and dogs 30 and in vitro using horse S9 liver fractions. 31 In addition, we performed the analysis of a positive equine urine sample which was previously confirmed using an accredited confirmatory analysis method and estimated at 50 pg/mL for 16β-hydroxy-stanozolol.…”

Estudio De Residuos De Boldenona en Carne De Bovinos Faenados en El Camal Metropolitano De Quito

“…[15] En la normativa ecuatoriana y en el Codex Alimentarius no se indican los Límites Máximos Permitidos de boldenona en carne de bovinos; existe autorización de uso solamente en equinos no destinados a consumo humano y en gatos con el fin de estimular el apetito, [10] sin embargo es prohibido su uso en varios deportes equinos [16] en los que se realiza constantes controles antidopaje. [17] Así mismo, su presencia en animales para consumo humano no es permitida, siendo prohibido en Argentina, Uruguay, Venezuela, Brasil, Paraguay, entre otros, mientras que en Colombia es autorizado con un tiempo de retiro de 30 días para carne. [2] Diversos estudios realizados en torno a la presencia de boldenona en animales de abasto exponen resultados de su presencia en varios niveles y utilizando diferentes metodologías analíticas.…”