Simple and Sensitive Analysis of Clenbuterol in Urine Matrices by UHPLC-MS/MS Method with Online-SPE Sample Preparation

Separations

article

Kristián Slíž¹  Dominika Olešová³  Juraj Piešťanský²  Peter Mikuš¹ 

[1] Department of Pharmaceutical Analysis and Nuclear Pharmacy, Faculty of Pharmacy, Comenius University Bratislava;Toxicologic and Antidoping Centre, Faculty of Pharmacy, Comenius University Bratislava;Slovak Academy of Sciences, Biomedical Research Center, Institute of Experimental Endocrinology;Department of Galenic Pharmacy, Faculty of Pharmacy, Comenius University Bratislava

关键词: clenbuterol;    human urine;    ultra-high performance liquid chromatography;    tandem mass spectrometry;    online SPE extraction;    antidoping analysis;   

DOI  :  10.3390/separations9120440

学科分类：社会科学、人文和艺术（综合）

来源: mdpi

【 摘 要 】

Clenbuterol is one of the most misused anabolic agents in professional sports. Therefore, the monitoring of clenbuterol in body fluids such as human urine is related to the development of rapid, selective and sensitive analytical methods that produce reliable results. In this work, these requirements were met by a two-dimensional separation method based on online solid-phase extraction coupled with ultra-high performance liquid chromatography–tandem mass spectrometry (SPE–UHPLC–MS/MS). The developed method provides favorable performance parameters, and it is characterized by minimum manual steps (only dilution and the addition of an internal standard) in the sample preparation. A limit of quantification (LOQ) of 0.1 ng/mL, excellent linearity (0.9999), remarkable precision (1.26% to 8.99%) and high accuracy (93.1% to 98.7%) were achieved. From a practical point of view, the analytical performance of the validated SPE–UHPLC–MS/MS method was demonstrated on blinded spiked urine samples from ten healthy volunteers. The estimated concentrations of clenbuterol were in accordance with their corresponding nominal values, as supported by the precision and accuracy data (relative standard deviation ≤5.4%, relative error ≤11%). The fulfillment of the World Anti-Doping Agency’s screening and confirmation criteria indicates that the proposed method is suitable for implementation in routine use in toxicologic and antidoping laboratories. Due to its high orthogonality and separation efficiency, the SPE–UHPLC–MS/MS method should also be easily adapted to the separation of structurally related compounds (such as clenbuterol metabolites). Thus, future antidoping applications could also include monitoring of clenbuterol metabolites, providing a longer detection widow.

【 授权许可】

CC BY