Journal of Chromatography B, 1036 (2016) 76–83 Contents lists available at ScienceDirect Journal of Chromatography B journal homepage: www.elsevier.com/locate/chromb Analysis of beta-agonist residues in bovine hair: Development of a UPLC–MS/MS method and stability study Luigi Giannetti a,∗ , Giovanni Ferretti b , Valentina Gallo a , Francesco Necci a , Andrea Giorgi a , Francesca Marini a , Elisa Gennuso a , Bruno Neri a a Istituto Zooprofilattico Sperimentale Regioni Lazio e Toscana Via Appia Nuova, 1411 - 00178 Rome, Italy b Agenzia Italiana del Farmaco Via del Tritone, 181 - 00187 Rome, Italy a r t i c l e i n f o Article history: Received 15 June 2016 Received in revised form 22 September 2016 Accepted 27 September 2016 Available online 28 September 2016 Keywords: -Agonist UPLC–MS/MS Validation Bovine hair Commission decision 2002/657/EC a b s t r a c t Beta-sympathomimetic compounds are widely used in therapy because of the bronchodilator, smooth muscle-relaxant and tocolytic properties. However, their growth promoting and performance enhanc- ing effects are often subject to illegal use. The present work describes the development of a fast and reliable analytical multiresidue method for the confirmation 20 -agonist compounds in animal hair. The procedure is based on alkaline digestion, LLE with organic solvents, SPE clean up and liq- uid chromatography–tandem mass spectrometry (UPLC–MS/MS) analysis, and is suitable for the public Official control of -agonist residues in hair sample. Validation was performed according to Commission Decision 2002/657/EC requirements. Independent samples spiked with the investigated compounds in the range 0.2–10.0 g kg −1 are showing intra-day and inter-day precision (RSD) lower than 17.8% and 19.7%, respectively. Linearity, measured in the range of 0.1–10.0 g kg −1 , resulted with a Pearson’s r > 0.996. The decision limits (CC) for the all investigated beta agonists resulted in the range 0.2–1.0 g kg −1 . Furthermore, the method was tested on real hair samples obtained from cattle, known as positive to clenbuterol, in order to check its effectiveness and the -agonists stability. Published by Elsevier B.V. 1. Introduction The -agonists are phenyl-ethanolamines derivative com- pounds related to catecholamines, having different substituents on the aromatic ring and on the terminal amino group, and endowed with a powerful pharmacological activity. They are mainly used in human and veterinary medicine as bronchodilator and tocolytic agents. Moreover, due to the anabolic effects exerted when higher doses are administered to the animals, these compounds are also used as growth promoters in livestock production: these molecules contribute to increase the animals’ ability to efficiently turn what they eat into lean muscle rather than fat. Hence, in order to ascer- tain the high quality of food products of animal origin and to protect consumers, the presence of such residues in animal food products is a critical public health concern: the European Union banned the use of -agonists in food-producing animals since 1996 Directive 96/22/EC [1], except for tocolysis in mares and cows and bronchodi- ∗ Corresponding author. E-mail address: luigi.giannetti@izslt.it (L. Giannetti). lation in horses. In other 22 countries, included USA, a -agonist belonging to the phenolic group, ractopamine, has been approved for finishing swine, furthermore zilpaterol has been licensed for zootechnical use in South Africa and Mexico. Several cases of intoxication causing distal tremors, palpitations, headache, facial erythema, tachipnoea-dyspnoea nausea, vomit and cramps were described [2]. The European Community requires that each unauthorized and forbidden substance be detected and confirmed according to Com- mission Decision 2002/657/EC [3]. The detection of veterinary drug residues is generally performed on urine, plasma or tissue samples (muscle, liver, kidney); however, in the last decade various litera- ture studies evidenced that hair could be considered a useful matrix for the determination of drugs. It has been widely reported that many doping agents, therapeutic compounds and drugs of abuse are subject to accumulation in hair (human and animal) [4–6], and some scientific papers described the application of hair samples for the detection of fraudulent uses of -agonists in animals, in particular for the analysis of clenbuterol [7,8]. Hair is a useful tissue in veterinary control because of the clear advantages over classical biological matrices (e.g. urine, plasma, http://dx.doi.org/10.1016/j.jchromb.2016.09.041 1570-0232/Published by Elsevier B.V.