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The aim of the study was to evaluate the adequacy of enzyme-linked immunosorbent assay (ELISA) in the post-exposure determination of the β-agonists clenbuterol and salbutamol in animal plasma and serum. Experimental guinea pigs (n = 20) were treated with two doses (0.25 and 2.5 mg/kg) of clenbuterol (n = 10) and salbutamol (n = 10) for seven days, whereas the control animal group (n = 10) was left untreated. Validation of the applied method yielded acceptable recovery (mean > 70%) and repeatability rates, showing ELISA to be applicable for the semi-quantitative determination of both analytes in both matrices, preferably in plasma. In both matrices, clenbuterol concentrations were proven to be significantly (14-fold) higher than those of salbutamol. Concentrations of both analytes were higher in plasma than in serum. The application of a 10-fold higher clenbuterol and salbutamol dose (2.5 mg/kg) resulted in concentrations 3- to 4-fold higher for clenbuterol and 2- to 3-fold higher for salbutamol, indicating a different release rate of these two β-agonists.

Hair has been shown to be an excellent site for the accumulation of different drugs including β-agonists, and therefore, it would be an appropriate matrix for surveillance for the presence of drug residues. The aim of this study was to determine concentrations and to compare accumulation of two different β-agonists in black and white mice hair by use of ELISA as a screening quantitative method. The study included 200 8-week-old white and black mice. One group of black mice and one group of white mice were treated with clenbuterol in a dose of 2.5 mg/kg body mass per os for 28 days. Other animals were treated in the same way with salbutamol. The highest (±SD) clenbuterol concentration of 631.4 ± 23.5 ng/g in black hair and 228.5 ± 156.2 ng/g in white hair was determined on day 1 of treatment withdrawal. Study results revealed the black-to-white hair ratio of clenbuterol accumulation to be 1:2-1:4 and of salbutamol accumulation 1:1.4. The mean (±SD) salbutamol concentrations determined on day 1 of treatment withdrawal was 23.9 ± 0.9 ng/g and 16.4 ± 1.1 ng/g in black and white hair samples, respectively. The study demonstrated that residues could be determined in hair samples even after a 30-day withdrawal period.

The aim of the present study was to assess the persistence of clenbuterol residues in retinal tissue of pigs after repeated administration in a growth-promoting dose, using enzyme-linked immunosorbent assay (ELISA) as a screening method for quantitative determination. A growth-promoting dose of clenbuterol (20 μg/kg body mass per day) was administered orally to the experimental group (n = 6) for 21 days, whereas control animals (n = 3) were left untreated. Clenbuterol-treated pigs were randomly sacrificed (n = 3) on days 0 and 45 of treatment discontinuation, and clenbuterol residues were determined in retinal tissue dissected from the eye. ELISA was found to be acceptable for quantitative determination of clenbuterol in retinal samples because previous method validation yielded mean recovery values of 84.3-96.5% with variation coefficients < 14%. The mean (± SD) retinal clenbuterol concentration was 1874 ± 114 ng/g immediately upon clenbuterol withdrawal (day 0) and 73 ± 4 ng/g on the last day post-withdrawal (day 45). Study results pointed to a very high potential of clenbuterol accumulation in retinal tissue and marked persistence of clenbuterol residues upon anabolic dose administration, suggesting retinal tissue to be a very useful matrix for effective control of residual clenbuterol in food-producing pigs.

The paper reviews the literature data on the β-adrenergic agonists as a group of substances with anabolic effect in animals. The chemical structures of individual compounds are displayed as their basic properties, mechanism of action, physiological effects and adverse effects in humans and animals caused by exposure to anabolic doses of these substances. β- agonists in the body achieve the effect by binding to specific β-adrenergic receptors located on cell membranes of target tissues. Their use in anabolic dose in animals results in a significant increase in muscle mass and a decrease in body fat mass, better utilization of food, increased growth of animals and improved organoleptic properties of meat produced. In the last two decades, in some countries of the European Union, β-agonists were illegally used during the fattening of animals, with the aim to achieve significant yield in meat production and higher economic profit. A misuse of the clenbuterol as the most important representative has caused adverse effects on human and animal health. In the European Union, as well as in Croatia, the use of these substances with anabolic purposes in animals for meat production is prohibited, and the control of abuse is carried out through national residue monitoring. [ABSTRACT FROM AUTHOR]