Analytica Chimica Acta 468 (2002) 43–52 Application of the luminol chemiluminescence reaction for the determination of nine corticosteroids Y. Iglesias a , C. Fente a , B.I. Vázquez a,∗ , C. Franco a , A. Cepeda a , S. Mayo b a Laboratorio de Higiene, Inspección y Control de Alimentos, Departamento de Qu´ ımica Anal´ ıtica, Nutrición y Bromatolog´ ıa, Facultad de Veterinaria. Universidad de Santiago de Compostela, E-27002 Lugo, Spain. b Laboratorio de Salud Pública de Lugo, Conseller´ ıa de Salud Pública de Lugo, Conseller´ ıa de Sanidade e Benestar Social, Xunta de Galicia, E-27071 Lugo, Spain Received 12 March 2002; received in revised form 1 July 2002; accepted 2 July 2002 Abstract A luminol chemiluminescence (CL) reaction was applied for the simultaneous determination of nine corticosteroids (triamci- nolone, prednisolone, prednisone, cortisone, betamethasone, dexamethasone, corticosterone, flumethasone and triamcinolone acetonide) using flow injection. The luminol solution was prepared in NaOH with hexacyanoferrate(II) and hexacyanofer- rate(III) was used as oxidant. Adequate isocratic chromatographic separation was obtained using a simple mobile phase (acetonitrile–water). The kinetic aspects of CL reactions are very important in the sense that maximum intensity occurs simul- taneous to the arrival of the substance in the detector cell. Here we determined the moment at which maximal luminescence is reached for the nine corticosteroids assayed. When optimal experimental conditions were used, the lowest range injected was 15 mg/l for prednisolone, prednisone and cortisone, and the highest was 2 mg/l for betamethasone. Reproducibility ranged from 2.59 to 7.59 and repeatability from 1.50 to 3.61 for the different corticosteroids. © 2002 Elsevier Science B.V. All rights reserved. Keywords: Chromatography; Chemiluminescence; Luminol; Corticosteroids; Kinetic studies 1. Introduction Glucocorticoids are compounds widely used in vet- erinary medicine. Their illicit use as feed additives in animal fattening is well known, despite the fact that they are illegal for such purposes. However, the reason for such fraudulent practice is that they affect water retention in meat and also lipid metabolism, facilitat- ing the effects of other growth promoters. Glucocorti- coids increase the number and affinity of -adrenergic ∗ Corresponding author. Tel.: +34-98-225-4592; fax: +34-98-225-4592. E-mail address: bvbelda@correo.lugo.usc.es (B.I. V´ azquez). receptors and prevent receptor down-regulation and tachyphylaxis [1]. The consumption of meat contain- ing residues of these substances is a threat to human health and hence highly sensitive analytical multi-drug methods designed to measure very low levels of glu- cocorticoids are necessary. Current analytical techniques available for the analysis of these substances include thin-layer chro- matography [2–6], high performance liquid chro- matography with diode-array detection [6–8], as well as techniques involving liquid chromatography and gas chromatography/mass spectrometry [9–14]. Spe- cific immunoassay methods have also been developed and improved [15–17]. 0003-2670/02/$ – see front matter © 2002 Elsevier Science B.V. All rights reserved. PII:S0003-2670(02)00598-6