ORIGINAL PAPER Determination of heterocyclic aromatic amines by capillary high-performance liquid chromatography with diode array detection in ready-to-eat cooked ham treated with electron-beam irradiation N. Rosales-Conrado & M. E. León-Gonzáles & L. V. Pérez-Arribas & L. M. Polo-Díez Received: 8 October 2007 / Revised: 14 December 2007 / Accepted: 20 December 2007 / Published online: 1 February 2008 # Springer-Verlag 2008 Abstract Heterocyclic aromatic amines (HAs) are a group of mutagenic and carcinogenic substances present in significant amounts in cooked meat and fish that can potentially be formed during food processing operations. This paper proposes a capillary liquid chromatography method with diode array detection for the trace-level determination of three HAs, namely, MeIQx (2-amino-3,8- dimethylimidazo[4,5-f]quinoxaline), norharman (9H-pyrido [3,4-b]indole) and harman (1-methyl-9H-pyrido[3,4-b]in- dole), in ready-to-eat (RTE) cooked ham processed by electron-beam (accelerated electrons) irradiation to elimi- nate pathogenic microorganisms and to extend its shelf-life. The HAs selected have frequently been detected and quantified in a wide range of food and could be potential markers to indicate the presence of these toxic compounds. The method is based on the separation in an Inertsil C 8 capillary column (150 mm×0.3-mm internal diameter, 3 μm) by gradient elution mode using a mixture of acetonitrile and 30 mM ammonium acetate pH 4.5 buffer as the mobile phase. Detection was at 250 and 265 nm and, to improve sensitivity, large injection volumes (20 μL) and on-column focusing techniques based on the injection of HA samples in low organic solvent strength solutions were employed. A simple and short solid-phase extraction and purification procedure was also optimized for sample preparation. Nonirradiated and irradiated RTE cooked ham samples at doses between 1 and 8 kGy were analyzed. HAs were not detected in any of the samples analyzed; so both types of samples were spiked at concentration levels in the range 5–25 ng g -1 , which may be found in meat products. The quality parameters of the method developed in the food matrix were established, and detection limits around 0.3 ng g -1 were obtained. Spiked recoveries between 70 and 79% (n =3 for each spiked level) relative standard deviations between 1 and 5% were also obtained, showing the effectiveness of the proposed method. Keywords Heterocyclic amines . Capillary liquid chromatography . Irradiation . Ready-to-eat foods . Cooked ham Introduction Heterocyclic aromatic amines (HAs) are potent mutagenic and carcinogenic compounds formed in protein-rich foods during their cooking under ordinary conditions [1]. The amount and variety of HAs present in foods depend on many factors, such as processing conditions (time and temperature), the presence of relative amounts of precur- sors, enhancers and inhibitors, lipids, antioxidants and water content [2, 3]. To date, more than 20 HAs have been isolated and characterized from different cooked food samples and several model systems [4, 5]; the population is continuously exposed to these compounds through diet, as demonstrated by their detection in a wide variety of commercial prepared food and homemade cooked food at levels between 1 and 100 ng g -1 [6]. On the other hand, the great change of the dietary habits of the population and the food quality demands of Anal Bioanal Chem (2008) 391:1433–1442 DOI 10.1007/s00216-007-1826-6 N. Rosales-Conrado (*) : M. E. León-Gonzáles : L. V. Pérez-Arribas : L. M. Polo-Díez Dpto. Química Analítica, Facultad de Ciencias Químicas, Universidad Complutense de Madrid, 28040 Madrid, Spain e-mail: noerosales@quim.ucm.es