RESUMEN
Urban and rural population exposure to ochratoxin A (OTA) in central zone of Portugal was investigated in three places: Coimbra, Verride and Ereira. The analytical method proposed for the determination of ochratoxin A involved extraction with chloroform-orthophosphoric acid, cleanup through an immunoaffinity column (IAC), high performance liquid chromatography (HPLC) with spectrofluorimetric detection (FD) for separation and identification of ochratoxin A, and confirmation with HPLC-FD after OTA methylation in serum. The limit of quantification of the proposed method was 0.1 microg/L for serum and 0.05 microg/L for blood. OTA recoveries in serum ranged from 70.3% to 115.3% for levels at 0.25 microg/L and 0.5 microg/L, respectively, with a within-day RSD between 8.0% and 16.2%. Ochratoxin A serum levels were evaluated in an hundred and four donors from Coimbra city, Verride, and Ereira. The study revealed a frequency of detection of 100%. The ratio of ochratoxin A level in serum to whole blood was 2.0+/-0.7. The overall concentrations range from 0.25 to 2.49 microg/L, 0.14 to 1.91 microg/L, and 0.19 to 0.96 microg/L, for samples of Verride, Ereira, and Coimbra, respectively. The mean concentration and standard deviation were 0.78+/-0.53 microg/L, 0.44+/-0.31 microg/L, and 0.42+/-0.18 microg/L for the same samples. A significant difference was found in Verride population (P-value=0.000). Levels of OTA are clearly higher in males from rural areas than in females. For all samples, a significant difference was found in Verride male population (P-value=0.014).
Asunto(s)
Exposición a Riesgos Ambientales , Micotoxinas/sangre , Ocratoxinas/sangre , Población Rural , Población Urbana , Cromatografía Líquida de Alta Presión , Humanos , Espectrometría de FluorescenciaRESUMEN
Ochratoxin A (OTA) produced by Aspergillus and Penicillium genera contaminates several foods. OTA is nephrotoxic to all animal species studied so far, and most likely to humans, who show the longest half-life for elimination of this toxin among all examined species. OTA has other toxic effects such as teratogenicity, immunotoxicity, genotoxicity, and is also mutagenic and carcinogenic, all of which lead to life-threatening pathologies through several molecular pathways. A sensitive, specific and rapid method applying high performance liquid chromatography coupled to a spectrofluorimeter for the determination of ochratoxin A in human serum was validated. Serum samples were extracted with chloroform-orthophosphoric acid, and cleaned-up through immunoaffinity column (IAC). The separation and identification was performed by HPLC coupled to a spectrofluorimeter, and, after OTA methylation, the confirmation was achieved. Chromatographic separation of the analyte was performed on a reverse phase column with a mobile phase of water:acetonitrile:glacial acetic acid (49.5:49.5:1.0). Linearity was established between the range of 1 and 10 ng/ml. Under the optimized conditions, the recoveries were higher than 83.0% for all fortification levels. The intra-day precision oscillated between 8.0 and 5.0% at levels of 0.25 and 0.5 microg/l, while the inter-day precision was in the range of 10.7-16.0%. The limit of quantification of the method was 0.05 microg/l. The method is appropriate for quantitative determination of OTA in human serum and has been successfully applied to the analysis of OTA in haemodialysis patients from two principal cities of Portugal, in order to evaluate its exposure degree. Levels of OTA in Coimbra were higher than in Aveiro, 0.50 microg/l versus 0.49 microg/l. In respect to gender, levels of OTA were higher in males from Aveiro than in females, 0.52 microg/l versus 0.44 microg/l, and in Coimbra were similar, 0.50 microg/l versus 0.51 microg/l. However, in none of the cases, significant statistical differences were found.