title: Determination of Aflatoxin B1-Lysine in Pig Serum and Plasma by Liquid Chromatography-Tandem Mass Spectrometry.
abstract: Aflatoxin B1 (AFB1) is a hepatocarcinogen produced by certain Aspergillus species growing on crops. After biotransformation in the liver, AFB1 generates several metabolites, one of which is AFB1 bound to lysine on serum albumin. AFB1-lysine (AFB1-lys) is a digest product of AFB1-albumin and is considered a biomarker of exposure to AFB1 in humans and animals. The objectives of this paper were to evaluate the performance characteristics of a new analytical method for determination of AFB1-lys levels in pig serum, heparinized and ethylenediaminetetraacetic acid (EDTA) plasma and to evaluate the interference of these anticoagulants in AFB1-lys quantification. Blank blood samples were obtained from eight crossbreed 91-day-old barrows fed AFB1-free diets. Pooled samples (n = 3) and individual samples of serum, EDTA and heparinized plasma collected from five pigs were enzymatically digested with pronase at 37°C for 4 h. AFB1-lys was isolated by solid-phase extraction and quantified by liquid chromatography coupled to tandem mass spectrometry. The analytical method was applied for determination of AFB1-lys in serum and EDTA plasma collected from five 49-day-old crossbreed barrows fed ad libitum diets containing 1.1 mg of AFB1 per kg of feed during 7 days (three animals) or 42 days (two animals). Samples of heparinized plasma were only available from animals intoxicated for 42 days. All animals had lower levels of AFB1-lys in EDTA plasma samples (24.78-37.40 ng/mL), when compared to serum (49.32-252.07 ng/mL-1) or heparinized plasma (176.81 and 264.24 ng/mL-1). EDTA did not interfere in AFB1-lys standard detection, but our findings suggest that EDTA should be avoided during blood collection since it affects the pronase activity in AFB1-albumin adduct digestion and, consequently, causes a reduction in the AFB1-lys levels. Hence, determination of AFB1-lys in serum and heparinized plasma is an approach to assess an individual's exposure of swine to AFB1.
authors.affiliation: Department of Food Engineering, Faculty of Animal Science and Food Engineering, University of São Paulo, Av. Duque de Caxias Norte 225, CEP 13635-900 Pirassununga, SP, Brazil.
authors.forename: Mayra C
authors.initials: MC
authors.lastname: Di Gregorio
authors.affiliation: Department of Physics and Chemistry, School of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Av. do Café s/n, CEP 14040-903 Ribeirão Preto, SP, Brazil.
authors.forename: Alessandra V
authors.initials: AV
authors.lastname: Jager
authors.affiliation: Department of Food Engineering, Faculty of Animal Science and Food Engineering, University of São Paulo, Av. Duque de Caxias Norte 225, CEP 13635-900 Pirassununga, SP, Brazil.
authors.forename: Aline A
authors.initials: AA
authors.lastname: Costa
authors.affiliation: Department of Food Engineering, Faculty of Animal Science and Food Engineering, University of São Paulo, Av. Duque de Caxias Norte 225, CEP 13635-900 Pirassununga, SP, Brazil.
authors.forename: Keliani
authors.initials: K
authors.lastname: Bordin
authors.affiliation: Veterinary Medical Diagnostic Laboratory, College of Veterinary Medicine, University of Missouri, 1600 East Rollins Street, Columbia, MO 65211, USA.
authors.forename: George E
authors.initials: GE
authors.lastname: Rottinhghaus
authors.affiliation: Department of Food Engineering, Faculty of Animal Science and Food Engineering, University of São Paulo, Av. Duque de Caxias Norte 225, CEP 13635-900 Pirassununga, SP, Brazil.
authors.forename: Tânia
authors.initials: T
authors.lastname: Petta
authors.affiliation: Department of Food Engineering, Faculty of Animal Science and Food Engineering, University of São Paulo, Av. Duque de Caxias Norte 225, CEP 13635-900 Pirassununga, SP, Brazil.
authors.forename: Pollyana C M C
authors.initials: PC
authors.lastname: Souto
authors.affiliation: Institute of Animal Science and Pastures, Department of Agriculture and Food Supply of the São Paulo State, R. Heitor Penteado 56, CEP 13460-000 Nova Odessa, SP, Brazil.
authors.forename: Fabio E L
authors.initials: FE
authors.lastname: Budiño
authors.affiliation: Department of Food Engineering, Faculty of Animal Science and Food Engineering, University of São Paulo, Av. Duque de Caxias Norte 225, CEP 13635-900 Pirassununga, SP, Brazil.
authors.forename: Carlos A F
authors.initials: CA
authors.lastname: Oliveira
mesh_terms: D016604:Aflatoxin B1 / Q000097:blood*; D000821:Animal Feed / Q000032:analysis; D000818:Animals; D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods*; D005506:Food Contamination / Q000032:analysis*; D057230:Limit of Detection; D008239:Lysine / Q000097:blood*; D011485:Protein Binding; D012015:Reference Standards; D012709:Serum Albumin / Q000378:metabolism*; D013552:Swine; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D012709:Serum Albumin; C066451:aflatoxin B1-lysine adduct; D016604:Aflatoxin B1; D008239:Lysine