Abstract
An analytical procedure for the determination of aflatoxin B1 in eggs was introduced and validated in laboratory 1. The method consisted of the extraction of aflatoxin B1 from a sample, purification of the extract with solvents, immunoaffinity column cleanup and the determination by liquid chromatography with post-column bromination and fluorescence detection at λ ex = 362 nm and λ em = 425 nm. The method was transferred to laboratory 2, where it was modified and validated. The limit of detection (LOD) and limit of quantification (LOQ) obtained in laboratory 1 were 2 and 6 ng/kg, respectively, and 2 and 5 ng/kg in laboratory 2, respectively. The repeatability of measurements in laboratory 1, represented by differences between results of duplicate measurements, was 10 ng/kg at the contamination level of 50 ng/kg. At the same concentration level, the standard deviation (s R) and the relative standard deviation (RSD R) for the within-laboratory reproducibility were 5.5 ng/kg and 11 %, respectively, and the measurement uncertainty was ±10 ng/kg. The mean recovery was 70 %. In laboratory 2, the repeatability of measurements at the contamination level of 20 ng/kg, represented by the standard deviation (s R), repeatability (r) and relative standard deviation (RSD R) was 4 ng/kg, 11 ng/kg and 20 %, respectively, and the recovery was 67 %. The results indicate that the procedures are suitable for the determination of aflatoxin B1 in eggs and can be implemented for the routine analysis. Using the procedure validated in laboratory 1, 25 samples from farms in Slovenia were analysed. In none of the analysed samples, aflatoxin B1 was detected.
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References
AOAC International (1995) Official methods of analysis, 16th Ed. Ch. 49, Arlington, pp 31–32
Baltacı C, İlyasoğlu H, Yüksel F (2013) Single-laboratory validation for the determination of aflatoxin B1, B2, G1, and G2 in foods based on immunoaffinity column and liquid chromatography with postcolumn derivatization and fluorescence detection. Food Anal Methods 6:36–44
Bintvihok A, Thiengnin S, Doi K, Kumagai S (2002) Residues of aflatoxins in the liver, muscle and eggs, of domestic fowles. J Vet Med Sci 64:1037–1039
Brera C, Debegnach F, De Santis B, Pannunzi E, Berdini C, Prantera E, Gregori E, Miraglia M (2011) Simultaneous determination of aflatoxins and ochratoxin A in baby foods and paprika by HPLC with fluorescence detection: A single-laboratory validation study. Talanta 83:1442–1446
Cavaliere C, Foglia P, Guarino C, Nazzari M, Samperi R, Laganà A (2007) Determination of aflatoxins in olive oil by liquid chromatography−tandem mass spectrometry. Anal Chim Acta 596:141–148
Chen D, Cao X, Tao Y, Wu Q, Pan Y, Huang L, Wang X, Wang Y, Peng D, Liu Z, Yuan Z (2012) Development of a sensitive and robust liquid chromatography coupled with tandem mass spectrometry and a pressurized liquid extraction for the determination of aflatoxins and ochratoxin A in animal derived foods. J Chromatogr A 1253:110–119
Di Mavungu JD, Monbaliu S, Scippo M-L, Maghuin-Rogister G, Schneider Y-J, Larondelle Y, Callebaut A, Robbens J, Van Peteghem C, De Saeger S (2009) LC-MS/MS multi-analyte method for mycotoxin determination in food supplements. Food Addit Contam 26:885–895
Dragacci S, Grosso F, Gilbert J (2001) Immunoaffinity column cleanup with liquid chromatography for determination of aflatoxin M1 in liquid milk: collaborative study. J AOAC Int 84:437–443
EA, European co-operation for Accreditation (2003) EA guidelines on the expression of uncertainty in quantitative testing. EA-4/16 G:2003. http://www.european-accreditation.org/publications.Accessed 5 Oct 2013
European Commission (2002) Decision 2002/657/EC of 12 August 2002 implementing Council Directive 96/23/EC concerning the performance of analytical methods and the interpretation of results. Off J Eur Union L221:8–36
European Commission (2006a) Regulation(EC) No 401/2006 of 23 February 2006 laying down the methods of sampling and analysis for the official control of the levels of mycotoxins in foodstuffs. Off J Eur Union L70:12–34
European Commission (2006b) Regulation (EC) No 1881/2006 of 19 December 2006 setting maximum levels for certain contaminants in foodstuffs. Off J Eur Union L364:5–24
European Commission (2010) Regulation (EU) No 165/2010 of 26 February 2010 amending Regulation (EC) No 1881/2006 setting maximum levels for certain contaminants in foodstuffs as regards aflatoxins. Off J Eur Union L50:8–12
European Commission (2011) Regulation (EU) No 574/2011 of 16 June 2011 amending Annex I to Directive 2002/32/EC of the European Parliament and of the Council as regards maximum levels for nitrite, melamine, Ambrosia spp. and carry over of certain coccidiostats and histomonostats and consolidating Annexes I and II thereto. Off J Eur Union L159:7–24
European Commission (2013) Rapid Alert System for Food and Feed (RASFF). https://webgate.ec.europa.eu/rasff-window/portal/index.cfm?event=notificationsList. Accessed 5 Oct 2013
European Parliament the Council (2002) Directive 2002/32/EC of 7 May 2002 on undesirable substances in animal feed. Off J Eur Union L140:10–21
European Parliament the Council (2004) Regulation (EC) No 882/2004 of 29 April 2004 on official controls performed to ensure the verification of compliance with feed and food law, animal health and animal welfare rules. Off J Eur Union L191:1–52
FAO, Food and Agriculture Organization of the United Nations (2004) Worldwide regulations for mycotoxins in food and feed in 2003, FAO Food and Nutrition paper 81. FAO, Rome
FEFAC, European Feed Manufacturers’ Federation (2013) Aflatoxin B1 in Maize from South-Eastern Europe. http://www.fefac.eu/positions.aspx. Accessed 5 Oct 2013
Han RW, Zheng N, Wang JQ, Zhen YP, Xu XM, Li SL (2013) Survey of aflatoxin in dairy cow feed and raw milk in China. Food Control 34:35–39
Hassan ZU, Khan MZ, Khan A, Javed I, Hussain Z (2012) Effects of individual and combined administration of ochratoxin A and aflatoxin B1 in tissues and eggs of White Leghorn breeder hens. J Sci Food Agric 92:1540–1544
Huang B, Han Z, Cai Z, Wu Y, Re Y (2010) Simultaneous determination of aflatoxins B1, B2, G1, G2, M1 and M2 in peanuts and their derivative products by ultra-high-performance liquid chromatography−tandem mass spectrometry. Anal Chim Acta 662:62–68
Hussein HS, Brasel JM (2001) Toxicity, metabolism, and impact of mycotoxins on humans and animals. Toxicology 167:101–134
Kokkonen M, Jestoi M, Rizzo A (2005) Determination of selected mycotoxins in mould cheeses with liquid chromatography coupled to tandem with mass spectrometry. Food Addit Contam 22:449–456
Miller JN, Miller JC (2000) Statistics and Chemometrics for Analytical Chemistry. Prentice Hall, Harlow
Muscarella M, Iammarino M, Nardiello D, Lo Magro S, Palermo C, Centonze D, Palermo D (2009) Validation of a confirmatory analytical method for the determination of aflatoxins B1, B2, G1 and G2 in foods and feed materials by HPLC with on-line photochemical derivatization and fluorescence detection. Food AdditContam A 26:1402–1410
Oliveira CAF, Kobashogawa E, Alves Reis T, Mestieri L, Albuquerque R, Corrêa B (2000) Aflatoxin B1 residues in eggs of laying hens fed a diet containing different levels of the mycotoxin. Food Addit Contam 17:459–462
Oliveira CAF, Rosmaninho JF, Castro AL, Butkeraitis P, Alves Reis T, Corrêa B (2003) Aflatoxin residues in eggs of laying Japanese quail after long-term administration of rations containing low levels of aflatoxin B1. Food Addit Contam 20:648–653
R-Biopharm Rhône (2001) Application of immunoaffinity columns for sample clean-up prior to HPLC analysis for aflatoxins. Instruction for use
Rodríguez Velasco ML, Calonge Delso MM, Ordónez Escudero D (2003) ELISA and HPLC determination of the occurrence of aflatoxin M1 in raw cow’s milk. Food Addit Contam 20:276–280
Romer (2013) Aflatoxins pose new risk to European maize production. http://www.romerlabs.com/en/company/latest-news. Accessed 5 Oct 2013
Rubert J, Soler C, Mañes J (2012) Occurrence of fourteen mycotoxins in tiger-nuts. Food Control 25:374–379
Shepard GS (2009) Aflatoxin analysis at the beginning of the twenty-first century. Anal Bioanal Chem 395:1215–1224
Shepard GS, Berthiller F, Burdaspal P, Crews C, Jonker MA, Krska R, MacDonald S, Malone B, Maragos C, Sabino M, Solfrizzo M, van Egmond HP, Whitaker TB (2011) Developments in mycotoxin analysis: an update for 2009-2010. World Mycotoxin J4:3–28
Siegel D, Babuscio T (2011) Mycotoxin management in the European cereal trading sector. Food Control 22:1145–1153
Sizoo EA, van Egmond HP (2005) Analysis of duplicate 24-hour diet samples for aflatoxin B1, aflatoxin M1 and ochratoxin A. Food Addit Contam 22:163–172
Stroka J, Anklam E, Jörissen U, Gilbert J (2000) Immunoaffinity column cleanup with liquid chromatography using post-column bromination for determination of aflatoxins in peanut butter, pistachio paste, fig paste, and paprika powder: collaborative study. J AOAC Int 83:320–340
Stroka J, Anklam E, Jörissen U, Gilbert J (2001) Determination of aflatoxin B1 in baby food (infant formula) by immunoaffinity column cleanup liquid chromatography with postcolumn bromination: collaborative study. J AOAC Int 84:1116–1123
Sulyok M, Krska R, Schuhmacher R (2007) A liquid chromatography/tandem mass spectrometric multi-mycotoxin method for the quantification of 87 analytes and its application to semi-quantitative screening of mouldy food samples. AnalBioanal Chem 389:1505–1523
Tavčar-Kalcher G, Vrtač K, Pestevšek U, Vengušt A (2007) Validation of the procedure for the determination of aflatoxin B1 in animal liver using immunoaffinity columns and liquid chromatography with postcolumn derivatisation and fluorescence detection. Food Control 18:333–337
Varga E, Glauner T, Berthiller F, Krska R, Schuhmacher R, Sulyok M (2013) Development and validation of a (semi)-quantitative UHPLC-MS/MS method for the determination of 191 mycotoxins and other fungal metabolites in almonds, hazelnuts, peanuts and pistachios. Anal Bioanal Chem 405:5087–5104
Zain ME (2011) Impact of mycotoxins on humans and animals. J Saudi Chem Soc 15:129–144
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Katarina Pavšič-Vrtač declares that she has no conflict of interest. Suvi Ojanperä declares that she has no conflict of interest. Juha Apajalahti declares that he has no conflict of interest. Karin Šrimpf declares that she has no conflict of interest. Gabrijela Tavčar-Kalcher declares that she has no conflict of interest. This article does not contain any studies with human or animal subjects.
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Pavšič-Vrtač, K., Ojanperä, S., Apajalahti, J. et al. Analytical Procedures for the Determination of Aflatoxin B1 in Eggs of Laying Hens Using Immunoaffinity Columns and Liquid Chromatography with Post-Column Derivatisation and Fluorescence Detection. Food Anal. Methods 7, 1917–1924 (2014). https://doi.org/10.1007/s12161-014-9836-4
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DOI: https://doi.org/10.1007/s12161-014-9836-4