Login | Register

Liquid Chromatography – High Resolution Mass Spectrometry Method for Monitoring of 17 Mycotoxins in Human Plasma for Exposure Studies


Liquid Chromatography – High Resolution Mass Spectrometry Method for Monitoring of 17 Mycotoxins in Human Plasma for Exposure Studies

Slobodchikova, Irina and Vuckovic, Dajana (2018) Liquid Chromatography – High Resolution Mass Spectrometry Method for Monitoring of 17 Mycotoxins in Human Plasma for Exposure Studies. Journal of Chromatography A . ISSN 00219673 (In Press)

[thumbnail of slobodchikova-2018.pdf]
Text (application/pdf)
slobodchikova-2018.pdf - Accepted Version
Available under License Spectrum Terms of Access.

Official URL: http://dx.doi.org/10.1016/j.chroma.2018.03.030


Mycotoxins are secondary metabolites produced by filamentous fungi. Primary route of human exposure to mycotoxins is the intake of the contaminated food. Minimizing mycotoxin exposure is important for population health, as their chronic toxic effects have been associated with kidney and liver diseases, some types of cancer and immunosuppression. The objective of this work was to develop and validate a multi-class mycotoxin method suitable for exposure monitoring of mycotoxins in human plasma. A sensitive liquid chromatography – mass spectrometry method was developed for 17 mycotoxins: nivalenol (NIV), deoxynivalenol, fusarenon X, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, T-2 toxin, HT-2 toxin, aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, zearalenone, α-zearalenol (α-ZOL), β-zearalenol, zearalanone, α-zeranoland, and β-zeranol. The method relies on three-step liquid-liquid extraction with ethyl acetate to eliminate the need for immunoaffinity extraction and minimize ionization matrix effects. Chromatographic separation of mycotoxins, including all isomers, was achieved with pentafluorophenyl column and water/methanol mobile phase. Mycotoxin detection and quantitation were performed using high-resolution mass spectrometry on LTQ Velos Orbitrap, in both positive and negative electrospray ionization (ESI(+) and (ESI(−)). The use of 0.02% acetic acid as mobile phase additive for ESI(−) resulted in significant increase in ionization efficiency ranging from 1.7 to 26 times for mycotoxins that ionize better in ESI(−). The optimized method was validated according to FDA guidance procedures. LOQs of all mycotoxins ranged from 0.1 to 0.5 ng/ml, except NIV which resulted in LOQ of 3 ng/ml because of low extraction recovery of this highly polar mycotoxin. Mean intra-day accuracy ranged from 85.8% to 116.4%, and intra-day precision (n = 6) ranged from 1.6% to 12.5% RSD for all mycotoxins except α-ZOL where mean accuracy ranged from 72.9% to 97.2%. Inter-day accuracy and precision were 85.6% to 111.5% and 2.7 to 15.6% RSD respectively, showing good analytical performance of the method for biomonitoring.

Divisions:Concordia University > Faculty of Arts and Science > Chemistry and Biochemistry
Item Type:Article
Authors:Slobodchikova, Irina and Vuckovic, Dajana
Journal or Publication:Journal of Chromatography A
Date:15 March 2018
  • Fonds du Recherche du Quebec
  • Centre for Biological Applications of Mass Spectrometry at Concordia University
Digital Object Identifier (DOI):10.1016/j.chroma.2018.03.030
Keywords:Mycotoxins; High-resolution mass spectrometry; Human plasma; Pentafluorophenyl; Biomonitoring; Validation
ID Code:983595
Deposited By: Michael Biron
Deposited On:21 Mar 2018 20:31
Last Modified:15 Mar 2020 00:00


A.L. Capriotti, G. Caruso, C. Cavaliere, P. Foglia, R. Samperi, A. Laganà Multiclass mycotoxin analysis in food, environmental and biological matrices with chromatography/mass spectrometry Mass Spectrom. Rev., 31 (2012), pp. 466–503

H. Fromme, M. Gareis, W. Völkel, C. Gottschalk Overall internal exposure to mycotoxins and their occurrence in occupational and residential settings – An overview Int. J. Hyg. Environ. Health, 219 (2016), pp. 143–165

G.A. Lombaert, P. Pellaers, V. Roscoe, M. Mankotia, R. Neil, P.M. Scott Mycotoxins in infant cereal foods from the Canadian retail market Food Addit.Contam., 20 (2003), pp. 494–504

V. Roscoe, G. Lombaert, V. Huzel, G. Neumann, J. Melietio, D. Kitchen, S. Kotello, T. Krakalovich, R. Trelka, P.M. Scott Mycotoxins in breakfast cereals from the Canadian retail market: a 3-year survey, Food Addit Contam. Part A. Chem. Anal. Control. Expo. Risk Assess., 25 (2008), pp. 347–355

G. Lombaert, P. Pellaers, G. Neumann, D. Kitchen, V. Huzel, R. Trelka, S. Kotello, P.M. Scott Ochratoxin A in dried vine fruits on the Canadian retail market Food Addit. Contam., 21 (2004), pp. 578–585

L. Ng, M. Mankotia, P. Pantazopoulos, R.J. Neil, P.M. Scott Ochratoxin A in wine and grape juice sold in Canada Food Addit. Contam., 21 (2004), pp. 971–981

S.T. Tran, T.K. Smith, G.N. Girgis A survey of free and conjugated deoxynivalenol inthe 2008 corn crop in Ontario, Canada J. Sci. Food Agric., 92 (2012), pp. 37–41

K. Muñoz, M. Blaszkewicz, G.H. Degen Simultaneous analysis of ochratoxin A and its major metabolite ochratoxin alpha in plasma and urine for an advanced biomonitoring of the mycotoxin J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 878 (2010), pp. 2623–2629

E. Märtlbauer, E. Usleber, R. Dietrich, E. Schneider Ochratoxin A in human blood serum – Retrospective long-term data Mycotoxin Res., 25 (2009), pp. 175–186

N.W. Turner, S. Subrahmanyam, S.A. Piletsky Analytical methods for determination of mycotoxins: A review Anal. Chim. Acta, 632 (2009), pp. 168–180

N.W. Turner, H. Bramhmbhatt, M. Szabo-Vezse, A. Poma, R. Coker, S.A. Piletsky Analytical methods for determination of mycotoxins: an update (2009–2014) Anal. Chim. Acta, 901 (2015), pp. 12–33

E. Heyndrickx, I. Sioen, M. Bellemans, M. De Maeyer, A. Callebaut, S. De Henauw, S. De Saeger Assessment of mycotoxin exposure in the Belgian population using biomarkers: Aim, design and methods of the BIOMYCO study Food Addit. Contam. – Part A, 31 (2014), pp. 924–931

N. Ali, K. Muñoz, G.H. Degen Ochratoxin A and its metabolites in urines of German adults—an assessment of variables in biomarker analysis Toxicol. Lett., 275 (2017), pp. 19–26

P.F. Scroll, J.D. Groopman Long-term stability of human aflatoxin B1 albumin adducts assessed by isotope dilution mass spectrometry and high-performance liquid chromatography-fluorescence Cancer Epidemiol Biomarkers Prev., 17 (2008), pp. 1436–1439

G. Shephard, P. Thiel, E. Sydenham Initial studies on the toxicokinetics of fumonisin B1 in rats Food Chem. Toxicol., 30 (1992), pp. 277–279

Z. Han, Z. Zhao, S. Song, G. Liu, J. Shi, J. Zhang, Y. Liao, D. Zhang, Y. Wu, S. De Saeger, A. Wu Establishment of an isotope dilution LC-MS/MS method revealing kinetics and distribution of co occurring mycotoxins in rats Anal. Methods, 4 (2012), p. 3708

B. Huybrechts, J.C. Martins, P. Debongnie, S. Uhlig, A. Callebaut Fast and sensitive LC?MS/MS method measuring human mycotoxin exposure using biomarkers in urine Arch. Toxicol., 89 (2015), pp. 1993–2005

E. Heyndrickx, I. Sioen, B. Huybrechts, A. Callebaut, S. De Henauw, S. De Saeger Human biomonitoring of multiple mycotoxins in the Belgian population: Results of the BIOMYCO study Environ. Int., 84 (2015), pp. 82–89

J. Gerding, N. Ali, J. Schwartzbord, B. Cramer, D.L. Brown, G.H. Degen, H.-U. Humpf A comparative study of the human urinary mycotoxin excretion patterns in Bangladesh, Germany, and Haiti using a rapid and sensitive LC-MS/MS approach Mycotoxin Res. (2015), pp. 127–136

M. Solfrizzo, L. Gambacorta, A. Visconti Assessment of multi-mycotoxin exposure in southern Italy by urinary multi-biomarker determination Toxins (Basel), 6 (2014), pp. 523–538

S. Wallin, L. Gambacorta, N. Kotova, E. Warensjö Lemming, C. Nälsén, M. Solfrizzo, M. Olsen Biomonitoring of concurrent mycotoxin exposure among adults in Sweden through urinary multi-biomarker analysis Food Chem. Toxicol., 83 (2015), pp. 133–139

E. Njumbe Ediage, J. Diana Di Mavungu, S. Song, A. Wu, C. Van Peteghem, S. De Saeger A direct assessment of mycotoxin biomarkers in human urine samples by liquid chromatography tandem mass spectrometry Anal. Chim. Acta, 741 (2012), pp. 58–69

B. Warth, M. Sulyok, P. Fruhmann, H. Mikula, F. Berthiller, R. Schuhmacher, C. Hametner, W.A. Abia, G. Adam, J. Fröhlich, R. Krska Development and validation of a rapid multi-biomarker liquid chromatography/tandem mass spectrometry method to assess human exposure to mycotoxins Rapid Commun. Mass Spectrom., 26 (2012), pp. 1533–1540

B. Warth, M. Sulyok, R. Krska LC-MS/MS-based multibiomarker approaches for the assessment of human exposure to mycotoxins Anal. Bioanal. Chem., 405 (2013), pp. 5687–5695

J. Rubert, N. León, C. Sáez, C.P.B. Martins, M. Godula, V. Yusà, J. Mañes, J.M. Soriano, C. Soler Evaluation of mycotoxins and their metabolites in human breast milk using liquid chromatography coupled to high resolution mass spectrometry Anal. Chim. Acta, 820 (2014), pp. 39–46

Z. Zhao, N. Liu, L. Yang, Y. Deng, J. Wang, S. Song, S. Lin, A. Wu, Z. Zhou, J. Hou Multi-mycotoxin analysis of animal feed and animal-derived food using LC-MS/MS system with timed and highly selective reaction monitoring Anal. Bioanal. Chem., 407 (2015), pp. 7359–7368

S. De Baere, A. Osselaere, M. Devreese, L. Vanhaecke, P. De Backer, S. Croubels Development of a liquid-chromatography tandem mass spectrometry chromatography high-resolution mass spectrometry method for the quantitative determination of zearalenone and its major metabolites in chicken and pig plasma Anal. Chim. Acta, 756 (2012), pp. 37–48

A.B. Serrano, A.L. Capriotti, C. Cavaliere, S. Piovesana, R. Samperi, S. Ventura, A. Laganà Development of a rapid LC-MS/MS method for the determination of emerging fusarium mycotoxins enniatins and beauvericin in human biological fluids Toxins (Basel), 7 (2015), pp. 3554–3571

M. Devreese, S. De Baere, P. De Backer, S. Croubels Quantitative determination of several toxicological important mycotoxins in pig plasma using multi-mycotoxin and analyte-specific high performance liquid chromatography-tandem mass spectrometric methods J. Chromatogr. A, 1257 (2012), pp. 74–80

B. De Santis, M.E. Raggi, G. Moretti, F. Facchiano, A. Mezzelani, L. Villa, A. Bonfanti, A. Campioni, S. Rossi, S. Camposeo, S. Soricelli, G. Moracci, F. Debegnach, E. Gregori, F. Ciceri, L. Milanesi, A. Marabotti, C. Brera Study on the association among mycotoxins and other variables in children with autism Toxins (Basel), 9 (2017), pp. 1–20

J. Tolosa, G. Font, J. Mañes, E. Ferrer Multimycotoxin analysis in water and fish plasma by liquid chromatography-tandem mass spectrometry Chemosphere, 145 (2016), pp. 402–408

B. Osteresch, S. Viegas, B. Cramer, H.U. Humpf Multi-mycotoxin analysis using dried blood spots and dried serum spots Anal. Bioanal. Chem., 409 (2017), pp. 3369–3382

G. Schatzmayr, E. Streit Global occurrence of mycotoxins in the food and feed chain: facts and figures World Mycotoxin J., 6 (2013), pp. 213–222

Food and Drug Administration Draft Guidance for Industry Bioanalytical Method Validation (2013)

A. Breidbach The Impact of Superficially Porous Particles and New Stationary-Phase Chemistries on the LC-MS Determination of Mycotoxins in Food and Feed LC GC, 34 (2016), pp. 10–14

D. Baker, C. Titman, N. Loftus, J. Horner Multi-residue analysis of 18 regulated mycotoxins by LC-MS/MS, ASMS 2017 TP-185 (2017), pp. 1–8

D. Qi, T. Fei, H. Liu, H. Yao, D. Wu, B. Liu Development of Multiple Heart-Cutting Two-Dimensional Liquid Chromatography Coupled to Quadrupole-Orbitrap High Resolution Mass Spectrometry for Simultaneous Determination of Aflatoxin B1, B2, G1, G2, and Ochratoxin A in Snus, a Smokeless Tobacco Product J. Agric. Food Chem., 65 (2017), pp. 9923–9929

B. De Santis, F. Debegnach, E. Gregori, S. Russo, F. Marchegiani, G. Moracci, C. Brera Development of a LC-MS/MS method for the multi-mycotoxin determination in composite cereal-based samples Toxins (Basel), 9 (2017), p. 169

S. Song, E.N. Ediage, A. Wu, S. De Saeger Development and application of salting-out assisted liquid/liquid extraction for multi-mycotoxin biomarkers analysis in pig urine with high performance liquid chromatography/tandem mass spectrometry J. Chromatogr. A, 1292 (2013), pp. 111–120

F. Soleimany, S. Jinap, F. Abas Determination of mycotoxins in cereals by liquid chromatography tandem mass spectrometry Food Chem., 130 (2012), pp. 1055–1060

U. Brezina, H. Valenta, I. Rempe, S. Kersten, H.U. Humpf, S. Dänicke Development of a liquid chromatography tandem mass spectrometry method for the simultaneous determination of zearalenone, deoxynivalenol and their metabolites in pig serum Mycotoxin Res., 30 (2014), pp. 171–186

Z. Wu, W. Gao, M.A. Phelps, D. Wu, D.D. Miller, J.T. Dalton Favorable Effects of Weak Acids on Negative-Ion Electrospray Ionization Mass Spectrometry Changes, 76 (2004), pp. 839–847

X. Zhang, M.R. Clausen, X. Zhao, H. Zheng, H.C. Bertram Enhancing the power of liquid chromatography-mass spectrometry-based urine metabolomics in negative ion mode by optimization of the additive Anal. Chem., 84 (2012), pp. 7785–7792

D.S. Wishart, D. Tzur, C. Knox, R. Eisner, A.C. Guo, N. Young, D. Cheng, K. Jewell, D. Arndt, S. Sawhney, C. Fung, L. Nikolai, M. Lewis, M.A. Coutouly, I. Forsythe, P. Tang, S. Shrivastava, K. Jeroncic, P. Stothard, G. Amegbey, D. Block, D.D. Hau, J. Wagner, J. Miniaci, M. Clements, M. Gebremedhin, N. Guo, Y. Zhang, G.E. Duggan, G.D. MacInnis, A.M. Weljie, R. Dowlatabadi, F. Bamforth, D. Clive, R. Greiner, L. Li, T. Marrie, B.D. Sykes, H.J. Vogel, L. Querengesser HMDB: The human metabolome database Nucleic Acids Res., 35 (2007), pp. 521–526

E. Lim, A. Pon, Y. Djoumbou, C. Knox, S. Shrivastava, A.C. Guo, V. Neveu, D.S. Wishart T3DB: A comprehensively annotated database of common toxins and their targets Nucleic Acids Res., 38 (2009), pp. 781–786

J. Schenzel, R.P. Schwarzenbach, T.D. Bucheli Multi-residue screening method to quantify mycotoxins in aqueous environmental samples J. Agric. Food Chem., 58 (2010), pp. 11207–11217

J. Diana Di Mavungu, S. Monbaliu, M.-L. Scippo, G. Maghuin-Rogister, Y.-J. Schneider, Y. Larondelle, A. Callebaut, J. Robbens, C. Van Peteghem, S. De Saeger LC-MS/MS multi-analyte method for mycotoxin determination in food supplements Food Addit. Contam. Part A. Chem. Anal. Control. Expo. Risk Assess., 26 (2009), pp. 885–895

M. Solfrizzo, L. Gambacorta, V.M.T. Lattanzio, S. Powers, A. Visconti Simultaneous LC-MS/MS determination of aflatoxin M 1, ochratoxin A, deoxynivalenol, de-epoxydeoxynivalenol, α and β-zearalenols and fumonisin B 1 in urine as a multi-biomarker method to assess exposure to mycotoxins Anal. Bioanal. Chem., 401 (2011), pp. 2831–2841

E. Anil, I.M. Alkis Ochratoxin A and Brewing Technology: A Review J. Inst. Brew., 116 (2010), pp. 23–32

L. Escrivá, L. Manyes, G. Font, H. Berrada Mycotoxin analysis of human urine by LC-MS/MS: a comparative extraction study Toxins (Basel), 9 (2017)

H. Belhassen, I. Jiménez-Díaz, R. Ghali, H. Ghorbel, J.M. Molina-Molina, N. Olea, A. Hedili Validation of a UHPLC-MS/MS method for quantification of zearalenone, α-zearalenol, β-zearalenol, α-zearalanol, β-zearalanol and zearalanone in human urine J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 962 (2014), pp. 68–74

P.D. Andrade, J.L.G. da Silva, E.D. Caldas Simultaneous analysis of aflatoxins B1, B2, G1, G2, M1 and ochratoxin A in breast milk by high-performance liquid chromatography/fluorescence after liquid-liquid extraction with low temperature purification (LLE-LTP) J. Chromatogr. A, 1304 (2013), pp. 61–68

N. Fabregat-Cabello, P. Zomer, J.V. Sancho, A.F. Roig-Navarro, H.G. Mol Comparison of approaches to deal with matrix effects in LC-MS/MS based determinations of mycotoxins in food and feed World Mycotoxin J., 9 (2016), pp. 149–161

P.A.M. Gonzalo, J. Diaz, M. Sandra, Cepeda Stability of aflatoxins in solution J. AOAC Int., 95 (2012), pp. 1084–1088

A. Santini, R. Ferracane, G. Meca, A. Ritieni Comparison and improvement of the existing methods for the determination of aflatoxins in human serum by LC-MS/MS Anal. Methods, 2 (2010), p. 884

X. Cao, X. Li, J. Li, Y. Niu, L. Shi, Z. Fang, T. Zhang, H. Ding Quantitative determination of carcinogenic mycotoxins in human and animal biological matrices and animal-derived foods using multi-mycotoxin and analyte-specific high performance liquid chromatography-tandem mass spectrometric methods J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 1073 (2018), pp. 191–200

Z.-Y. Chen, S. Ying, J.-H. Liu, P.-P. Zhan, Y.-G. Zhao PRiME pass-through cleanup for the fast determination of aflatoxins in human serum by using LC-MS/MS Anal. Methods., 8 (2016), pp. 1457–1462

L.A. Corcuera, M. Ibáñez-Vea, A. Vettorazzi, E. González-Peñas, A.L. de Cerain Validation of a UHPLC-FLD analytical method for the simultaneous quantification of aflatoxin B1 and ochratoxin a in rat plasma, liver and kidney J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 879 (2011), pp. 2733–2740

S. De Baere, J. Goossens, A. Osselaere, M. Devreese, V. Vandenbroucke, P. De Backer, S. Croubels Quantitative determination of T-2 toxin, HT-2 toxin, deoxynivalenol and deepoxy-deoxynivalenol in animal body fluids using LC-MS/MS detection J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 879 (2011), pp. 2403–2415

Y. Sun, G. Zhang, H. Zhao, J. Zheng, F. Hu, B. Fang Liquid chromatography-tandem mass spectrometry method for toxicokinetics, tissue distribution, and excretion studies of T-2 toxin and its major metabolites in pigs J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 958 (2014), pp. 75–82

N. Broekaert, M. Devreese, T. De Mil, S. Fraeyman, S. De Baere, S. De Saeger, P. De Backer, S. Croubels Development and validation of an LC-MS/MS method for the toxicokinetic study of deoxynivalenol and its acetylated derivatives in chicken and pig plasma J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 971 (2014), pp. 43–51

P. Songsermsakul, G. Sontag, M. Cichna-Markl, J. Zentek, E. Razzazi-Fazeli Determination of zearalenone and its metabolites in urine, plasma and faeces of horses by HPLC-APCI-MS J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 843 (2006), pp. 252–261

A. Mally, M. Solfrizzo, G.H. Degen Biomonitoring of the mycotoxin Zearalenone: current state-of-the art and application to human exposure assessment Arch. Toxicol., 90 (2016), pp. 1281–1292
All items in Spectrum are protected by copyright, with all rights reserved. The use of items is governed by Spectrum's terms of access.

Repository Staff Only: item control page

Downloads per month over past year

Research related to the current document (at the CORE website)
- Research related to the current document (at the CORE website)
Back to top Back to top