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Determination of Ochratoxin A and Its Metabolite Ochratoxin Alpha in Different Food Matrices After Enzymatic Biotransformation

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Abstract

Ochraxotina A (OTA) is a mycotoxin which has carcinogenic, nephrotoxic, and teratogenic effects, and it is found at different levels and matrices. As consequence, the development of methods capable of reducing this contamination to levels permitted by legislation is encouraged, with emphasis on the enzymatic process that enables the hydrolysis of OTA to a nontoxic metabolite (OTα) under mild reaction conditions. Therefore, the objectives of this work were to optimize the DLLME (dispersive liquid–liquid microextraction) for OTA and OTα determination and to study the kinetic of OTA enzymatic hydrolyses intending its application in the food chain. First, the optimization of the DLLME extraction technique for OTA and OTα was performed, obtaining as an ideal condition the use of 100 μL of 1-octanol (extractor solvent) and 2000 μL of ethyl acetate (dispersant solvent). After the validation, the proposed method was successfully applied during the optimization of OTA biotransformation into OTα. The ideal conditions for OTA hydrolysis were the use of pancreatin (ratio 1 ng OTA:2.7 mg protein) at 50 °C. At these conditions, a kinetic study was conducted and a prediction model for OTA degradation was established based on logarithmic curves. In this study, an innovative method for OTA and OTα detection was developed and with this method, it was possible to study the kinetic of OTA hydrolysis in a fast, reliable, safe, and environmentally friendly manner. Through this kinetic study, the enzymatic hydrolyses developed can be used as a guidance to assure the biotransformation of OTA in different food matrices.

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References

  • Abrunhosa L, Paterson RR, Venancio A (2010) Biodegradation of ochratoxin a for food and feed decontamination. Toxins 2(5):1078–1099

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Amezqueta S, Schorr-Galindo S, Murillo-Arbizu M, Gonzalez-Peñas E, De Cerain AL, Guiraud J (2012) OTA-producing fungi in foodstuffs: a review. Food Control 26(2):259–268

    Article  CAS  Google Scholar 

  • Andrade MA, Lanças FM (2017) Determination of Ochratoxin A in wine by packed in-tube solid phase microextraction followed by high performance liquid chromatography coupled to tandem mass spectrometry. J Chromatogr A 1493:41–48

    Article  CAS  PubMed  Google Scholar 

  • ANVISA ANdVS (2017) ANVISA. Resolução da Diretoria Colegiada-RDC No. 166, de 24 de julho de 2017. Dispõe sobre a validação de métodos analíticos e dá outras providências. In: Diário Oficial da União

  • AOAC (2000) Official Methods of Analysis International, 17. ed., CD- ROM

  • Arroyo-Manzanares N, García-Campaña AM, Gámiz-Gracia L (2011) Comparison of different sample treatments for the analysis of ochratoxin A in wine by capillary HPLC with laser-induced fluorescence detection. Anal Bioanal Chem 401(9):2987

    Article  CAS  PubMed  Google Scholar 

  • Arroyo-Manzanares N, Gámiz-Gracia L, García-Campaña AM (2012) Determination of ochratoxin A in wines by capillary liquid chromatography with laser induced fluorescence detection using dispersive liquid–liquid microextraction. Food Chem 135(2):368–372

    Article  CAS  PubMed  Google Scholar 

  • Bui-Klimke TR, Wu F (2015) Ochratoxin A and human health risk: a review of the evidence. Crit Rev Food Sci Nutr 55(13):1860–1869

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Caldas SS, Gonçalves FF, Primel EG, Prestes OD, Martins ML, Zanella R (2011) Modern techniques of sample preparation for pesticide residues determination in water by liquid chromatography with detection by diode array and mass spectrometry. Quim Nova 34(9):1604–1617

    Article  CAS  Google Scholar 

  • Campone L, Piccinelli AL, Rastrelli L (2011) Dispersive liquid–liquid microextraction combined with high-performance liquid chromatography–tandem mass spectrometry for the identification and the accurate quantification by isotope dilution assay of Ochratoxin A in wine samples. Anal Bioanal Chem 399(3):1279–1286

    Article  CAS  PubMed  Google Scholar 

  • Campone L, Piccinelli AL, Celano R, Rastrelli L (2012) pH-controlled dispersive liquid–liquid microextraction for the analysis of ionisable compounds in complex matrices: Case study of ochratoxin A in cereals. Anal Chim Acta 754:61–66

    Article  CAS  PubMed  Google Scholar 

  • Campone L, Piccinelli AL, Celano R, Pagano I, Russo M, Rastrelli L (2018) Rapid and automated on-line solid phase extraction HPLC–MS/MS with peak focusing for the determination of ochratoxin A in wine samples. Food Chem 244:128–135

    Article  CAS  PubMed  Google Scholar 

  • Chen F, Luan C, Wang L, Wang S, Shao L (2017) Simultaneous determination of six mycotoxins in peanut by high-performance liquid chromatography with a fluorescence detector. J Sci Food Agric 97(6):1805–1810

    Article  CAS  PubMed  Google Scholar 

  • de Almeida ÂB, Corrêa IP, Furuie JL, de Fariasdo RocioDalzoto PTP, Pimentel IC (2019) Inhibition of growth and ochratoxin A production in Aspergillus species by fungi isolated from coffee beans. Braz J Microbiol 50(4):1091–1098

    Article  PubMed  PubMed Central  Google Scholar 

  • EC (2006a) COMMISSION REGULATION (EC) No 401/2006a of 23 February 2006a laying down the methods of sampling and analysis for the official control of the levels of mycotoxins in foodstuffs. In

  • EC (2006b) Regulation (EC) No 1881/2006b of the European Parliament and the Council of 19 December 2006b setting maximum levels for certain contaminants in foodstuffs. In p 5–24

  • Escrivá L, Manyes L, Font G, Berrada H (2017) Mycotoxin analysis of human urine by LC-MS/MS: a comparative extraction study. Toxins 9(10):330

    Article  PubMed Central  Google Scholar 

  • Hendriks G, Uges D, Franke J (2007) Reconsideration of sample pH adjustment in bioanalytical liquid–liquid extraction of ionisable compounds. J Chromatogr B 853(1–2):234–241

    Article  CAS  Google Scholar 

  • Kabak B, Dobson AD, Var I (2006) Strategies to prevent mycotoxin contamination of food and animal feed: a review. Crit Rev Food Sci Nutr 46(8):593–619

    Article  CAS  PubMed  Google Scholar 

  • Karami-Osboo R, Miri R, Javidnia K, Kobarfard F, AliAbadi MHS, Maham M (2015) A validated dispersive liquid-liquid microextraction method for extraction of ochratoxin A from raisin samples. J Sci Food Agric 52(4):2440–2445

    CAS  Google Scholar 

  • Kupski L, Badiale-Furlong E (2015) Principal components analysis: an innovative approach to establish interferences in ochratoxin A detection. Food Chem 177:354–360

    Article  CAS  PubMed  Google Scholar 

  • Kupski L, Alves CL, Garda-Buffon J, Badiale-Furlong E (2013) Aplication of carboxypeptidase from Rhizopus on ochratoxin A degradation. Biochem Biotechonol Rep 2(4):30–36

    Article  Google Scholar 

  • Kupski L, Freitas M, Ribeiro D, Badiale-Furlong E, Fernandes E (2016) Ochratoxin A activates neutrophils and kills these cells through necrosis, an effect eliminated through its conversion into ochratoxin a. Toxicology 368:91–102

    Article  PubMed  Google Scholar 

  • Kupski L, Queiroz MI, Badiale-Furlong E (2018) Application of carboxypeptidase A to a baking process to mitigate contamination of wheat flour by ochratoxin A. Process Biochem 64:248–254

    Article  CAS  Google Scholar 

  • Lai X, Ruan C, Liu R, Liu C (2014a) Application of ionic liquid-based dispersive liquid–liquid microextraction for the analysis of ochratoxin A in rice wines. Food Chem 161:317–322

    Article  CAS  PubMed  Google Scholar 

  • Lai XW, Sun DL, Ruan CQ, Zhang H, Liu CL (2014b) Rapid analysis of aflatoxins B1, B2, and ochratoxin A in rice samples using dispersive liquid–liquid microextraction combined with HPLC. J Sep Sci 37(1–2):92–98

    Article  CAS  PubMed  Google Scholar 

  • Lavilla I, Gil S, Costas M, Bendicho C (2012) Dispersive liquid–liquid microextraction combined with microvolume spectrophotometry to turn green the 5530 APHA standard method for determining phenols in water and wastewater. Talanta 98:197–202

    Article  CAS  PubMed  Google Scholar 

  • Leitch AC, Abdelghany TM, Probert PM, Dunn MP, Meyer SK, Palmer JM, Cooke MP, Blake LI, Morse K, Rosenmai AK (2020) The toxicity of the methylimidazolium ionic liquids, with a focus on M8OI and hepatic effects. Food Chem Toxicol 136:111069

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li P, Pei F, Liu Q, Fang Y (2018) Magnetic solid-phase extraction for the determination of ochratoxin A in wine and beer by HPLC-FLD. Curr Anal Chem 14(2):129–134

    Article  CAS  Google Scholar 

  • Luan C, Wang L, Chen F, Wang S, Zhao L, Shao L (2016) Determination of ochratoxin A in pig muscle using dispersive liquid-liquid microextraction combined with high-performance liquid chromatography. Food Anal Methods 9(6):1490–1494

    Article  Google Scholar 

  • Marube LC, Caldas SS, Santos EOd, Michaelsen A, Primel EG (2018) Multi-residue method for determination of thirty-five pesticides, pharmaceuticals and personal care products in water using ionic liquid-dispersive liquid-liquid microextraction combined with liquid chromatography-tandem mass spectrometry. J Braz Chem Soc 29(6):1349–1359

    CAS  Google Scholar 

  • Merwe vdK (1965) Mycotoxins. Part II. The constitution of ochratoxins A, B, and C, metabolites of Aspergillus ochraceus wilh. J Chem Soc 1965:7083–7088

    Article  Google Scholar 

  • Pallarés N, Font G, Mañes J, Ferrer E (2017) Multimycotoxin LC–MS/MS analysis in tea beverages after dispersive liquid–liquid microextraction (DLLME). J Agric Food Chem 65(47):10282–10289

    Article  PubMed  Google Scholar 

  • Peng W-X, Marchal J, Van der Poel A (2018) Strategies to prevent and reduce mycotoxins for compound feed manufacturing. Anim Feed Sci Tech 237:129–153

    Article  CAS  Google Scholar 

  • Pfohl-Leszkowicz A, Manderville RA (2007) Ochratoxin A: an overview on toxicity and carcinogenicity in animals and humans. Mol Nutr Food Res 51(1):61–99

    Article  CAS  PubMed  Google Scholar 

  • Pham TPT, Cho C-W, Yun Y-S (2010) Environmental fate and toxicity of ionic liquids: a review. Water Res 44(2):352–372

    Article  CAS  PubMed  Google Scholar 

  • Pleadin J, Frece J, Markov K (2019) Mycotoxins in food and feed. Adv Food Nutr Res 89:297–345

    Article  CAS  PubMed  Google Scholar 

  • Primel EG, Caldas SS, Marube LC, Escarrone ALV (2017) An overview of advances in dispersive liquid–liquid microextraction for the extraction of pesticides and emerging contaminants from environmental samples. Trends Environ Anal Chem 14:1–18

    Article  CAS  Google Scholar 

  • Rezaee M, Assadi Y, Hosseini M-RM, Aghaee E, Ahmadi F, Berijani S (2006) Determination of organic compounds in water using dispersive liquid–liquid microextraction. J Chromatogr A 1116(1–2):1–9

    Article  CAS  PubMed  Google Scholar 

  • Spietelun A, Marcinkowski Ł, de la Guardia M, Namieśnik J (2014) Green aspects, developments and perspectives of liquid phase microextraction techniques. Talanta 119:34–45

    Article  CAS  PubMed  Google Scholar 

  • Stander MA, Steyn PS, van der Westhuizen FH, Payne BE (2001) A kinetic study into the hydrolysis of the ochratoxins and analogues by carboxypeptidase A. Chem Res Toxicol 14(3):302–304

    Article  CAS  PubMed  Google Scholar 

  • Temba BA, Sultanbawa Y, Kriticos DJ, Fox GP, Harvey JJ, Fletcher MT (2016) Tools for Defusing a Major Global Food and Feed Safety Risk: Nonbiological Postharvest Procedures To Decontaminate Mycotoxins in Foods and Feeds. J Agric Food Chem 64(47):8959–8972

    Article  CAS  PubMed  Google Scholar 

  • Tolcha T, Merdassa Y, Megersa N (2013) Low-density extraction solvent based solvent-terminated dispersive liquid–liquid microextraction for quantitative determination of ionizable pesticides in environmental waters. J Sep Sci 36(6):1119–1127

    Article  CAS  PubMed  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  • Zhao Z, Yang X, Zhao X, Bai B, Yao C, Liu N, Wang J, Zhou C (2017) Vortex-assisted dispersive liquid-liquid microextraction for the analysis of major Aspergillus and Penicillium mycotoxins in rice wine by liquid chromatography-tandem mass spectrometry. Food Control 73:862–868

    Article  CAS  Google Scholar 

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Acknowledgments

The authors acknowledge the financial support by the Brazilian agencies FAPERGS, CNPq and FINEP. Part of this study was financially supported by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES)—Finance Code 001.

Funding

Larine Kupski received a postdoctoral fellowship from CNPq (150439/2017–2). Ednei Gilberto Primel received a productivity research fellowship from the Brazilian Agency CNPq (DT 305716/2020-4). Eliana Badiale Furlong received a Scholarship in Research Productivity from the Brazilian Agency CNPq (PQ 307817/2020-2).

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Authors and Affiliations

  1. Departamento de Tecnologia, Universidade Estadual de Maringá, Av. Ângelo Moreira da Fonseca, Umuarama, Paraná, 1800, Brazil

    Larine Kupski

  2. Escola de Química E Alimentos, Laboratório de Análise de Compostos Orgânicos E Metais (LACOM), Universidade Federal Do Rio Grande, Av Itália, km 8, s/n, Rio Grande, Rio Grande do Sul, Brazil

    Duane Volpato, Caroline B. Rocha, Sergiane C. Barbosa & Ednei G. Primel

  3. Escola de Química E Alimentos, Laboratório de Micotoxinas E Ciência de Alimentos (LAMCA), Universidade Federal Do Rio Grande, Av Itália, km 8, s/n, Rio Grande, Rio Grande do Sul, Brazil

    Eliana B. Furlong

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  1. Larine Kupski
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  2. Duane Volpato
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  5. Sergiane C. Barbosa
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  6. Ednei G. Primel
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Correspondence to Larine Kupski.

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Larine Kupski declares that she has no conflict of interest. Duane Volpato declares that she has no conflict of interest. Caroline B. Rocha declares that she has no conflict of interest. Eliana B. Furlong declares that she has no conflict of interest. Sergiane C. Barbosa declares that she has no conflict of interest. Ednei G. Primel declares that he has no conflict of interest.

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Kupski, L., Volpato, D., Rocha, C.B. et al. Determination of Ochratoxin A and Its Metabolite Ochratoxin Alpha in Different Food Matrices After Enzymatic Biotransformation. Food Anal. Methods 15, 3003–3012 (2022). https://doi.org/10.1007/s12161-022-02349-5

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