Health, Toxicology and Mutagenesis; Toxicology; Food supplement; Algae; spirulina; Aphanizomenon flos-aquae; Microcystin; UHPLC-TandemMS; health risks
Abstract :
[en] Food supplements are gaining popularity worldwide. However, harmful natural compounds can contaminate these products. In the case of algae-based products, the presence of toxin-producing cyanobacteria may cause health risks. However, data about the prevalence of algal food supplements on the Belgian market and possible contaminations with cyanotoxins are scarce. Therefore, we optimized and validated a method based on Ultra High Performance Liquid Chromatography-Tandem Mass Spectrometry to quantify eight microcystin congeners and nodularin in algal food supplements. Our analytical method was successfully validated and applied on 35 food supplement samples. Nine out of these samples contained microcystin congeners, of which three exceeded 1 µg g−1, a previously proposed guideline value. Additionally, the mcyE gene was amplified and sequenced in ten products to identify the taxon responsible for the toxin production. For seven out of these ten samples, the mcyE gene could be amplified and associated to Microcystis sp. EFSA and posology consumption data for algal-based food supplements were both combined with our toxin prevalence data to establish different toxin exposure scenarios to assess health risks and propose new guideline values.
Research center :
InBios - Integrative Biological Sciences - ULiège
Disciplines :
Microbiology Food science
Author, co-author :
Van Hassel, Wannes ; Université de Liège - ULiège > Integrative Biological Sciences (InBioS)
Council for Responsible Nutrition Growth of Dietary Supplement Use over the Last Decade 2017 Available online: www.crnusa.org/survey (accessed on 1 June 2021)
Brocatus L. De Ridder K. Verrijkte voeding en voedingssupplementen Voedselconsumptiepeiling 2014–2015 Bel S. Tafforeau J. Rapport 4 WIV-ISP Brussels, Belgium 2016
Sánchez-Parra E. Boutarfa S. Aboal M. Are Cyanotoxins the Only Toxic Compound Potentially Present in Microalgae Supplements? Results from a Study of Ecological and Non-Ecological Products Toxins 2020 12 552 10.3390/toxins12090552
Muys M. Sui Y. Schwaiger B. Lesueur C. Vandenheuvel D. Vermeir P. Vlaeminck S.E. High Variability in Nutritional Value and Safety of Commercially Available Chlorella and Spirulina Biomass Indicates the Need for Smart Production Strategies Bioresour. Technol. 2019 275 247 257 10.1016/j.biortech.2018.12.059
Nelissen B. Wilmotte A. Neefs J.-M. De Wachter R. Phylogenetic Relationships Among Filamentous Helical Cyanobacteria Investigated on the Basis of 16S Ribosomal RNA Gene Sequence Analysis Syst. Appl. Microbiol. 1994 17 206 210 10.1016/S0723-2020(11)80009-3
Delrue F. Alaux E. Moudjaoui L. Gaignard C. Fleury G. Perilhou A. Richaud P. Petitjean M. Sassi J.-F. Optimization of Arthrospira platensis (Spirulina) Growth: From Laboratory Scale to Pilot Scale Fermentation 2017 3 59 10.3390/fermentation3040059
El-Sayed A. El-Sheekh M. Outdoor Cultivation of Spirulina platensis for Mass Production Not. Sci. Biol. 2018 10 38 10.15835/nsb10110177
Costa J.A. Freitas B.C. Santos T. Mitchell B. Morais M. Open pond systems for microalgal culture Biomass, Biofuels Biochemicals Elsevier Amsterdam, The Netherlands 2019 199 223 9780444641922
Furmaniak M.A. Misztak A.E. Franczuk M.D. Wilmotte A. Waleron M. Waleron K.F. Edible Cyanobacterial Genus Arthrospira: Actual State of the Art in Cultivation Methods, Genetics, and Application in Medicine Front. Microbiol. 2017 8 2541 10.3389/fmicb.2017.02541
Carmichael W.W. Drapeau C. Anderson D.M. Harvesting of Aphanizomenon flos-aquae Ralfs Ex Born. & Flah. var. flos-aquae (Cyanobacteria) from Klamath Lake for Human Dietary Use J. Appl. Phycol. 2000 12 585 895 10.1023/A:1026506713560
Ciferri O. Spirulina, the Edible Microorganism Microbiol. Rev. 1983 47 551 578 10.1128/mr.47.4.551-578.1983
Heussner A.H. Mazija L. Fastner J. Dietrich D.R. Toxin Content and Cytotoxicity of Algal Dietary Supplements Toxicol. Appl. Pharmacol. 2012 265 263 271 10.1016/j.taap.2012.10.005 23064102
Roy-Lachapelle A. Solliec M. Bouchard M.F. Sauvé S. Detection of Cyanotoxins in Algae Dietary Supplements Toxins 2017 9 76 10.3390/toxins9030076
Gilroy D.J. Kauffman K.W. Hall R.A. Huang X. Chu F.S. Assessing Potential Health Risks from Microcystin Toxins in Blue-Green Algae Dietary Supplements Environ. Health Perspect. 2000 108 435 439 10.1289/ehp.00108435
Rellán S. Osswald J. Saker M. Gago-Martinez A. Vasconcelos V. First Detection of Anatoxin-a in Human and Animal Dietary Supplements Containing Cyanobacteria Food Chem. Toxicol. 2009 47 2189 2195 10.1016/j.fct.2009.06.004
Gutiérrez-Praena D. Jos Á. Pichardo S. Moreno I.M. Cameán A.M. Presence and Bioaccumulation of Microcystins and Cylindrospermopsin in Food and the Effectiveness of Some Cooking Techniques at Decreasing Their Concentrations: A Review Food Chem. Toxicol. 2013 53 139 152 10.1016/j.fct.2012.10.062
Vichi S. Lavorini P. Funari E. Scardala S. Testai E. Contamination by Microcystis and Microcystins of Blue-Green Algae Food Supplements (BGAS) on the Italian Market and Possible Risk for the Exposed Population Food Chem. Toxicol. 2012 50 4493 4499 10.1016/j.fct.2012.09.029 23036452
Testai E. Buratti F.M. Funari E. Manganelli M. Vichi S. Arnich N. Fessard V. Sialehaamoa A. Review and Analysis of Occurrence, Exposure and Toxicity of Cyanobacteria Toxins in Food EFSA Extern. Sci. Rep. 2016 13 998E 10.2903/sp.efsa.2016.EN-998
Lawrence J.F. Niedzwiadek B. Menard C. Lau B.P. Lewis D. Kuper-Goodman T. Carbone S. Holmes C. Comparison of Liquid Chromatography/Mass Spectrometry, ELISA, and Phosphatase Assay for the Determination of Microcystins in Blue-Green Algae Products J. AOAC Int. 2001 84 1035 1044 10.1093/jaoac/84.4.1035
Ortelli D. Edder P. Cognard E. Jan P. Fast Screening and Quantitation of Microcystins in Microalgae Dietary Supplement Products and Water by Liquid Chromatography Coupled to Time of Flight Mass Spectrometry Anal. Chim. Acta 2008 617 230 237 10.1016/j.aca.2008.03.033
Parker C.H. Stutts W.L. Degrasse S.L. Development and Validation of a Liquid Chromatography-Tandem Mass Spectrometry Method for the Quantitation of Microcystins in Blue-Green Algal Dietary Supplements J. Agric. Food Chem. 2015 63 10303 10312 10.1021/acs.jafc.5b04292
Mantzouki E. Lürling M. Fastner J. de Senerpont Domis L. Wilk-Wo’zniak E. Temperature Effects Explain Continental Scale Distribution of Cyanobacterial Toxins Toxins 2018 10 156 10.3390/toxins10040156 29652856
Giménez-Campillo C. Pastor-Belda M. Campillo N. Arroyo-Manzanares N. Hernández-Córdoba M. Viñas P. Determination of Cyanotoxins and Phycotoxins in Seawater and Algae-Based Food Supplements Using Ionic Liquids and Liquid Chromatography with Time-Of-Flight Mass Spectrometry Toxins 2019 11 610 10.3390/toxins11100610
Svirčev Z. Lalić D. Bojadzija Savic G. Tokodi N. Drobac Backovic D. Chen L. Meriluoto J. Codd G. Global Geographical and Historical Overview of Cyanotoxin Distribution and Cyanobacterial Poisonings Arch. Toxicol. 2019 93 2429 2481 10.1007/s00204-019-02524-4
Bouaïcha N. Miles C.O. Beach D.G. Labidi Z. Djabri A. Benayache N.Y. Nguyen-Quang T. Structural Diversity, Characterization and Toxicology of Microcystins Toxins 2019 11 714 10.3390/toxins11120714
Turner A.D. Dhanji-Rapkova M. O’Neill A. Coates L. Lewis A. Lewis K. Analysis of Microcystins in Cyanobacterial Blooms from Freshwater Bodies in England Toxins 2018 10 39 10.3390/toxins10010039
Van Hassel W.H.R. Andjelkovic M. Durieu B. Marroquin V.A. Masquelier J. Huybrechts B. Wilmotte A. A Summer of Cyanobacterial Blooms in Belgian Waterbodies: Microcystin Quantification and Molecular Characterizations Toxins 2022 14 61 10.3390/toxins14010061
Fischer A. Hoeger S.J. Stemmer K. Feurstein D.J. Knobeloch D. Nussler A. Dietrich D.R. The Role of Organic Anion Transporting Polypeptides (OATPs/SLCOs) in the Toxicity of Different Microcystin Congeners in Vitro: A Comparison of Primary Human Hepatocytes and OATP-Transfected HEK293 Cells Toxicol. Appl. Pharmacol. 2010 245 9 20 10.1016/j.taap.2010.02.006
Altaner S. Jaeger S. Fotler R. Zemskov I. Wittmann V. Schreiber F. Dietrich D.R. Machine Learning Prediction of Cyanobacterial Toxin (Microcystin) Toxicodynamics in Humans ALTEX 2020 37 24 36 10.14573/altex.1904031e
Feurstein D. Stemmer K. Kleinteich J. Speicher T. Dietrich D.R. Microcystin Congener- and Concentration-Dependent Induction of Murine Neuron Apoptosis and Neurite Degeneration Toxicol. Sci. 2011 124 424 431 10.1093/toxsci/kfr243
Steiner K. Zimmermann L. Hagenbuch B. Dietrich D. Zebrafish Oatp-Mediated Transport of Microcystin Congeners Arch. Toxicol. 2016 90 1129 1139 10.1007/s00204-015-1544-3
Hooser S.B. Fulminant Hepatocyte Apoptosis in Vivo Following Microcystin-LR Administration to Rats Toxicol. Pathol. 2000 28 726 733 10.1177/019262330002800513
Hooser S.B. Beasley V.R. Waite L.L. Kuhlenschmidt M.S. Carmichael W.W. Haschek W.M. Actin Filament Alterations in Rat Hepatocytes Induced In Vivo and In Vitro by Microcystin-LR, a Hepatotoxin from the Blue-Green Alga, Microcystis aeruginosa Vet. Pathol. 1991 28 259 266 10.1177/030098589102800401 1949504
Falconer I.R. Burch M.D. Steffensen D.A. Choice M. Coverdale O.R. Toxicity of the Blue-green Alga (Cyanobacterium) Microcystis aeruginosa in Drinking Water to Growing Pigs, as an Animal Model for Human Injury and Risk Assessment Environ. Toxicol. Water Qual. 1994 9 131 139 10.1002/tox.2530090209
Yoshida T. Makita Y. Nagata S. Tsutsumi T. Yoshida F. Sekijima M. Tamura S.I. Ueno Y. Acute Oral Toxicity of Microcystin-LR, a Cyanobacterial Hepatotoxin, in Mice Nat. Toxins 1997 5 91 95 10.1002/nt.1
World Health Organization Cyanobacterial Toxins: Microcystins. Background Document for Development of WHO Guidelines for Drinking-Water Quality and Guidelines for Safe Recreational Water Environments WHO Geneva, Switzerland 2020
Miles C.O. Sandvik M. Haande S. Nonga H. Ballot A. LC-MS Analysis with Thiol Derivatization to Differentiate [Dhb7]- from [Mdha7]-Microcystins: Analysis of Cyanobacterial Blooms, Planktothrix Cultures and European Crayfish from Lake Steinsfjorden, Norway Environ. Sci. Technol. 2013 47 4080 4087 10.1021/es305202p 23531156
Park H. Namikoshi M. Brittain S.M. Carmichael W.W. Murphy T. [D-Leu1] Microcystin-LR, a New Microcystin Isolated from Waterbloom in a Canadian Prairie Lake Toxicon 2001 39 855 862 10.1016/S0041-0101(00)00224-5
Botes D.P. Kruger H. Viljoen C.C. Isolation and Characterization of Four Toxins from the Blue-Green Alga, Microcystis aeruginosa Toxicon 1982 20 945 954 10.1016/0041-0101(82)90097-6
Welker M. Von Dohren H. Cyanobacterial Peptides: Nature’s Own Combinatorial Biosynthesis FEMS Microbiol. Rev. 2006 30 530 563 10.1111/j.1574-6976.2006.00022.x
Puddick J. Prinsep M.R. Wood S.A. Cary S.C. Hamilton D.P. Holland P.T. Further Characterization of Glycine-Containing Microcystins from the McMurdo Dry Valleys of Antarctica Toxins 2015 7 493 515 10.3390/toxins7020493
Foss A.J. Miles C.O. Samdal I.A. Løvberg K.E. Wilkins A.L. Rise F. Jaabæk J.A.H. McGowan P.C. Aubel M.T. Analysis of Free and Metabolized Microcystins in Samples Following a Bird Mortality Event Harmful Algae 2018 80 117 129 10.1016/j.hal.2018.10.006 30502804
Sivonen K. Namikoshi M. Evans W.R. Fardig M. Carmichael W.W. Rinehart K.L. Three New Microcystins, Cyclic Heptapeptide Hepatotoxins, from Nostoc sp. Strain 152 Chem. Res. Toxicol. 1992 5 464 469 10.1021/tx00028a003
Namikoshi M. Rinehart K.L. Sakai R. Stotts R.R. Dahlem A.M. Beasley V.R. Carmichael W.W. Evans W.R. Identification of 12 Hepatotoxins from a Homer Lake Bloom of the Cyanobacteria Microcystis aeruginosa, Microcystis viridis, and Microcystis wesenbergii: Nine New Microcystins J. Org. Chem. 1992 57 866 872 10.1021/jo00029a016
Namikoshi M. Rinehart K.L. Sakai R. Sivonen K. Carmichael W.W. Structures of Three New Cyclic Heptapeptide Hepatotoxins Produced by the Cyanobacterium (Blue-Green Alga) Nostoc sp. Strain 152 J. Org. Chem. 1990 55 6135 6139 10.1021/jo00312a019
Namikoshi M. Yuan M. Sivonen K. Carmichael W.W. Rinehart K.L. Rouhiainen L. Sun F. Brittain S. Otsuki A. Seven New Microcystins Possessing Two L-Glutamic Acid Units, Isolated from Anabaena Sp. Strain 186 Chem. Res. Toxicol. 1998 11 143 149 10.1021/tx970120t 9511906
Namikoshi M. Sivonen K. Evans W.R. Carmichael W.W. Rouhiainen L. Luukkainen R. Rinehart K.L. Structures of Three New Homotyrosine-Containing Microcystins and a New Homophenylalanine Variant from Anabaena sp. Strain 66 Chem. Res. Toxicol. 1992 5 661 666 10.1021/tx00029a011
Turner A.D. Waack J. Lewis A. Edwards C. Lawton L. Development and Single-Laboratory Validation of a UHPLC-MS/MS Method for Quantitation of Microcystins and Nodularin in Natural Water, Cyanobacteria, Shellfish and Algal Supplement Tablet Powders J. Chromatogr. B Analyt. Technol. Biomed. Life. Sci. 2018 1074–1075 111 123 10.1016/j.jchromb.2017.12.032 29358154
Tillett D. Dittmann E. Erhard M. von Döhren H. Börner T. Neilan B.A. Structural Organization of Microcystin Biosynthesis in Microcystis aeruginosa PCC7806: An Integrated Peptide–Polyketide Synthetase System Chem. Biol. 2000 7 753 764 10.1016/S1074-5521(00)00021-1
Jungblut A.-D. Neilan B.A. Molecular Identification and Evolution of the Cyclic Peptide Hepatotoxins, Microcystin and Nodularin, Synthetase Genes in Three Orders of Cyanobacteria Arch. Microbiol. 2006 185 107 114 10.1007/s00203-005-0073-5
Saker M.L. Jungblut A.D. Neilan B.A. Rawn D.F.K. Vasconcelos V.M. Detection of Microcystin Synthetase Genes in Health Food Supplements Containing the Freshwater Cyanobacterium Aphanizomenon flos-aquae Toxicon 2005 46 555 562 10.1016/j.toxicon.2005.06.021 16098554
Publications Office of the European Union CELEX1, 2002/657/EC: Commission Decision of 12 August 2002 Implementing Council Directive 96/23/EC Concerning the Performance of Analytical Methods and the Interpretation of Results (Text with EEA Relevance) (Notified under Document Number C(2002) 3044) Available online: http://op.europa.eu/en/publication-detail/-/publication/ed928116-a955-4a84-b10a-cf7a82bad858/language-en (accessed on 30 November 2020)
Van Loco J. Elskens M. Croux C. Beernaert H. Linearity of Calibration Curves: Use and Misuse of the Correlation Coefficient Accredit. Qual. Assur. 2002 7 281 285 10.1007/s00769-002-0487-6
Marsan D.W. Conrad S.M. Stutts W.L. Parker C.H. Deeds J.R. Evaluation of Microcystin Contamination in Blue-Green Algal Dietary Supplements Using a Protein Phosphatase Inhibition-Based Test Kit Heliyon 2018 4 e00573 10.1016/j.heliyon.2018.e00573 29862339
Miller T.R. Xiong A. Deeds J.R. Stutts W.L. Samdal I.A. Løvberg K.E. Miles C.O. Microcystin Toxins at Potentially Hazardous Levels in Algal Dietary Supplements Revealed by a Combination of Bioassay, Immunoassay, and Mass Spectrometric Methods J. Agric. Food Chem. 2020 68 8016 8025 10.1021/acs.jafc.0c02024
Saker M.L. Welker M. Vasconcelos V.M. Multiplex PCR for the Detection of Toxigenic Cyanobacteria in Dietary Supplements Produced for Human Consumption Appl. Microbiol. Biotechnol. 2007 73 1136 1142 10.1007/s00253-006-0565-5
Yakes B.J. Handy S.M. Kanyuck K.M. DeGrasse S.L. Improved Screening of Microcystin Genes and Toxins in Blue-Green Algal Dietary Supplements with PCR and a Surface Plasmon Resonance Biosensor Harmful Algae 2015 47 9 16 10.1016/j.hal.2015.05.001
Dietrich D. Hoeger S. Guidance Values for Microcystins in Water and Cyanobacterial Supplement Products (Blue-Green Algal Supplements): A Reasonable or Misguided Approach? Toxicol. Appl. Pharmacol. 2005 203 273 289 10.1016/j.taap.2004.09.005
EFSA Scientific Committee Guidance on Selected Default Values to Be Used by the EFSA Scientific Committee, Scientific Panels and Units in the Absence of Actual Measured Data EFSA J. 2012 10 32 10.2903/j.efsa.2012.2579
Dietrich D.R. Fischer A. Michel C. Hoeger S. Toxin Mixture in Cyanobacterial Blooms–A Critical Comparison of Reality with Current Procedures Employed in Human Health Risk Assessment Adv. Exp. Med. Biol. 2008 619 885 912 10.1007/978-0-387-75865-7_39 18461795
Van Hassel W.H.R. Huybrechts B. Masquelier J. Wilmotte A. Andjelkovic M. Development, Validation and Application of a Targeted LC-MS Method for Quantification of Microcystins and Nodularin: Towards a Better Characterization of Drinking Water Water 2022 14 1195 10.3390/w14081195
European Food Safety Authority Use of the EFSA Comprehensive European Food Consumption Database in Exposure Assessment EFSA J. 2011 9 34 10.2903/j.efsa.2011.2097
