Acute oral toxicity study on Wistar rats fed microalgal protein hydrolysates from Bellerochea malleus

  • Ines Barkia
  • Hanen Ketata Bouaziz
  • Tahiya Sellami Boudawara
  • Lotfi Aleya
  • Ali Faouzi Gargouri
  • Nazamid Saari
Nanotechnology, Nanopollution, Nanotoxicology and Nanomedicine (NNNN)


Protein hydrolysates and bioactive peptides from various protein sources have demonstrated their effectiveness for the prevention of illness and the improvement of symptoms from several diseases. In particular, the use of microalgae to generate bioactive peptides has received a growing interest because of their potential to be cultivated on non-arable land and high nutritional value. However, scant research is available on the toxicity of peptide-based preparations. The present study aims to evaluate the toxicity of microalgal protein hydrolysates (MPH) from one marine species of microalgae (Bellerochea malleus) to determine the feasibility of their use for functional food applications. Results showed that the oral administration of MPH at three doses (D1, 100 mg kg−1 BW; D2, 400 mg kg−1 BW; and D3, 2000 mg kg−1 BW) to male Wistar rats did not induce any adverse effects or mortality up to13 days of treatment. Data analysis of relative organ weights and biochemical and hematological parameters did not show any significant differences between control and treated groups at the three doses investigated. Data from histopathological observations did not reveal any signs of major toxicity at the doses D1 and D2. However, mild signs of inflammation and necrosis were observed in the kidney of rats fed MPH at D3. All together, these results reveal the overall safety of MPH and provide new evidence for advocating their use for functional food or nutraceutical applications.


Microalgal protein hydrolysates Toxicity Liver Kidney Rats 



This work is part of the PhD thesis of Ines Barkia. The authors would like to thank M. D Choumous Kallel for her help in the biochemical analysis and Souad Eljoudi for her contribution in the preparation of the histological slides. We express our appreciation to the editor, Dr. Philippe Garrigues, and to the anonymous reviewers for helping to improve our paper.


  1. Abdel-Daim MM, Aleya L, Najda AB, Abo-El-Sooud K, Bungǎu SG, Bungau SG, Saluja R (2018) Alleviation of drugs and chemicals toxicity: biomedical value of antioxidants. Oxidative Med Cell Longev 2.
  2. Anadón A, Martínez MA, Ares I, Ramos E, Martínez-Larrañaga MR, Contreras MM, Ramos M, Recio I (2010) Acute and repeated dose (4 weeks) oral toxicity studies of two antihypertensive peptides, RYLGY and AYFYPEL, that correspond to fragments (90–94) and (143–149) from αs1-casein. Food Chem Toxicol 48(7):1836–1845CrossRefGoogle Scholar
  3. Bailey SA, Zidell RH, Perry RW (2004) Relationships between organ weight and body/brain weight in the rat: what is the best analytical endpoint? Toxicol Pathol 32(4):448–466CrossRefGoogle Scholar
  4. Barkia I, Zadjali F, Saari N, Zakaria MH, Li CL, Witkowski A, Al-Haj L (2018a) Isolation and identification of indigenous marine diatoms (Bacillariophyta) for biomass production in open raceway ponds. Aquac Res 49:928–938CrossRefGoogle Scholar
  5. Barkia I, AlHaj L, Abdul Hamid A, Zakaria M, Saari N, Zadjali F (2018b) Indigenous marine diatoms as novel sources of bioactive peptides with antihypertensive and antioxidant properties. Int J Food Sci Technol.
  6. Bisar GH, Youssef M, Saadany KE, El-Kholy W, Kheadr E (2017) Effect of lentil and buffalo protein hydrolysates on histopathology of liver and kidney in diabetic rats. J Cytol Histol 8:489Google Scholar
  7. Caldwell GS (2009) The influence of bioactive oxylipins from marine diatoms on invertebrate reproduction and development. Mar Drugs 7(3):367–400CrossRefGoogle Scholar
  8. Cam A, Sivaguru M, De Mejia EG (2013) Endocytic mechanism of internalization of dietary peptide lunasin into macrophages in inflammatory condition associated with cardiovascular disease. PLoS One 8(9):e72115CrossRefGoogle Scholar
  9. Caporgno MP, Mathys A (2018) Trends in microalgae incorporation into innovative food products with potential health benefits. Front Nutr 5:58CrossRefGoogle Scholar
  10. Chay SY, Salleh A, Sulaiman NF, Abidin NZ, Hanafi MA, Zarei M, Saari N (2018) Blood-pressure lowering efficacy of winged bean seed hydrolysate in spontaneously hypertensive rats, peptide characterization and a toxicity study in Sprague-Dawley rats. Food Funct 9(3):1657–1671CrossRefGoogle Scholar
  11. Csapo J, Albert C, Csapó-Kiss Z (2009) The D-amino acid content of foodstuffs (a review). Acta Univ Sapientiae, Aliment 1:5–30Google Scholar
  12. Cudennec B, Fouchereau-Peron M, Ferry F, Duclos E, Ravallec R (2012) In vitro and in vivo evidence for a satiating effect of fish protein hydrolysate obtained from blue whiting (Micromesistius poutassou) muscle. J Funct Foods 4:271–277CrossRefGoogle Scholar
  13. de Almeida Vaucher R, Gewehr CDCV, Correa APF, Sant’Anna V, Ferreira J, Brandelli A (2011) Evaluation of the immunogenicity and in vivo toxicity of the antimicrobial peptide P34. Int J Pharm 421(1):94–98CrossRefGoogle Scholar
  14. García JL, de Vicente M, Galán B (2017) Microalgae, old sustainable food and fashion nutraceuticals. Microb Biotechnol 10(5):1017–1024CrossRefGoogle Scholar
  15. Giknis MLA, Clifford CB (2006) Clinical laboratory parameters for Crl: CD (SD) rats. Charles River Laboratories, Wilmington, pp 1–14Google Scholar
  16. Gowda S, Desai PB, Kulkarni SS, Hull VV, Math AA, Vernekar SN (2010) Markers of renal function tests. N Am J Med Sci 2(4):170–173Google Scholar
  17. Guo B, Liu B, Yang B, Sun P, Lu X, Liu J, Chen F (2016) Screening of diatom strains and characterization of Cyclotella cryptica as a potential fucoxanthin producer. Mar Drugs 14(7):125CrossRefGoogle Scholar
  18. Heimbach JT, Sebestyen G, Semjen G, Kennepohl E (2007) Safety studies regarding a standardized extract of fermented wheat germ. Int J Toxicol 26(3):253–259CrossRefGoogle Scholar
  19. Jung EY, Suh HJ, Kim SY, Hong YS, Kim MJ, Chang UJ (2008) Appetite suppressive effects of yeast hydrolysate on nitric oxide synthase (NOS) expression and vasoactive intestinal peptide (VIP) immunoreactivity in hypothalamus. Phytother Res 22(11):1417–1422CrossRefGoogle Scholar
  20. Jung EY, Park SS, Kim JH, Chang UJ, Bae SH, Choi JW, Suh HJ (2011) Safety study of yeast hydrolysate with below 10 kDa molecular weight in animal models. J Health Sci 57(6):532–539CrossRefGoogle Scholar
  21. Kim KM, Chang UJ, Kang DH, Kim JM, Choi YM, Suh HJ (2004) Yeast hydrolysate reduces body fat of dietary obese rats. Phytother Res 18(11):950–953CrossRefGoogle Scholar
  22. Kiriyama Y, Nochi H (2016) D-amino acids in the nervous and endocrine systems. Scientifica 2016:1–9CrossRefGoogle Scholar
  23. Kruger CL, Marano KM, Morita Y, Takada Y, Kawakami H, Kobayashi T, Sunaga M, Furukawa M, Kawamura K (2007) Safety evaluation of a milk basic protein fraction. Food Chem Toxicol 45(7):1301–1307CrossRefGoogle Scholar
  24. Lauritano C, Andersen JH, Hansen E, Albrigtsen M, Escalera L, Esposito F, Helland K, Hanssen KØ, Romano G, Ianora A (2016) Bioactivity screening of microalgae for antioxidant, anti-inflammatory, anticancer, anti-diabetes, and antibacterial activities. Front Mar Sci 3:68. CrossRefGoogle Scholar
  25. Levin ME, Blackhurst DM, Kirstein F, Kok D, Van der Watt GF, Marais AD (2017) Residual allergenicity of amino acid-based and extensively hydrolysed cow’s milk formulas. S Afr Med J 107(9):763–767CrossRefGoogle Scholar
  26. Mohanty DP, Mohapatra S, Misra S, Sahu PS (2016) Milk derived bioactive peptides and their impact on human health—a review. Saudi J Biol Sci 23(5):577–583CrossRefGoogle Scholar
  27. Nicklas W, Baneux P, Boot R, Decelle T, Deeny AA, Fumanelli M, Illgen-Wilcke B (2002) FELASA (Federation of European Laboratory Animal Science Associations Working Group on health monitoring of rodent and rabbit colonies). Recommendations for the health monitoring of rodent and rabbit colonies in breeding and experimental units. Lab Anim 36(1):20–42CrossRefGoogle Scholar
  28. Nunn PB, Bell EA, Watson AA, Nash RJ (2010) Toxicity of non-protein amino acids to humans and domestic animals. Nat Prod Commun 5(3):485–504Google Scholar
  29. Organisation for Economic Cooperation Development (1987) Guidelines for the testing of chemicals OECD, vol 401. Acute Oral Toxicity. Organisation for Economic Cooperation and Development, ParisGoogle Scholar
  30. Pesta DH, Samuel VT (2014) A high-protein diet for reducing body fat: mechanisms and possible caveats. Nutr Metab 11(1):53CrossRefGoogle Scholar
  31. Phelan M, Aherne A, FitzGerald RJ, O'Brien NM (2009) Casein-derived bioactive peptides: biological effects, industrial uses, safety aspects and regulatory status. Int Dairy J 19(11):643–654CrossRefGoogle Scholar
  32. Schilter B, Andersson C, Anton R, Constable A, Kleiner J, O'Brien J, Renwick AG, Korver O, Smit F, Walker R, Natural Toxin Task Force of the European Branch of the International Life Sciences Institute (2003) Guidance for the safety assessment of botanicals and botanical preparations for use in food and food supplements. Food Chem Toxicol 41(12):1625–1649CrossRefGoogle Scholar
  33. Taha S, El Abd M, De Gobba C, Abdel-Hamid M, Khalil E, Hassan D (2017) Antioxidant and antibacterial activities of bioactive peptides in buffalo’s yoghurt fermented with different starter cultures. Food Sci Biotechnol 26(5):1325–1332CrossRefGoogle Scholar
  34. Torres-Duran PV, Ferreira-Hermosillo A, Juarez-Oropeza MA (2007) Antihyperlipemic and antihypertensive effects of Spirulina maxima in an open sample of Mexican population: a preliminary report. Lipids Health Dis 6(1):33CrossRefGoogle Scholar
  35. Vaughn N, Rizzo A, Doane D, Beverly JL, de Mejia EG (2008) Intracerebroventricular administration of soy protein hydrolysates reduces body weight without affecting food intake in rats. Plant Food Hum Nutr 63(1):41–46CrossRefGoogle Scholar
  36. Waghmare AG, Salve MK, LeBlanc JG, Arya SS (2016) Concentration and characterization of microalgae proteins from Chlorella pyrenoidosa. Bioresour Bioprocess 3(1):16CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Ines Barkia
    • 1
  • Hanen Ketata Bouaziz
    • 2
  • Tahiya Sellami Boudawara
    • 3
  • Lotfi Aleya
    • 4
  • Ali Faouzi Gargouri
    • 5
  • Nazamid Saari
    • 1
  1. 1.Department of Food ScienceUniversiti Putra MalaysiaSerdangMalaysia
  2. 2.Animal Physiology Laboratory, Faculty of SciencesUniversity of Sfax,SfaxTunisia
  3. 3.Histopathology LaboratoryUniversity of SfaxSfaxTunisia
  4. 4.Laboratoire Chrono-Environnement, UMR 6249 CNRSBesançonFrance
  5. 5.Laboratory of Molecular Biotechnology of Eucaryotes, Centre of Biotechnology of Sfax (CBS)SfaxTunisia

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