Plant Foods for Human Nutrition

, Volume 71, Issue 2, pp 174–182 | Cite as

Antithrombotic and Antioxidant Activity of Amaranth Hydrolysate Obtained by Activation of an Endogenous Protease

  • Ana Clara Sabbione
  • Sabrina M. Ibañez
  • E. Nora Martínez
  • María Cristina Añón
  • Adriana A. Scilingo
Original Paper


Ingestion of diets with antithrombotic and antioxidant components offer a convenient and effective way to prevent and reduce the incidence of cardiovascular diseases. The aim of the present work was to obtain an amaranth hydrolysate by the activation of an endogenous aspartic protease, to establish adequate experimental conditions, and to evaluate its antithrombotic and antioxidant activity in order to assess its potential application as an ingredient in functional foods. The results obtained not only confirmed the presence of an endogenous protease in the amaranth isolate, but also allowed us to select an adequate incubation conditions (pH 2, 40 °C, 16 h). The hydrolysate obtained (degree of hydrolysis 5.3 ± 0.4 %) showed potential antithrombotic activity (IC50 = 5.9 ± 0.1 mg soluble protein/mL) and had more antioxidant activity than the isolate, indicating that the activation of the protease released bioactive peptides from amaranth proteins. Decreasing the pH is a simple and cheap process and is another way to obtain potential functional ingredients with bioactive compounds.


Amaranth proteins Endogenous aspartic protease Bioactive peptides Antithrombotic and antioxidant activity 



Degree of hydrolysis


Concentration of isolate or hydrolysate that inhibits 50 % of the thrombus formation or produces 50 % radical neutralization


Hydrolysate prepared by activation of endogenous protease



This work was supported by PIP - CONICET 11220110101109.

Compliance with Ethical Standards

Conflict of Interest

Authors, A.C. Sabbione, S.M. Ibañez, E.N. Martínez, M.C. Añón, and A. Scilingo, declare that they have no conflict of interest.

Supplementary material

11130_2016_540_MOESM1_ESM.docx (8.1 mb)
ESM 1 (DOCX 8291 kb)
11130_2016_540_MOESM2_ESM.docx (1.9 mb)
ESM 2 (DOCX 1953 kb)


  1. 1.
    Hasler CM, Bloch AS, Thomson CA, Enrione E, Manning C (2004) Position of the American Dietetic Association: functional foods. J Am Diet Assoc 104:814–826CrossRefGoogle Scholar
  2. 2.
    Martínez-Maqueda D, Miralles B, Recio I, Hernández-Ledesma B (2012) Antihypertensive peptides from food proteins: a review. Food Funct 3:350–361CrossRefGoogle Scholar
  3. 3.
    Madureira AR, Tavares T, Gomes AMP, Pintado ME, Malcata FX (2010) Invited review: physiological properties of bioactive peptides obtained from whey proteins. J Dairy Sci 93:437–455CrossRefGoogle Scholar
  4. 4.
    Kim SK, Wijesekara I (2010) Development and biological activities of marine-derived bioactive peptides: a review. J Funct Foods 2:1–9CrossRefGoogle Scholar
  5. 5.
    Udenigwe CC, Aluko RE (2012) Food protein-derived bioactive peptides: production, processing, and potential health benefits. J Food Sci 77:11–24CrossRefGoogle Scholar
  6. 6.
    Silva-Sánchez C, Barba de la Rosa AP, León-Galván MF, De Lumen BO, De León-Rodríguez A, González de Mejía E (2008) Bioactive peptides in amaranth (Amaranthus hypochondriacus) seed. J Agric Food Chem 56:1233–1240CrossRefGoogle Scholar
  7. 7.
    López VRL, Razzeto GS, Escudero NL, Giménez MS (2013) Biochemical and molecular study of the influence of Amaranthus hypochondriacus flour on serum and liver lipids in rats treated with ethanol. Plant Foods Hum Nutr 68:396–402CrossRefGoogle Scholar
  8. 8.
    Tironi V, Añón MC (2010) Amaranth as a source of antioxidant peptides: effect of proteolysis. Food Res Int 43:315–322CrossRefGoogle Scholar
  9. 9.
    Sabbione AC, Scilingo A, Añón MC (2015) Potential antithrombotic activity detected in amaranth proteins and its hydrolysates. LWT-Food Sci Technol 60:171–177CrossRefGoogle Scholar
  10. 10.
    Ventureira JL, Martínez EN, Añón MC (2012) Effect of acid treatment on structural and foaming properties of soy amaranth protein mixtures. Food Hydrocoll 29:272–279CrossRefGoogle Scholar
  11. 11.
    Hemalatha KPJ, Siva Prasad D (2003) Changes in the metabolism of protein during germination of sesame (Sesamum indicum L.) seeds. Plant Foods Hum Nutr 58:1–10CrossRefGoogle Scholar
  12. 12.
    Schaller A (2004) A cut above the rest: the regulatory function of plant proteases. Planta 220:183–197CrossRefGoogle Scholar
  13. 13.
    Martínez EN, Añón MC (1996) Composition and structural characterization of amaranth proteins isolates. An electrophoretic and calorimetric study. J Agric Food Chem 44:2523–2530CrossRefGoogle Scholar
  14. 14.
    Paredes-López O (1994) Amaranth: biology, chemistry and technology. CRC Press, Boca Raton, FLGoogle Scholar
  15. 15.
    Martínez EN, Castellani OF, Añón MC (1997) Common molecular features among amaranth storage proteins. J Agric Food Chem 45:3832–3839CrossRefGoogle Scholar
  16. 16.
    Schägger H (2006) Tricine-SDS-PAGE. Nat Protoc 1:16–22CrossRefGoogle Scholar
  17. 17.
    Nielsen PM, Petersen D, Dambmann C (2001) Improved method for determining food protein degree of hydrolysis. J Food Sci 66:642–646CrossRefGoogle Scholar
  18. 18.
    Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the folin phenol reagent. J Biol Chem 193:265–275Google Scholar
  19. 19.
    Yang WG, Wang Z, Xu SY (2007) A new method for determination of antithrombotic activity of egg white protein hydrolysate by microplate reader. Chin Chem Lett 18:449–451CrossRefGoogle Scholar
  20. 20.
    Zhang S, Wang Z, Xu SY (2008) Antioxidant and antithrombotic activities of rapeseed peptides. JAOCS 85:521–527Google Scholar
  21. 21.
    Orsini Delgado MC, Galleano M, Añón MC, Tironi VA (2015) Amaranth peptides from simulated gastrointestinal digestion: antioxidant activity against reactive species. Plant Foods Hum Nutr 70:27–34CrossRefGoogle Scholar
  22. 22.
    Siddhuraju P (2006) The antioxidant activity and free radical-scavenging capacity of phenolics of raw and dry heated moth bean (Vigna aconitifolia) (jacq.) marechal seed extracts. Food Chem 99:149–157CrossRefGoogle Scholar
  23. 23.
    Atanassov A, Tchorbanov B (2009) Synthetic and natural peptides as antithrombotic agents. A view on the current development. Biotechnol Biotechnol Equip 23:1109–1114CrossRefGoogle Scholar
  24. 24.
    Abugoch LE, Martínez EN, Añón MC (2010) Influence of pH on structure and function of amaranth (A. hypochondriacus) protein isolates. Cereal Chem 87:448–453CrossRefGoogle Scholar
  25. 25.
    Walsh R, Martin E, Darvesh S (2010) A method to describe enzyme-catalyzed reactions by combining steady state and time course enzyme kinetic parameters. Biochim Biophys Acta 1800:1–5CrossRefGoogle Scholar
  26. 26.
    Condés MC, Scilingo A, Añón MC (2009) Characterization of amaranth proteins modified by trypsin proteolysis. Structural and functional changes LWT 42:963–970Google Scholar
  27. 27.
    Marcone MF, Kakuda Y (1999) A comparative study of functional properties of amaranth and soybean globulins isolates. Nahrung 43:368–373CrossRefGoogle Scholar
  28. 28.
    Quiroga AV, Martínez EN, Rogniaux H, Geairon A, Añón MC (2010) Amaranth (Amaranthus hypochondriacus) vicilin subunit structure. J Agric Food Chem 58:12957–12963CrossRefGoogle Scholar
  29. 29.
    Laudano AP, Doolittle RF (1978) Synthetic peptide derivatives that bind to fibrinogen and prevent the polymerization of fibrin monomers. Proc Natl Acad Sci USA 75:3085–3089CrossRefGoogle Scholar
  30. 30.
    Lee KA, Kim SH (2005) SSGE and DEE, new peptides isolated from a soy protein hydrolysate that inhibit platelet aggregation. Food Chem 90:389–394CrossRefGoogle Scholar
  31. 31.
    Hyun KW, Jeong SC, Lee DH, Park JS, Lee JS (2006) Isolation and characterization of a novel platelet aggregation inhibitory peptide from the medicinal mushroom, Inonotus obliquus. Peptides 27:1173–1178CrossRefGoogle Scholar
  32. 32.
    Jo HY, Jung WK, Kim SK (2008) Purification and characterization of a novel anticoagulant peptide from marine echiuroid worm Urechis unicinctus. Process Biochem 43:179–184CrossRefGoogle Scholar
  33. 33.
    Gan ZR, Gould RJ, Jacobs JW, Freidman PA, Polokoff MA (1988) Echistatin, a potent platelet aggregation inhibitor from venom of viper, Echis carinatus. J Biol Chem 263:19827–19832Google Scholar
  34. 34.
    Re R, Pellegrini A, Proteggente A, Pannala M, Yang C, Rice-Evans C (1999) Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic Biol Med 26:1231–1237CrossRefGoogle Scholar
  35. 35.
    Orsini Delgado MC, Tironi VA, Añón MC (2011) Antioxidant activity of amaranth protein or their hydrolysates under simulated gastrointestinal digestion. LWT 44:1752–1760CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Ana Clara Sabbione
    • 1
    • 2
    • 3
  • Sabrina M. Ibañez
    • 2
  • E. Nora Martínez
    • 1
    • 2
    • 3
  • María Cristina Añón
    • 1
    • 2
    • 3
  • Adriana A. Scilingo
    • 1
    • 2
  1. 1.Centro de Investigación y Desarrollo en Criotecnología de Alimentos (CIDCA)La PlataArgentina
  2. 2.Facultad de Ciencias ExactasUniversidad Nacional de La PlataLa PlataArgentina
  3. 3.CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), CCT-La PlataLa PlataArgentina

Personalised recommendations