Food Analytical Methods

, Volume 9, Issue 8, pp 2174–2181 | Cite as

Optimisation of the Supercritical Fluid Extraction of Antioxidants from Broccoli Leaves

  • Esther Arnáiz
  • José Bernal
  • María Teresa Martín
  • Juan Carlos Diego
  • José Luis Bernal
  • Laura Toribio RecioEmail author


This work presents the application of supercritical fluid extraction (SFE), an environmentally friendly methodology, to obtain broccoli leaves extracts with a high antioxidant activity. The following processing variables: pressure, temperature, percentage of ethanol and dynamic extraction time were optimised by means of a central composite design, combined with response surface methodology. The optimised conditions were applied to the extraction of broccoli leaves from different cultivars (Naxos, Nubia, Marathon, Parthenon and Viola). The optimal values of the extraction variables were 150 bars, 35 °C, 20 % ethanol and 10 min of dynamic extraction time. Percentage of ethanol and the combined effect of pressure and time were the factors that affected the extraction the most. The results obtained from different cultivars showed that the Naxos variety provided the extracts with the highest antioxidant activity and total phenolic content; moreover, DPPH (EC 50) values were higher than those previously reported for water extracts. The extracts were recovered in a small quantity of ethanol that could be easily eliminated, obtaining a residue free of contaminants.


SFE Broccoli leaves Antioxidant activity Total phenolic content DPPH ABTS 



The authors wish to thank the Junta de Castilla y León for financial support (GIR 127) and Dra. Cristina García Viguera (CEBAS-CSIC, Murcia) for the samples supplied. M.T. Martín would like to thank the Spanish Ministry of Science and Innovation for her “Ramón y Cajal” contract.

Compliance with Ethical Standards


This study was financially supported by Junta de Castilla y León (GIR 127).

Conflict of Interest

Esther Arnáiz declares that she has no conflict of interest. José Bernal declares that he has no conflict of interest. María Teresa Martín declares that she has no conflict of interest. Juan Carlos Diego declares that he has no conflict of interest. José Luis Bernal declares that he has no conflict of interest. Laura Toribio declares that she has no conflict of interest.

Ethical Approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Informed Consent

Informed consent is not applicable for this study.


  1. Ares AM, Nozal MJ, Bernal JL, Bernal J (2014) Optimized extraction, separation and quantification of twelve intact glucosinolates in broccoli leaves. Food Chem 152:66–74CrossRefGoogle Scholar
  2. Arnáiz E, Bernal J, Martín MT, García-Viguera C, Bernal JL, Toribio L (2011) Supercritical fluid extraction of lipids from broccoli leaves. Eur J Lipid Sci Technol 113:479–486CrossRefGoogle Scholar
  3. Bandoniene D, Pukalskas A, Venskutomis PR, Gruzdiene D (2000) Preliminary screening of antioxidant of some plant extracts in rapeseed oil. Food Res Int 23:785–791CrossRefGoogle Scholar
  4. Bao J, Cai Y, Sun M, Wang G, Corke H (2005) Anthocyanins, flavonols, and free radical scavenging activity of Chinese bayberry (Myrica rubra) extracts and their color properties and stability. J Agric Food Chem 53:2327–2332CrossRefGoogle Scholar
  5. Bhandari SR, Kwak JH (2015) Chemical composition and antioxidant activity in different tissues of Brassica vegetables. Molecules 20:1228–1243CrossRefGoogle Scholar
  6. Brand-Williams W, Cuvelier ME, Berset C (1995) Use of a free radical method to evaluate antioxidant activity. Food Sci Technol LEB 28:25–30CrossRefGoogle Scholar
  7. Cao G, Sofic E, Prior RL (1996) Antioxidant capacity of tea and common vegetables. J Agric Food Chem 44:3426–3431CrossRefGoogle Scholar
  8. Cavero S, García-Risco MR, Marín FR, Jaime L, Santoyo S, Señoráns FJ, Reglero G, Ibañez E (2006) Supercritical fluid extraction of antioxidant compounds from oregano. Chemical and functional characterization via LC–MS and in vitro assays. J Supercrit Fluids 38:62–69CrossRefGoogle Scholar
  9. Chaitanya KV, Rama-Krishna CH, Khasim-Beebi SK, Divya K (2015) Supercritical fluid extraction of functional ingredients from plants: a review. Curr Biochem Eng 2:24–32CrossRefGoogle Scholar
  10. Chu YF, Sun J, Wu X, Liu RH (2002) Antioxidant and antiproliferative activities of common vegetables. J Agric Food Chem 50:6910–6916CrossRefGoogle Scholar
  11. Floegel A, Kim DO, Chung SJ, Koo SI, Chun OK (2011) Comparison of ABTS/DPPH assays to measure antioxidant capacity in popular antioxidant-rich US foods. J Food Compos Anal 24:1043–1048CrossRefGoogle Scholar
  12. Gawlik-Dziki U (2008) Effect of hydrothermal treatment on the antioxidant properties of broccoli (Brassica oleracea var. botrytis italica) florets. Food Chem 109:393–401CrossRefGoogle Scholar
  13. Hall C III (2001) Sources of natural antioxidants: oilseeds, nuts, cereals, legumes, animal products and microbial sources. In: Pokorny J, Yanishlieva N, Gordon M (eds) Antioxidants in food. CRC Press, New York, pp 159–209CrossRefGoogle Scholar
  14. Herrero M, Mendiola JA, Cifuentes A, Ibañez E (2010) Supercritical fluid extraction: recent advances and applications. J Chromatogr A 1217:2495–2511CrossRefGoogle Scholar
  15. Jang HW, Moon JK, Shibamoto T (2015) Analysis and antioxidant activity of extracts from broccoli (Brassica oleracea L.) sprouts. J Agric Food Chem 63:1169–1174CrossRefGoogle Scholar
  16. Jokić S, Cvjetko M, Božić Ð, Fabek S, Toth N, Vorkapić-Furač J, Radojčić-Redovniković I (2012) Optimisation of microwave-assisted extraction of phenolic compounds from broccoli and its antioxidant activity. Int J Food Sci Technol 47:2613–2619CrossRefGoogle Scholar
  17. Kähkönen MP, Hopia AI, Vuorela HJ, Rauha JP, Pihlaja K, Kujala TS, Heinonen M (1999) Antioxidant activity of plant extracts containing phenolic compounds. J Agric Food Chem 47:3954–3962CrossRefGoogle Scholar
  18. Kelsey NA, Wilkins HM, Linseman DA (2010) Nutraceutical antioxidants as novel neuroprotective agents. Molecules 15:7792–7814CrossRefGoogle Scholar
  19. Kitrytè V, Šaduikis A, Venskutonis PR (2015) Assessment of antioxidant capacity of brewer’s spent grain and its supercritical carbon dioxide extract as sources of valuable dietary ingredients. J Food Eng 167:18–24CrossRefGoogle Scholar
  20. Krishnaiah D, Sarbatly R, Nithyanandam R (2011) A review of the antioxidant potential of medicinal plant species. Food Bioprod Process 89:217–233CrossRefGoogle Scholar
  21. Kumar Roy M, Raj Juneja L, Isobe S, Tsushida T (2009) Steam processed broccoli (Brassica oleracea) has higher antioxidant activity in chemical and cellular assay systems. Food Chem 114:263–269CrossRefGoogle Scholar
  22. Kurilich AC, Jeffery EH, Juvik JA, Wallig MA, Klein BP (2002) Antioxidant capacity of different broccoli (Brassica oleracea) genotypes using the oxygen radical absorbance capacity (ORAC) assay. J Agric Food Chem 50:5053–5057CrossRefGoogle Scholar
  23. Menoci-Gonçalves R, Ortega-Terra Lemos C, Correa-Ramos Leal I, Vataru-Nakamura C, Garcia-Cortez DA, Da Silva EA, Ferreira-Cabral V, Cardozo-Filho L (2013) Comparing conventional and supercritical extraction of (−)-Mammea A/BB and the antioxidant activity of Calophyllum brasiliense extracts. Molecules 18:6215–6229CrossRefGoogle Scholar
  24. Moreno DA, Carvajal M, López-Berenguer C, García-Viguera C (2006) Chemical and biological characterisation of nutraceutical compounds of broccoli. J Pharm Biomed Anal 41:1508–1522CrossRefGoogle Scholar
  25. Podsędek A (2007) Natural antioxidants and antioxidant capacity of Brassica vegetables: a review. Food Sci Technol LEB 40:1–11CrossRefGoogle Scholar
  26. Re R, Pellegrini N, Proteggente A, Pannala A, Yang M, Rice-Evans C (1999) Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic Biol Med 26:1231–1237CrossRefGoogle Scholar
  27. Soengas P, Cartea ME, Francisco M, Sotelo T, Velasco P (2012) New insights into antioxidant activity of Brassica crops. Food Chem 134:725–733CrossRefGoogle Scholar
  28. Solana M, Boschiero I, Dall’acqua S, Bertucco A (2015) A comparison between supercritical fluid and pressurized liquid extraction methods for obtaining phenolic compounds from Asparagus officinalis L. J Supercrit Fluids 100:201–208CrossRefGoogle Scholar
  29. Sun T, Powers JR, Tang J (2007) Evaluation of the antioxidant activity of asparagus, broccoli and their juices. Food Chem 105:101–106CrossRefGoogle Scholar
  30. Zhang D, Hamauzu Y (2004) Phenolics, ascorbic acid, carotenoids and antioxidant activity of broccoli and their changes during conventional and microwave cooking. Food Chem 88:503–509CrossRefGoogle Scholar
  31. Zhou SH, Fang ZX, Lü Y, Chen JC, Liu DH, Ye XQ (2009) Phenolics and antioxidant properties of bayberry (Myrica rubra Sieb. et Zucc.) pomace. Food Chem 112:394–399CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Esther Arnáiz
    • 1
  • José Bernal
    • 1
  • María Teresa Martín
    • 1
  • Juan Carlos Diego
    • 1
  • José Luis Bernal
    • 1
  • Laura Toribio Recio
    • 1
    Email author
  1. 1.I.U. CINQUIMA, Analytical Chemistry GroupUniversity of ValladolidValladolidSpain

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