Contents lists available at ScienceDirect The Journal of Supercritical Fluids journal homepage: www.elsevier.com/locate/supflu Selective fractionation and isolation of allelopathic compounds from Helianthus annuus L. leaves by means of high-pressure techniques F. Fuentes-Gandara a,b , A. Torres b , M.T. Fernández-Ponce c, ⁎ , L. Casas c , C. Mantell c , R. Varela b , E.J. Martínez de la Ossa-Fernández c , F.A. Macías b a Department of Exact and Natural Sciences, Universidad de la Costa, Calle 58 Nº 55-66, Barranquilla, Colombia b Allelopathy Group, Department of Organic Chemistry, Institute of Biomolecules (INBIO), University of Cadiz, Av. República Saharahui, nº 7, 11510, Puerto Real, Cadiz, Spain c Chemical Engineering and Food Technology Department, Institute of Viticulture and Agri-Food Research (IVAGRO), University of Cadiz, Av. República Saharahui s/n, 11510, Puerto Real, Cadiz, Spain GRAPHICAL ABSTRACT ARTICLE INFO Keywords: Helianthus annus Fractionation Allelopathy Supercritical fluids Enhanced solvent extraction ABSTRACT The allelopathic potential of Helianthus annuus L. leaves was study based on bio-directed chemical fractionation approach. Aerial parts of H. annuus were extracted by means of SFE using supercritical carbon dioxide (scCO 2 ) and ESE using CO 2 +50% EtOH/H 2 O (varying ethanol in water from 0 to 100%). Extractions were carried out at 400 bar, 55 °C, 20 g/min and for 4 h. Then, extracts were fractionated in three separators at the following conditions: S1: 200 bar/45 °C; S2: 90 bar/40 °C; and S3: 1 atm/30 °C. ESE obtained higher overall yields than scCO 2 and the use of water as cosolvent (CO 2 +50% H 2 O) resulted in a S3 fraction free from chlorophylls and rich in bioactive compounds. 14 compounds, including fatty acids, terpenes, flavonoids and heliannuols, were isolated from this fraction. After performing the bioassay on pure compounds, heliannuol D, tambulin, pinor- esinol and sesquiterpene 10-oxo-isodauc-3-en-15-al showed the most effective inhibitor profiles. 1. Introduction Plants and their constituents stand as a key factor in the progression of modern studies and the knowledge on the biological activity of active substances. Plants are rich in bioactive compounds, defined as sec- ondary metabolites, which are products from metabolism themselves, and they may have a therapeutic or toxic effect on humans, animals and plants [1]. Among functional properties of plants, allelopathy is the https://doi.org/10.1016/j.supflu.2018.08.004 Received 22 June 2018; Received in revised form 2 August 2018; Accepted 3 August 2018 ⁎ Corresponding author. E-mail address: teresafernandez.ponce@uca.es (M.T. Fernández-Ponce). The Journal of Supercritical Fluids 143 (2019) 32–41 Available online 04 August 2018 0896-8446/ © 2018 Elsevier B.V. All rights reserved. T