New possibilities for the valorization of olive oil by-products.

In this contribution, the capabilities of pressurized liquid extraction (PLE) using food-grade solvents, such as water and ethanol, to obtain antioxidant extracts rich on polyphenolic compounds from olive leaves are studied. Different extraction conditions were tested, and the PLE obtained extracts were characterized in vitro according to their antioxidant capacity (using the DPPH radical scavenging and the TEAC assays) and total phenols amounts. The most active extracts were obtained with hot pressurized water at 200 °C (EC(50) 18.6 μg/mL) and liquid ethanol at 150 °C (EC(50) 27.4 μg/mL), attaining at these conditions high extraction yields, around 40 and 30%, respectively. The particular phenolic composition of the obtained extracts was characterized by LC-ESI-MS. Using this method, 25 different phenolic compounds could be tentatively identified, including phenolic acids, secoiridoids, hydroxycinnamic acid derivatives, flavonols and flavones. Among them, hydroxytyrosol, oleuropein and luteolin-glucoside were the main phenolic antioxidants and were quantified on the extracts together with other minor constituents, by means of a UPLC-MS/MS method. Results showed that using water as extracting agent, the amount of phenolic compounds increased with the extraction temperature, being hydroxytyrosol the main phenolic component on the water PLE olive leaves extracts, reaching up to 8.542 mg/g dried extract. On the other hand, oleuropein was the main component on the extracts obtained with ethanol (6.156-2.819 mg/g extract). Results described in this work demonstrate the good possibilities of using PLE as a useful technique for the valorization of by-products from the olive oil industry, such as olive leaves.

Herrero M ，Temirzoda TN ，Segura-Carretero A ，Quirantes R ，Plaza M ，Ibañez E ... -  《-》

Enrichment of antioxidant compounds from lemon balm (Melissa officinalis) by pressurized liquid extraction and enzyme-assisted extraction.

In this work enzyme-assisted extraction (EAE) and pressurized liquid extraction (PLE) are applied for extraction of natural compounds from lemon balm (Melissa officinalis). Cellulase, endo-β-1,4 xylanase and pectinase were studied in order to degrade cell wall of lemon balm leaves and to release phenolic compounds. On the other hand, in order to compare the performance obtained with EAE, PLE using water and ethanol was employed maintaining 150°C as extraction temperature. The obtained extracts were characterized in terms of antioxidant capacity by using DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging and trolox equivalent antioxidant capacity (TEAC) in vitro assays, whereas the Folin-Ciocalteu procedure was employed to estimate the total phenols content. On the other hand, extracts were chemically characterized by liquid chromatography tandem mass spectrometry (LC-MS/MS). The results showed that EAE enhanced the total phenolic content and the antioxidant capacity compared to a non-enzymatic control. PLE extracts presented higher amount of phenols and antioxidant capacity than enzyme-assisted extracts, reaching the highest values on water extracts (193.18mggallicacid/gextract and EC50=6.81μg/mL). Among the bioactive phenolic compounds identified in lemon balm, rosmarinic acid was the main component, although other important compounds were also identified, such as caffeic acid derivatives (salvianolic acids, lithospermic acid) and rosmarinic acid derivatives (rosmarinic acid hexoside, sagerinic acid, sulfated rosmarinic acid). The present study confirms that EAE and PLE can be considered alternative methods for the extraction of natural compounds with biological activity from natural sources.

Miron TL ，Herrero M ，Ibáñez E 《-》

HPLC-ESI-QTOF-MS as a powerful analytical tool for characterising phenolic compounds in olive-leaf extracts.

Olea europaea L. leaves may be considered a cheap, easily available natural source of phenolic compounds. In a previous study we evaluated the possibility of obtaining bioactive phenolic compounds from olive leaves by pressurised liquid extraction (PLE) for their use as natural anti-oxidants. The alimentary use of these kinds of extract makes comprehensive knowledge of their composition essential. To undertake a comprehensive characterisation of two olive-leaf extracts obtained by PLE using high-performance liquid chromatography coupled to electrospray ionisation and quadrupole time-of-flight mass spectrometry (HPLC-ESI-QTOF-MS). Olive leaves were extracted by PLE using ethanol and water as extraction solvents at 150°C and 200°C respectively. Separation was carried out in a HPLC system equipped with a C₁₈-column working in a gradient elution programme coupled to ESI-QTOF-MS operating in negative ion mode. This analytical platform was able to detect 48 compounds and tentatively identify 31 different phenolic compounds in these extracts, including secoiridoids, simple phenols, flavonoids, cinnamic-acid derivatives and benzoic acids. Lucidumoside C was also identified for the first time in olive leaves. The coupling of HPLC-ESI-QTOF-MS led to the in-depth characterisation of the olive-leaf extracts on the basis of mass accuracy, true isotopic pattern and tandem mass spectrometry (MS/MS) spectra. We may conclude therefore that this analytical tool is very valuable in the study of phenolic compounds in plant matrices.

Quirantes-Piné R ，Lozano-Sánchez J ，Herrero M ，Ibáñez E ，Segura-Carretero A ，Fernández-Gutiérrez A ... -  《-》

Use of advanced techniques for the extraction of phenolic compounds from Tunisian olive leaves: phenolic composition and cytotoxicity against human breast cancer cells.

A comparison among different advanced extraction techniques such as microwave-assisted extraction (MAE), supercritical fluid extraction (SFE) and pressurized liquid extraction (PLE), together with traditional solid-liquid extraction, was performed to test their efficiency towards the extraction of phenolic compounds from leaves of six Tunisian olive varieties. Extractions were carried out at the best selected conditions for each technique; the obtained extracts were chemically characterized using high-performance liquid chromatography (HPLC) coupled to electrospray time-of-flight mass spectrometry (ESI-TOF-MS) and electrospray ion trap tandem mass spectrometry (ESI-IT-MS(2)). As expected, higher extraction yields were obtained for PLE while phenolic profiles were mainly influenced by the solvent used as optimum in the different extraction methods. A larger number of phenolic compounds, mostly of a polar character, were found in the extracts obtained by using MAE. Best extraction yields do not correlate with highest cytotoxic activity against breast cancer cells, indicating that cytotoxicity is highly dependent on the presence of certain compounds in the extracts, although not exclusively on a single compound. Therefore, a multifactorial behavior is proposed for the anticancer activity of olive leaf compounds.

Taamalli A ，Arráez-Román D ，Barrajón-Catalán E ，Ruiz-Torres V ，Pérez-Sánchez A ，Herrero M ，Ibañez E ，Micol V ，Zarrouk M ，Segura-Carretero A ，Fernández-Gutiérrez A ... -  《-》

Methods for preparing phenolic extracts from olive cake for potential application as food antioxidants.

Suárez M ，Romero MP ，Ramo T ，Macià A ，Motilva MJ ... -  《-》