title: Analysis of free hydroxytyrosol in human plasma following the administration of olive oil.
abstract: Hydroxytyrosol (HT) from olive oil, a potent bioactive molecule with health benefits, has a poor bioavailability, its free form (free HT) being undetectable so far. This fact leads to the controversy whether attained HT concentrations after olive oil polyphenol ingestion are too low to explain the observed biological activities. Due to this, an analytical methodology to determine free HT in plasma is crucial for understanding HT biological activity. Plasma HT instability and low concentrations have been major limitations for its quantification in clinical studies. Here, we describe a method to detect and quantify free HT in human plasma by using liquid chromatography coupled to tandem mass spectrometry. The method encompasses different steps of sample preparation including plasma stabilization, protein precipitation, selective derivatization with benzylamine, and purification by solid-phase extraction. A high sensitivity (LOD, 0.3ng/mL), specificity and stability of HT is achieved following these procedures. The method was validated and its applicability was demonstrated by analyzing human plasma samples after olive oil intake. A pharmacokinetic comparison was performed measuring free HT plasma concentrations following the intake of 25mL of ordinary olive oil (nearly undetectable concentrations) versus an extra-virgin olive oil (Cmax=4.40ng/mL). To our knowledge, this is the first time that an analytical procedure for quantifying free HT in plasma after olive oil dietary doses has been reported. The present methodology opens the door to a better understanding of the relationship between HT plasma concentrations and its beneficial health effects.
authors.affiliation: Integrative Pharmacology and Systems Neuroscience, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; School of Medicine, Universitat Autònoma de Barcelona, Bellaterra, Spain; CIBER de Fisiopatología Obesidad y Nutrición, Santiago de Compostela 15706, Spain. Electronic address: apastor@imim.es.
authors.forename: Antoni
authors.initials: A
authors.lastname: Pastor
authors.affiliation: Integrative Pharmacology and Systems Neuroscience, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; CIBER de Fisiopatología Obesidad y Nutrición, Santiago de Compostela 15706, Spain; Universitat Pompeu Fabra (CEXS-UPF), Dr. Aiguader 88, 08003 Barcelona, Spain. Electronic address: jrodriguez1@imim.es.
authors.forename: Jose
authors.initials: J
authors.lastname: Rodríguez-Morató
authors.affiliation: Integrative Pharmacology and Systems Neuroscience, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; Universitat Pompeu Fabra (CEXS-UPF), Dr. Aiguader 88, 08003 Barcelona, Spain. Electronic address: eolesti@imim.es.
authors.forename: Eulàlia
authors.initials: E
authors.lastname: Olesti
authors.affiliation: Integrative Pharmacology and Systems Neuroscience, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; CIBER de Fisiopatología Obesidad y Nutrición, Santiago de Compostela 15706, Spain. Electronic address: mpujadas@imim.es.
authors.forename: Mitona
authors.initials: M
authors.lastname: Pujadas
authors.affiliation: Integrative Pharmacology and Systems Neuroscience, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; School of Medicine, Universitat Autònoma de Barcelona, Bellaterra, Spain. Electronic address: cperez@imim.es.
authors.forename: Clara
authors.initials: C
authors.lastname: Pérez-Mañá
authors.affiliation: Integrative Pharmacology and Systems Neuroscience, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; CIBER de Fisiopatología Obesidad y Nutrición, Santiago de Compostela 15706, Spain. Electronic address: okhymenets@gmail.com.
authors.forename: Olha
authors.initials: O
authors.lastname: Khymenets
authors.affiliation: CIBER de Fisiopatología Obesidad y Nutrición, Santiago de Compostela 15706, Spain; Cardiovascular Risk and Nutrition Research Group, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain. Electronic address: mfito@imim.es.
authors.forename: Montserrat
authors.initials: M
authors.lastname: Fitó
authors.affiliation: CIBER de Fisiopatología Obesidad y Nutrición, Santiago de Compostela 15706, Spain; Cardiovascular Risk and Nutrition Research Group, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; NUPROAS (Nutritional Project Assessment), Handesbolag (NUPROAS HB), Nacka, Sweden. Electronic address: Maria.nuproas@gmail.com.
authors.forename: María-Isabel
authors.initials: MI
authors.lastname: Covas
authors.affiliation: NFOC Group, CIBERDEM, St. Joan de Reus University Hospital, IISPV, Facultat de Medicina i Ciències de la Salut, Universitat Rovira i Virgili, Reus, Spain. Electronic address: rosa.sola@urv.cat.
authors.forename: Rosa
authors.initials: R
authors.lastname: Solá
authors.affiliation: Food Technology Department, Agrotecnio Research Center, Escuela Técnica Superior de Ingeniería Agraria, University of Lleida, Lleida, Spain. Electronic address: motilva@tecal.udl.cat.
authors.forename: María-José
authors.initials: MJ
authors.lastname: Motilva
authors.affiliation: Integrative Pharmacology and Systems Neuroscience, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; School of Medicine, Universitat Autònoma de Barcelona, Bellaterra, Spain; Hospital Universitari Germans Trias i Pujol (IGTP), Badalona, Spain. Electronic address: Mfarre.germanstrias@gencat.cat.
authors.forename: Magí
authors.initials: M
authors.lastname: Farré
authors.affiliation: Integrative Pharmacology and Systems Neuroscience, IMIM (Hospital del Mar Research Institute), Dr. Aiguader 88, 08003 Barcelona, Spain; CIBER de Fisiopatología Obesidad y Nutrición, Santiago de Compostela 15706, Spain; Universitat Pompeu Fabra (CEXS-UPF), Dr. Aiguader 88, 08003 Barcelona, Spain. Electronic address: rtorre@imim.es.
authors.forename: Rafael
authors.initials: R
authors.lastname: de la Torre
mesh_terms: D001682:Biological Availability; D001774:Blood Chemical Analysis; D002853:Chromatography, Liquid; D004042:Dietary Fats, Unsaturated / Q000097:blood / Q000378:metabolism; D006801:Humans; D031658:Olea / Q000737:chemistry; D000069463:Olive Oil / Q000378:metabolism*; D010626:Phenylethyl Alcohol / Q000031:analogs & derivatives* / Q000097:blood; D059808:Polyphenols; D052616:Solid Phase Extraction; D053719:Tandem Mass Spectrometry
chemical_list: D004042:Dietary Fats, Unsaturated; D000069463:Olive Oil; D059808:Polyphenols; C005975:3,4-dihydroxyphenylethanol; D010626:Phenylethyl Alcohol
keywords: Benzylamine derivatization; Health; Humans; Hydroxytyrosol; LC–MS/MS; Virgin olive oil