Olive leaf phenolics and cardiovascular risk reduction: Physiological effects and mechanisms of action

Olive oil, an important component of the Mediterranean diet, is rich in polyphenols and is known to possess positive health effects relative to other dietary fats. In addition, the leaves of the olive plant (Olea europaea) contain similar phenolics (oleuropein, luteolin-7-glucoside, apigenin-7-glucoside, verbascoside and hydroxytyrosol) to those of olives and olive oil, although at higher concentrations. For example, the most abundant is the secoiridoid, oleuropein, representing 1–14% of olive leaf weight vs. 0.005–0.12% in olive oil. Although currently considered a waste product of the olive oil industry, recent research has suggested beneficial effects of phenolic-rich olive leaf extracts (OLE) in modifying cardiovascular risk biomarkers such as blood pressure, hyperglycaemia, oxidative stress and inflammation, as well as improving vascular function and lipid profiles. Despite this, data regarding the biological actions of OLE has mostly derived from animal, in vitro and ex vivo studies, with limited evidence deriving from human trials. Although the absorption and metabolism of olive oil phenolics has been investigated, less is known about the bioavailability of phenolics from OLE, limiting the interpretation of existing in vitro and ex vivo data. The current review will begin by describing the phenolic composition of olive leaves in comparison with that of the better studied olive oil. It will then review the effects of OLE on cardiovascular risk factors, covering both animal and human studies and will end by considering potential mechanisms of action

Authorised EU health claims for polyphenols in olive oil

There is evidence that various phenolic compounds (such as oleuropein, tyrosol and hydroxytyrosol) found in virgin olive oil may be responsible for the beneficial effects on cardiovascular disease. In the EU there is an authorized health claim that‘olive oil polyphenols contribute to the protection of blood lipids from oxidative stress’ on the basis of human studies showing significantly reduced levels of oxidized LDL in plasma after virgin olive oil consumption. The claim may be used only for olive oil that contains at least 5 mg of hydroxytyrosol and its derivatives per 20 g of olive oil. Other claims proposed (including maintenance of normal blood pressure and HDL cholesterol concentration, and anti-inflammatory properties) were rejected.

Secoiridoids delivered as olive leaf extract induce acute improvements in human vascular function and reduction of an inflammatory cytokine: a randomised, double-blind, placebo-controlled, cross-over trial

The leaves of the olive plant (Olea europaea) are rich in polyphenols, of which oleuropein and hydroxytyrosol (HT) are most characteristic. Such polyphenols have been demonstrated to favourably modify a variety of cardiovascular risk factors. The aim of the present intervention was to investigate the influence of olive leaf extract (OLE) on vascular function and inflammation in a postprandial setting and to link physiological outcomes with absorbed phenolics. A randomised, double-blind, placebo-controlled, cross-over, acute intervention trial was conducted with eighteen healthy volunteers (nine male, nine female), who consumed either OLE (51 mg oleuropein; 10mg HT), or a matched control (separated by a 4-week wash out) on a single occasion. Vascular function was measured by digital volume pulse (DVP), while blood collected at baseline, 1, 3 and 6 h was cultured for 24 h in the presence of lipopolysaccharide in order to investigate effects on cytokine production. Urine was analysed for phenolic metabolites by HPLC. DVP-stiffness index and ex vivo IL-8 production were significantly reduced (P < 0.05) after consumption of OLE compared to the control. These effects were accompanied by the excretion of several phenolic metabolites, namely HT and oleuropein derivatives, which peaked in urine after 8-24 h. The present study provides the first evidence that OLE positively modulates vascular function and IL-8 production in vivo, adding to growing evidence that olive phenolics could be beneficial for health.

LC-DAD/ESI-MS/MS study of phenolic compounds in ash (Fraxinus excelsior L. and F. americana L.) heartwood. Effect of toasting intensity at cooperage

The phenolic composition of heartwood extracts from Fraxinus excelsior L. and F. americana L., both before and after toasting in cooperage, was studied using LC-DAD/ESI-MS/MS. Low-molecular weight (LMW) phenolic compounds, secoiridoids, phenylethanoid glycosides, dilignols and oligolignols compounds were detected, and 48 were identified, or tentatively characterized, on the basis of their retention time, UV/Vis and MS spectra, and MS fragmentation patterns. Some LMW phenolic compounds like protocatechuic acid and aldehyde, hydroxytyrosol and tyrosol, were unlike to those for oak wood, while ellagic and gallic acid were not found. The toasting of wood resulted in a progressive increase in lignin degradation products with regard to toasting intensity. The levels of some of these compounds in medium-toasted ash woods were much higher than those normally detected in toasted oak, highlighting vanillin levels, thus a more pronounced vanilla character can be expected when using toasted ash wood in the aging wines. Moreover, in seasoned wood, we found a great variety of phenolic compounds which had not been found in oak wood, especially oleuropein, ligstroside and olivil, along with verbascoside and isoverbascoside in F. excelsior, and oleoside in F. americana. Toasting mainly provoked their degradation, thus in medium-toasted wood, only four of them were detected. This resulted in a minor differentiation between toasted ash and oak woods. The absence of tannins in ash wood, which are very important in oak wood, is another peculiar characteristic that should be taken into account when considering its use in cooperage. Copyright (C) 2012 John Wiley & Sons, Ltd.

Individuazione e caratterizzazione dei prodotti di degradazione ossidativa dei secoiridoidi nell'olio vergine di oliva mediante tecniche analitiche innovative

Lo scopo di questo lavoro di tesi è la caratterizzazione dei prodotti di ossidazione di diversi fenoli idrofili contenuti nell’olio vergine d’oliva come idrossitirosolo, tirosolo e la forma dialdeidica dell’acido decarbossimetil elenolico legato all’idrossitirosolo, e la loro identificazione nel prodotto durante la conservazione. L’obiettivo della ricerca è trovare degli indici analitici che possono essere usati sia come marker di “freschezza” dell’olio vergine di oliva sia nella valutazione della “shelf life” del prodotto stesso. Due sistemi di ossidazione sono stati usati per ossidare le molecole sopracitate: ossidazione enzimatica e ossidazione di Fenton. I prodotti di ossidazione sono stati identificati come chinoni, dimeri e acidi.

Valorization of pomaces from the mechanical extraction of virgin olive oils in dairy animal feeding

This research work is aimed at the valorization of two types of pomace deriving from the extra virgin olive oil mechanical extraction process, such as olive pomace and a new by-product named “paté”, in the livestock sector as important sources of antioxidants and unsaturated fatty acids. In the first research the suitability of dried stoned olive pomace as a dietary supplement for dairy buffaloes was evaluated. The effectiveness of this utilization in modifying fatty acid composition and improving the oxidative stability of buffalo milk and mozzarella cheese have been proven by means of the analysis of qualitative and quantitative parameters. In the second research the use of paté as a new by-product in dietary feed supplementation for dairy ewes, already fed with a source of unsaturated fatty acids such as extruded linseed, was studied in order to assess the effect of this combination on the dairy products obtained. The characterization of paté as a new by-product was also carried out, studying the optimal conditions of its stabilization and preservation at the same time. The main results, common to both researches, have been the detection and the characterization of hydrophilic phenols in the milk. The analytical detection of hydroxytyrosol and tyrosol in the ewes’ milk fed with the paté and hydroxytyrosol in buffalo fed with pomace showed for the first time the presence in the milk of hydroxytyrosol, which is one of the most important bioactive compounds of the oil industry products; the transfer of these antioxidants and the proven improvement of the quality of milk fat could positively interact in the prevention of some human cardiovascular diseases and some tumours, increasing in this manner the quality of dairy products, also improving their shelf-life. These results also provide important information on the bioavailability of these phenolic compounds.

Molecular basis of Osteoarthritis and aspects of cellular senescence in disease

The aim of this study is to investigate on some molecular mechanisms contributing to the pathogenesis of osteoarthritis (OA) and in particular to the senescence of articular chondrocytes. It is focused on understanding molecular events downstream GSK3β inactivation or dependent on the activity of IKKα, a kinase that does not belong to the phenotype of healthy articular chondrocytes. Moreover, the potential of some nutraceuticals on scavenging ROS thus reducing oxidative stress, DNA damage, and chondrocyte senescence has been evaluated in vitro. The in vitro LiCl-mediated GSK3β inactivation resulted in increased mitochondrial ROS production, that impacted on cellular proliferation, with S-phase transient arrest, increased SA-β gal and PAS staining, cell size and granularity. ROS are also responsible for the of increased expression of two major oxidative lesions, i.e. 1) double strand breaks, tagged by γH2AX, that associates with activation of GADD45β and p21, and 2) 8-oxo-dG adducts, that associate with increased IKKα and MMP-10 expression. The pattern observed in vitro was confirmed on cartilage from OA patients. IKKa dramatically affects the intensity of the DNA damage response induced by oxidative stress (H2O2 exposure) in chondrocytes, as evidenced by silencing strategies. At early time point an higher percentage of γH2AX positive cells and more foci in IKKa-KD cells are observed, but IKKa KD cells proved to almost completely recover after 24 hours respect to their controls. Telomere attrition is also reduced in IKKaKD. Finally MSH6 and MLH1 genes are up-regulated in IKKαKD cells but not in control cells. Hydroxytyrosol and Spermidine have a great ROS scavenging capacity in vitro. Both treatments revert the H2O2 dependent increase of cell death and γH2AX-foci formation and senescence, suggesting the ability of increasing cell homeostasis. These data indicate that nutraceuticals represent a great challenge in OA management, for both therapeutical and preventive purposes.

Studio ed ottimizzazione di una via di sintesi di idrossitirosolo a ridotto impatto ambientale

In questo progetto di tesi abbiamo lavorato all’ottimizzazione di un nuovo processo di sintesi di idrossitirosolo eco-compatibile e di possibile applicazione industriale. Il processo, che risulta una via molto competitiva per ottenere idrossitirosolo, si compone di due step: una prima sintesi di un intermedio acetalico e la successiva riduzione di questo al prodotto di interesse. In particolare il lavoro di tesi ha riguardato lo studio di catalizzatori e delle condizioni per le due reazioni al fine di aumentare le selettività dei prodotti target, analizzando la formazione dei sottoprodotti. Si è cercato, inoltre, di intraprendere una strada alternativa, provando la sintesi di un intermedio acetalico ciclico (possibilmente più stabile dell’altro intermedio acetalico) e, quindi, la sua riduzione, ritenendo di poter così evitare la formazione dei sottoprodotti. In this project we worked on the optimization of a new and industrially applicable process for the synthesis of hydroxytyrosol. Furthermore, we devised a process according to the principles of green chemistry. The process, that is a competitive way to obtain hydroxytyrosol, consists of two steps: first the synthesis of an acetalic precursor and then its reduction. Particularly the work was focused on the study of the catalysts and the conditions for the two reactions in order to increase the selectivity of target products, also analyzing the formation of the by-products. At last, we tried to take an alternative route with the synthesis of another acetalic precursor: a cyclic one. This because we thought that a cyclic acetal should be more stable than not cyclic one and maybe the reduction on this second precursor could not pass by the formation of the by-products already saw.

In vivo and in vitro effects of antioxidant compounds and PPAR γ modulators on rainbow trout oxidative stress and lipid metabolism.

The aquafeed use of raw plant materials, as protein and lipid sources, has been considered and approved as a sustainable alternative to fish products (fish meal and oils) because the current trend to use high-lipid diets has been shown to induce undesirable increase in fat depots or further physiological alterations, such as induction of oxidative stress. In the aquaculture perspective, the addition of natural substances with antioxidant properties is an emerging strategy for protecting biological systems and foodstuffs from oxidative damage. Among natural substances, hydroxytyrosol (HT) and caffeic acid (CA) have attracted considerable attention as food antioxidant additives and modulators of physiological and molecular pathways involved in energy metabolism and adiposity. The aim of this study was to evaluate the effects of CA and HT on lipid metabolism and oxidative stress of rainbow trout (Oncorhynchus mykiss). In vitro results showed the potential anti-obesogenic effects of the compounds CA and HT on the adipose tissue of the rainbow trout. To support these data, in vitro assays performed (MTT, ORO, immunofluorescence) resulted in accordance among them; only results from proliferating cell nuclear antigen (PCNA) assay were not significant. In vivo results showed a possible anti-obesogenic effect of CA in liver and HT in adipose tissue. Regarding oxidative stress, we could hypothesize a possible anti-oxidant role of CA in liver.

Radiation metabolomics. 5. Identification of urinary biomarkers of ionizing radiation exposure in nonhuman primates by mass spectrometry-based metabolomics

Mass spectrometry-based metabolomics has previously demonstrated utility for identifying biomarkers of ionizing radiation exposure in cellular, mouse and rat in vivo radiation models. To provide a valuable link from small laboratory rodents to humans, γ-radiation-induced urinary biomarkers were investigated using a nonhuman primate total-body-irradiation model. Mass spectrometry-based metabolomics approaches were applied to determine whether biomarkers could be identified, as well as the previously discovered rodent biomarkers of γ radiation. Ultra-performance liquid chromatography-electrospray ionization quadrupole time-of-flight mass spectrometry analysis was carried out on a time course of clean-catch urine samples collected from nonhuman primates (n = 6 per cohort) exposed to sham, 1.0, 3.5, 6.5 or 8.5 Gy doses of (60)Co γ ray (∼0.55 Gy/min) ionizing radiation. By multivariate data analysis, 13 biomarkers of radiation were discovered: N-acetyltaurine, isethionic acid, taurine, xanthine, hypoxanthine, uric acid, creatine, creatinine, tyrosol sulfate, 3-hydroxytyrosol sulfate, tyramine sulfate, N-acetylserotonin sulfate, and adipic acid. N-Acetyltaurine, isethionic acid, and taurine had previously been identified in rats, and taurine and xanthine in mice after ionizing radiation exposure. Mass spectrometry-based metabolomics has thus successfully revealed and verified urinary biomarkers of ionizing radiation exposure in the nonhuman primate for the first time, which indicates possible mechanisms for ionizing radiation injury.