Antioxidant effect of phenolic compounds, alpha-tocopherol, and other minor components in virgin olive oil

The effect of acidity, squalene, hydroxytyrosol, aldehydic form of oleuropein aglycon, hydroxytyrosyl acetate, tyrosol, homovanillic acid, luteolin, apigenin, alpha-tocopherol, and the mixtures hydroxytyrosol/hydroxytyrosyl acetate, hydroxytyrosol/tyrosol, and hydroxytyrosol/alpha-tocopherol on the oxidative stability of an olive oil matrix was evaluated. A purified olive oil was spiked with several concentrations of these compounds and, then, subjected to an accelerated oxidation in a Rancimat apparatus at 100 degrees C. Acidity, squalene, homovanillic acid, and apigenin showed negligible effect. At the same millimolar concentrations, the different o-diphenolic compounds yielded similar and significant increases of the induction time, alpha-tocopherol a lesser increase, and tyrosol a scarce one. At low concentrations of o-diphenols and alpha-tocopherol, a linear relationship between induction time and concentration was found, but at high concentrations the induction time tended toward constant values. To explain this behavior, a kinetic model was applied. The effect of the mixtures hydroxytyrosol/hydroxytyrosyl acetate was similar to that of a single o-diphenol at millimolar concentration equal to the sum of millimolar concentrations of both compounds. Concentrations of tyrosol >0.3 mmol/kg increase the induction time by 3 h. The mixtures hydroxytyrosol/alpha-tocopherol showed opposite effects depending on the relative concentrations of both antioxidants; so, at hydroxytyrosol concentrations <0.2 mmol/kg, the addition of alpha-tocopherol increased the induction time, whereas at higher hydroxytyrosol concentrations, the alpha-tocopherol diminished the stability.     

Evolution of minor polar compounds and antioxidant capacity during storage of bottled extra virgin olive oil

We characterized "Olivastra Seggianese" extra virgin olive oil (EVOO) and evaluated its chemical and sensory characteristics and antioxidant and antiradical activities during storage under novel conditions. Two oils (A and B) were analyzed for the commodity characteristics at blending (t0) and after 9, 12, and 18 months; panel tests were performed and minor polar compounds (MPC) content was assessed at blending (t0) and after 6, 9, 12, and 18 months. Antioxidant and antiradical activities in vitro were evaluated at t0 and after 12 months, by human low density lipoprotein (LDL) and 1,1-diphenyl-2-picrylhydrazil radical (DPPH*) tests. Oil A, which had an initially higher MPC content, possessed "harder" organoleptic characteristics than oil B, which had a lower MPC content and was endowed with a "smoother" taste profile. Statistical analyses showed that secoiridoids, particularly deacetoxy-oleuropein aglycone, should be quantified to evaluate EVOO stability during storage. The antioxidant activity toward human LDL was linked to MPC content and to storage time. The tests on the stable free radical DPPH* confirmed the results on human LDL. We propose this as an additional parameter to evaluate olive oil quality and stability over time.     

Monovarietal extra virgin olive oils: correlation between thermal properties and chemical composition

Thermal properties of monovarietal extra virgin olive oils were evaluated by means of differential scanning calorimetry (upon cooling) and related to their chemical composition (triacylglycerols, diacylglycerols, total and free fatty acids, oxidation status). The overall crystallization enthalpy did not significantly differ among samples and did not account for the differences observed in chemical compositions. On the contrary, a higher degree of unsaturation in the lipid profile induced a shift of the crystallization onset towards lower temperatures and narrowing of the crystallization temperature range. The presence of triacylglycerol lysis and lipid oxidation products shifted the crystallization towards higher temperatures and the phase transition developed over a larger temperature range. Differential scanning calorimetry thermograms were deconvoluted into three constituent exothermic peaks for all samples. The area of the two lower-temperature exotherms was found to be statistically correlated with the amount of triunsaturated and monosaturated triacylglycerols present in the oil. Thermal properties of extra virgin olive oil were found to be affected by oil chemical composition.     

alpha-tocopherol content of Greek virgin olive oils

alpha-Tocopherol, the main tocopherol homologue found in olive oil, was determined using normal phase HPLC. Ninety Greek virgin olive oils, selected according to a designed sampling protocol from different cultivars and regions all over Greece for three successive crop years, were analyzed. For a specific olive cultivar, which is widely used for the production of olive oil in Greece, additional measurements were made to study the effect of milling conditions on alpha-tocopherol concentration. Finally, a significant number of commercial olive oil samples (25) obtained from the retail market were analyzed. High concentrations of alpha-tocopherol were observed in most of the samples selected from various regions. Values ranging between 98 and 370 mg/kg were found (>200 mg/kg in 60% of samples). Extraction conditions were not found to influence alpha-tocopherol level. alpha-Tocopherol content of retail market samples was high, ranging from 120 to 250 mg/kg of oil (>180 mg/kg in 60% of samples). Storage of samples under domestic conditions for two years showed that good handling is quite important for retaining high alpha-tocopherol levels and for increasing, thus, the storage life and nutritional value of this exquisite oil.     

Comparison of the amounts of volatile compounds in French protected designation of origin virgin olive oils

Headspace solid-phase microextraction (HS-SPME) -gas chromatography using flame ionization detection and multivariate analysis were applied to the study of the specificity of protected designation of origin (PDO) virgin olive oils produced in a southern French region (Alpes-Maritimes) based on their volatile compounds. A total of 35 PDO olive oils from Nice, 6 commercial oils, and 12 other French PDO olive oils were analyzed. Recorded data were subjected to principal component analysis (PCA) and soft independent modeling of class analogy (SIMCA). The method developed here was able to perfectly distinguish different qualities of olive oils. Representative samples from each class obtained by chemometric treatment were analyzed by HS-SPME and GC-MS. PCA and SIMCA of chromatographic data were related to sensory analysis and led to a better understanding of the chemical features and observed sensory effects of olive oils.     

Effect of olive stoning on the volatile and phenolic composition of virgin olive oil

Olive stoning during the virgin olive oil (VOO) mechanical extraction process was studied to show the effect on the phenolic and volatile composition of the oil. To study the impact of the constitutive parts of the fruit in the composition of olive pastes during processing, the phenolic compounds and several enzymatic activities such as polyphenoloxidase (PPO), peroxidase (POD), and lipoxygenase (LPO) of the olive pulp, stone, and seed were also studied. The olive pulp showed large amounts of oleuropein, demethyloleuropein, and lignans, while the contribution of the stone and the seed in the overall phenolic composition of the fruit was very low. The occurrence of crushed stone in the pastes, during malaxation, increased the peroxidase activity in the pastes, reducing the phenolic concentration in VOO and, at the same time, modifying the composition of volatile compounds produced by the lipoxygenase pathway. The oil obtained from stoned olive pastes contained higher amounts of secoiridoid derivatives such as the dialdehydic forms of elenolic acid linked to (3,4-dihydroxyphenyl)ethanol and (p-hydroxyphenyl)ethanol (3,4-DHPEA-EDA and p-HPEA-EDA, respectively) and the isomer of the oleuropein aglycon (3,4-DHPEA-EA) and, at the same time, did not show significant variations of lignans. The stoning process modified the volatile profile of VOO by increasing the C6 unsaturated aldehydes that are strictly related to the cut-grass sensory notes of the oil.     

Comparison of the antioxidant activities of extra virgin olive oils

The phenol content and antioxidant activity of extra virgin olive oils (EVOOs) differing in their origins and degradation degrees were studied. The o-diphenolic compounds typical of olive oil, namely, the oleuropein derivatives hydroxytyrosol (3',4'-dihydroxyphenylethanol, 3',4'-DHPEA), the dialdehydic form of elenolic acid linked to 3',4'-DHPEA (3',4'-DHPEA-EDA), and an isomer of oleuropein aglycon (3',4'-DHPEA-EA), were analyzed by HPLC. The antioxidant activity was studied by (a) the xanthine oxidase (XOD)/xanthine system, which generates superoxide radical and hydrogen peroxide; (b) the diaphorase (DIA)/NADH/juglone system, which generates superoxide radical and semiquinonic radical; and (c) the 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) test. Results showed that EVOOs with a low degradation level (as evaluated by acidity, peroxide number, and spectroscopic indices K(232), K(270), and deltaK according to the EU Regulation) had a higher content of 3',4'-DHPEA-EDA and a lower content of 3',4'-DHPEA than oils having intermediate and advanced degradation levels. EVOOs with a low degradation degree were 3-5 times more efficient as DPPH scavengers and 2 times more efficient as inhibitors of the XOD-catalyzed reaction than oils with intermediate and advanced degradation levels. The DIA-catalyzed reaction was inhibited by EVOOs having low or intermediate degradation levels but not by the most degraded oils.     

Solid-phase microextraction in the analysis of virgin olive oil volatile fraction: characterization of virgin olive oils from two distinct geographical areas of northern Italy

SPME was employed to characterize the volatile profile of virgin olive oils produced in two geographical areas of northern Italy: the region of the Gulf of Trieste and the area near Lake Garda. There are as yet no data on the headspace composition of virgin olive oils from these regions, characterized by particular conditions of growth for Olea europaea. Using the SPME technique coupled to GC-MS and GC-FID, the volatile components of 42 industrially produced virgin olive oil samples were identified and the principal compounds quantitatively analyzed. Significant differences in the proportion of volatile constituents from oils of different varieties and geographical origins were detected. The results suggest that besides the genetic factor, environmental conditions influence the volatile formation.     

Effects of olive fruit quality and oil storage practices on the diacylglycerol content of virgin olive oils

The evolution of 1,3- and 1,2-isomers of diacylglycerols (DGs) in olive oils obtained from healthy olives and the influence of the olive quality was studied. Healthy olive fruits yielded oils containing almost exclusively 1,2-isomers whereas altered olives produced oils with significant amounts of 1,3-isomers. Virgin olive oils obtained from various olive cultivars and stored at different temperatures showed triacylglycerol hydrolysis and diacylglycerol isomerization depending on the acidity and temperature. The results indicated that the relationship between acidity and total diacylglycerol content has scarce utility for detecting mild refined oil in virgin olive oil. On the other hand, the 1,3-/1,2-DG isomers ratio is useful for assessing the genuineness of virgin olive oils with low acidities during the early stages of storage.     

Irrigation effects on quality, phenolic composition, and selected volatiles of virgin olive oils cv. Leccino

Field-grown olive trees (Olea europaea L. cv. Leccino) were used over two growing seasons to determine the effect of deficit irrigation regimes on virgin olive oil (VOO) quality. Drip irrigation was managed to maintain a predawn leaf water potential (PLWP): (a) higher than -1.1 MPa (full irrigation: FI); (b) between -1.0 and -3.3 MPa (deficit irrigation: DI); (c) higher than -4.2 MPa (severe deficit irrigation: SI). The fruit yield and oil yield of DI trees were over 90% of those of FI treatments in both years, respectively, whereas yields of SI trees ranged from 61 to 76%. The irrigation regime had minor effects on the free acidity, peroxide value, and fatty acid composition of VOO. The concentrations of phenols and o-diphenols in VOO were negatively correlated with PLWP. The concentrations of the dialdehydic form of decarboxymethyl elenolic acid linked to (3,4-dihydroxyphenyl)ethanol (3,4-DHPEA-EDA), the isomer of the oleuropein aglycon (3,4-DHPEA-EA), and the dialdehydic form of decarboxymethyl elenolic acid linked to (p-hydroxyphenyl)ethanol (p-HPEA-EDA) were lower in FI than in SI treatments. The concentrations of lignans (+)-1-acetoxipinoresinol and (+)-1-pinoresinol were unaffected by the irrigation regime. The tree water status had a marked effect on the concentration of volatile compounds, such as the C(6)-saturated and unsaturated aldehydes, alcohols, and esters.     

Monovarietal extra virgin olive oils. Correlation between thermal properties and chemical composition: heating thermograms

Extra virgin olive oils from drupes of three Sicilian varieties (Biancolilla, Cerasuola, and Nocellara del Belice) collected at three different harvesting periods were analyzed upon heating by means of DSC, and thermal properties were related to the chemical composition of the samples. All thermograms exhibited multiple transitions with a minor exothermic peak, followed by a major endothermic event. Cerasuola samples showed higher overall enthalpy and narrower range of transition at all harvesting periods, as compared to the other oils. A more ordered crystal structure originating from a more uniform chemical composition, with higher triolein content, in Cerasuola may be hypothesized. At different harvesting periods, thermal transitions started at lower temperatures and developed over a narrower range in all cultivars, probably due to the insertion of molecules derived from triacylglycerol lysis (diacylglycerols and free fatty acids) and lipid oxidation products into the triacylglycerol crystal lattice. All heating thermograms were deconvoluted into one exothermic and five endothermic constituent peaks, and the effect of chemical components on thermal properties of the peaks was evaluated. DSC application upon heating appears to be very promising in discriminating among oil samples from olives of different cultivars and/or harvesting periods.     

Influence of the decrease in oxygen during malaxation of olive paste on the composition of volatiles and phenolic compounds in virgin olive oil

The sensory and health properties of virgin olive oil (VOO) are highly related to its volatile and phenolic composition. Oxygen control in the pastes during malaxation may be a new technological parameter to regulate enzymatic activities, such as polyphenoloxidase, peroxidase, and lipoxygenase, which affect the phenolic and volatile composition of VOO. In this work, we monitored CO2 and O2 concentrations during industrial-scale olive paste malaxation with various initial O2 concentrations within the malaxer headspace. Results show that the O2 concentration in the malaxer headspace did not affect CO2 production during processing, whereas a strong influence was observed on the changes of the phenolic composition of olive pastes and VOOs, with high correlation coefficient for the total phenols (R = 0.94), especially for oleuropein and demethyloleuropein derivatives (R = 0.81). In contrast, aroma production during malaxation was minimally affected by the O2 concentration in the malaxer headspace.     

Novel approach to study oxidative stability of extra virgin olive oils: importance of alpha-tocopherol concentration

The stability of extra virgin olive oils is mainly due to their relatively low fatty acids unsaturation and to the antioxidant activity of some of the unsaponifiable components. We studied the activity of alpha-tocopherol in extra virgin olive oil in its natural state, using new and simple oxidizing conditions in "thin layer" and "bulk phase". Oxidation time course was monitored at 37 degrees C or 75 degrees C. A storage test was also performed. Two parameters were evaluated: depletion of alpha-tocopherol and formation of PUFA hydroperoxides, measured as conjugated dienes (CD) and peroxide value. The value of complex polyphenols was also measured. alpha-Tocopherol concentration decreased in function of time and temperature and showed a strong inverse correlation with the increase of CD. When alpha-tocopherol reached a "threshold value" of 60-70 mg/kg, a significant increase of CD formation was observed, together with a good correlation between CD and peroxide value. The initial amount of alpha-tocopherol is one of the components that influences oil oxidative susceptibility.     

Influence of harvest date and crop yield on the fatty acid composition of virgin olive oils from cv. Picual

In this study was analyzed the effect of crop year and harvesting time on the fatty acid composition of cv. Picual virgin olive oil. The study was carried out during the fruit ripening period for three crop seasons. The mean fatty acid composition of Picual oils was determined. The oils contained palmitic acid (11.9%), oleic acid (79.3%), and linoleic acid (2.95%). The content of palmitic acid and saturated fatty acids decreased during fruit ripening while oleic and linoleic acids increased. The amount of stearic and linolenic acids decreased. The amount of saturated acids, palmitic and stearic, and the polyunsaturated acids linoleic and linolenic was dependent on the time of harvest, whereas the amount of oleic acid varied with the crop year. The differences observed between crop years for both palmitic and linoleic acid may be explained by the differences in the temperature during oil biosynthesis and by the amount of summer rainfall for oleic acid content. A significant relationship was observed between the MUFA/PUFA ratio and the oxidative stability measured by the Rancimat method.     

Phenolic compounds profile of cornicabra virgin olive oil

This study presents the phenolic compounds profile of commercial Cornicabra virgin olive oils from five successive crop seasons (1995/1996 to 1999/2000; n = 97), determined by solid phase extraction reversed phase high-performance liquid chromatography (SPE RP-HPLC), and its relationship with oxidative stability, processing conditions, and a preliminary study on variety classification. The median of total phenols content was 38 ppm (as syringic acid), although a wide range was observed, from 11 to 76 ppm. The main phenols found were the dialdehydic form of elenolic acid linked to tyrosol (p-HPEA-EDA; 9 +/- 7 ppm, as median and interquartile range), oleuropein aglycon (8 +/- 6 ppm), and the dialdehydic form of elenolic acid linked to hydroxytyrosol (3,4-DHPEA-EDA; 5 +/- 8 ppm). In many cases the correlation with oxidative stability was higher when the sum of the dialdehydic form of elenolic acid linked to hydroxytyrosol (3,4-DHPEA-EDA) and oleuropein aglycon (r (2) = 0.91-0.96) or the sum of these two and hydroxytyrosol (r (2) = 0.90-0.97) was considered than was observed with HPLC total phenols (r (2)= 0.91-0.95) and especially with colorimetric determination of total polyphenols and o-diphenols (r (2) = 0.77-0.95 and 0.78-0.92, respectively). 3,4-DHPEA-EDA, p-HPEA-EDA, the aglycons of oleuropein and ligstroside, and HPLC total phenols content presented highly significant differences (p = 0.001-0.010) with respect to the dual- and triple-phase extraction systems used, whereas colorimetric total polyphenols content did not (p = 0.348) and o-diphenols showed a much lower significant difference (p = 0.031). The five variables that most satisfactorily classified the principal commercial Spanish virgin olive oil varieties were 1-acetoxypinoresinol, 4-(acetoxyethyl)-1,2-dihydroxybenzene (3,4-DHPEA-AC), ligstroside aglycon, p-HPEA-EDA, and RT 43.3 contents.     

Protective effect of the phenolic fraction from virgin olive oils against oxidative stress in human cells

This paper reports the protective effect of the phenolic fraction extracted from extra virgin olive oils (OOPEs) against the cytotoxic effects of reactive oxygen species in human erythrocytes and Caco-2 cells, employed as model systems. Pretreatment of cells with various OOPEs, indeed, provides a remarkable protection against oxidative damages: this effect was strictly dependent on the o-diphenolic content of the extracts. Moreover, the protective effects observable in cellular systems were compared with in vitro antioxidant properties, measured by using the FRAP (ferric reducing/antioxidant power) assay; the reducing ability of OOPEs strictly parallels their o-phenolic content. The linear relationship demonstrated between biological effects and antioxidant capacity measured by the FRAP assay allows us to propose the use of this rapid colorimetric method in assessing and certifying the antioxidant power of extra virgin olive oil.     

Influence of olive maturity stage and geographical origin on some minor components in virgin olive oil of the chemlali variety

Minor compounds such as sterols, aliphatic alcohols, tocopherols, and chlorophylls in virgin olive oil from Chemlali cultivar were analyzed during the maturity process. The study concerns oils from the following three different olive growing areas of Tunisia: Sfax, Sidi Bouzid, and Enfidha. Analytical results showed that both the maturity process and the olive provenances influence the evolution of the content of these compounds.     

Qualitative and semiquantitative analysis of phenolic compounds in extra virgin olive oils as a function of the ripening degree of olive fruits by different analytical techniques

Capillary electrophoresis (CE) can be effectively used as a fast screening tool to obtain qualitative and semiquantitative information about simple and complex phenolic compounds of extra virgin olive oil. Three simple phenols (tyrosol, hydroxytyrosol, and vanillic acid), a secoiridoid derivative (deacetoxy oleuropein aglycon), and two lignans (pinoresinol and acetoxypinoresinol) were detected as the main compounds in extra virgin olive oils by high-performance liquid chromatography (HPLC) and capillary zone electrophoresis (CZE). Spectrophotometric indices, radical scavenging activity, and oxidative stability of extra virgin olive oil samples obtained from olives hand-picked at different ripening degrees were statistically correlated with the CZE and HPLC quantification. The concentration of phenols in extra virgin olive oil decreased with ripeness of olive fruits. The high correlations found between CZE and the other analytical results indicate that CE can be applied as a rapid and reliable tool to routinely determine phenolic compounds in extra virgin olive oils.     

Rapid quantitative assessment of the adulteration of virgin olive oils with hazelnut oils using Raman spectroscopy and chemometrics

The authentication of extra virgin olive oil and its adulteration with lower-priced oils are serious problems in the olive oil industry. In addition to the obvious effect on producer profits, adulteration can also cause severe health and safety problems. A number of techniques, including chromatographic and spectroscopic methods, have recently been employed to assess the purity of olive oils. In this study Raman spectroscopy together with multivariate and evolutionary computational-based methods have been employed to assess the ability of Raman spectroscopy to discriminate between chemically very closely related oils. Additionally, the levels of hazelnut oils used to adulterate extra virgin olive oil were successfully quantified using partial least squares and genetic programming.     

Determination of the diglyceride content in greek virgin olive oils and some commercial olive oils by employing (31)P NMR spectroscopy

In this study, the diglyceride contents of 96 samples of virgin olive oils from the regions of Crete, Lesvos, Messinia, Pilion, Zakynthos, Halkidiki, and Ilia, 15 samples of commercial extra virgin and pure olive oils, and 3 samples each of refined olive oils and pomace oils were determined by a facile method introduced in a previous publication. This method is based on the phosphitylation of the free hydroxyls of the diglycerides with 2-chloro-4,4,5,5-tetramethyldioxaphospholane and the integration of the appropriate peaks in the (31)P NMR spectra. This preliminary study showed interesting trends in the diglyceride content of the virgin olive oils from the various regions of Greece that can be used as simple criteria to assess the olive oil characteristics. Analysis of variance has been carried out for the diglyceride content of each region in an attempt to detect possible differences in the diglyceride levels among the various regions. Finally, the relationship between the ratio of 1,2-diglycerides to the total amount of diglycerides and the total amount of diglycerides has been used to monitor the quality of virgin olive oils, commercial olive oils, refined olive oils, and pomace oils.