Cluster analysis applied to the exploratory analysis of commercial spanish olive oils by means of excitation-emission fluorescence spectroscopy

Olive oil fluorescence is related to oil composition. Here it is shown that the natural clustering of different types of commercial Spanish olive oils depends on their fluorescence excitation-emission matrices (EEMs). Fifty-six commercial samples of olive oil (29 virgin olive oils, 20 pure olive oils, and 7 olive-pomace oils) were used. The clustering method was hierarchical agglomerative clustering using the Euclidean distance as a similarity measure and the average linkage. Two spectral ranges were considered (which either contained the fluorescence peak of the chlorophylls or did not), and various methods for preprocessing the fluorescence spectra were compared. The oils were clearly distinguished using the unfolded EEMs measured between lambda(ex) = 300-400 nm and lambda(em) = 400-600 nm. The optimal preprocessing was normalization of the unfolded spectra followed by column autoscaling. Also shown are the advantages of using second-order data (EEMs) instead of first-order data (a single fluorescence spectrum) for each sample.     

Origin of French virgin olive oil registered designation of origins predicted by chemometric analysis of synchronous excitation-emission fluorescence spectra

The authentication of virgin olive oil samples requires usually the use of sophisticated and very expensive analytical techniques, so there is a need for fast and inexpensive analytical techniques for use in a quality control methodology. Virgin olive oils present an intense fluorescence spectra. Synchronous excitation-emission fluorescence spectroscopy (SEEFS) was assessed for origin determination of virgin olive oil samples from five French registered designation of origins (RDOs) (Nyons, Vallée des Baux, Aix-en-Provence, Haute-Provence, and Nice). The spectra present bands between 600 and 700 nm in emission due to chlorophylls a and b and pheophytins a and b. The bands between 275 and 400 nm in emission were attributed to alpha-, beta-, and gamma-tocopherols and to phenolic compounds, which characterize the virgin olive oils compared to other edible oils. The chemometric treatment (PLS1) of synchronous excitation-emission fluorescence spectra allows one to determine the origin of the oils from five French RDOs (Baux, Aix, Haute-Provence, Nice, and Nyons). Results were quite satisfactory, despite the similarity between two denominations of origin (Baux and Aix) that are composed by some common cultivars (Aglandau and Salonenque). The interpretation of the regression coefficients shows that RDOs are correlated to chlorophylls, pheophytins, tocopherols, and phenols compounds, which are different for each origin. SEEFS is part of a global analytic methodology that associates spectroscopic and chromatographic techniques. This approach can be used for traceability and vindicates the RDOs.     

Simultaneous fluorometric determination of chlorophylls a and B and pheophytins a and B in olive oil by partial least-squares calibration

The resolution of quaternary mixtures of chlorophylls a and b and pheophytins a and b has been accomplished by partial least-squares (PLS) multivariate calibration, applied to the fluorescence signals of these pigments. The total luminescence information of the compounds has been used to optimize the spectral data set to perform the calibration. After preliminary studies, a method is described in acetone media, to avoid emulsions with the olive oil samples. Different scanning paths have been selected for each method. For the simultaneous determination of the pigments in olive oil samples, a comparative study of the results found by using excitation, emission, and synchronous spectral data, as analytical signal, was performed. The excitation spectra were selected as the better analytical signals for the determination of the pigments in olive oil samples. The optimum wavelength range to record the excitation spectra (lambda(em) = 662 nm) was selected to minimize the contribution of pheophytin a and to maximize the contribution of the other pigments, which are the minor constituents in olive oil. Determination of these pigments in olive oil samples was effected from the excitation spectra of dissolutions o suitable aliquots in acetone. Recovery values from olive oil, spiked with chlorophylls a and b and pheophytins a and b, were in the ranges of 70-112, 71-111, 76-105, and 82-109%, respectively.     

Detection of the presence of refined hazelnut oil in refined olive oil by fluorescence spectroscopy

The fluorescence spectroscopy technique has been tested as regards its ability to differentiate between refined hazelnut and olive oils. Classification of these oils based on their excitation-emission fluorescence spectra data (spectral range 300-500 nm of the excitation spectra at lambdaem=655 and spectral range 650-900 of the emission spectra at lambdaex=50 nm) was performed using principal component analysis and artificial neural networks. Both methods provided good discrimination between the refined hazelnut and olive oils. The results have also pointed out the possibilities of a spectrofluorimetric method joined to multivariate analysis, to differentiate refined oils, and even to detect the presence of refined hazelnut oils in refined olive oils at percentages higher than 9%.     

Phenolic compounds in olive oils intended for refining: formation of 4-ethylphenol during olive paste storage

The phenolic composition of "lampante olive oil", "crude olive pomace oil", and "second centrifugation olive oil" was characterized by high-performance liquid chromatography with UV, fluorescence, and mass spectrometry detection. The phenolic profile of these olive oils intended for refining was rather similar to that previously reported for virgin olive oil. However, a new compound was found in these oils, which is mainly responsible of their foul odor. It was identified as 4-ethylphenol by comparison of its UV and mass spectra with those of a commercial standard. Although 4-ethylphenol was discovered in all oils intended for refining, its presence was particularly significant in "second centrifugation olive oils", its concentration increasing with time of olive paste storage. Similar trends were observed for hydroxytyrosol, hydroxytyrosol acetate, tyrosol, and catechol, the concentration of these substances reaching values of up to 600 mg/kg of oil, which makes their recovery for food, cosmetic, or pharmaceutical purposes attractive.     

Fluorescence of cereal flours.

Front-surface fluorescence spectra of cereal flours are easily measured with a commercial spectrofluorometer and a simple cell. The emission spectra are intense in the range 290-600 nm in which three emission bands are active. The most intense is due to the aromatic amino acid residues present in the proteins of flours. At least two other fluorophores are active in the above spectral range and compete for light with the chromophores present in the flours. Four absorption bands are revealed in the same spectral region by reflectance spectra, the most intense being due to the amino acidic fluorophores. Thus, the measured emission is the result of absorption, scattering, emission, and reabsorption processes. Information on the microenvironment of some fluorophores can be obtained. The possibility to recognize different species of cereals, even of different cultivars, is shown.     

Interpretation of Fourier transform Raman spectra of the unsaponifiable matter in a selection of edible oils.

The unsaponifiable matter of edible oils is a source of information for their characterization and authentication. FT-Raman spectroscopy has been applied with success to the determination of the spectra of the unsaponifiable matter of varietal olive oils as well as other refined and crude edible oils. The spectra of the major unsaponifiable series of compounds (squalene, sterolic, and terpenic alcoholic fractions), together with beta-carotene and lutein, have been used to explain the most prominent bands found in the spectra of the unsaponifiable matter of 15 edible oil samples. The order of the scattering intensities of the varietal olive oils agrees with the results obtained by chromatography. An unsupervised multivariate statistical analysis of selected bands points out differences between olive oils and the other seed oils and also among varietal virgin olive oils.     

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.     

Nondestructive evaluation of anthocyanins in olive (Olea europaea) fruits by in situ chlorophyll fluorescence spectroscopy

Anthocyanins (Anths) in olive (Olea europaea L.) fruits at different degrees of pigmentation were assessed nondestructively by measuring chlorophyll fluorescence (ChlF). The method is based on the comparison of the ChlF excitation spectra from olives with different pigmentation from green to green-red, reddish-purple, and purple. The logarithm of the ratio between the fluorescence excitation spectra (logFER) from two different colored zones gave the difference in the absorption spectrum between them. The absorbance spectrum derived from the logFER between a red olive and the same olive devoid of the skin showed the typical Anth green band (at 550 nm). It matched that recorded by microspectrophotometry on a single pulp cell and the in vitro absorbance spectrum of the olive skin extract. As expected, the in vivo Anths absorption maximum increased in intensity going from less to more mature olives and was higher in the sun-exposed olive side with respect to the sun-shaded side. Absolute quantitative nondestructive determination of Anths for each olive sample was obtained by the logFER calculated for two excitation wavelengths, 550 and 625 nm, of ChlF at 740 nm. Going from green to purple skin colors, the Log[ChlF(625)/ChlF(550)] was fairly well-correlated to the extract Anths concentration. Finally, the relationship between the Anths and the other main phenolics present in the olives analyzed by high-performance liquid chromatography was evaluated. The main result was a net increase of verbascoside with increasing Anths content. On the basis of our results, the development of a new rapid and noninvasive method for the monitoring of olive development and ripening can be envisaged.     

Spectrophotometric and adsorptive stripping square wave voltammetric determination of iron in olive oils,as complex with 5,5-dimethylcyclohexane-1,2,3-trione 1,2-dioxime 3-thiosemicarbazone (DCDT)

Two methods for the determination of iron in olive oil by spectrophotometry and by adsorptive stripping square wave voltammetry (Ad-SSWV) have been developed. These two methods are based on the formation of a 5,5-dimethylcyclohexane-1,2,3-trione 1,2-dioxime 3-thiosemicarbazone (DCDT)-iron(II) complex in strongly acid media. In both, iron is extracted from the olive oil by using HCl. Spectrophotometric determination of iron with DCDT is based on the feature that the DCDT-Fe complex shows an absorbance maximum at 550 nm. A calibration graph has been constructed from 0 to 4000 ng mL(-)(1), and the detection limit was 115 ng mL(-)(1) (57 ng g(-)(1) in olive oil). On the other hand, the voltammetric determination of the metal is based on the appearance of a peak due to an adsorptive reductive process of the complex that it is observed when the Ad-SSWV technique is used. A calibration graph has been constructed from 0 to 30 ng mL(-)(1), and the detection limit was 0.55 ng mL(-)(1) (13.75 ng g(-)(1) in olive oil according to the proposed procedure).     

Alkyl esters of fatty acids a useful tool to detect soft deodorized olive oils

Fatty acid alkyl esters (FAAEs) are a family of natural neutral lipids present in olive oils and formed by esterification of free fatty acids (FFAs) with low molecular alcohols. Inappropriate practices during the olive oil extraction process and bad quality of the olive fruits promote their formation. Quantification can be done by isolation with a silica gel solid phase extraction cartridge followed by analysis on a gas chromatograph equipped with a programmed temperature vaporizer injector using a polar capillary column. The application of the method to more than 100 Spanish olive oils from different categories, varieties, and geographical origin allowed for establishing the average content of FAAEs and distinguishing the Spanish protected denomination of origin (PDO) and extra virgin olive oils from other categories of olive oils. Those other categories of oils can be subjected to a mild refining process, which leads to blending with extra virgin olive oils. Studies on low quality oils subjected to mild refining showed that FAAEs remain after that process. Thereby, blends of extra virgin olive and mildly refined low quality olive oils can be detected by their alkyl ester concentrations.     

Detection of vinegary defect in virgin olive oils by metal oxide sensors

An array of metal oxide sensors has been set up for detecting the vinegary defect in virgin olive oil. The optimization process was carried out evaluating the variables affecting the process by three desirability functions. Repeatability studies for 6 months and within day were done to evaluate the sensor responses and remove those with high relative standard deviation. The sensor responses were preprocessed applying five weight functions previously to build a regression model. Samples of Spanish Arbequina and Picual virgin olive oil varieties spiked with different amounts of acetic acid (15-200 mg/L) were used as a training set for the regression model. The test set was composed of samples of Italian Coratina virgin olive oil spiked with the vinegary standard at five percentages (10, 25, 40, 50, and 75). A fine-adjusted regression coefficient (R(adj)(2) = 0.98) was computed with the test set.     

Antithrombotic lipid minor constituents from vegetable oils. Comparison between olive oils and others

Many epidemiological studies suggest that vegetable oils and especially olive oil present a protective effect against atherosclerosis. In this study, total lipids (TL) of Greek olive oils and seed oils of four kinds, namely, soybean, corn, sunflower, and sesame oil, were separated into total polar lipids (TPL) and total neutral lipids (TNL) via a novel extraction procedure. TPL and TNL of olive oil were fractionated by HPLC for further study. Each lipid fraction from HPLC separation along with TL, TPL, and TNL lipid samples from oils were tested in vitro for their capacity to induce or to inhibit washed rabbit platelet aggregation. Comparison between olive and seed oils supports the superiority of olive oil as high levels of platelet activating factor (PAF) antagonists have been detected, mainly in TPL. In addition, the structure of the most active fraction from olive oil was elucidated, as a glycerol-glycolipid. Because it has already been reported that PAF plays a pivotal role in atherogenesis, the existence of PAF agonists and antagonists in vegetable oils may explain their protective role against atherosclerosis.     

Rapid quantification of the phenolic fraction of Spanish virgin olive oils by capillary electrophoresis with UV detection

A rapid and reliable capillary zone electrophoresis method was used as a tool to obtain both qualitative and quantitative information about simple phenols, lignans, complex phenols (isomeric forms of secoiridoids), phenolic acids, and flavonoids in the solid-phase separation extracts from different Spanish extra-virgin olive oil in a short time (less than 6 min). Peak identification was done by using commercial and HPLC-isolated standards, studying the information of the electropherograms obtained at several wavelengths and also using the information previously reported. For the quantification of lignans and complex phenols (secoiridoid derivatives), we used a reference compound (oleuropein glucoside) at two different wavelengths (200 and 240 nm) and for the quantification of tyrosol and flavonoids, we used their commercially available standards.     

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.     

Quality and stability of edible oils enriched with hydrophilic antioxidants from the olive tree: the role of enrichment extracts and lipid composition

Phenolic extracts from olive tree leaves and olive pomace were used to enrich refined oils (namely, maize, soy, high-oleic sunflower, sunflower, olive, and rapeseed oils) at two concentration levels (200 and 400 μg/mL, expressed as gallic acid). The concentration of characteristic olive phenols in these extracts together with the lipidic composition of the oils to be enriched influenced the mass transfer of the target antioxidants, which conferred additional stability and quality parameters to the oils as a result. In general, all of the oils experienced either a noticeable or dramatic improvement of their quality-stability parameters (e.g., peroxide index and Rancimat) as compared with their nonenriched counterparts. The enriched oils were also compared with extra virgin olive oil with a natural content in phenols of 400 μg/mL. The healthy properties of these phenols and the scarce or nil prices of the raw materials used can convert oils in supplemented foods or even nutraceuticals.     

基于前表面荧光光谱鉴别新鲜与冻融大黄鱼

为研究反复冻融对水产品品质的影响,通过理化方法检测了不同冻融次数处理对大黄鱼解冻损失、pH 值、色泽、硫代巴比妥酸值、羰基含量等指标的影响,并采用前表面荧光光谱结合主成分分析（principal component analysis,PCA）和 Fisher 线性判别分析法（Fisher linear discriminant analysis,FLDA）对不同冻融次数的大黄鱼进行区分。结果显示随着冻融次数增加,大黄鱼的解冻损失显著增加（P<0.05）;pH 值呈先上升后下降的趋势;L*（亮度）值、b*（黄度）值均有不同程度的增加（P<0.05）,a*（红度）值下降（P<0.05）;羰基含量和硫代巴比妥酸反应物值（thiobarbituric acid reactive substances,TBARS）增加（P<0.05）,反复冻融导致大黄鱼的品质下降。色氨酸和烟酰胺腺嘌呤二核苷酸磷酸（nicotinamide adenine dinucleotide,NADH）的荧光光谱分别结合 PCA 和 FLDA 对不同冻融处理组进行分析,结果表明 FLDA 识别效果优于 PCA。通过 FLDA 建立了新鲜大黄鱼与冻融大黄鱼荧光光谱判别模型,发现色氨酸原始判别的准确率和交叉验证的准确率分别为68.3%和66.7%,NADH 原始判别的准确率和交叉验证的准确率均达到100%。由此可见,利用 NADH荧光光谱结合化学计量分析可以鉴别不同冻融处理的大黄鱼。研究结果为水产品新鲜度的快速评价提供参考。     

Molecular Analysis of Olive Cultivars in the Molise Region of Italy

The Italian region of Molise is characterised by a remarkable variety of microenvironments, where some olive cultivars have settled in the course of many centuries. The study of the varieties cultivated in Molise, and of their relationships with varieties of close regions, is very important in order to clarify the entity of the native olive germplasm, well adapted to the local conditions, and to characterise the oil with regional origin. Eighteen cultivars from Molise have been examined using AFLP markers. They were compared with other cultivars, widely cultivated in Italy and France, in order to evaluate their similarities and to test some hypotheses on their origin and spreading among different areas of cultivation. Numerous varieties showed a high similarity and one case of synonymy (Frantoio-Paesana Bianca) was observed. The results provide useful information in order to distinguish the native germplasm and evaluate the consistency of the olive heritage of the region.     

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 geographical origin of olive oils using 13C nuclear magnetic resonance spectroscopy. I - Classification of olive oils of the Puglia region with denomination of protected origin

13C nuclear magnetic resonance spectroscopy was used to classify olive oils from the three production areas of the Puglia region labeled with the "denomination of protected origin" (DPO) Terra di Bari, Colline di Brindisi, and Dauno. High resolution (13)C spectra of 173 olive oil samples were measured, and the intensity data of triacylglycerol resonances were processed by using linear discriminant analysis, which was carried out stepwise for variable selection. The olive oil samples from the DPOs Colline di Brindisi and Terra di Bari were 90% correctly classified, whereas only 74% of "Dauno" DPO oils were classified in the true group. The performance of the discriminant model was verified by applying the cross-validation procedure based on the "leave one out" formalism. The discriminant model was evaluated against a blind test set of olive oils from the three DPO areas. All the oils used for the purpose were correctly assigned to their respective groups, with the exception of the Dauno oil samples based on the Coratina cv. They were misclassified as Terra di Bari oils because of a common monovarietal composition.