Free and Esterified 4,4′-dimethylsterols in Hazelnut Oil and their Retention During Refining Processes

Free and esterified forms of sterols provide detailed information on the identity and the quality of vegetable oils. In this study, 4,4′-dimethylsterols in free and esterified forms were investigated in hazelnut and virgin olive oils. Moreover, a sample of solvent-extracted hazelnut oil was refined at the laboratory to monitor the effects of processing on the levels of 4,4′-dimethylsterols. Generally, the level of total 4,4′-dimethyslterols was higher in the esterified form (49–68%) compared with that in free form (32–51%) of these compounds in the hazelnut oil. In virgin olive oil samples, cycloartenol and 24-methylenecycloartanol were present in higher amounts in free forms (70–80%) than in esterified forms (20–30%). Among the refining processes, degumming, deodorization, neutralization and bleaching, only neutralization and bleaching considerably reduced 4,4′-dimethylsterols. In fully refined hazelnut oil, 18 and 37% of lupeol and an unknown compound X in the esterified form were lost, respectively. The loss of these two compounds in the free form was considerably higher, 26 and 72%, respectively. GC–MS analysis showed that adulteration of olive oil with a sample of fully refined hazelnut oil could be detected at a level as low as 2% by tracing lupeol in total or only in esterified forms of 4,4′-dimethylsterols. Further studies on the levels of free and esterified 4,4′-dimethylsterols and their retention during refining processes are anticipated in hazelnut cultivars from different origins.     

Detection of adulterated and misbranded olive oil products

The Food and Drug Administration has been examining bulk and packaged olive oil products in a continuing program to detect adulteration of olive oil products. Thirteen of 20 products collected in 1983–84 labeled as olive oil contained undeclared esterified (synthetic) olive oil and four contained undeclared olive-residue oil (derived from olive pomace and pits). Seven of 13 brands of imported olive oil contained undeclared esterified oil, suggesting that considerable quantities of esterified oil have been shipped to the United States identified as olive oil.     

Detection of adulteration of olive oil with seed oils by a combination of column and gas liquid chromatography

Samples of virgin olive oil and refined seed oils, as well as mixtures of olive oil with 10 and 5% seed oils were fractionated by column chromatography on silicic acid impregnated with ammoniacal silver nitrate. It was possible to isolate a characteristic fraction enriched in polyunsaturated triglycerides. Its linoleic acid content in pure olive oil never exceeds 9.3%, whereas in pure seed oils, it varies between 38.1 and 70.1%; in mixtures of olive oil with 10 and 5% of seed oils, the respective values are 22.3–38.2% and 15.6–32.1%. The oleic-to-linoleic acid ratios of the same fraction are more than 7.6 (olive oil), 0.2–0.8 (seed oils), 1.1–2.0 (olive oil with 10% seed oils) and 1.4–3.6 (olive oil with 5% seed oils). These analytical values may be used as a safe criterion for the eventual adulteration of olive oil with seed oils. This work was taken in part from the doctoral dissertation of S. Passaloglou-Emmanouilidou.     

Sterol fractions in hazelnut and virgin olive oils and 4,4′-dimethylsterols as possible markers for detection of adulteration of virgin olive oil

Reports on the methylsterol fractions of hazelnut oils are scarce. The objectives of this study were to characterize methylsterols in hazelnut and virgin olive oils and to study the possibility of detection of adulteration of virgin olive oils. In hazelnut oils, 4-desmethylsterols were present in higher proportions (86 to 91%) than in virgin olive oils where this fraction was ca. 50% of the total sterol. In the 4-monomethylsterol fraction, citrostadienol was the major component in both kinds of oils followed by cycloeucalenol and obtusifoliol in virgin olive oils, and obtusifoliol in hazelnut oils. 24-Methylenecycloartanol was predominant in both kinds of oils in the 4,4′-dimethylsterols. For the first time, δ-amyrin was tentatively identified by comparing published mass spectral data in the analyzed samples of both kinds of oils. An unknown compound X (containing a lupane skeleton) and lupeol were detected only in the 4,4′-dimethylsterols fraction of hazelnut oils at a level of 2–8 and 6–10%, respectively. GC-MS analysis showed that adulteration of virgin olive oil by hazelnut oil could be detected at a level less than 4% by using these two compounds as possible potential markers.     

Identification of adulterants in olive oils

The application of discriminant analysis for identifying and quantifying adulterants in extra virgin olive oils is presented. Three adulterants were used (sunflower oil, rapessed oil, and soybean oil) and were present in the range 5–95%. Near-infrared spectroscopy and principal components analysis were used to develop a discriminant analysis equation that could identify correctly the type of seed oil present in extra virgin olive oil in 90% of cases. Partial least squares analysis was used to develop a calibration equation that could predict the level of adulteration. Cross validation suggested that it was possible to measure the level of adulteration to an accuracy of ±0.9%. External validation of the derived calibation equation gave a standard error of performance of ±2.77%.     

Triglyceride composition of olive oil,cottonseed oil and their mixtures by low temperature crystallization and gas liquid chromatography

Crystallization and gas liquid chromatography (GLC) have been used to characterize the triglyceride composition of olive and cottonseed oil and their precipitates from acetone or methanol/acetone (10:90, v/v) at −2 C. The precipitate obtained after a 24 hr crystallization of a 5% (w/v) solution of the sample in acetone or methanol/acetone (10:90, v/v) at −2 C was named Precipitate I (P-I); that isolated after 2 successive crystallizations under identical conditions was named Precipitate II (P-II). In each case, the ratio of oleic to linoleic acid (O/L) was calculated and proved to be a useful index for detecting adulteration of olive oil with cottonseed oil. In olive oil, the ratio O/L increased from the original sample to its precipitates, whereas in cottonseed oil and the adulterated samples the ratio O/L was lower in the precipitates than in the original sample. For olive oil P-II, the lowest value of the ratio O/L was 8.4; for the adulterated samples it was 7.6. On the basis of this index, adulteration of olive with cottonseed oil as low as 10% can be detected. Hydrolysis of P-1 by porcine pancreatic lipase and analysis of the fatty acids of the sn-2 position showed that the enrichment factor of linoleic acid varied between 1.11–1.30 for olive oil and between 1.55–1.90 for the adulterated samples. Even for adulteration with 5% cottonseed oil, the enrichment factor appears to increase (1.55–1.57) and can be used as a criterion for adulteration.     

Frying Quality Characteristics of French Fries Prepared in Refined Olive Oil and Palm Olein

The objective of this study was to compare two oils with different polyunsaturated/saturated (P/S) fatty acid ratios, refined olive oil (P/S 0.75) and palm olein (P/S 0.25), in frying French fries. The chemical qualities of the oil residues extracted from the French fries were assayed for five consecutive batches fried at 1-h intervals. The levels of total polar compounds, free fatty acids, p-anisidine value and phytosterol oxidation products (POPs) were elevated in French fries fried in both oils. The level of total polar compounds increased from 4.6 in fresh refined olive oil to 7.3% in final batches of French fries. The corresponding figures for palm olein were 9.8–13.8%. The level of free fatty acid in fresh refined olive oil increased from 0.06 to 0.11% in final products. These figures for palm olein were 0.04–0.13%. The p-anisidine value increased from 3.7 to 32.8 and 2.5 to 53.4 in fresh oils and in final batches of French fries in refined olive oil and palm olein, respectively. The total amount of POPs in fresh refined olive oil increased from 5.1 to 9.6 μg/g oil in final products. These figures were 1.9 to 5.3 μg/g oil for palm olein.     

Detection of pressed hazelnut oil in admixtures with virgin olive oil by analysis of polar components

Analysis of the polar fraction from virgin olive oil and pressed hazelnut oil by high-performance liquid chromatography showed marked differences in the chromatograms of the polar components in the two oils. Six commercial samples of pressed hazelnut oil and 12 samples of virgin olive oil (or blended olive oil including virgin olive oil) were analyzed. The phenolic content of the pressed hazelnut oil samples was 161±6 mg·kg⁻¹. Inspection of the chromatograms showed that the pressed hazelnut oil extracts contained a component that eluted in a region of the chromatogram that was clear in the olive oil samples, and consequently this component could be used to detect adulteration of virgin olive oil by pressed hazelnut oil. The component had a relative retention time of 0.9 relative to 4-hydroxybenzoic acid added to the oil as an internal standard. The ultraviolet spectrum of the component showed a maximum at 293.8 nm, but the component could not be identified. Analysis of blends of oils showed that adulteration of virgin olive oil by commercial pressed hazelnut oil could be detected at a level of about 2.5%.     

Recognition of Adulterated Oils by Direct Analysis of the Minor Components

Recognition of adulteration by other oils via direct analysis of the minor components (“sterol fraction”) is shown for olive oil. 10 % of various oils were admixed, the free alcohols silylated and the minor components analyzed by on-line coupled LC-GC-FID. For most oils, even smaller additions can be recognized. Admixed oils can no longer be determined, however, if their minor components have been removed by strong raffination. Bleaching of rapeseed oil with 7% of earth at 180 °C, in fact, completely removed free and esterified sterols.     

Evaluation of MALDI-TOF/MS Technology in Olive Oil Adulteration

Adulteration of extra virgin olive oil (EVOO) by addition of other vegetable oils or lower-grade olive oils is a common problem of the oil market worldwide. Therefore, we developed a fast protocol for detection of EVOO adulteration by mass spectrometry fingerprinting of triacylglycerol (TAG) profiles based on MALDI-TOF/MS. For that purpose, EVOO TAG profiles were compared with those of edible sunflower oil and olive oil composed of refined olive oil and virgin olive oils. Adulteration of EVOO was simulated by addition of sunflower and mixture of refined olive oil and virgin olive oils at 1, 10 and 20% w/w. Results of mass spectrometry TAG profiling were compared with routinely assessed K values for identification of adulteration. MALDI-TOF/MS technology coupled with statistical analysis was proven as useful for detection of adulteration in EVOO at a rate down to 1%. In contrast, standard spectrophotometric methods failed to identify minor adulterations. In addition, the ability of MALDI-TOF/MS in detection of adulteration was tested on EVOO samples from different geographical regions. Results demonstrated that MALDI-TOF/MS technology coupled with statistical analysis is able to distinguish adulterated oils from other EVOO.     

Assessment of olive oil adulteration by reversed-phase high-performance liquid chromatography/amperometric detection of tocopherols and tocotrienols

A method involving reversed-phase high-performance liquid chromatography with amperometric detection has been developed for the analysis of tocopherols and tocotrienols in vegetable oils. The sample preparation avoids saponification. Recoveries of α-tocotrienol and γ-tocotrienol in extra virgin olive oil were 97.0 and 102.0%, respectively. No tocotrienols were detected in olive, hazelnut, sunflower, and soybean oils, whether virgin or refined. However, relatively high levels of tocotrienols were found in palm and grapeseed oils. This method could detect small quantities (1–2%) of palm and grapeseed oils in olive oil or in any tocotrienol-free vegetable oil and might, therefore, help assess authenticity of vegetable oils.     

Chemical Composition of Turkish Olive Oil––Ayvalik

Although large amounts of olive oil are produced in Turkey, not much information on its chemical composition is available in the literature to date. The aim of this study was to evaluate the chemical composition of commercial olive oils produced from the Ayvalik olive cultivar in Canakkale, Turkey. Five different samples corresponding to the olive oil categories of extra virgin (conventional, extra virgin olive oil (EVOO), and organic extra virgin olive oil (OGOO) production), virgin olive oil (OO-1), ordinary virgin olive oil (OO-2) and refined olive oil (RFOO) were evaluated. Olive oils were collected from two consecutive production years. According to the free fatty acids, the absorbance values (K₂₃₂ and K₂₇₀), and peroxide values of all the samples conformed to the European standards for olive oil. The level of oleic acid was in the range of 68–73%; while the linoleic acid content was significantly lower in the refined olive oils. The tocopherol and polyphenol content was in the lower range of some European olive oils. However, pinoresinol was a major phenolic compound (5–77 mg/kg depending on the oil category). Its content was markedly higher than in many other oils, which would be a useful finding for olive oil authentication purposes.     

Use of high-resolution 13C nuclear magnetic resonance spectroscopy for the screening of virgin olive oils

¹³C Nuclear magnetic resonance (NMR) spectra of 104 oil samples were obtained and analyzed in order to study the use of this technique for routine screening of virgin olive oils. The oils studied included the following: virgin olive oils from different cultivars and regions of Europe and north Africa, and refined olive, “lampante” olive, refined olive pomace, high-oleic sunflower, hazelnut, sunflower, corn, soybean, rapeseed, grapeseed, and peanut oils, as well as mixtures of virgin olive oils from different geographical origins and mixtures of 5–50% hazelnut oil in virgin olive oil. The analysis of the spectra allowed us to distinguish among virgin olive oils, oils with a high content of oleic acid, and oils with a high content of linoleic acid, by using stepwise discriminant analysis. This parametric method gave 97.1% correct validated classifications for the oils. In addition, it classified correctly all the hazelnut oil samples and the mixtures of hazelnut oil in virgin olive oil assayed. All of these results suggested that ¹³C NMR may be used satisfactorily for discriminating some specific groups of oils, but to obtain 100% correct classifications for the different oils and mixtures, more information than that obtained from the direct spectra of the oils is needed.     

Analysis of free and esterified sterols in vegetable oils

In vegetable oils, phytosterols occur as free sterols or as steryl esters. Few analytical methods report the quantification of esterified and free sterols in vegetable oils. In this study, esterified and free sterols were separated by silica gel column chromatography upon elution with n-hexane/ethyl acetate (90∶10 vol/vol) followed by n-hexane/diethyl ether/ethanol (25∶25∶50 by vol). Both fractions were saponified separately and the phytosterol content was quantified by GC. The analytical method for the analysis of esterified and free sterols had a relative standard deviation of 1.16% and an accuracy of 93.6–94.1%, which was comparable to the reference method for the total sterol analysis. A large variation in the content and distribution of the sterol fraction between different vegetable oils can be observed. Corn and rapeseed oils were very rich in phytosterols, which mainly occurred as steryl esters (56–60%), whereas the majority of the other vegetable oils (soybean, sunflower, palm oil, etc.) contained a much lower esterified sterol content (25–40%). No difference in the relative proportion of the individual sterols among crude and refined vegetable oils was observed.     

Detection of olive oil adulteration with canola oil from triacylglycerol analysis by reversed-phase high-performance liquid chromatography

The objective of this study was to explore the use of reversed-phase high-performance liquid chromatography (RP-HPLC) as a means to detect adulteration of olive oil with less expensive canola oil. Previously this method has been shown to be useful in the detection of some other added seed oils; however, the detection of adulteration with canola oil might be more difficult due to similarities in fatty acid composition between canola oil and olive oil. Various mixtures of canola oil with olive oils were prepared, and RP-HPLC profiles were obtained. Adulteration of olive oil samples with less than 7.5% (w/w) canola oil could not be detected.     

Evaluation of Authenticity of Iranian Olive Oil by Fatty Acid and Triacylglycerol Profiles

For evaluation of the authenticity of Iranian olive oil, samples from many Iranian olive oil producers especially north of Iran in the production year 2007 were collected. The fatty acid and triacylglycerol compositions were measured. The most recent calculation methods including ∆ECN the difference between the actual and theoretical ECN42 (equivalent carbon number), triglyceride content and R of olive oils according to IOOC methods were applied. On the basis of our results, we were able to classify the olive oils into the extra virgin, virgin olive and olive oil categories. The important fatty acids are oleic, palmitic and linoleic acids and their main triacylglycerols are OOO, POO, OOL, PLO, SOS plus POP, and OLL, respectively. On the basis of the triacylglycerol results, experimental ECN48, ECN46, ECN50, ECN44 and ECN42 were obtained. By using the fatty acids results and a computer program, the theoretical ECN42 and ECN44 were calculated. Then R values, being the ratio of r ECN42/r ECN44 for authenticity of all olive oils and ∆ECN for determining categories of olive oils, were defined. The results of olive oil samples were in the accepted limits of Codex and IOOC. Finally we suggest that the R and ∆ECN can be used in identification of adulteration of olive oils and also they are useful from the point of view of authenticity and classification.     

Low molecular weight polypeptides in virgin and refined olive oils

Twenty-eight virgin olive oils—from different regions of Spain and prepared from olive drupes of different varieties—and six refined olive oils were analyzed to determine the presence of proteins in these oils. All oils studied showed the presence of proteins in the range of 7–51 μ/100 g of oil. There were no significant differences in protein content in oils from different varieties or between virgin or refined oils. In addition, all oils exhibited analogous amino acid patterns, suggesting a similarity among protein fractions obtained from different oils. A polypeptide with an apparent M.W. of 4600 Da was common to the isolated protein fractions. These results suggest that this polypeptide is a previously unknown minor component in olive oils. No clear influence of this component on oil stability was observed when oil stabilities were estimated as a function of phenol, tocopherol, phosphorus, and protein contents of the oils.     

Characterization of Turkish Virgin Olive Oils Produced from Early Harvest Olives

Olives were collected from various districts of Turkey (North and South Aegean sub-region, Bursa-Akhisar, South East Anatolia region) harvested over seven (2001–2007) seasons. The aim of this study was to characterize the chemical profiles of the oils derived from single variety Turkish olives including Ayvalik, Memecik, Gemlik, Erkence, Nizip Yaglik and Uslu. The olive oils were extracted by super press and three phase centrifugation from early harvest olives. Chosen quality indices included free fatty acid content (FFA), peroxide value (PV) and spectrophotometric characteristics in the ultraviolet (UV) region. According to the FFA results, 46% (11 out of 24 samples) were classified as extra virgin olive oils; whereas using the results of PV and UV, over 83% (over 19 of the 24 samples) had the extra virgin olive oil classification. Other measured parameters included oil stability (oxidative stability, chlorophyll pigment, pheophytin-α), cis–trans fatty acid composition and color index. Oxidative stability among oils differed whereas the cis–trans fatty acid values were within the national and international averages. Through the application of two multivariate statistical methods, Principal component and hierarchical analyses, early harvest virgin olive oil samples were classified according to the geographical locations categorized in terms of fatty acid profiles. Such statistical clustering gave rise to defined groups. These data provide evidence of the variation in virgin olive oil quality, especially early harvest and cis–trans isomers of fatty acid profiles from the diverse agronomic conditions in the olive growing regions of Turkey.     

Comparison of near-infrared, fourier transform-infrared, and fourier transform-raman methods for determining olive pomace oil adulteration in extra virgin olive oil

The adulteration of extra virgin olive oil with cheaper oils is a major problem in the olive oil market. In this study, near-infrared, mid-infrared, and Raman spectroscopic techniques were used to quantify the amount of olive pomace oil adulteration in extra virgin olive oil. The concentration of olive pomace oil in extra virgin olive oil was in the range between 0 and 100% in 5% increments by weight. Of the methods studied, Fourier transform-Raman spectroscopy gave the highest correlation with a correlation coefficient of 0.997 and a standard error of prediction of 1.72%. The spectroscopic techniques have the potential to become a tool for rapid determination of adulteration in extra virgin olive oil, because they are casy to use and cost-effective.     

Antioxidant capacity of extra-virgin olive oils

In this study, the oxygen radical absorbance capacity (ORAC) of vegetable oils was investigated using a spectrofluorometric method, which measures the protection of the phenolic substances of the oil on the β-phycoerythrin fluorescence decay in comparison with Trolox. More than 97% of the phenolic substances was extracted from the oil using methanol, and the methanolic extract was then used for the ORAC and the total phenolics assay. We found a significant correlation between ORAC values of different olive oils and the total amount of phenolics. For extra-virgin olive oils, maximal ORAC values reached 6.20±0.31 μmol Trolox equivalent/g, while refined and seed oils showed values in the 1–1.5 μmol Trolox equivalent/g range. Our method is useful to assess the quality of olive oils and to predict, in combination with the rancidity tests, their stability against oxidation.