Different Effects of Vitamin C-Based Supplements on the Advance of Linseed Oil Component Oxidation and Lipolysis during In Vitro Gastrointestinal Digestion.

Although widely consumed, dietary supplements based on Vitamin C contain high doses of this compound, whose impact on lipid oxidation during digestion needs to be addressed. Therefore, the effect of seven commercial supplements and of pure l-ascorbic acid and ascorbyl palmitate on linseed oil during in vitro gastrointestinal digestion was tackled. The advance of lipid oxidation was studied through the generation of oxidation compounds, the degradation of polyunsaturated fatty acyl chains and of gamma-tocopherol, by employing Proton Nuclear Magnetic Resonance. Supplements containing exclusively l-ascorbic acid enhanced the advance of linseed oil oxidation during digestion. This was evidenced by increased formation of linolenic-derived conjugated hydroxy-dienes and alkanals and by the generation of conjugated keto-dienes and reactive alpha,beta-unsaturated aldehydes, such as 4,5-epoxy-2-alkenals; moreover, gamma-tocopherol was completely degraded. Conversely, supplements composed of mixtures of ascorbic acid/salt with citric acid and carotenes, and of ascorbyl palmitate, protected linseed oil against oxidation and reduced gamma-tocopherol degradation. The study through Solid Phase Microextraction-Gas Chromatography/Mass Spectrometry of the volatile compounds of the digests corroborated these findings. Furthermore, a decreased lipid bioaccessibility was noticed in the presence of the highest dose of l-ascorbic acid. Both the chemical form of Vitamin C and the presence of other ingredients in dietary supplements have shown to be of great relevance regarding oxidation and hydrolysis reactions occurring during lipid digestion.

Assessment of Soybean Oil Oxidative Stability from Rapid Analysis of its Minor Component Profile.

The minor components of vegetable oils are important for their oxidative stability. In order to know to what extent they can influence oil behaviour under oxidative conditions, two commercial soybean oils, one virgin and the other refined, both with very similar compositions in acyl groups but differing in their minor component profiles, were subjected to accelerated storage conditions. They were characterized by 1H nuclear magnetic resonance (NMR) and direct immersion solid-phase microextraction coupled to gas chromatography/mass spectrometry (DI-SPME-GC/MS), while oil oxidation was monitored by 1H-NMR. The lower levels of tocols and sterols in the virgin oil, together with its higher free fatty acid content when compared to the refined one, result in a lower oxidative stability. This is deduced from faster degradation of acyl groups and earlier generation of hydroperoxides, epoxides, and aldehydes in the virgin oil. These findings reveal that commercial virgin soybean oil quality is not necessarily higher than that of the refined type, and that a simple and rapid analysis of oil minor components by DI-SPME-GC/MS would enable one to establish quality levels within oils originating from the same plant species and similar unsaturation level regarding composition in potentially bioactive compounds and oxidative stability.

Effect of adding alpha-tocopherol on the oxidation advance during in vitro gastrointestinal digestion of sunflower and flaxseed oils.

Few in vitro studies have tackled the effect of alpha-tocopherol on lipid oxidation during digestion, and discrepant results have been reported. As a result, the aim of this study was to elucidate whether the addition of alpha-tocopherol enhances or slows down the advance of oxidation that occurs during in vitro gastrointestinal digestion of polyunsaturated lipids. For this purpose, commercial sunflower and flaxseed oils (as models of omega-6 and omega-3 rich lipid systems, respectively) were in vitro digested in the absence or in the presence of this tocol at different concentrations (0.02%, 0.2% and 2%). Proton Nuclear Magnetic Resonance (1H NMR) and Solid Phase Microextraction followed by Gas Chromatography/Mass Spectrometry (SPME-GC/MS) were used to investigate in detail potential differences among the digests regarding lipolysis and oxidation level. Alpha-tocopherol addition did not affect the advance of lipolysis, whereas lipid oxidation was enhanced in a dose-dependent manner. In this regard, the increased degradation of polyunsaturated lipids and greater generation of primary and secondary oxidation products observed at higher concentrations of alpha-tocopherol confirmed this observation. Among the formed oxidation products, hydroperoxy-, hydroxy- and keto-dienes, as well as oxygenated alpha,beta-unsaturated aldehydes are worth mentioning. The in vitro bioaccessibility of added tocopherol was estimated to be very low, suggesting a notable transformation under the assayed conditions. Further in vivo studies are necessary to confirm this prooxidant activity of alpha-tocopherol during gastrointestinal digestion.

Influence of minor components on lipid bioaccessibility and oxidation during in vitro digestion of soybean oil.

BACKGROUND: Minor components of edible oils could influence their evolution during in vitro digestion. This might affect the bioaccessibility of lipid nutrients and the safety of the ingested food. Bearing this in mind, the evolution of virgin and refined soybean oils, which are very similar in acyl group composition, has been studied throughout in vitro digestion using 1 H nuclear magnetic resonance (NMR) and solid-phase microextraction-gas chromatography /mass spectrometry, focusing on lipolysis and oxidation reactions. The fate of γ-tocopherol, the main antioxidant present in soybean oil, has also been analyzed with 1 H NMR. RESULTS: There were no noticeable differences in lipolysis between the two oils that were studied. The extent of oxidation during digestion, which was very low in both cases, was slightly higher in the virgin type, which showed lower tocopherols and squalene concentrations than the refined one, together with a considerable abundance of free fatty acids. This can be deduced both from the appearance after digestion of conjugated hydroperoxy- and hydroxy-dienes only in the virgin oil, and from its higher levels of volatile aldehydes and 2-pentyl-furan. Under in vitro digestion conditions, the formation of epoxides seemed to be favored over other oxidation products. Finally, although some soybean oil essential nutrients like polyunsaturated fatty acids exhibited no significant degradation after digestion, γ-tocopherol concentration diminished during this process, especially in the virgin oil. CONCLUSION: Although the minor component composition of the soybean oils did not affect lipolysis during in vitro digestion, it influenced the extent of their oxidation and γ-tocopherol bioaccessibility. © 2019 Society of Chemical Industry.

Monitoring of minor compounds in corn oil oxidation by direct immersion-solid phase microextraction-gas chromatography/mass spectrometry. New oil oxidation markers.

The aim of this study is to shed light on the evolution of the minor compounds in the corn oil oxidation process, through the information provided by direct immersion-microextraction in solid phase followed by gas chromatography/mass spectrometry (DI-SPME-GC/MS). This methodology enables one, in a single run, to establish the identity and abundance both of original oil minor components, some with antioxidant capacity, and of other compounds coming from both main and minor oil components oxidation. For the first time, some of the compounds formed from oil minor components degradation are proposed as new markers of oil incipient oxidation. Although the study refers to corn oil, the methodology can be applied to any other edible oil and constitutes a new approach to characterizing the oxidation state of edible oils.

A thorough insight into the complex effect of gamma-tocopherol on the oxidation process of soybean oil by means of 1H Nuclear Magnetic Resonance. Comparison with alpha-tocopherol.

The effect of γ-tocopherol in proportions between 0.02 and 2% by weight on the accelerated storage process of refined soybean oil is studied by 1H NMR, and compared with that of α-tocopherol. Whereas the lowest γ-tocopherol enrichment level does not affect oil evolution, at higher concentrations both γ- and α-tocopherols initially accelerate acyl groups degradation and hydroperoxides generation, more as higher is the tocopherol concentration, this effect being less marked for γ-tocopherol. However, after this initial stage, the rates of acyl groups degradation and hydroperoxides formation decrease with tocopherol concentration. Furthermore, in the case of γ-tocopherol, the higher the enrichment degree, the later hydroperoxides decomposition occurs, so that, unlike α-tocopherol, γ-tocopherol delays the generation of most secondary oxidation products (aldehydes, (E,E)-keto-dienes, epoxy-keto-enes, (E)-epoxystearates and alcohols) with the exception of some epoxides. Similarly to α-tocopherol, γ-tocopherol modifies the oil oxidation pathway at the highest addition level, promoting the formation of compounds with (Z,E)-isomerism, although less noticeably than α-tocopherol.

Prooxidant effect of α-tocopherol on soybean oil. Global monitoring of its oxidation process under accelerated storage conditions by 1H nuclear magnetic resonance.

The effect of adding α-tocopherol in proportions ranging from 0.002 to 5% in weight on the oxidative stability of soybean oil was studied. For the first time, the oxidation process under accelerated storage conditions including evolution of the molar percentages of the several types of oil acyl groups, and formation and evolution of various kinds of oxidation products comprising hydroperoxides, hydroxy-dienes and other alcohols, epoxides, aldehydes and keto-dienes, was followed by 1H nuclear magnetic resonance. It is proved that, except in the lowest proportion, α-tocopherol not only exerts a prooxidant effect on soybean oil but also modifies its oxidation pathway, affecting the oxidation products generation rate, their nature, relative proportions and concentrations. It is noticeable that the highest α-tocopherol concentrations induce the generation of some toxic compounds at earlier stages of the thermoxidation process and sometimes in higher concentration, such as certain oxygenated α,ß-unsaturated aldehydes and monoepoxides derived from linoleic groups.

Behaviour of non-oxidized and oxidized flaxseed oils, as models of omega-3 rich lipids, during in vitro digestion. Occurrence of epoxidation reactions.

Fresh and partially oxidized flaxseed oil, as models of omega-3 rich lipids, were submitted to in vitro gastrointestinal digestion. Hydrolysis level, lipid composition and oxidative status of the samples before and after digestion were studied by Proton Nuclear Magnetic Resonance (1H NMR) and Solid Phase Microextraction-Gas Chromatography/Mass Spectrometry (SPME-GC/MS). Although a great degree of lipolysis was reached in both kinds of samples after digestion, it was somewhat lower in the digests of oxidized flaxseed oil. The occurrence of lipid oxidation during digestion was evidenced by decreased unsaturated lipids and increased primary and secondary oxidation products, especially in oxidized samples. In these latter, linolenic-derived monoepoxy-octadecadienoates were the main oxidation products generated. SPME-GC/MS study showed the highest abundances of highly reactive alkadienals (C5-C10), alkatrienals (C9-C10) and linolenic-derived 4,5-epoxy-2-heptenals in the headspace of oxidized flaxseed oil digests. Volatile markers of Maillard-type reactions were also detected.

Polyunsaturated lipids and vitamin A oxidation during cod liver oil in vitro gastrointestinal digestion. Antioxidant effect of added BHT.

The extent of cod liver oil hydrolysis and oxidation during in vitro gastrointestinal digestion was investigated by Proton Nuclear Magnetic Resonance (1H NMR) and Solid Phase Microextraction-Gas Chromatography/Mass Spectrometry (SPME-GC/MS). These techniques evidenced the degradation of polyunsaturated ω-3 and ω-6 lipids and, for the first time, that of vitamin A, naturally present in cod liver oil. Cis,trans-conjugated dienes associated with hydroperoxides, as well as monoepoxides, cis,trans-2,4-alkadienals, 4-hydroperoxy- and 4-hydroxy-2-alkenals, and several vitamin A derived metabolites were generated. Moreover, the effect of the addition of the synthetic antioxidant 2,6-di-tert-butyl-hydroxytoluene (BHT) at 20 and 800ppm was tackled. Both techniques showed BHT to be efficient in limiting oxidation reactions during digestion, almost inhibiting them at 800ppm. Therefore, the simultaneous intake of antioxidants with cod liver oil should be considered, in order to increase polyunsaturated lipid and vitamin A bioaccessibility and avoid formation of toxic oxidation compounds like oxygenated alpha,beta-unsaturated aldehydes.

Direct study of minor extra-virgin olive oil components without any sample modification. 1H NMR multisupression experiment: A powerful tool.

Proton Nuclear Magnetic Resonance (1H NMR) was employed to study monovarietal commercial Spanish extra-virgin olive oils (EVOO) (Arbequina, Arroniz, Cornicabra, Hojiblanca and Picual). Each sample was analyzed by a standard pulse and by an experiment suppressing the main lipid signals, enabling the detection of signals of minor components. The aim was to determine the possibilities of both 1H NMR approaches to characterize EVOO composition, focusing on acyl groups, squalene, sterols, triterpene acids/esters, fatty alcohols, wax esters and phenols (lignans, tyrosol, hydroxytyrosol, oleocanthal, oleacein, oleokoronal, oleomissional, ligstrodials and oleuropeindials), and to determine hydrolysis and oxidation levels. The signal assignments (in deuterated chloroform) are thoroughly described, identifying for the first time those of the protons of esters of phytol and of geranylgeraniol. Correct signal assignment is fundamental for obtaining sound results when interpreting statistical data from metabolomic studies of EVOO composition and adulteration, making it possible to differentiate and classify oils.

1H NMR and SPME-GC/MS study of hydrolysis, oxidation and other reactions occurring during in vitro digestion of non-oxidized and oxidized sunflower oil. Formation of hydroxy-octadecadienoates.

Both fresh and slightly oxidized sunflower oils, as models of omega-6 rich lipids, were submitted to in vitro gastrointestinal digestion and studied by 1H NMR and SPME-GC/MS. Changes in lipolysis degree, lipid composition and oxidative level were studied by 1H NMR. Three quantitative approaches were used and several equations were newly developed. In oxidized oil digestates slightly lower hydrolysis and a higher advance of oxidation took place during digestion. This latter was evidenced by a greater decrease of lipid unsaturation degree and enhanced generation of oxidation products (cis,trans-hydroperoxy-octadecadienoates, cis,trans- and trans,trans-hydroxy-octadecadienoates). For the first time, the generation of hydroxy-octadecadienoates during in vitro digestion is reported. Furthermore, SPME-GC/MS study of non-digested and digested samples headspaces confirmed that lipid oxidation occurred: abundances of volatile markers increased (including potentially toxic alpha,beta-unsaturated aldehydes), especially in oxidized oils digestates. Markers of Maillard-type and esterification reactions were also detected in the digestates.

A new methodology capable of characterizing most volatile and less volatile minor edible oils components in a single chromatographic run without solvents or reagents. Detection of new components.

The possibilities offered by a new methodology to determine minor components in edible oils are described. This is based on immersion of a solid-phase microextraction fiber of PDMS/DVB into the oil matrix, followed by Gas Chromatography/Mass Spectrometry. It enables characterization and differentiation of edible oils in a simple way, without either solvents or sample modification. This methodology allows simultaneous identification and quantification of sterols, tocols, hydrocarbons of different natures, fatty acids, esters, monoglycerides, fatty amides, aldehydes, ketones, alcohols, epoxides, furans, pyrans and terpenic oxygenated derivatives. The broad information provided by this methodology is useful for different areas of interest such as nutritional value, oxidative stability, technological performance, quality, processing, safety and even the prevention of fraudulent practices. Furthermore, for the first time, certain fatty amides, gamma- and delta-lactones of high molecular weight, and other aromatic compounds such as some esters derived from cinnamic acid have been detected in edible oils.

¹H Nuclear Magnetic Resonance monitoring of the degradation of margarines of varied compositions when heated to high temperature.

In this study, (1)H Nuclear Magnetic Resonance was used to monitor the evolution of three margarines of varied compositions when submitted to heating at 180°C in an oven with aeration. Heating causes degradation of polyunsaturated acyl groups and this depends not only on their unsaturation degree, but also on the concentration of the different acyl groups. The evolution of monounsaturated groups varies depending on the disappearance rate of the groups with higher unsaturation degree. Heat treatment also causes hydrolysis reactions that lead to a reduction in 1-monoglycerides and an increase in 1,2-diglycerides, especially in the margarines with higher water content, as well as degradation of some vegetable sterols. Different types of aldehydes and epoxides were identified and quantified, above all in the margarine with the highest proportion of polyunsaturated groups, especially linoleic; some of these are toxic, such as 4-hydroxy- and 4,5-epoxy-2-alkenals.

Fourier transform infrared spectroscopy as a tool to study farmed and wild sea bass lipid composition.

BACKGROUND: The lipids of 16 farmed and wild European sea bass (Dicentrarchus labrax) samples were studied by Fourier transform infrared (FTIR) spectroscopy. The spectroscopic parameters which would be useful when distinguishing between both fish origins were analysed. RESULTS: It was shown, for the first time, that the frequency and the ratio between the absorbance of certain bands are efficient and reliable authentication tools for the origin of sea bass. Furthermore, relationships between infrared data and fish lipids composition referring to the molar percentage or concentration of certain acyl groups were also studied. It was proved that some infrared spectroscopic data (the frequency of certain bands or the ratio of the absorbance of others), are very closely related to the composition of sea bass lipids. It was shown for the first time that certain infrared spectroscopic data could predict, with a certain degree of approximation, the molar percentage, or concentration, of omega-3, docosahexaenoic (DHA) and di-unsaturated omega-6 (linoleic) in sea bass lipids. CONCLUSION: The consistency of the results confirms the usefulness of FTIR spectroscopy to detect frauds regarding sea bass origin, and to provide important compositional data about sea bass lipids from the nutritional and technological point of view.

Relationships between the evolution of the percentage in weight of polar compounds and that of the molar percentage of acyl groups of edible oils submitted to frying temperature.

The evolution of the molar percentage of several kinds of acyl groups of extra virgin olive, sunflower and virgin linseed oils was monitored throughout heating at frying temperature by means of (1)H nuclear magnetic resonance. Likewise, the evolution of the percentage in weight of the polar compounds of the same oils under the same heating conditions was also determined. Relationships between both sets of parameters, in each oil and in the oils as a group, were studied. An equation which is able to accurately predict the percentage in weight of the polar compounds, throughout the heating at frying temperature, of any one of these three oils, from the molar percentage of triunsaturated, diunsaturated and monounsaturated acyl groups, was obtained. In this way both molar percentage of acyl groups and percentage in weight of polar compounds can be obtained in a few minutes that registration of the (1)H NMR spectrum of the oil takes, in addition to the rest of information provided by this technique. The study reveals the close relationships between percentage in weight of polar compounds and the composition expressed in terms of molar percentages of acyl groups in edible oils heated at frying temperature.

Volatile components of several virgin and refined oils differing in their botanical origin.

BACKGROUND: The volatile components of virgin and refined oils whose compositions were very different in terms of oleic, linoleic and linolenic acyl groups were studied by solid phase microextraction (SPME) followed by gas chromatography/mass spectrometry (GC/MS), and their composition of acyl groups was determined by proton nuclear magnetic resonance (1) H NMR. The virgin oils studied were olive, linseed and walnut oils, while the refined oils were sunflower and rapeseed oils. RESULTS: Large differences were found among the headspace compositions of the virgin oils. These were due in part to compounds that could be considered as markers of different vegetable origins and manufacturing processes, but also to compounds resulting from degradative oxidation of the main acyl group components. By virtue of their being refined, fewer components were present in the refined oils, although differences in their headspaces were found owing to the presence of compounds resulting from oxidation of the determined acyl groups. CONCLUSION: The studied oils show specific characteristics regarding not only their main components, which are well known, but also their volatile components. The latter provide information on some of the processes to which oils have been submitted, on their oxidation level, on their botanical origin and on the compounds responsible for their odour.

Analysis of hydroperoxides, aldehydes and epoxides by 1H nuclear magnetic resonance in sunflower oil oxidized at 70 and 100 degrees C.

A global study of sunflower oil oxidation at two different temperatures (100 and 70 degrees C) with aeration was carried out by (1)H nuclear magnetic resonance ((1)H NMR), paying attention not only to the degradation of the main components but also to the formation of others. The functional groups of compounds monitored simultaneously during the oxidation were, in addition to acyl groups, the following: hydroperoxides, conjugated dienic systems of hydroperoxy acyl groups, aldehydes including the genotoxic and cytotoxic oxygenated alpha,beta-unsaturated aldehydes, and mono- and diepoxides. Chemical shifts of protons of all mentioned groups were given. Differences between both oxidation processes were found from the initial stages to the end. The formation of diepoxides in the oxidation process of edible oils was shown for the first time, as was the influence of temperature on the formation of epoxy acyl groups, some of these latter related to leukotoxin and isoleukotoxin.

A study of the toxic effect of oxidized sunflower oil containing 4-hydroperoxy-2-nonenal and 4-hydroxy-2-nonenal on cortical TrkA receptor expression in rats.

Sunflower oil at a specific oxidation stage (when several oxygenated alpha,beta-unsaturated aldehydes are generated, mainly 4-hydroperoxy-trans-2-alkenals and 4-hydroxy-trans-2-alkenals), caused at 70 degrees C with aeration for 7 days, was administered intraperitoneally to rats. This oil was studied by means of solid phase micro-extraction followed by gas chromatography/mass spectrometry (SPME-GC/MS) and by proton nuclear magnetic resonance ((1)H-NMR). Oxidized sunflower oil (3 ml/kg/day) was administered to male Sprague-Dawley rats for 21 days. The control group was administered non-oxidized sunflower oil in the same volume and for the same duration as the experimental group. A significant decrease in the number of neural cells positively immunostained for TrkA receptor was detected in the frontal cortex of the experimental group, with respect to controls, suggesting both neuronal damage as well as a deficit in neuronal survival signalling at this level. This could lead to apoptosis of cholinergic neurons, which play a key role in memory and attention function. These results indicate that toxic substances present in the oxidized sunflower oil, among them 4-hydroxy-trans-2-nonenal (HNE) and 4-hydroperoxy-trans-2-nonenal (HPNE), could disrupt survival signalling of frontal cortex cholinergic neurons, which could lead to apoptosis and neurodegenerative diseases. In the case of humans, this fact reinforces the necessity of avoiding the re-utilization of oxidized sunflower oil, in order to contribute to long-term neurodegenerative diseases prevention.

Contribution to further understanding of the evolution of sunflower oil submitted to frying temperature in a domestic fryer: study by 1H nuclear magnetic resonance.

A study is made of the evolution of the composition of sunflower oil kept over prolonged periods of time at high temperature (190 degrees C) in a domestic fryer. The technique used is (1)H NMR spectroscopy. The degradation rate of linoleic acyl groups is determined in this process, as well as the proportions of monounsaturated, of saturated plus modified acyl groups and the iodine values. Intermediate oxidation compounds having hydroperoxide groups and conjugated dienic systems were not detected; however, some secondary oxidation compounds such as aldehydes are generated very early, among them, the genotoxic and cytotoxic 4-hydroxy-trans-2-alkenals. Both concentrations of each kind of aldehyde at different heating times and changes in their concentration were also determined. Simultaneously, the level of oil degradation corresponding to a content of 25% of polar compounds measured by Viscofrit test was analyzed in function of the (1)H NMR spectra derived data.

Use of an in vitro digestion model to study the bioaccessibility of 4-hydroxy-2-nonenal and related aldehydes present in oxidized oils rich in omega-6 acyl groups.

Mixtures of either sunflower oil or thermodegraded sunflower oil and a standard meal were submitted to an in vitro digestion model. The same experiment was carried out with fluid deep-frying fat and thermodegraded fluid deep-frying fat. The thermodegradation of the oil and fat was provoked by submitting them to 190 degrees C with aeration in a convection oven, and the presence in the headspace of the thermodegraded oil and fat of oxygenated alpha,beta-unsaturated aldehydes (OalphabetaUAs), such as 4-hydroxy-2-nonenal (HNE), 4-oxo-2-nonenal (ONE), and 4,5-epoxy-2-decenal (EDE), was monitored by solid phase microextraction (SPME) followed by gas chromatography-mass spectrometry (GC-MS). The digestion products were separated by centrifugation in a lipidic phase, an aqueous phase, and a pellet phase. The headspace of these three phases was also studied by SPME/GC-MS to check if the toxic and very reactive OalphabetaUAs above-mentioned remained unaltered after the in vitro digestion process or if they had reacted with the various compounds present in the digestion products, so disappearing from the samples. With the same aim the extract in ethyl acetate of the aqueous and pellet phases, and of the lipidic phase after dilution, were studied by GC-MS. All results obtained showed that a certain proportion of the toxic OalphabetaUAs remains unaltered after digestion, dispersed in the three phases above-mentioned, and thus are bioaccessible in the gastrointestinal tract and so could reach the systemic circulation. Compounds that may originate in Maillard type reactions (2-pentylpyridine) are found among digestion products, proving that these reactions are possible in this process if adequate substrates are present. In addition, it has been shown that toxic metabolites from the synthetic antioxidant BHT, present in fat before digestion, remain unaltered after this process and could reach the systemic circulation.