The tocopherol pattern in human serum is markedly influenced by intake of vitamin E drugs—Results of the german national health surveys

During national and regional health surveys, done from 1984–1995 in Germany, consumption data for all drugs used by the participants—approximately 18,000 persons—in the last 7 d before the examination were monitored with a detailed drug-usage questionnaire. The groups examined are representative for the national and regional German inhabitant population aged 25–69 yr. In serum samples of subsamples of the study participants, all tocopherols were measured by isocratic high-performance liquid chromatography (Si 60 column, fluorescence detection). Consumption data for tocopherol-containing drugs showed that up to 5% of females and up to 3% of males of the study population used those drugs. During the study period, the serum content of α-tocopherol (mean values ± SD) rose from 7.5 ± 2.6 mg/L serum to 11.8 ± 2.8 mg/L serum for nonusers and from 11.9 ± 4.3 mg/L serum to 15.3 ± 4.9 mg/L serum in tocopherol-drug users. Throughout all studies, it could be shown that β- and γ-tocopherol were heavily reduced in those persons taking daily doses ≥50 mg α-tocopherol. The reduction of the two tocopherols is dose-dependent and especially pronounced in females using high-dose α-tocopherol drugs. Owing to the emerging evidence of the physiological importance concerning the balance of the different tocopherols in biological systems, the possible benefits of using natural tocopherol mixtures from plant origin instead of pure RRR-α-tocopherol, gained from permethylation procedures, as vitamin supplements in human nutrition should be considered.     

Analysis of tocopherols in vegetable oils by high-performance liquid chromatography: Comparison of fluorescence and evaporative light-scattering detection

A comparison of the responses of an evaporative light-scattering detector (ELSD) and a fluorescence detector for tocopherols in vegetable oils by high-performance liquid chromatography is presented. The tocopherols were separated from acylglycerols by gel-permeation chromatography (GPC). The tocopherol fraction was collected off a set of four GPC columns with a mobile phase of methylene chloride before separation on a normal-phase silica column with a mobile phase of hexane/isopropanol, 99.7∶0.3 (vol/vol). An internal standard of 5,7 dimethyltocol, which was detected by both the ELSD and fluorescence detector, was used to obtain quantitative data. The fluorescence detector was ten times more sensitive than the ELSD. γ-Tocopherol was the major tocopherol detected in the vegetable oils studied and ranged from 24.1–93.3 mg/100 g. The amounts of tocopherols found in the vegetable oils agreed favorably with the literature values.     

Optimal tocopherol concentrations to inhibit soybean oil oxidation

The optimal concentration for tocopherols to inhibit soybean oil oxidation was determined for individual tocopherols (α-, γ-, and δ-tocopherol) and for the natural soybean oil tocopherol mixture (tocopherol ratio of 1∶13∶5 for α-, γ-, and δ-tocopherol, respectively). The concentration of the individual tocopherols influenced oil oxidation rates, and the optimal concentrations were unique for each tocopherol. For example, the optimal concentrations for α-tocopherol and γ-tocopherol were ∼100 and ∼300 ppm, respectively, whereas δ-tocopherol did not exhibit a distinct concentration optimum at the levels studied (P<0.05). The optimal concentration for the natural tocopherol mixture ranged between 340 and 660 ppm tocopherols (P<0.05). The antioxidant activity of the tocopherols diminished when the tocopherol levels exceeded their optimal concentrations. Above their optimal concentrations, the individual tocopherols and the tocopherol mixture exhibited prooxidation behavior that was more pronounced with increasing temperature from 40 to 60°C (P<0.05). A comparison of the antioxidant activity of the individual tocopherols at their optimal concentrations revealed that α-tocopherol (∼100 ppm) was 3–5 times more potent than γ-tocopherol (∼300 ppm) and 16–32 times more potent than δ-tocopherol (∼1900 ppm).     

Separation and quantitation of cholesterol oxides by HPLC with an evaporative light scattering detector in a model system

A method to analyze cholesterol and 10 of its oxidation products, ranging from the weakly polar cholest-4-ene-3,6-dione to moderately polar cholest-5-ene-3β,7α-diol, in a single run is described. The separation was achieved by normal-phase gradient high-performance liquid chromatography with an evaporative light-scattering detector. This universal mass detector does not detect changes in solvent composition; this makes it possible to employ gradients, an essential technique whenever a wide range of compounds with diverse characteristics is to be separated. Standards at concentrations from 0.1–1.0 μg were separated within 37 min on an alumina/silica column with a gradient elution system that contained dichloromethane, acetonitrile, and water.     

Antioxidant activities of α- and γ-tocopherols in the oxidation of rapeseed oil triacylglycerols

Antioxidant properties of 5 to 500 μg/g levels of α-and γ-tocopherols, in the oxidation of rapeseed oil triacylglycerols (RO TAG), were studied at 40°C in the dark. Each tocopherol alone and in a mixture was studied for its stability in oxidizing RO TAG. Also the effects of tocopherols on the formation of primary and secondary oxidation products of RO TAG were investigated. Both tocopherols significantly retarded the oxidation of RO TAG. At low levels (≤50 μg/g), α-tocopherol was more stable and was a more effective antioxidant than γ-tocopherol. At higher α-tocopherol levels (>100 μg/g), there was a relative increase in hydroperoxide formation parallel to consumption of α-tocopherol, which was not found with γ-tocopherol. Therefore, γ-tocopherol was a more effective antioxidant than α-tocopherol at levels above 100 μg/g. As long as there were tocopherols present, the hydroperoxides were quite stable and no volatile aldehydes were formed. In a mixture, α-tocopherol protected γ-tocopherol from being oxidized at the addition levels of 5+5 and 10+10 μg/g but no synergism between the tocopherols was found. α-Tocopherol was less stable in the 500+500 μg/g mixture than when added alone to the RO TAG. No prooxidant activity of either tocopherol or their mixture was found.     

Oxidation kinetics of β-carotene in oleic acid solvent with addition of an antioxidant, α-tocopherol

Oxidation experiments with β-carotene were performed in oleic acid solvent with addition of an antioxidant, α-tocopherol. A kinetic model was proposed based on a reaction mechanism consisting of the oxidation of β-carotene, oleic acid, and α-tocopherol; the antioxidation reactions of β-carotene and oleic acid by α-tocopherol; the cross-reaction of β-carotene and oleic acid; and the radical-exchange reaction of β-carotene and α-tocopherol. The model quantitatively described the oxidation behavior of β-carotene over a wide range of temperatures, oxygen compositions, and initial antioxidant concentrations. The model simulated well the time over which β-carotene was almost totally consumed under practical storage conditions at room temperature in air.     

Rapid analysis of oxidized cholesterol derivatives by high-performance liquid chromatography combined with diode-array ultraviolet and evaporative laser light-scattering detection

Extensive evidence of the deleterious biological effects of oxidized 5-cholesten-3β-ol (cholesterol) derivatives has led to great interest in their detection. We observed that known oxidized cholesterol derivatives can be rapidly quantitated by combining reversed-phase high-performance liquid chromatography (HPLC) with ultraviolet (UV) absorption and evaporative laser light-scattering (ELSD) detection. Using a 20 × 0.46 cm C18 HPLC column and methanol/acetonitrile (60:40, vol/vol) as the mobile phase at 1.0 mL/min, 10 species of oxidized cholesterol derivative were measured by UV (205, 234, and 280 nm) while 5-cholestan-5α,6α-epoxy-3β-ol (5α-epoxycholesterol), 5-cholestan-5β,6β-epoxy-3β-ol (5β-epoxycholesterol), and 5-cholestan-3β,5α,6β-triol (cholestanetriol) were detected by only ELSD. The minimal limits of detection ranged from 100 to 500 ng depending on sterol and detector. The response was linear in the range 0–1000 or 0–2000 ng depending on detector. These oxidized cholesterol derivatives were also identified by HPLC/mass spectrometry analysis combined with UV detector. Heated tallow contained cholestanetriol, 5-cholesten-3β,7α-diol (7α-hydroxycholesterol), 5-cholesten-3β,7β-diol (7β-hydroxycholesterol), 5-cholesten-3β-ol-7-one (7-ketocholesterol), 5α- and 5β-epoxycholesterols under the developed analysis condition. Photooxidized cholesterol had cholestanetriol, 7α- and 7β-hydroxycholesterols and 3,5-cholestadien-7-one. On the other hand, 7α- and 7β-hydroxycholesterols, 7-ketocholesterol, 5α- and 5β-epoxycholesterols and 3,5-cholestadien-7-one were observed in copper-oxidized low-density lipoprotein. Thus, this developed HPLC analysis method could be applied to identification of oxidized cholesterol derivatives in food and biological specimen.     

Alteration of steryl ester content and positional distribution of fatty acids in triacylglycerols by chemical and enzymatic interesterification of plant oils

Steryl ester content of refined and interesterified corn, soybean, and rapeseed oils has been measured via clean-up on a short silica gel column, followed by high performance liquid chromatography with evaporative light-scattering mass detector. Chemical interesterification, catalyzed by sodium methoxide, led to random positional distribution of fatty acids in triacylglycerols and some increase in the steryl ester content of all three oils. Enzymatic interesterification, catalyzed by the immobilized lipase from Rhizomucor miehei (Lipozyme), resulted in a distinct reduction in steryl ester content, but essentially no alteration in positional distribution of fatty acids in triacylglycerols occurred. Formation of steryl esters during chemical and enzymatic interesterification was also examined by radioactive tracer technique with [4-¹⁴C]β-sitosterol added as marker to refined rapeseed oil and measurement of the radioactive steryl esters formed. Chemical interesterification of rapeseed oil resulted in moderate formation (10% of total radioactivity) of radioactive β-sitosteryl esters. Enzymatic interesterification of the oil, catalyzed by Lipozyme, led to little formation of radioactive β-sitosteryl esters, whereas with the lipase from Candida cylindracea high proportions (>90% of total radioactivity) of ¹⁴C-labeled β-sitosteryl esters were formed. Part of doctoral thesis of Roseli Ap. Ferrari to be submitted to Faculdade de Engenharia de Alimentos, Universidade de Campinas, Campinas, Brazil.     

β-Apo-8′-Carotenoic acid and its esters in sunflower oil oxidation

The oxidation kinetics of sunflower oil (SO) and pure triacylglycerols of sunflower oil (TGSO) in the presence of different concentrations (0.0008–0.02%, 1.9–32.7×10⁻⁵ M) of β-apo-8′-carotenoic acid (CA), ethyl β-apo-8′-carotenoate (EC), and β-apo-8′-carotenoylglycerol (CG) were studied. The process was performed at high (kinetic regime) and low (diffusion regime) oxygen concentrations at room temperature and at 100°C and in the dark and in daylight. CA, EC, and CG were not antioxidants in TGSO systems. However, the carotenoid derivatives, especially CA, increased the stability of tocopherol-containing SO at room temperature and in daylight. The stabilization effect was more evident in a kinetic regime of oxidation. The synergism between the carotenoids and tocopherols was characterized by the increase of the stabilization factor F and activity A. F and A were highest for CA (F=1.2–5.5, A=2.4–78.6), followed by EC (F=1.2–3.5, A=1.7–14.6) and CG (F=1.1–2.1, A=1.6–5.5) in the kinetic regime for SO exposed to daylight at room temperature. Presented at the 91st AOCS Annual Meeting & Expo, April 25–28, 2000, San Diego, California.     

The influence of carotenoids and tocopherols on the stability of safflower seed oil during heat-catalyzed oxidation

The stability and antioxidant effects of carotenoids and tocopherols in safflower seed oil were evaluated under thermal (75°C) and oxidative conditions and the oxidative stability index (OSI) determined. The antioxidant capability of butylated hydroxytoluene (BHT) was also compared with that of β-carotene in a model system. Lycopene and β-carotene (1 to 2000 ppm) were heated (75°C) and exposed to air (2.5 psi) in an oxidative stability instrument. β-Carotene had no antioxidant effect at concentrations below 500 ppm, because it did not alter the induction time. Lycopene increased the induction time only slightly at low concentrations. However, at concentrations greater than 500 ppm, both β-carotene and lycopene acted as prooxidants, significantly decreasing the induction period. At the highest concentration, 2000 ppm, lycopene was more prooxidative than β-carotene. α- and γ-Tocopherol (concentration, 1000 ppm) delayed the induction time by 16 and 26 h, respectively. There was no cooperative interaction between α-tocopherol and β-carotene in delaying the onset of oxidation. Furthermore, BHT was significantly more antioxidative than β-carotene. Thus, under thermal and oxidative conditions, β-carotene could not delay the onset of oxidation. The tocopherols and BHT were effective in suppressing the onset of oxidation, as determined by the oxidative stability measurement.     

Obtaining Butter Oil Triacylglycerols Free from β-Carotene and α-Tocopherol via Activated Carbon Adsorption and Alumina-Column Chromatography Treatments

It is difficult to remove β-carotene from oils with alumina-column chromatography, because β-carotene is even less-polar than triacylglycerols (TAGs) are. The objective of this study was to obtain butter oil TAGs free from β-carotene and antioxidants via sequential treatments with activated carbon (AC) adsorption and alumina column chromatography. The AC used was prepared from waste apricots. The effects of AC dosages, temperatures and time courses on β-carotene adsorption were studied. The Langmuir and Freundlich isotherms were used to describe the adsorption of β-carotene onto AC, and it was found to be more consistent with the Freundlich isotherm with a higher R ² value (0.9784). Adsorption kinetics of β-carotene was analyzed by pseudo-first order and pseudo-second order models. The pseudo-second order model was found to explain the kinetics of β-carotene adsorption more effectively (R ² = 0.9882). The highest β-carotene reduction was achieved (from 31.9 to 1.84 mg/kg) at an AC dosage of 10 wt%, temperature of 50 °C, and adsorption time of 240 min. A considerable amount of α-tocopherol was also adsorbed during the AC treatment. Remaining portions of α-tocopherol were completely removed with alumina adsorption chromatography. The method described may be used for purification of vegetable oil TAGs, which will be used as model compounds in model oxidation studies.     

Lipid Components of North American Wild Rice (<Emphasis Type="Italic">Zizania palustris</Emphasis>)

The content and composition of fatty acids, sterols, tocopherols, and γ-oryzanol in wild rice (Zizania palustris) grown in North America were compared with those in regular brown rice (Oryza sativa L.). The lipid content of wild rice ranged from 0.7 to 1.1%, compared with 2.7% in regular brown rice. The lipids of wild rice comprised mainly linoleic (35–37%) and linolenic (20–31%) acids. Other fatty acids included palmitic (14.1–18.4%), stearic (1.1–1.3%), and oleic (12.8–16.2%). Wild rice lipids contained very large amounts of sterols, ranging from 70 g/kg for a Saskatchewan sample to 145 g/kg for Minnesota Naturally Grown Lake and River Rice. The main sterols found in an unsaponified fraction were: campesterol (14–52%), β-sitosterol (19–33%), Δ⁵-avenasterol (5–12%), and cycloartenol (5–12%). Some of sterols, γ-oryzanols, were present as the phenolic acid esters; the amount ranged from 459 to 730 mg/kg in wild rice lipids. The largest amounts of tocopherols and tocotrienols, 3682 and 9378 mg/kg, were observed in North Western Ontario wild rice samples, whereas the lowest were 251 mg/kg in an Athabasca Alberta sample and 224 mg/kg in regular long-grain brown rice. The α isomer was the most abundant among tocopherols and tocotrienols. The results of this study showed that wild rice lipids contain large amounts of nutraceuticals with proven positive health effects.     

Genotype and growing location effects on phytosterols in canola oil

There is little information available about phytosterols in canola (Brassica napa L.) oil and the effects of genotype and growing locations from Virginia and the mid-Atlantic region of the United States, a potential area for the establishment of domestic production to provide edible oil. Our objectives were to characterize the phytosterols, phospholipids, unsaponifiable matter, and FA in oil from Virginia-grown canola. Among 11 canola genotypes grown at two locations during 1995–1996 significant variations existed for oil content and FA profiles, but not for contents of phospholipids, unsaponifiable matter, total phytosterols, campesterol, stigmasterol, and β-sitosterol, Total phytosterol content in the oil of Virginia-grown canola varied from 0.7 to 0.9% with a mean of 0.8%. This concentration compared favorably with oil from Canadian canola, which typically contains 0.5 to 1.1% total phytosterols. The mean contents of brassicasterol, campesterol, stigmasterol, β-sitosterol, Δ⁵-avenasterol, and Δ⁷-stigmatenol as percentages of total phytosterols in Virginia-grown canola were: 9.7, 32.0, 0.6, 49.3, 4.99, and 3.5%, respectively. Growing location did not affect phytosterols in Virginia-grown canola oil but had significant effects on contents of phospholipids, and saturated (myristic, stearic, and arachidic) and unsaturated (palmitoleic, linoleic, linolenic, eicosenoic, and erucic) FA.     

Side-chain autoxidation of stigmasterol and analysis of a mixture of phytosterol oxidation products by chromatographic and spectroscopic methods

Although the structure of phytosterols is closely related to cholesterol, there is a gap in knowledge concerning the formation and occurrence of oxidation products from phytosterols. The main objective of this study was to isolate and characterize some side-chain oxidation products formed after autoxidation of stigmasterol. Another objective was to highlight the difficulties in the analysis of phytosterol and a mixture of their oxidation products by GC. Pure stigmasterol was oxidized at 120°C for 72 h in an air-ventilated oven. Preparative TLC separated the oxidation products, and the products were characterized with GC-MS and NMR. In addition to the common ring-structure oxidation compounds, three semipolar oxidation products—24-ethylcholest-5,22-dien-3β,25-diol, 24-ethylcholest-5,22-dien-3β,24-diol, and 24-ethyl-5,22-choladien-3β-ol-24-one—were characterized for the first time by TLC, GC-MS, and NMR. Moreover, the results of the analysis of a large number of oxidation products from sitosterol, campesterol, and stigmasterol by capillary column GC indicated that further efforts and optimization are required in this area.     

Effect of α- and γ-tocopherols on thermal polymerization of purified high-oleic sunflower triacylglycerols

The antipolymerization effects of α- and γ-tocopherols were compared in model systems composed of purified high-oleic sunflower triacylglycerols at 180°C. γ-Tocopherol was much more effective as an antipolymerization inhibitor than α-tocopherol, partly due to lower oxidizability/disappearance. Purified triacylglycerols of sunflower, rapeseed, and high-oleic sunflower oils were less stable than their nonpurified forms containing tocopherols. Results confirmed that tocopherols per se can act as antipolymerization agents in high-oleic oils at frying temperatures. No synergism was observed when α- and γ-tocopherols were present together although larger amounts of residuals were left for both tocols. Results suggested that high-oleic/high-γ-tocopherol oils (such as high-oleic canola and high-oleic soybean oils) may provide better frying oils than high-oleic/high-α-tocopherol oils (such as high-oleic sunflower oil).     

Role of minor constituents in the photooxidation of virgin olive oil

Virgin olive oil was photooxidized at 2 and 40°C and at fluorescent light intensities of 0, 620, 2710, and 5340 lux. As expected, higher fluorescent light intensities induced higher peroxide formation in the oil. The thiobarbituric acid reactive substances (TBARS) were found to be good indicators of photooxidation during the early stage of the reaction. Pheophytin A and β-carotene were light- and temperature-sensitive, whereas α-tocopherol and total polyphenols were mostly affected by light. Pheophytin A functioned as a photosensitizer, resulting in rapid oxidation of the oil. β-Carotene was a strong natural inhibitor of photooxidation for all light intensities at 2°C, suggesting quenching properties for singlet oxygen. However, β-carotene antioxidant activity was reduced at 40°C because of its rapid destruction.     

The effect of the N-linked glycans on structural features and physicochemical functions of soybean β-conglycinin homotrimers

β-Conglycinin is a trimeric protein consisting of three subunits, α,α′,and β, which are N-glycosylated. The α and α′ subunits contain extension regions in addition to core regions common to all subunits. We purified homogeneous trimers consisting of only α, α′, or β from mutant soybean cultivars containing β-conglycinin lacking one or two subunits: α homotrimers from an α′-lacking mutant, α′ homotrimers from an α-lacking mutant, and β homotrimers from an α-and α′-lacking mutant. Structural features and physicochemical functions of the three homotrimers were examined and compared with those of recombinant homotrimers having no N-linked glycans. The native homotrimers have secondary structures very similar to those of the recombinant ones. In analogy with the recombinant homotrimers, the native ones exhibit different thermal stabilities from one another (β>α′>α), and the native α and α′ homotrimers exhibit better solubility, emulsifying ability, and heat-induced association than the native β homotrimer. Further, the N-linked glycans contribute to solubilities of the three subunits at low ionic strength (μ=0.08) and to the emulsifying ability of the native β homotrimer. N-Linked glycans also prevent heat-induced associations of the native α and α′ homotrimers but do not contribute to the secondary structure and the thermal stability of β-conglycinin.     

A rigorous kinetic model for β-carotene oxidation in the presence of an antioxidant, α-tocopherol

Oxidation of β-carotene in an inert solvent, n-decane, in the presence of various concentrations of the antioxidant α-tocopherol was studied. The progress of carotene oxidation was suppressed as long as the tocopherol remained in the system. A rigorous kinetic model for carotene oxidation in the presence of an antioxidant was proposed based on a reaction mechanism in which not only the antioxidation but also the co-oxidation and radical-exchange reaction of tocopherol with carotene were incorporated. The model quantitatively described the oxidation behavior of carotene over a wide range of temperatures, oxygen compositions, and initial antioxidant concentrations.     

Rheological properties of sodium caprate-induced β-lactoglobulin gel and changes in secondary structure during gelation

In this study, we found that transparent gels of β-lactoglobulin (β-LG) were formed by adding different concentrations of sodium caprate to protein solutions at ambient temperature. We investigated changes in the dynamic viscoelasticity of the mixture with time at 25°C and found that more than 12% β-LG induced the formation of a viscoelastic gel with a suitable amount of sodium caprate (for example, 12% β-LG and 3.6% sodium caprate). Furthermore, we analyzed the changes in the secondary structure of proteins during the gelation step by FHR spectroscopy. Dissociation of the β-LG dimer was first observed just after mixing with sodium caprate. Furthermore, in the β-LG protein in which the original contents were predominantly β-sheets, intermolecular β-sheets attributable to aggregation increased with a decrease in the content of intramolecular β-sheets. Sodium caprate-induced gel was heated at 80°C for 30 min after the gel was formed, and a large increase in the intermolecular β-sheet bands was observed by heat treatment. These results suggest that the formation of sodium caprate-induced gels of β-LG was accompanied by less marked changes in the protein conformation than those in heat-induced gels.     

Shape-selective alkylation of isopropylnaphthalene on MCM-22 zeolite

An MCM-22 zeolite with an Si/Al ratio 13 was found to exhibit higher selectivity to β,β-diisopropylnaphthalene in the alkylation of isopropylnaphthalene with isopropylalcohol, as compared with other zeolites such as mordenite, Y and beta having similar Si/Al ratios. This high selectivity is ascribed to the specific pore shape of MCM-22 zeolite. The gradual increase in the selectivity with reaction time is due to its pore modification by coke deposit.