Sterol Fraction Research Articles

Characterization of Total, Free and Esterified Phytosterols in Tetraploid and Hexaploid Wheats

Dietary plant sterols have received increasing attention in recent years due to their favorable health benefits. The present research focused on quantification of phytosterols as free, esterified and total forms in different tetraploid (5 cultivars of Triticum durum Desf., 9 cultivars of Triticum dicoccon Schrank) and hexaploid (5 cultivars of T. aestivum L., 12 cultivars of Triticum spelta L.) wheats. Tetraploid wheats showed the highest content of total sterol (79.4 and 79.5 mg of sterols /100 g dry weight for T. durum and T. dicoccon, respectively). Hexaploid cultivars were the best source of esterified sterols (40.7% and 37.3% of total sterols for Triticum aestivum and T. spelta, respectively). Significant amounts of free sterols (65.5% and 60.7% of total sterols for T. durum and T. dicoccon, respectively) were found in the tetraploid cultivars. The most abundant phytosterol in all wheat samples was sitosterol accounting for 45.1-59.1, 46.6-57.4 and 38.6-59.5% of total, free and esterified sterol fraction, respectively. These results demonstrate that although the sterol profile present in tetraploid and hexaploid wheat species is the same, differences in their relative amounts and distribution allow statistical differentiation between hexaploids and tetraploids, and between soft and durum wheats.

Polysaccharides and sterols from green algae Caulerpa lentillifera and C. sertularioides

Sterols and polysaccharides of green alga Caulerpa lentillifera grown under laboratory conditions and in mariculture and polysaccharides of green alga C. sertularioides grown under natural conditions were studied. The sterol fraction consisted of C27-C29 steroidal alcohols with Δ5-unsaturation in the steroid core regardless of the growth conditions. The dominant (79.9%) steroid component of the sterol fraction was clionasterol. The water-soluble fraction of C. lentillifera grown under laboratory conditions was a mixture of 1,4-α- and 1,3-β-D-glucans and protein. The same fraction isolated from C. lentillifera grown in mariculture contained only protein. The water-soluble fraction of C. sertularioides grown under natural conditions contained 1,3;1,6-β-D-galactan sulfated at C2. The principal components of the base-soluble polysaccharide fractions from all algae samples were 1,4-α-D-glucans.

Lipid fraction constituents and evaluation of anti-anaphylactic activity of <i>Prunus mahaleb</i> l. Kernels

The lipid fraction constituents as well as evaluation of anti-anaphylactic activity of Prunus mahaleb L. kernels were studied. Prunus mahaleb L. kernels were obtained from the local market in Cairo, Egypt. Investigation of the fatty acids revealed that oleic and linoleic acids are the major constituents. 12 compounds were identified from the hydrocarbon fraction. The sterol fraction comprises of cholesterol, stigmasterol, beta-sitosterol and campesterol. The pharmacotoxicity studies were carried out on total and defatted ethanolic extracts as well as the oil fraction. The oil fraction proved to be extremely safe and free from any acute lethal toxicity in intraperitoneal (i.p.) and oral doses up to 100 ml/kg. In vivo assessment of prophylactic efficacy was afforded by 7 days course of daily medication schedule of sensitized adult male guinea pigs against ovalbumin bronchospasm. The prophylactic anti-inflammatory activity of the total ethanolic extract was higher than that of the defatted ethanolic extract. In addition, the lipid fraction of Prunus mahaleb L. kernels evoked complete anti-inflammatory efficacy among the survival animals receiving low and medium doses.

Multivariate analysis of NMR fingerprint of the unsaponifiable fraction of virgin olive oils for authentication purposes

A new approach to the geographical characterisation of virgin olive oils (VOOs) based on the 1H NMR fingerprint of the unsaponifiable matter is presented. The 1H NMR spectra of the unsaponifiable fraction of virgin olive oils from Spain, Italy, Greece, Tunisia, Turkey, and Syria were analysed by several pattern recognition techniques (LDA, PLS-DA, SIMCA, and CART). PLS-DA (PLS-1 approach) obtained the best classification results for all classes. Moreover, 1H NMR spectra of the bulk oil, and its corresponding unsaponifiable fraction, as well as the subfractions of the unsaponifiable fraction (alcohol, sterol, hydrocarbon, and tocopherol fractions) were studied in the search for the markers that multivariate techniques revealed to be related to the geographical origin of olive oils. Additionally, a preliminary study regarding 1H NMR data of the bulk oil and the corresponding unsaponifiable fraction of VOOs suggested that these spectral data contained complementary information for the geographical characterisation of VOOs.

Leaf surface compounds of the potato (Solanum tuberosum) and their influence on Colorado potato beetle (Leptinotarsa decemlineata) feeding

The sampling behavior of the Colorado potato beetle Leptinotarsa decemlineata Say (CPB) involves examination of the surface of potato leaves. It has been suggested that leaf surface compounds (volatiles and cuticular waxes) may be involved in host-plant recognition, acceptance or discrimination. Here we report on the effect of leaf surface extracts of six Polish commercial potato varieties on CPB feeding. We tested the influence of potato leaf surface extracts on CPB adult and larval feeding, then separated the extracts with HPLC, and finally tested the effect of the HPLC-separated fractions on CPB feeding. The bioassays were performed using potato leaf discs deprived of their original surface compounds. Applied to test discs at concentrations ten times higher than natural (10 leaf area equivalent), the extracts deterred CPB adults and larvae from feeding. HPLC-separated fractions composed of alkanes, sesquiterpene hydrocarbons, wax esters, benzoic acid esters, fatty acid methyl, ethyl, isopropyl and phenylethyl esters, aldehydes, ketones, methyl ketones, fatty acids, primary alcohols, β-amyrin and sterols did not affect adult CPB feeding. Similarly, alkanes, sesquiterpene hydrocarbons, wax esters, methyl ketones, sesquiterpene alcohols and secondary alcohols had no effect on larval CPB feeding. The sterol fraction (cholesterol, β-sitosterol and stigmasterol) acted as a phagostimulant to CPB larvae. We isolated a fraction demonstrating a phagodeterrent effect on CPB adults and larvae. The qualitative composition of the deterrent fraction was quite similar in all potato extracts, but there were quantitative differences between the varieties. Much further work is needed to identify the compounds that can produce the deterrent effect.

New Hemiketal Steroid from the Introduced Soft Coral Chromonephthea braziliensis is a Chemical Defense against Predatory Fishes

Recent studies show that chemical defenses in the exotic soft coral Chromonephthea braziliensis Ofwegen (Nephtheidae, Alcyonacea) can be one of the reasons for the success of this introduced species. We report for the first time the detailed composition of the monohydroxylated sterol fraction and a new hemiketal steroid, 23-keto-cladiellin-A, isolated from the unpalatable hexane extract from C. braziliensis. Bioassay-guided fractionation of this extract revealed that this hemiketal steroid exhibits potent feeding deterrent properties against a natural assemblage of fishes at the natural concentration. The major sterol fraction, containing the monohydroxylated sterols, was inactive in the bioassay. The results suggest that this active molecule may be driving the observed success of the invasion of this soft coral along the Brazilian Atlantic coast.

Cholesterol Substitution Increases the Structural Heterogeneity of Caveolae

Caveolin-1 binds cholesterol and caveola formation involves caveolin-1 oligomerization and cholesterol association. The role of cholesterol in caveolae has so far been addressed by methods that compromise membrane integrity and abolish caveolar invaginations. To study the importance of sterol specificity for the structure and function of caveolae, we replaced cholesterol in mammalian cells with its immediate precursor desmosterol by inhibiting 24-dehydrocholesterol reductase. Desmosterol could substitute for cholesterol in maintaining cell growth, membrane integrity, and preserving caveolar invaginations. However, in desmosterol cells the affinity of caveolin-1 for sterol and the stability of caveolin oligomers were decreased. Moreover, caveolar invaginations became more heterogeneous in dimensions and in the number of caveolin-1 molecules per caveola. Despite the altered caveolar structure, caveolar ligand uptake was only moderately inhibited. We found that in desmosterol cells, Src kinase phosphorylated Cav1 at Tyr(14) more avidly than in cholesterol cells. Taken the role of Cav1 Tyr(14) phosphorylation in caveolar endocytosis, this may help to preserve caveolar uptake in desmosterol cells. We conclude that a sterol C24 double bond interferes with caveolin-sterol interaction and perturbs caveolar morphology but facilitates Cav1 Src phosphorylation and allows caveolar endocytosis. More generally, substitution of cholesterol by a structurally closely related sterol provides a method to selectively modify membrane protein-sterol affinity, structure and function of cholesterol-dependent domains without compromising membrane integrity.

Sterolic composition of Chétoui virgin olive oil: Influence of geographical origin

The sterol profile of Tunisian virgin olive oils produced from Chétoui cultivar, the second main variety cultivated in the north of the country, grown under different environmental conditions, was established by gas chromatography using a flame ionisation detector. More than ten compounds were identified and characterised. As expected for virgin olive oil, the main sterols found in all Chétoui olive oils were β-sitosterol, Δ5-avenasterol, campesterol and stigmasterol. Cholesterol, 24-methylenecholesterol, clerosterol, campestanol, sitostanol, Δ7-stigmastenol, Δ5,24-stigmastadienol, and Δ7-avenasterol were also found in all samples, but in lower amounts. Most of these compounds are significantly affected by the geographical origin. The majority of the Chétoui virgin olive oils analysed respected EC Regulation No. 2568, and in all cases total sterol amounts were higher than the minimum limit set by legislation, ranging from 1017 to 1522mg/kg.Two triterpenic dialcohols (erythrodiol and uvaol), were also detected besides the sterolic components. Their content was below the upper legal limit of 4% in all analysed samples, with a range from 1.2% to 3.2%. These results suggest that, besides the genetic factor, environmental conditions influence the sterolic fraction.

High Performance Thin Layer Chromatographic Analysis of Neutral Lipids in the Feces of BALB/c Mice Infected with Echinostoma caproni

High performance thin layer chromatography was used to determine neutral lipid profiles in the feces of BALB/c mice infected with adults of Echinostoma caproni (Trematoda). An approximate 25 worm burden per host induced changes in the neutral lipid profile of the mice at 1 to 5 weeks post infection (PI) relative to the uninfected controls. Infection caused a significant decrease (Student's t‐test, P<0.05) in the triacylglycerol fraction at 3 weeks PI and a significant increase (Student's t‐test, P<0.05) in the free sterol fraction at 2 weeks PI relative to the uninfected controls. Neutral lipid profiles of feces may serve as an indicator of infection by intestinal trematodes in animals and humans.

Sterols and related metabolites from five species of sponges

Free sterol fractions were isolated from the marine sponges Phyllospongia madagascarensis, Scalarispongia sp., Oceanapia sp., Monanchora clathrata and studied by GLC, GLC–MS, and spectroscopy NMR. P. madagascarensis and Scalarispongia sp. contained common Δ 5-sterols; cholesterol was shown to be a main sterol of both the sponges. Oceanapia sp. contained stanols and minor Δ 5-sterols with 24R-24,25-methylene-5α-cholestan-3β-ol as a main constituent. Many free sterols from M. clathrata were Δ 7-series compounds, and latosterol was a main sterol. Δ 4-3-Ketosteroids and Δ 5-sterol esters were found in the Antarctic sponge Haliclona sp., but free sterols were practically absent except for trace amount of cholesterol. A chemotaxonomic application of sterols in relation to the genera Phyllospongia, Oceanapia and the family Crambeidae is provided. The known cases of the absence of sterols in sponges and probable reasons of the phenomenon are discussed.

Rosemary extract (Rosmarinus officinalis L.) supplementation into the diet of Nero Siciliano pigs: effects on lipid oxidation

During the growing-fattening period (93 days; ILW 33.5±6 kg to FLW 67±3 kg), 30 Nero Siciliano pigs were fed on a basal diet supplemented with (ROX group) or not (CTR group) a rosemary extract (1g?kg-1). At 1, 3 and 5 days after slaughtering, the oxidative stability of the Longissimus dorsi muscle was determined by using TBArs test. Moreover, on the individual samples of the muscle the lipid content, the fatty acid and the sterol composition were determined; the acidic composition of the lard, removed from backfat, was also studied. Data were subjected to ANOVA. The fat content of the muscle was unaffected by the dietary treatment as well as the oxidative stability whereas, significant differences (P≤0.01) were observed for the polyunsaturated fatty acid content which showed the highest values in the meat as well as in the lard of the ROX group; PUFA/SFA ratio was also significantly highest in the muscle (P=0.004) and in the lard (P=0.017) of the ROX group, testifying a possible antioxidative activity of the rosemary extract. The sterol fraction (cholesterol, cholestanol, stigmasterol, beta-sitosterol and delta 5-avenasterol) of the Longissimus dorsi muscle was unaffected by the rosemary supplementation; a significant difference was observed only for the campesterol (CTR group=1.08, ROX group=0.90; P = 0.021).

Cholestanol metabolism in patients with cerebrotendinous xanthomatosis: absorption, turnover, and tissue deposition

To study the metabolism of cholestanol in patients with cerebrotendinous xanthomatosis (CTX), we measured the cholestanol absorption, the cholesterol and cholestanol turnover, and the tissue content of sterols in two patients. Cholestanol absorption was approximately 5.0%. The rapid exchangeable pool of cholestanol was 233 mg, and the total exchangeable pool was 752 mg. The production rate of cholestanol in pool A was 39 mg/day. [4-14C]cholestanol was detected in the xanthomas, but neither [4-14C]cholestanol nor [4-14C]cholesterol was detected in peripheral nerves biopsied at 49 and 97 days after [4-14C]cholesterol given intravenously. Of the 18 tissues analyzed at biopsy and autopsy, the cholestanol content varied from 0.09 mg/g in psoas muscle to 76 mg/g in a cerebellar xanthoma. With the assumption that the cholestanol-to-cholesterol ratio is 1.0, the relative cholestanol-to-cholesterol ratio varied from 1.0 in plasma and liver to 30.0 in the cerebellar xanthoma; cholestanol was especially high in nerve tissue. Our data indicate that CTX patients absorb cholestanol from the diet. They have a higher than normal cholestanol production rate. Cholestanol was derived from cholesterol. In CTX patients, the blood-brain barrier was intact to the passage of [4-14C]cholesterol and [4-14C]cholestanol. The deposition of large amounts of cholestanol (up to 30% of total sterols in cerebellum) in nerve tissues must have an important role in the neurological symptoms in CTX patients. In view of the intact blood-brain barrier, several other explanations for the large amounts of cholestanol in the brain were postulated.

Lipid composition of the liver oil of the ray, Himantura bleekeri

If rays are traditionally fished for their caudal fins, the rest of the body is wasted, except part of the skin that can be transformed into leather. Liver oil of the ray, Himantura bleekeri, was characterized in terms of lipid class composition and fatty acid profile. Liver oil content was high and represented 54% of the liver weight (w/w). Neutral lipids were predominant (92%), major components being triacylglycerols (63%). Other neutral lipids identified were hydrocarbons, sterol esters, mono- and di-acylglycerols, free fatty acids, sterols and glyceryl ethers. Polyunsaturated fatty acids of the n−3 series, namely eicosapentaenoic and docosahexaenoic acids, were high (4% and 16%, respectively). Cholesterol was the major component in the sterol fractions (free or esterified). Hydrocarbons were up to 30 carbons and squalene was present at the level of 22%. Thus, this liver oil proved to be an adequate source of n−3 fatty acids and other valuable lipidic compounds.

Chemistry of Pinus sylvestris cones

The chemical composition of the individual organs (needles, needleless runners, buds, cones) of common pine (Pinus sylvestris L.) was studied in order to develop and improve industrial reprocessing of green wood. Second-year cones have not been studied previously. We attempted to find the dominant components of the extract. Raw material (195 g calc. for absolute dry material) was collected on the banks of Lake Baikal in Bol′shie Koty village during July 2003 and was preserved in ethanol at the collection site in order to prevent fermentation. The concentrated ethanol extract (54 g) was extracted exhaustively with CHCl3 and then butanol. The CHCl3 extract (10 g) was repeatedly chromatographed over silica gel to isolate dehydroabietic acid (1, 60 mg) and a sterol fraction (15 mg). The butanol extract (21 g) afforded (-)-catechin (2, 100 mg), shikimic acid (3, 700 mg), and β-sitosterol glucoside (4, 54 mg). The pure compounds were identified by spectral methods including 2D NMR. Signals for C atoms were assigned to the corresponding signals for H atoms using 1H—13C HETCOR. Through-space C—H pairings were found using 1H—13C HMBC. Free sterols comprised β-sitosterol (M+ 414), stigmasterol (M+ 412), and campesterol (M+ 400) according to GC—MS (Agilent HP 6890-5973 N). Dehydroabietinic acid (1): amorphous powder, [α]546 +74° (c 1.2, CHCl3) [1]. The 13C NMR spectrum was identical to that in the literature [2]. (-)-Catechin (2): mp 184-186°C, [α]546 -10° (c 0.8, CH3OH) [3]. The 13C NMR spectrum agreed the that in the literature for 2,3-trans-isomers of flavan-3-ols [4]: chemical shifts were 81.41 ppm for C-2 and 67.38 ppm for C-3. The spin—spin coupling constants for H-2 and H-3 were 7.5 Hz, which confirmed the trans-position of the substituents on C-2 and C-3 [5]. The negative rotation angle of the plane of polarized light for 2 enabled one of the two optical isomers of catechin to be selected. Shikimic acid (3): mp 184°C (ethanol), [α]546 -206° (c 2.1, ethanol) [1]. β-Sitosterol glucopyranoside (4): mp 267-270°C (CH3OH), [α]546 -38° (c 0.9, pyridine) [6]. Compound 4 was identified by comparing its physical chemical constants with those in the literature and by its chromatographic behavior. All compounds were isolated from common pine cones for the first time.

Quantification of free and esterified sterols in Portuguese olive oils by solid-phase extraction and gas chromatography–mass spectrometry

A simple and accurate method based on solid-phase extraction (SPE), transesterification and gas chromatography–mass spectrometry (GC–MS) was developed for the quantitative analysis of free and esterified sterols of olive oil. In order to achieve better separation of esterified and free sterols, silica and alumina SPE adsorbents were tested. Separations by silica provided more reproducible results. The transesterification of both sterol fractions was found to be more user friendly than saponification as a method to liberate the sterols from the respective esters. The free sterols were then silylated with N, O-bis-trimethylsilyltrifluoroacetamide (BSTFA) with 1% of trimethylchlorosilane (TMCS). The most favourable conditions for exploitation of this reagent were established. The optimized methodology was suitable for evaluation of free and esterified sterols in Protected Designation of Origin (PDO) olive oils and monovarietal olive oils with different maturation indices. The prevailing phytosterols in all olive oils were β-sitosterol and campesterol. The free sterols predominated, although they seemed to decrease with the maturation of the olive fruits.

Composition of total sterols (4-desmethyl-sterols) in extravirgin olive oils obtained with different extraction technologies and their influence on the oil oxidative stability

The composition and antioxidant activity of total sterols in extravirgin olive oils obtained with different extraction technologies from olives harvested at two ripening stages, were studied. The antioxidant activity was evaluated with an oxidative stability instrument (OSI), by using a model system (made of a mixture of treated/untreated commercial refined peanut oil) enriched with the total sterol fractions of the extravirgin olive oils. No correlation was found between the OSI time and the extraction technologies, the ripening stages or the actual amount of sterols added. No significant differences were observed in the percent composition of sterols of extravirgin olive oils produced with different technologies during the same harvesting period. The latter, however, had a significant effect on the percent of β-sitosterol and 5-avenasterol in extravirgin olive oils produced with the same technology.

The use of the Dhcr7 knockout mouse to accurately determine the origin of fetal sterols

Mice with a targeted mutation of 3beta-hydroxysterol Delta(7)-reductase (Dhcr7) that cannot convert 7-dehydrocholesterol to cholesterol were used to identify the origin of fetal sterols. Because their heterozygous mothers synthesize cholesterol normally, virtually all sterols found in a Dhcr7 knockout fetus having a Delta(7) or a Delta(8) double bond must have been synthesized by the fetus itself but any cholesterol had to have come from the mother. Early in gestation, most fetal sterols were of maternal origin, but at approximately E13-14, in situ synthesis became increasingly important, and by birth, 55-60% of liver and lung sterols had been made by the fetus. In contrast, at E10-11, upon formation of the blood-brain barrier, the brain rapidly became the source of almost all of its own sterols (90% at birth). New, rapid, de novo sterol synthesis in brain was confirmed by the observation that concentrations of C24,25-unsaturated sterols were low in the brains of all very young fetuses but increased rapidly beginning at approximately E11-12. Reduced activity of sterol C24,25-reductase (Dhcr24) in brain, suggested by the abundance of C24,25-unsaturated compounds, seems to be the result of suppressed Dhcr24 expression. The early fetal brain also appears to conserve cholesterol by keeping cholesterol 24-hydroxylase expression low until approximately E18.

Phytosterol biosynthesis pathway in Mortierella alpina

The Zygomycetes fungus Mortierella alpina was cultured to growth arrest to assess the phytosterol biosynthesis pathway in a less-advanced fungus. The mycelium was found to produce 13 sterols, but no ergosterol. The sterol fractions were purified to homogeneity by HPLC and their identifies determined by a combination of GC–MS and 1H NMR spectroscopy. The principal sterol of the mycelium was cholesta-5, 24-dienol (desmosterol) (83%), with lesser amounts of 24β-methyl-cholesta-5,25(27)-dienol (codisterol) (2%), 24-methyldesmosterol (6%), 24(28)-methylene cholesterol (3%) and lanosterol (3%) and several other minor compounds (3%). The total sterol accounted for approximately 0.07% of the mycelial dry wt. Mycelium fed methionine- methyl- 2H 3 for 6 days, generated 3 2H-24-methyl(ene) sterols, [C28- 2H 2]24(28)-methylenecholesterol, [C28- 2H 3]24-methylcholesta-5,24-dienol and [C28- 2H 3]24β-methyl-cholesta-5,25(27)-dienol. The formation of the 24-methyl sterols seems to be catalyzed by the direct methylation of a common Δ 24-acceptor sterol thereby bypassing the intermediacy of an isomerization step for rearrangement of the Δ 24(28)-bond to Δ 25(25)-position as operates in Ascomycetes fungi and all plants.

Effect of cobalt ions on lipid and sterol metabolism in the marine invertebrates Mytilus galloprovincialis and Actinia equina

The lipid and sterol fractions of the mussel Mytilus galloprovincialis and the anemone Actinia equina were analyzed before and after incubation with cobalt ions. There were significant changes in the lipid and sterol composition, strongly depending on the cobalt ions concentrations. The changes in the lipid composition of both invertebrates were different for neutral lipids and phospholipids. These changes concern the length of the fatty acid chains as well as their unsaturation. The cobalt ions inhibited the oxidation of sterols and their alkylation at C-24. Because of these results we can assume that some of the changes caused by cobalt ions might possess adaptive value.

Refining of high oleic safflower oil: Effect on the sterols and tocopherols content

The purpose of this work was to study the influence of the industrial process steps on free fatty acids, peroxide value (PV), p-anisidine value (PAV), trans fatty acids, tocopherols and sterols (free, esterified and total) in high oleic safflower oil. Degumming, bleaching and deodorization steps removed 91.4% of free fatty acids, 96.31% of oxidation primary products (PV), and 54.57% of oxidation secondary products (PAV), from crude high oleic safflower oil. Degumming neither affected the content of sterified sterols nor its proportion with respect to the crude oil. A significant increment (p<0.05) in the content of free sterols was observed during degumming and bleaching due to the acid-catalyzed hydrolysis of steryl esters. A significant reduction (p<0.05) in the content of total sterols during bleaching was observed, which is attributed to a reduction in the sterified sterol fraction. During deodorization, free sterols were distilled from oil, with a gradual reduction in the total sterol content as a function of the deodorization temperature. α- and γ-tocopherols represented 93.3% of the total tocopherols in high oleic safflower crude oil. The refining process removed 28.5% of the tocopherols. Deodorization was the main step which increased the level of trans fatty acids as an effect of temperature and heating time.