Isolation and identification of mosquito bite deterrent terpenoids from leaves of American (Callicarpa americana) and Japanese (Callicarpa japonica) beautyberry

Essential oil extracts from Callicarpa americana and Callicarpa japonica were investigated. Bioassay-guided fractionation of C. americana extracts using the yellow fever mosquito, Aedes aegypti, led to the isolation of alpha-humulene, humulene epoxide II, and intermedeol and a newly isolated terpenoid (callicarpenal). Similar work involving C. japonica resulted in the isolation of an additional compound, spathulenol, as well as the four compounds isolated from C. americana. Structure elucidation was performed on all isolated compounds using a combination of gas chromatography-mass spectrometry-electron ionization, high-resolution liquid chromatography-MS-electrospray ionization, and one- and two-dimensional NMR experiments. Heretofore, 13,14,15,16-tetranorclerodane, callicarpenal, has never been identified from natural sources. Complete (1)H and (13)C NMR assignment data are provided for this compound. In bite deterrent studies, spathulenol, intermedeol, and callicarpenal showed significant repellent activity against A. aegypti and Anopheles stephensi.     

Variation of chemical composition of the lipophilic extracts from yellow birch (Betula alleghaniensis) foliage

The occurrence of biologically active compounds identified for the first time in the lipophilic extracts of yellow birch (Betula alleghaniensis Britt.) foliage led to the quantification of the seasonal variation of their concentrations. Yellow birch foliage was collected from late June until late September 2003 in two different regions of Quebec. The extraction yields using hexane as a solvent were determined, and the extracts were analyzed by GC-MS to identify their molecular composition. In terms of both extraction yields and the concentration of the targeted molecules present in the extracts, mid-September has been determined as the best time to collect foliage samples. A total of 14 constituents were identified in these extracts. This is the first report of the presence of all of these constituents in yellow birch foliage and of some of them in the genus Betula. The most important compounds identified in yellow birch foliage extracts are triterpene squalene and aliphatic hydrocarbon tetracosan, aliphatic alcohol phytol, fatty acids hexadecanoic and octadecanoic, pentacyclic triterpenes alpha- and beta-amyrin, and phytosterol stigmast-5-en-3-ol.     

Phytochemical and nutrient composition of the freeze-dried amazonian palm berry, Euterpe oleraceae mart. (acai)

Euterpe oleraceae is a large palm tree indigenous to the Amazon River and its tributaries and estuaries in South America. Its fruit, known as acai, is of great economic value to native people. In this study, a standardized freeze-dried acai fruit pulp/skin powder was used for all analyses and tests. Among many findings, anthocyanins (ACNs), proanthocyanidins (PACs), and other flavonoids were found to be the major phytochemicals. Two ACNs, cyandin 3-glucoside and cyanidin 3-rutinoside were found to be predominant ACNs; three others were also found as minor ACNs. The total content of ACNs was measured as 3.1919 mg/g dry weight (DW). Polymers were found to be the major PACs. The concentration of total PACs was calculated as 12.89 mg/g DW. Other flavonoids, namely, homoorientin, orientin, isovitexin, scoparin, and taxifolin deoxyhexose, along with several unknown flavonoids, were also detected. Resveratrol was found but at a very low concentration. In addition, components including fatty acids, amino acids, sterols, minerals, and other nutrients were analyzed and quantified. Total polyunsaturated fatty acid, total monounsaturated fatty acid, and total saturated fatty acids contributed to 11.1%, 60.2%, and 28.7% of total fatty acid. Oleic acid (53.9%) and palmitic acid (26.7%) were found to be the two dominant fatty acids. Nineteen amino acids were found; the total amino acid content was determined to be 7.59% of total weight. The total sterols accounted for 0.048% by weight of powder. The three sterols B-sitosterol, campesterol, and sigmasterol were identified. A complete nutrient analysis is also presented. Microbiological analysis was also performed.     

Dichloromethane as a solvent for lipid extraction and assessment of lipid classes and fatty acids from samples of different natures

The usefulness of the solvent mixture dichloromethane/methanol for lipid extraction and the determination of lipid classes and fatty acids in samples of different natures was conducted. Two different extraction methods were compared, one containing chloroform/methanol, another containing dichloromethane/methanol. Total lipid extraction showed some minor differences but no variation in the lipid classes. Regarding the fatty acid profile, in Echium virescens seeds, 17 major fatty acids could be identified and quantified, and all were equally extracted when either solvent system was employed. In Echium acanthocarpum hairy roots, 17 major fatty acids were quantified, showing some statistical differences for one cell line in favor of chloroform. The data obtained from the liquid nutrient medium were also comparable. The cod roe sample showed 31 major fatty acids, showing no statistical differences between the two solvent systems. Contrarily, the CH 2Cl 2 method was able to extract 31 main fatty acids found in European seabass dorsal muscle more efficiently than the CHCl 3 method. The results indicate that, for lipid extraction and fatty acid assessment, dichloromethane/methanol can readily replace the commonly employed chloroform/methanol, thus avoiding the major health, security, and regulatory problems associated with the use of chloroform.     

Isolation and structure elucidation of antioxidant polyphenols from quince (Cydonia vulgaris) peels

Thirteen new compounds, as well as 16 already known, have been isolated from organic extracts of peels of Cydonia vulgaris, a fruit of a shrub belonging to the same tribe as the apple. All of the structures were elucidated by EI- or ESI-MS and (1)H and (13)C NMR after purification of individual compounds by HPLC. Thirteen fatty acid esters of cinnamyl alcohols, three fatty acid esters of hydroxybenzoic acid, three fatty acid esters of hydroxybenzaldehyde, three glucosides of aromatic acids, four chlorogenic acids, two flavonols, and a benzylamine have been identified. The fatty acid moieties have been identified by GC-MS analysis of the methanolysis products. All of the compounds were tested for their radical scavenging and antioxidant activities by measuring their capacity to scavenge the 2,2'-diphenyl-1-picrylhydrazyl (DPPH) radical and anion superoxide radical and to induce the reduction of Mo(VI) to Mo(V). The chlorogenic acids and the flavonols exhibited more antioxidant and radical scavenger capacity than the positive standards alpha-tocopherol and ascorbic acid. The results of the tests were analyzed by cluster analysis that grouped all of the compounds on the basis of the substituents on the aromatic ring.     

Evaluation of fatty acid extraction methods for Thraustochytrium sp. ONC-T18

Various extraction methods were assessed in their capacity to extract fatty acids from a dried biomass of Thraustochytrium sp. ONC-T18. Direct saponification using KOH in ethanol or in hexane:ethanol was one of the most efficient techniques to extract lipids (697 mg g(-1)). The highest amount of fatty acids (714 mg g(-1)) was extracted using a miniaturized Bligh and Dyer extraction technique. The use of ultrasonics to break down cell walls while extracting with solvents (methanol:chloroform) also offered high extraction yields of fatty acids (609 mg g(-1)). Moreover, when the transesterification mixture used for a direct transesterification method was doubled, the extraction of fatty acids increased approximately 77% (from 392 to 696 mg g(-1)). This work showed that Thraustochytrium sp. ONC-T18 has the ability to produce over 700 mg g(-1) of lipids, including more than 165 mg g(-1) of docosahexaenoic acid, which makes this microorganism a potential candidate for the commercial production of polyunsaturated fatty acids. Finally, other lipids, such as myristic, palmitic, palmitoleic, and oleic acids, were also produced and recovered in significant amounts (54, 196, 123, and 81 mg g(-1)), respectively.     

Polyunsaturated fatty acids (PUFAs) content of the fungus Mortierella alpina isolated from soil

Twenty-five isolates of Mortierella spcies were prepared, which can be used for the production of polyunsaturated fatty acids (PUFAs)-rich oil for nutritional supplements. The fatty acid contents were determined after heterotrophic fermentation. The content of total fatty acids (TFAs) in the cell dry weight of all isolates including two commercially purchased Mortierella alpina strains ranged from 207.51 to 370.11 mg/g, whereas PUFAs were the dominant fatty acid type. The highest PUFA-containing strain, M. alpina SC9, was identified and confirmed as a new strain of M. alpina through comparison analysis of the sequences of internal transcribed spacers 1 and 2 (ITS1 and ITS2) and the 5.8S rDNA region. During a 7-day fermentation, the PUFAs content of M. alpina SC9 varied between 189.83 and 240.00 mg/g, with a remarkable correlation between the oleic acid (C18:1, OA) and arachidonic acid (C20:4n-6, AA) contents and between the linoleic acid (C18:2n-6, LA) and AA contents, suggesting the PUFA content in the fungus is tightly regulated. This study provides a framework of isolation, identification, and characterization of an important PUFA-producing species, M. alpina.     

Fatty acids and fat-soluble vitamins in salted herring (Clupea harengus) products

The fatty acid composition and contents of fat and fat-soluble vitamins of three salted products prepared from Icelandic herring were analyzed. The effects of storage on the products over their shelf life, 6 or 12 months, were investigated. The average oil content of salted, gutted herring and salted fillets in vacuum remained constant, 17 and 12% of wet weight, respectively. In the pickled product the oil content decreased during the 12 months of storage from 13 to 12%. The composition of the products was typical for herring, the most abundant fatty acids being oleic (18:1n-9), palmitic (16:0), cetoleic (22:1n-11), and gadoleic (20:1n-9) acids. Monounsaturated acids constituted clearly the main group with a proportion of >50% of all fatty acids. Eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n-3) comprised together >12% of all fatty acids. During storage, some hydrolysis of triacylglycerol (TAG) occurred, causing a slight reduction in practically all esterified fatty acids. In none of the three products was the loss of polyunsaturated fatty acids from TAG greater than the loss of saturated ones, indicating that the loss of EPA and DHA was not due to oxidation. After packing, the average content of vitamins A, D, and E in the products varied between 27 and 87 microg/100 g (wet weight), between 17-28 microg/100 g (wet weight), and between 77-120 microg/100 g (wet weight), respectively. During storage, the level of vitamin A decreased significantly, whereas no loss of vitamin D was observed. The content of vitamin E was low in all products and showed wide variation. When compared to the recommended daily intake, it could be concluded that the products investigated were good and stable sources of long-chain n-3 fatty acids (EPA, DHA) and vitamin D.     

Lipophilic (hydroxy)phenylacetates by solvent-free lipase-catalyzed esterification and transesterification in vacuo

Various long-chain alkyl (hydroxy)phenylacetates were prepared in high yield by lipase-catalyzed transesterification of the corresponding short-chain alkyl hydroxyphenylacetates and fatty alcohols in equimolar ratios. The reactions were performed in vacuo at moderate temperatures in the absence of solvents and drying agents in direct contact with the reaction mixture. Immobilized lipase B from Candida antarctica (Novozym 435) was the most effective biocatalyst for the various transesterification reactions. Generally, Novozym 435-catalyzed transesterifications of short-chain alkyl (hydroxy)phenylacetates with long-chain alcohols led to higher conversions and enzyme activities than the corresponding esterifications. For example, the transesterification activity was up to 4-fold higher than the esterification activity for the formation of oleyl 4-hydroxy-3-methoxyphenylacetate using Novozym 435 as a biocatalyst. The relative transesterification activities were as follows: phenylacetate > 3-methoxyphenylacetate approximately 4-methoxyphenylacetate > 4-hydroxy-3-methoxyphenylacetate > 3-hydroxyphenylacetate approximately 4-hydroxyphenylacetate > 2-methoxyphenylacetate > 3,4-dihydroxyphenylacetate. With respect to the position of methoxy and hydroxy substituents, the transesterification activity of Novozym 435 decreased in the order meta approximately para > ortho. Compounds with inverse chemical structures, for example, tyrosyl oleate, were obtained by Novozym 435-catalyzed esterification and transesterification of fatty acids and their methyl esters, respectively, with 2-phenylethan-1-ols. In contrast to the transesterifications of short-chain alkyl (hydroxy)phenylacetates with fatty alcohols, higher conversions and enzyme activities were observed for the Novozym 435-catalyzed esterifications of (hydroxy)phenylethanols with long-chain fatty acids than the corresponding transesterifications with fatty acid methyl esters.     

Analysis of polar lipids in the serum from rats fed shiitake by liquid chromatography-mass spectrometry/mass spectrometry

Consumption of a shiitake mushroom diet has been reported to have effects on serum phospholipids. However, much less is known about the effect on serum polar lipids including lysophospholipids and free fatty acids. In the present study, the effects of a shiitake diet were evaluated on the basis of identification and quantification of individual polar lipid components in rat serum using liquid chromatography-mass spectrometry/mass spectrometry. By comparison with standards and published data, 50 lysophospholipids and 32 free fatty acids were identified, and the concentrations of 27 polar lipids in rat serum were determined. Shiitake diets decreased the levels of all individual polar lipid components in the serum of male rat. The total level of serum polar lipids in males fed 4% shiitake diets (1365.71 mol/L) was significantly lower than that of the control (2270.26 mol/L). However, shiitake diets did not significantly affect the levels of serum polar lipids in female rats.     

Volatile fatty acid production with alfalfa (Medicago sativa) and glucose in denitrifying soil at different temperatures

Summary A laboratory study was done to determine how, if at all, temperatures affect the production of volatile fatty acids and the rate of denitrification in soil. Glucose and alfalfa were compared as C substrates at temperatures of -2, 10, and 25°C under anaerobic conditions. At -2°C (soil not frozen), the denitrification rate was slow but was as rapid with alfalfa as glucose. This indicated that the production of volatile fatty acids by fermenters or other C substrates from alfalfa were adequate to sustain denitrifiers. No volatile fatty acids were apparently produced with glucose at -2°C whereas acetate and propionate were produced with alfalfa during the 26-day incubation period. During the 8-day incubation period at 10°C, there were also greater accumulations of acetate and propionate with alfalfa than with glucose. At 25°C, there was no major difference in the denitrification rate between glucose and alfalfa over a 4-day period. In a contrast to the other temperatures, more butyrate than propionate was produced at 25°C, especially with alfalfa. Acetate was the dominant volatile fatty acid produced and generally increased with temperature, especially after NO₃ exhaustion at 10 and 25°C. This indicated that acetate was a source of C for denitrifiers.     

Microbial diversity in three floodplain soils at the Elbe River (Germany)

Microbial communities in floodplain soils are exposed to periodical flooding. A long-term submerged Eutric Gleysol (GLe), an intermediate flooded Eutric Fluvisol (FLe), and a short-time flooded Mollic Fluvisol (FLm) at the Elbe River (Germany) with similar organic carbon contents (C_org) between 8.1% and 8.9% were selected to test the quality of phospholipid fatty acids (PLFA), soil microbial carbon (C_mic), basal respiration (BR), metabolic quotient (qCO₂), and C_mic/C_org ratio to characterize and discriminate these soils with microbial parameters.The three floodplain soils can be differentiated by C_mic and by total PLFA-biomass. Due to the different flooding durations and the time since the soils were last flooded C_mic and PLFA-biomass increase in the order GLe<FLe<FLm. Both parameters correlate significantly (r=0.999;p<0.05). The C_mic/C_org ratios are low in comparison to terrestrial soils and revealed the same ranking over the three soils like C_mic. Contrary, qCO₂ and BR are highest in GLe and lowest in FLm according to inundation regime. The diminished C_mic, high BR, and high qCO₂ values in GLe seem to be an unspecific response of aerobic soil microorganisms on the long flooding period and the resulting short time for developing after last flooding as well as the low pH value. Different plant communities and their residues may influence the microbial diversity additionally.The PLFA profiles were dominated by the group of saturated fatty acids that together constituted almost 62-72% of the total fatty acids identified in the soils. In GLe all groups of PLFA, inclusive monounsaturated fatty acids, are lowest and in FLm highest, while in FLe the PLFA fractions show an intermediary amount of the three soils. The FLm had most of the time aerobic conditions and revealed therefore the highest C_mic, PLFA-biomass, especially monounsaturated fatty acids, C_mic/C_org ratio as well as relatively low BR and qCO₂ value. These indicate that microorganisms in FLm are more efficiently in using carbon sources than those in GLe and FLe.All 26 identified PLFA were found in FLe and FLm, while the polyunsaturated fungi biomarker 18:2ω6,9c could not be detected in GLe. In this long-time submerged soil the environmental conditions which microorganisms are exposed might be disadvantageous for fungi.     

Fatty acid composition of traditional and novel forages

Managing the fatty acid composition of grazing ruminant diets could lead to meat and milk products that have higher conjugated linoleic acid (CLA) concentrations, but forage fatty acid dynamics must be more fully understood for a range of forages before grazing systems can be specified. The fatty acid profiles of 13 different forages, including grasses, legumes, and forbs, grown under greenhouse conditions, were determined. Three separate harvests, at 3-week intervals, were made of each plant material. alpha-Linolenic [C18:3, 7.0-38.4 mg g(-1) of dry matter (DM)], linoleic (C18:2, 2.0-10.3 mg g(-1) of DM), and palmitic (C16:0, 2.6-7.5 mg g(-1) of DM) acids were the most abundant fatty acids in all species at each harvest, together representing approximately 93% of the fatty acids present. Concentrations of fatty acids declined as plants developed, but the fractional contribution of each fatty acid to total fatty acids remained relatively stable over time. Grasses had a uniform composition across species with a mean of 66% of total fatty acids provided by C18:3, 13% by C18:2, and 14% by C16:0. The fractional contribution of C18:3 to total fatty acids was lower and more variable in forbs than in grasses. Intake of fatty acid by grazing ruminants would be affected by the forage species consumed.     

Variability in coconut (Cocos nucifera L.) germplasm and hybrids for fatty acid profile of oil

Coconut oil, the main product of coconut fruit, is the richest source of glycerol and lauric acid and hence is called lauric oil. This paper reports the fatty acid profile of oil from 60 Talls, 14 Dwarfs, and 34 hybrids. These include collections from 13 countries covering a large coconut-growing area of the world, apart from the indigenous ones. Capillary gas chromatography analysis of oil indicated a wider variation for the fatty acid profile than earlier reported. Apart from this, for the first time other fatty acids such as behenic and lignoceric acids were detected. Oil from cultivars and hybrids of coconut has significantly differed, particularly for commercially important fatty acids such as lauric acid and unsaturated fatty acids. However, coconut oil seems to have a conserved fatty acid profile, mainly because of low unsaturated fatty acids, indicating the possibility of grouping cultivars on the basis of their fatty acid profiles. The cluster analysis based on fatty acid profile indicated grouping together of geographically and typically closely related cultivars. Cultivars with high concentrations of specific fatty acids can be of potential use for industrial exploitation, whereas those with high concentrations of short- and medium-chain fatty acids and unsaturated fatty acids are more suitable for human consumption. Cultivars and hybrids with high and low values for each of the fatty acids are also identified.     

Quantitation of fatty acids, sterols, and tocopherols in turpentine (Pistacia terebinthus Chia) growing wild in Turkey

The chemical composition (fatty acids, tocopherols, and sterols) of the oil from 14 samples of turpentine (Pistacia terebinthus L.) fruits is presented in this study. The oil content of the samples varied in a relatively small range between 38.4 g/100 g and 45.1 g/100 g. The dominating fatty acid of the oil is oleic acid, which accounted for 43.0 to 51.3% of the total fatty acids. The total content of vitamin E active compounds in the oils ranged between 396.8 and 517.7 mg/kg. The predominant isomers were alpha- and gamma-tocopherol, with approximate equal amounts between about 110 and 150 mg/kg. The seed oil of P. terebinthus also contained different tocotrienols, with gamma-tocotrienol as the dominate compound of this group, which amounted to between 79 and 114 mg/kg. The total content of sterols of the oils was determined to be between 1341.3 and 1802.5 mg/kg, with beta-sitosterol as the predominent sterol that accounted for more than 80% of the total amount of sterols. Other sterols in noteworthy amounts were campesterol, Delta5-avenasterol, and stigmasterol, which came to about 3-5% of the total sterols.     

Selected nutrient contents, fatty acid composition, including conjugated linoleic acid, and retention values in separable lean from lamb rib loins as affected by external fat and cooking method

Proximate composition and fatty acid profile, conjugated linoleic acid (CLA) isomers included, were determined in separable lean of raw and cooked lamb rib loins. The cooking methods compared, which were also investigated for cooking yields and true nutrient retention values, were dry heating of fat-on cuts and moist heating of fat-off cuts; the latter method was tested as a sort of dietetic approach against the more traditional former type. With significantly (P < 0.05) lower cooking losses, dry heating of fat-on rib-loins produced slightly (although only rarely significantly) higher retention values for all of the nutrients considered, including CLA isomers. On the basis of the retention values obtained, both techniques led to a minimum migration of lipids into the separable lean, which was higher (P < 0.05) in dry heating than in moist heating, and was characterized by the prevalence of saturated and monounsaturated fatty acids. On the whole, the response to cooking of the class of CLA isomers (including that of the nutritionally most important isomer cis-9,trans-11) was more similar to that of the monounsaturated than the polyunsaturated fatty acids.     

Accumulation of Oxygenated Fatty Acids in Oat Lipids During Storage

Oxygenated fatty acids were identified in oat grain by gas chromatography‐mass spectrometry. We hypothesized that most of these were the results of lipoxygenase activity. This hypothesis was tested by measuring concentrations of these compounds after hydrothermal treatments and storage of oat groats or oat flour for 22 weeks at 37°C and 65% relative humidity. Steam treatments inactivated lipases, whereas roasting at 106°C did not. Free fatty acids accumulated quickly in untreated or roasted flour, but not in steamed flour or groats. A total of six hydroxy and epoxy fatty acids were identified. Oxidized fatty acids were found in both esterified lipids and free fatty acids, indicating that lipase action was not necessary for lipid oxidation. More oxidation products were found in flour than in groats, and less were found in the steamed treatments. Lipoperoxygenase appeared to be involved in the formation of oxidation products, although nonenzymatic mechanisms may also operate. Hydroxy‐fatty acids are associated with strongly bitter flavors and are undesirable. Results indicate the importance of enzyme inactivation before storage of processed oat products.     

Fatty acid profile of table olives and its multivariate characterization using unsupervised (PCA) and supervised (DA) chemometrics

The fatty acid composition of 67 commercial presentations of table olives was determined. The most abundant fatty acids, in decreasing order of presence, were C18:1, C16:0, C18:2 n-6, and C18:0. The ranges, expressed as grams of fatty acids per 100 g of edible portion, for the different nutritional fractions were as follows: saturated fatty acids, 2.07-5.99; monounsaturated fatty acids, 5.67-19.42; polyunsaturated fatty acids, 0.52-3.87; and trans-fatty acids, 0.08-0.44. Principal component analysis of the matrix of the fatty acid composition led to the deduction of new factors. The first accounted for 55.10% of the total variance and was mainly related to C16:10, C18:0, C20:0, C22:0, C24:0, C18:1, C18:1t, and C20:1. The second factor accounted for 10.33% of variance and was related to C16:1 and C18:2 n-6. They did not permit differentiation among elaboration types or cultivars. However, discriminant analysis was successfully applied for this objective. The fatty acids that most contributed to discriminate among elaboration styles were C17:1, C18:1, C16:0, C17:0, and C18:0 (function 1) and C17:0, C17:1, C20:0, C16:0, C18:1, and C24:0 (function 2). In the case of cultivars, they were C20:0, C18:1, C17:1, C18:2 n-6, C18:1t, and C18:2t (function 1); C18:2 n-6, C18:1, C16:0, C20:0, C18:0, and C18:2t (function 2); and C17:0, C18:3 n-3, and C17:1 (function 3). Results from this study have shown differences among the fatty acid composition and fat content of the diverse commercial presentations of table olives, which can be applied in predictive and classification discriminant analysis.     

Compartive study of volatile components and fatty acids of plants and in vitro cultures of parsley (Petroselinum crispum (Mill) nym ex hill).

Volatile compounds from plants, callus tissue cultures, and cell suspensions of parsley (Petroselinum crispum) were captured during the growth cycle using a dynamic headspace extraction and were identified by gas chromatography-mass spectrometry. Parsley plants were found to produce mainly monoterpenes, and the compound of major abundance was p-1,3,8-menthatriene, followed by beta-phellandrene and apiole. Callus cultures and cell suspensions produced aldehydes (nonanal and decanal) that were also detected in parsley plant. The former also produced limonene, acetophenone, and benzotiazol; these were not observed in the plants. The production of volatiles in plants, callus tissue, and cell suspensions was found to be time-dependent. Free and bound fatty acids were also monitored by an in situ method. Palmitic (C16:0) and stearic (C18:0) acids were the most abundant fatty acids in all materials; however, higher levels were found in plants. On the other hand, the unsaturated C16:1 and C16:3 were not detected in the in vitro cultures.