Characteristic profiles of lipid classes,fatty acids and triacylglycerol molecular species of peas (Pisum sativum L.)

Seed oils from four legume cultivars of Pisum sativum, grown in Japan, were extracted and classified by thin‐layer chromatography (TLC) into seven fractions: hydrocarbons (HC; 0.5–0.9 wt‐%), steryl esters (SE; 0.8–2.4 wt‐%), triacylglycerols (TAG; 31.2–40.3 wt‐%), free fatty acids (FFA; 1.3–2.7 wt‐%), 1,3‐diacylglycerols (1,3‐DAG; 1.0–1.8 wt‐%), 1,2‐diacylglycerols (1,2‐DAG; 1.0–2.2 wt‐%) and phospholipids (PL; 52.2–61.3 wt‐%). All lipid samples had high amounts of total unsaturated fatty acids, representing 75.0–84.3 wt‐% for TAG and PL. Molecular species and fatty acid distributions of TAG, isolated from the total lipids in the peas, were analyzed by a combination of argentation‐TLC and GC. Eighteen different molecular species were detected. With a few exceptions, the main TAG components were SMD (7.5–10.3 wt‐%), M₂D (8.0–8.9 wt‐%), SD₂ (12.0–18.3 wt‐%), SMT (9.8–11.0 wt‐%), MD₂ (12.0–20.3 wt‐%), SDT (9.7–10.8 wt‐%), M₂T (2.5–7.3 wt‐%) and D₃ (14.5–15.2 wt‐%) (where S denotes a saturated fatty acid, M denotes a monoene, D denotes a diene, and T denotes a triene). It seems that the four cultivars were highly related to each other based on the fatty acid composition of the TAG as well as the distribution profiles in the different TAG molecular species. In general, these results suggest that there are no essential differences (p >0.05) in the oil components among the four cultivars.     

Positional distribution of fatty acids in triacylglycerols and phospholipids from adzuki beans (Vigna angularis)

The fatty acid distributions of triacylglycerols (TAG) and major phospholipids (PL) obtained from adzuki beans (Vigna angularis) were investigated. The total lipids extracted from the beans were separated by thin‐layer chromatography (TLC) into eight fractions. The major lipid components were PL (63.5 wt‐%), TAG (21.2 wt‐%), steryl esters (7.5 wt‐%) and hydrocarbons (5.1 wt‐%), while free fatty acids, diacylglycerols (1,3‐DAG and 1,2‐DAG) and monoacylglycerols were also present in minor proportions (0.2–1.1 wt‐%). The major PL components isolated from the beans were phosphatidylcholine (45.3 wt‐%), phosphatidylethanolamine (25.8 wt‐%) and phosphatidylinositol (21.5 wt‐%). Phosphatidylinositol was unique in that it had the highest saturated fatty acid content among the three PL. With a few exceptions, however, the principal characteristics of the fatty acid distribution in the TAG and three PL were evident in the beans: Unsaturated fatty acids were predominantly concentrated in the sn‐2 position while saturated fatty acids primary occupied the sn‐1 or sn‐3 position in the oils of the adzuki beans. In general, these results could be useful to both consumers and producers for the manufacture of traditional adzuki foods in Japan.     

Regiospecific distribution of fatty acids in triacylglycerols and phospholipids from broad beans (Vicia faba)

Regiospecific distributions of fatty acids of triacylglycerols (TAG) and phospholipids (PL) separated from broad beans (Vicia faba) of four cultivars (Minpo, Sanuki, Nintoku and Sanren) were investigated. The major lipid components were PL (47.5–50.5 wt‐%) and TAG (47.7–50.1 wt‐%), while steryl esters, hydrocarbons, free fatty acids, diacylglycerols and monoacylglycerols were present in minor proportions (1.6–2.4 wt‐%). The PL components isolated from the four cultivars were phosphatidylcholine (56.4–58.4 wt‐%), phosphatidylethanolamine (20.3–21.7 wt‐%) and phosphatidylinositol (16.6–18.6 wt‐%). Phosphatidylinositol was unique in that it had the highest saturated fatty acid content among these PL. The principal characteristics of the fatty acid distribution in the TAG and PL were evident in the beans: Unsaturated fatty acids were predominantly concentrated in the sn‐2 position while saturated fatty acids primarily occupied the sn‐1 or sn‐3 position in these lipids. The lipid components and fatty acid distributions were almost the same in the four cultivars and were not influenced by genetic variability and planting location. These results could be useful information to both consumers and producers for the manufacture of traditional broad bean foods in Japan.     

Triacylglycerols from viper bugloss (Echium vulgare L.) seed bio‐oil

Triacylglycerols (TAG) in viper bugloss oil were isolated from raw pressed oil by silicic acid column chromatography. The obtained blend of TAG was separated by silver ion thin‐layer chromatography (TLC Ag⁺) into nine fractions, varying in terms of unsaturation level and molecular polarity. The composition of TAG in viper bugloss oil was determined by HPLC coupled with a diode‐array detector and an evaporative light‐scattering detector. The results showed that the first three fractions were combinations of TAG containing palmitic, oleic and linoleic acids. Fractions 4 and 6 contained TAG of a similar acid composition as above, but with the addition of γ‐linolenic acid. The remaining fractions (7–9) were the most varied in acid composition. They were found to contain 26–39% palmitic acid, 12–15% oleic acid, 13–41% linoleic acid 8–24% γ‐linolenic acid, 1.5–5.5% α‐linolenic acid and 1–5% stearidonic acid. The analysis of fatty acid allocation in TAG of viper bugloss lipids revealed that linoleic acid (ranging from 2 to 100%) was the only acid found in all isolated fractions. In the investigated oil, the predominant TAG included: LnLnG (11.38%), LnLnSt (11.17%), LnGSt (7.71%), LnStSt (6.19%) and LnLnLn (5.44%). Almost 86% of the TAG contained α‐linolenic acid, while γ‐linolenic and stearidonic acids amounted to 49 and 38%, respectively.     

Characteristic distributions of fatty acids in different lipids from Jack beans (Canavalia gladiata DC.)

Extracted lipids obtained from Jack beans (white and red) were fractionated by TLC into nine subfractions. The major components were TAGs (TAG: 43.8–45.7 wt%) and phospholipids (PL: 46.7–47.0 wt%), while other components were also present in minor proportions (0.3–2.7 wt%). The principal fatty acids (FA) are generally palmitic (18.8–28.8%), stearic (0.7–6.8%), oleic (42.0–51.8%), linoleic (16.2–22.8%), and α‐linolenic (3.0–8.2%) acids, the distribution of which differs according to these lipid classes. There were no significant differences (p>0.05) in the positional distribution of FA in the TAG; unsaturated FA (97.5%) were predominantly concentrated in the sn‐2 position while saturated FA (33.3%) primarily occupied the sn‐1 position or sn‐3 position. However, significant differences (p<0.05) in FA distribution existed when the individual PL were compared between the white and red beans. Based on the FA composition of these lipids, it seems that the two cultivars of Jack beans are very similar to each other with a few exceptions. The results could be useful to both producers and consumers for our daily diet to improve value of the Japanese diet. Practical applications : The lipid composition suggests that these beans could be a good source of nutraceuticals with providing heath benefits. The white and red beans may be well incorporated into our daily Japanese diets to improve nutritional value. The data obtained in this study provide valuable information for manufacturing functional drinks such as Jack bean tea in Japan.     

Triacylglycerol composition and structure in genetically modified sunflower and soybean oils

Changes in composition were examined in oils extracted from genetically modified sunflower and soybean seeds. Improvements were made to the analytical methods to accomplish these analyses successfully. Triacylglycerols (TAG) were separated on two 300 mm × 3.9 mm 4μ Novapak C18 high-performance liquid chromatography (HPLC) columns and detected with a Varex MKIII evaporative light-scattering detector. Peaks were identified by coelution with known standards or by determining fatty acid composition of eluted TAG by capillary gas chromatography (GC). Stereospecific analysis (fatty acid position) was accomplished by partially hydrolyzing TAG with ethyl magnesium bromide and immediately derivatizing the resulting diacylglycerols (DAG) with (S)-(+)-1-(1-naphthyl)ethyl isocyanate. The derivatized sn-1,2-DAG were completely resolved from the sn-2,3-DAG on two 25 mm × 4.6 mm 3 μ silica HPLC columns. The columns were chilled to −20°C to obtain baseline resolution of collected peaks. The distribution of fatty acids on each position of the glycerol backbone was derived from the fatty acid compositions of the two DAG groups and the unhydrolyzed oil. Results for the sn-2 position were verified by hydrolyzing oils with porcine pancreatic lipase, isolating the resulting sn-2 monoacylglycerols by TLC, and determining the fatty acid compositions by GC. Results demonstrated that alterations in the total fatty acid composition of these seed oils are determined by the concentration of TAG species that contain at least one of the modified acyl groups. As expected, no differences were found in TAG with fatty acid quantities unaffected by the specific mutation. In lieu of direct metabolic or enzymatic assay evidence, the authors’ positional data are nevertheless consistent with TAG biosynthesis in these lines being driven by the mass action of available acyl groups and not by altered specificity of the acyltransferases, the compounds responsible for incorporating fatty acids into TAG.     

Optimization of the synthesis of lower glycerides rich in unsaturated fatty acid residues obtained via enzymatic ethanolysis of sesame oil

The lipases Novozym 435, Lipozyme TL IM and Lipozyme RM IM were employed in the production of lower acylglycerols (LG), i.e. mono‐ (MAG) and diacylglycerols (DAG), rich in unsaturated fatty acids from sesame oil in batch reactors. The effect of the molar ratio of ethanol to fatty acids on the reusability of these immobilized lipases was studied in detail. The effects of pretreatment on lipase activity for ethanolysis were investigated. Glycerol had a strong product inhibition effect on the ethanolysis reaction, and a relatively large excess of ethanol was necessary to remove the glycerol adsorbed on these biocatalysts. The enzymatic activity was drastically reduced by addition of water to the reaction medium. The presence of organic solvents (hexane and acetone) did not favor the production of LG. For the Novozym 435‐catalyzed reaction, optimum conditions were a molar ratio of ethanol to fatty acid residues of 5 : 1, 15 wt‐% lipase and 50 °C. For Lipozyme TL IM, the optimum conditions were a molar ratio of ethanol to fatty acid residues of 5 : 1, 20 wt‐% biocatalyst, and 30 °C. Novozym 435 and Lipozyme TL IM produced LG with molar ratios of unsaturated to saturated fatty acids of 20.4 in 1 h and 25.3 in 5 h, respectively. In the original oil, this ratio was 5. For trials conducted under optimum conditions, the products from the Novozym 435 trials contained 21.8 wt‐% triacylglycerols (TAG), 24 wt‐% DAG and 54.2 wt‐% MAG. The products of the Lipozyme TL IM trials consisted of 12.9 wt‐% DAG and 87.1 wt‐% MAG. No TAG species were detected.     

Chemical and enzymatic interesterification of a beef tallow and rapeseed oil equal‐weight blend

A mixture of beef tallow and rapeseed oil (1:1, wt/wt) was interesterified using sodium methoxide or immobilized lipases from Rhizomucor miehei (Lipozyme IM) and Candida antarctica (Novozym 435) as catalysts. Chemical interesterifications were carried out at 60 and 90 °C for 0.5 and 1.5 h using 0.4, 0.6 and 1.0 wt‐% CH₃ONa. Enzymatic interesterifications were carried out at 60 °C for 8 h with Lipozyme IM or at 80 °C for 4 h with Novozym 435. The biocatalyst doses were kept constant (8 wt‐%), but the water content was varied from 2 to 10 wt‐%. The starting mixture and the interesterified products were separated by column chromatography into a pure triacylglycerol fraction and a nontriacylglycerol fraction, which contained free fatty acids, mono‐, and diacylglycerols. It was found that the concentration of free fatty acids and partial acylglycerols increased after interesterification. The slip melting points and solid fat contents of the triacylglycerol fractions isolated from interesterified fats were lower compared with the nonesterified blends. The sn‐2 and sn‐1,3 distribution of fatty acids in the TAG fractions before and after interesterification were determined. These distributions were random after chemical interesterification and near random when Novozym 435 was used. When Lipozyme IM was used, the fatty acid composition at the sn‐2 position remained practically unchanged, compared with the starting blend. The interesterified fats and isolated triacylglycerols had reduced oxidative stabilities, as assessed by Rancimat induction times. Addition of 0.02% BHA and BHT to the interesterified fats improved their stabilities.     

Acylglycerols Containing Trihydroxy Fatty Acids in Castor Oil and the Regiospecific Quantification of Triacylglycerols

Ricinoleate, a monohydroxy fatty acid in castor oil, has many industrial uses. Dihydroxy and trihydroxy fatty acids can also be used in industry. We report here the identification of diacylglycerols (DAG) and triacylglycerols (TAG) containing trihydroxy fatty acids in castor oil. The C₁₈ HPLC fractions of castor oil were used for mass spectrometry of the lithium adducts of acylglycerols to identify trihydroxy fatty acids and the acylglycerols containing trihydroxy fatty acids. Two DAG identified were triOH18:1–diOH18:1 and triOH18:0–OH18:1. Four TAG identified were triOH18:1–OH18:1–OH18:1, triOH18:0–OH18:1–OH18:1, triOH18:1–OH18:1–diOH18:1 and triOH18:0–OH18:1–diOH18:1. The structures of these two newly identified trihydroxy fatty acids were proposed as 11,12,13-trihydroxy-9-octadecenoic acid and 11,12,13-trihydroxyoctadecanoic acid. The locations of these trihydroxy fatty acids on the glycerol backbone were almost 100% at the sn-1,3 positions or at trace levels at the sn-2 position. The content of these acylglycerols containing trihydroxy fatty acids was at the level of about 1% or less in castor oil.     

<Superscript>1</Superscript>H- and <Superscript>13</Superscript>C-NMR Characterization of the Molecular Components of the Lipid Fraction of Pecorino Sardo Cheese

In this work the molecular fatty components of Pecorino Sardo Protected Designation of Origin (PS PDO) cheese were characterized through an exhaustive investigation of the ¹H- and ¹³C-NMR spectra of the extracted lipids. Several fatty acids (FA), such as long chain saturated, oleic, linoleic, linolenic, butyric, capric, caprylic, caproic, trans vaccenic, conjugated linoleic acid (cis9, trans11–18:2), and caproleic (9–10:1) were unambiguously detected. The positional isomery of some acyl groups in the glycerol backbone of triacylglycerols (TAG) was assessed. Furthermore, the NMR signals belonging to sn-1,2/2,3, sn-1,3 diacylglycerols (DAG), and free fatty acids (FFA) were analysed as a measure of lipolytic processes on cheese. Lastly, ¹H-NMR resonances of saturated aldehydes and hydroperoxides were detected, their very low intensity indicating that the lipid oxidation process can be considered to be of minor relevance in Pecorino Sardo cheese.     

Preparation of stearidonic acid‐enriched triacylglycerols from Echium plantagineum seed oil

HPLC analysis of Echium plantagineum seed oil shows a complex triacylglycerol (TAG) profile. TAG species were separated on an analytical scale by HPLC and their fatty acid (FA) composition is reported. GLC analyses showed that some TAG fractions reached a stearidonic acid (SDA, 18:4n‐3) percentage significantly higher than that in the original oil. TAG separation on a bigger scale was also essayed, by means of a gravimetric normal‐phase chromatographic column, using silver ion‐silica gel as stationary phase. Gradient elution with solvents of increasing polarity was applied, allowing the separation of valuable TAG species containing γ‐linolenic acid (GLA, 18:3n‐6), α‐linolenic acid (ALA, 18:3n‐3) and SDA as the main constituents (more than 85% of the total FA). An enzymatic hydrolysis reaction showed the distribution of FA in the isolated species of TAG. SDA was the major FA in the sn‐2 position (more than 50% of total FA), followed by ALA (19%) and GLA (18.5%).     

Changes in olive oil composition due to microwave heating

The effects of microwave heating on some components of extra-virgin olive oil were studied. Traditional parameters, including free acidity, peroxide value and ultraviolet absorbance values at 232 and 268 nm, were determined in six extra-virgin olive oil samples before and after the microwave treatment. Significant differences (P<0.01) were detected for free acidity, peroxide, and ultraviolet absorbance at 268 nm; also, the absorbances at 232 nm showed significant differences (P<0.05) between treated and untreated samples. The glycerolic fractions, triacylglycerols (TAG), diacylglycerols (DAG), and monoacylglycerols (MAG), were isolated by thin-layer chromatography. The respective percentage fatty acid (FA) composition and percentage amount were obtained by high-resolution gas chromatography with an internal standard. For the most abundant TAG fraction, the stereospecific analysis was carried out to obtain the FA percentage compositions of the three sn-positions. Small but significant modifications were observed regarding the decrease in the TAG percentage and increases in the DAG and MAG percentage amounts. No significant changes were observed for the FA compositions of TAG, DAG, and MAG fractions before and after the treatment. Nevertheless, the results of TAG stereospecific analysis showed losses of unsaturated FA in all sn-positions. Higher percentage changes in the sn-1- than in sn-2-position of TAG were observed. Regarding the volatile fraction, different profiles were obtained after the treatment.     

Molecular species of triacylglycerols in the hulls of sunflower seeds (Helianthus annuus L.) following microwave treatment

Whole sunflower seeds (Helianthus annuus L.) were exposed to microwaves for 6, 12, 20 or 30 min at a frequency of 2450 MHz. The hulls were then stripped from the seeds. Molecular species and fatty acid distributions of triacylglycerols (TAGs), isolated from total lipids in the hulls, were analyzed by a combination of argentation thin‐layer chromatography (TLC) and gas chromatography. A modified argentation TLC procedure, developed to optimize the separation of the TAGs, provided 10 different groups of TAGs, based on both the degree of unsaturation and the total fatty acid chain‐length. Dilinoleolein (29.5—30.2 wt‐%), trilinolein (18.2—24.2 wt‐%), dilinoleopalmitin and dilinoleostearin (17.0—18.1 wt‐%), palmitoleolinolein and stearoleolinolein (11.4—14.0 wt‐%) and dioleolinolein (7.5—8.6 wt‐%) were the main TAGs detected after microwave roasting. However, roasting caused a significant decrease (p < 0.05), not only in TAG molecular species containing more than four double bonds, but also in the amounts of diene species present in TAGs. These results suggest that microwaves should affect TAGs in the hulls more significantly (p < 0.05) than those in the sunflower kernels.     

Ratios of Regioisomers of the Molecular Species of Triacylglycerols in Lesquerella (Physaria fendleri) Oil Estimated by Mass Spectrometry

The ratios of regioisomers of 72 molecular species of triacylglycerols (TAG) in lesquerella oil were estimated using the electrospray ionization mass spectrometry of the lithium adducts of TAG in the HPLC fractions of lesquerella oil. The ratios of ion signal intensities (or relative abundances) of the fragment ions from the neutral losses of fatty acids (FA) as α‐lactones at the sn‐2 position (MS³) of the molecular species of TAG were used as the ratios of the regioisomers. The order of the preference of FA incorporation at the sn‐2 position of the molecular species of TAG in lesquerella was as: normal FA > OH18 (monohydroxy FA with 18 carbon atoms) > diOH18 > OH20 > diOH20, while in castor was as: normal FA > OH18 > OH20 > diOH18 > triOH18. Elongation (from C₁₈ to C₂₀) was more effective than hydroxylation in lesquerella to incorporate hydroxy FA at the sn‐1/3 positions. The block of elongation in lesquerella may be used to increase the content of hydroxy FA, e.g., ricinoleate, at the sn‐2 position of TAG and to produce triricinolein (or castor oil) for industrial uses. The content of normal FA at the sn‐2 position was about 95 %, mainly oleate (38 %), linolenate (31 %) and linoleate (23 %). This high normal FA content (95 %) at the sn‐2 position was a big space for the replacement of ricinoleate to increase the hydroxy FA content in lesquerella oil. The content of hydroxy FA at the sn‐1/3 positions was 91 % mainly lesquerolic acid (85 %) and the content of normal FA was 6.7 % at the sn‐1/3 position in lesquerella oil.     

Positional‐Species Composition of Triacylglycerols from the Arils of Mature Euonymus Fruits

Positional‐species composition (PSC) of 1,2,3‐triacyl‐sn‐glycerols (TAG) from the arils of mature fruits of 13 species of Euonymus L. genus was established. The residues of six major fatty acids (FA), palmitic, stearic, hexadecenoic (H), octadecenoic (O), linoleic (L), and linolenic, were present in the TAG. PSC of TAG was determined by their partial lipase hydrolysis. By using hierarchical cluster and principal component analyses, it was definitely demonstrated that separate taxonomic units forming this genus were significantly distinguished as regards PSC of TAG. In particular, the Euonymus subgenus greatly exceeded the Kalonymus subgenus in both total content of L in TAG and in the rate of its incorporation into their mid‐position, while TAG of Kalonymus were marked by a prevalence of O‐TAG and sn‐2‐O isomers. Thus, these subgenera were significantly distinct in the rate of incorporation of O and L residues in the sn‐2 position of TAG molecules. Meanwhile, the TAG from the Euonymus section species were marked by an enhanced concentration of H and the incorporation of H in UUU TAG was much more active than in other TAG types. As for positional‐type composition of TAG, saturated FA were always virtually absent in the sn‐2 position of Euonymus aril TAG.     

Variations in the composition of acyl lipids and triacylglycerol molecular species of pumpkin seeds (Cucurbita spp.) following microwave treatment

Whole pumpkin seeds (Cucurbita spp.) of two cultivars were exposed to microwaves for 6, 12, 20 or 30 min at a frequency of 2450 MHz. The kernels were separated from the whole seeds, and were investigated not only for the different acyl lipids and their fatty acid compositions, but also for the molecular species of triacylglycerols (TAGs). A modified argentation TLC procedure, developed to optimize the separation of the complex mixture of total TAGs, provided 11 different groups of TAGs, based on both the degree of unsaturation and the chain‐length of fatty acid groups. With a few exceptions, dioleopalmitin (5.8–18.8 wt‐%), dipalmitolinolein (8.1–8.8 wt‐%), triolein (6.3–20.5 wt‐%), palmitoleolinolein (15.0–16.1 wt‐%), dioleolinolein (16.7–23.0 wt‐%), dilinoleopalmitin (4.6–15.4 wt‐%) and dilinoleolein (6.7–19.4 wt‐%) were the main TAG components. When pumpkin seeds were microwaved for 20 min or more, significant differences (p <0.05) occurred in the acyl lipids as well as their fatty acid distributions with a few exceptions. Therefore, microwave roasting caused a significant decrease (p <0.05), not only in TAGs molecular species containing more than 4 double bonds, but also in the amounts of diene species present in triacylglycerols. These results contribute to the study of the functional properties of pumpkin seed products.     

Preparation of Infant Formula Fat Analog Containing Capric Acid and Enriched with DHA and ARA at the sn-2 Position

An infant formula fat analog with capric acid mostly esterified at the sn‐1,3 positions, and substantial amounts of palmitic, docosahexaenoic (DHA), and arachidonic (ARA) acids at the sn‐2 position, was prepared by physically blending enzymatically synthesized structured lipids (SL) with vegetable oils. The components of the blend included high sn‐2 palmitic acid SL enriched with capric acid (SL_CA), canola oil (CAO), corn oil (CO), high sn‐2 DHA (DHAO_m), and high sn‐2 ARA (ARAO_m) enzymatically modified oils. Each component was proportionally blended to match the fatty acid profile of commercial fat blends used for infant formula. The infant formula fat analog (IFFA₁) was characterized for total and positional fatty acids (FA), triacylglycerol (TAG) molecular species, thermal behavior, and tocopherol content. IFFA₁ contained 17.37 mol% total palmitic acid of which nearly 35 % was located at the sn‐2 position. The total capric acid content was 13.93 mol%. The content of DHA and ARA were 0.49 mol% (48.18 % at sn‐2) and 0.57 mol% (35.80 % at sn‐2), respectively. The predominant TAG were OPO (24.09 %), POP (15.70 %), OOO (11.53 %), and CLC (7.79 %). The melting completion and crystallization onset temperatures were 18.65 and ?2.19 °C, respectively. The total tocopherol content was 566.45 μg/g. This product might be suitable for commercial production of infant formulas.     

Quantification of the Molecular Species of TAG and DAG in Lesquerella (Physaria fendleri) Oil by HPLC and MS

Ten diacylglycerols (DAG) and 74 triacylglycerols (TAG) in the seed oil of Physaria fendleri were recently identified by high‐performance liquid chromatography (HPLC) and mass spectrometry (MS). These acylglycerols (AG) were quantified by HPLC with evaporative light scattering detector and electrospray ionization mass spectrometry of the lithium adducts of the AG in the HPLC fractions of lesquerella oil. The MS¹ ion signal intensities of molecular ions [M + Li]⁺ in HPLC fractions of an HPLC peak were used to estimate the ratios of AG in the HPLC peak. The ratios of TAG with the same mass in HPLC fractions were estimated by the ratios of the sums of MS² ion signal intensities from the neutral loss of the three fatty acids [M + Li ? FA]⁺. The ratio of DAG with the same mass were estimated by the ratio of the sums of two MS² ion signal intensities [M + Li ? FA]⁺ and [FA + Li]⁺ from the two different FA of a DAG. We have estimated the contents of ten molecular species of DAG and 74 molecular species of TAG in P. fendleri oil using this new method. The content of ten DAG combined was about 1 % and 74 TAG was about 98 %. The contents of DAG in decreasing order were: LsLs (0.25 %), LsLn (0.25 %), LsO (0.24 %), and LsL (0.11 %); and the contents of TAG in decreasing order were: LsLsO (31.3 %), LsLsLn (24.9 %), LsLsL (15.8 %), LsL‐OH20:2 (4.3 %), LsO‐OH20:2 (2.8 %), and LsLn‐OH20:2 (2.5 %).     

Analysis of regioisomers of triacylglycerols of northern currant seed oil by tandem mass spectrometry

Currant oils have special health properties due to their moderate contents of α‐linolenic, γ‐linolenic and stearidonic acids. The distribution of fatty acids (FA) in the triacylglycerols (TAG) may affect the beneficial effects. Seed oils of wild northern red currant (NRC) (Ribes spicatum L.) from Northern Finland and of wild alpine currant (AC) (R. alpinum L.) from the South‐West coast of Finland were investigated. The purified TAG were analysed by tandem mass spectrometry by applying the ammonia negative ion chemical ionisation – collision‐induced dissociation method. Molecular weight fractions rich in C18:3 FA and C18:4 FA were investigated. Of the total oil, the molecular weight species 54:7 (ACN:DB), 54:8 and 54:9 were more abundant in NRC than in AC, being 21.0%, 15.8%, 7.4% and 16.2%, 11.2%, 4.8%, respectively (p <0.05). The species 52:6 was more abundant in AC (3.1%) than in NRC (2.6%) (p <0.05). The preferential order of FA to be in the sn‐2 position in both berries was typically C18:1 > C18:2 > C18:4 > C18:3. No difference was observed between relative locations of C16:0 FA and C18:3 FA in either of the oils. Within the TAG consisting of FA combinations C18:3/C18:3/C18:1 (54:7), C18:1 was more preferentially in the sn‐2 position (p <0.05) in AC (93.2%) than in NRC (74.6%), and in the case of C18:3, the preference was vice versa. Within the molecular weight species 54:9, FA combination C18:4/C18:3/C18:2, linoleic acid preferentially occupied the secondary position (p <0.005) in both berries, and the proportion of the TAG regioisomer pair sn‐C18:3‐C18:4‐C18:2 + sn‐C18:2‐C18:4‐C18:3 was more abundant (30.2%) in NRC than in AC (15.3%). Within the TAG species 52:6, proportions of all the existing combinations, C16:0/C18:3/C18:3, C16:0/C18:4/C18:2 and C16:1/C18:3/C18:2, varied between the two berry species (p <0.005).     

Positional distribution of Δ5-olefinic acids in triacylglycerols from conifer seed oils: General and specific enrichment in the sn-3 position

Triacylglycerols (TAG) were purified from the storage lipids extracted from the seeds of several conifer species (Taxus baccata, Larix decidua, Sciadopytis verticillata, and Juniperus communis), each species belonging to one of the four families Taxaceae, Pinaceae, Taxodiaceae, and Cupressaceae, respectively. Each species was characterized by a high content of 5,9-18:2, 5,9,12-18:3, 5,11,14-20:3, or 5,11,14,17-20:4 acids, respectively. TAG were partially deacylated with ethylmagnesium bromide, and the resulting 1,2-, 2,3-diacylglycerols (DAG), and 2-monoacylglycerols (MAG) were purified by thin-layer chromatography. 1,2- and 2,3-DAG were further fractionated by chiral column high-performance liquid chromatography of the 3,5-dinitrophenylurethane derivatives. Alternately, TAG were subjected to porcine pancreatic lipase, and the resulting 2-MAG were purified for further analysis. Gas-liquid chromatography of fatty acid methyl esters prepared from the separated DAG and MAG, coupled with appropriate calculations, indicated that the Δ5-olefinic acids, irrespective of the species, chainlengths and number of ethylenic bonds, were considerably enriched in the sn-3 position of TAG where they accounted for ca. 35 to 74 mole% of fatty acids esterified to this position (depending on the initial level of total Δ5-olefinic acids in TAG), which corresponded to 79–94% of Δ5-olefinic acids esterified to the three positions. On the other hand, Δ5-olefinic acids were less than 10% in the sn-2 position and less than 6% in the sn-1 position of TAG. This specific enrichment of Δ5-olefinic acids in the sn-3 position thus appears to be a general characteristic of conifer seed TAG. These results were extended to TAG from the seeds of two pine species (Pinus koraiensis and P. pinaster) that are rich in Δ5-olefinic acids and available commercially on a ton-scale.