Triacylglycerols (TAGs) containing positional isomers of hypogeic (Hy), palmitoleic (Po), and palmitvaccenic (Pv) acids from three microorganisms (top-fermenting brewer's yeast Saccharomyces cerevisiae, green alga Coccomyxa elongata, and arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis) were analyzed. Dozens of regioisomers and enantiomers of TAGs containing one, two or three hexadecenoic acids have been identified by means of reversed phase chromatography/mass spectrometry (RP-HPLC/MS). The regioisomers of TAGs containing two palmitic acids and any hexadecenoic acid were separated. Analysis of regioisomers of TAGs having one Pv residue showed that asymmetric molecular species such as PvPP or PPPv were dominant in Rhizophagus. TAGs were also analyzed on a chiral phase column and nine molecular species of TAGs containing two palmitic and any of three hexadecenoic acids were separated and identified. In the case of TAGs containing one palmitic and two hexadecenoic acids, the separation was successful only if the hexadecenoic acids were identical. Separation of TAGs containing three hexadecenoic acids was successful only if all three hexadecenoic acids were identical. Regardless of the type of TAG, it was found that TAGs in the AM fungus and containing palmitvaccenic acid bound at the sn-1 position of the glycerol backbone were dominant, suggesting similarity in the biosynthesis of the different TAGs. The covalent adduct chemical ionization method was used for identification of TAGs as adduct with (1-methyleneimino)-1-ethenyl ion, which reacted with double bond of the unsaturated fatty acid. Tandem MS thus makes it possible to identify TAGs containing various hexadecenoic acids.
Membrane phospholipids, including phosphatidylcholine (PC) and phosphatidylethanolamine (PE), consist of distinct fatty acids occupying the sn-1 and sn-2 positions, reflecting the highly regulated nature of lipid biosynthesis. However, little is known about the influence of dietary lipids on the positional nature of fatty acids in tissues, including the enrichment of omega-3 polyunsaturated fatty acid (PUFA) in chicken egg yolk phospholipids. This study was undertaken to characterize the PC and PE species in egg lipids derived from Lohmann hens (n=10/treatment) randomly allocated to either a control (no supplementation), a flaxseed oil (FO) or a marine algal oil (MA) diet. Each of the FO or MA diets supplied three levels of total omega-3 PUFA (0.20, 0.40 and 0.60% of diet) that were provided for 6 weeks. A combination of multiplexed mass spectrometry (MS) experiments are used to determine total, isobaric, and position molecules for PC and PE in egg yolk. The distribution of phospholipids in the yolk was predominantly PC over PE (~72 vs. 23%, respectively) across treatments. The longer chain PUFA existed in the sn-2 position in the PC and PE. Although docosahexaenoic acid (22:6) formed isomers with fatty acids 16:0, 18:0 and 18:1; it was preferentially enriched in the egg in combination with 16:0 with both the FO and MA-fed groups in both lipid pools. All 22:6-containing isomers were enriched by ~2-fold more (P < 0.0001) with MA than FO, however, all isomers exhibited a plateau with the FO-fed group. In addition, the MS analyses of PCs revealed several isobaric species containing eicosapentaenoic acid (EPA, 20:5), however, in the PE, EPA formed only one isomer (i.e. in combination with 16:0). These results may assist to elucidate potential aspects regulating the limited enrichment of omega-3 PUFA, particularly EPA and docosahexaenoic acid (22:6) in chicken eggs.
- MeSH
- Biomass * MeSH
- Phosphatidylcholines metabolism MeSH
- Phosphatidylethanolamines metabolism MeSH
- Animal Feed * MeSH
- Chickens metabolism MeSH
- Lipidomics MeSH
- Linseed Oil pharmacology MeSH
- Fatty Acids, Omega-3 pharmacology MeSH
- Egg Yolk metabolism MeSH
- Animals MeSH
- Check Tag
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
Triacylglycerols (TAGs) containing less common fatty acids (FAs) were isolated from the seeds of three plants (Santalum album, Crepis foetida, and Leucas aspera). These FAs had allenic (laballenic acid, Lb) and acetylenic (crepenynic, C; ximenynic acids, Xi) bonds. TAGs were analyzed on reversed-phase and chiral columns. High-resolution tandem mass spectrometry identified TAGs by positive electrospray ionization (ESI+). Twenty-two molecular species of TAGs isolated from the seed oil of Santalum album were separated by RP-HPLC and chiral HPLC methods and identified by positive electrospray ionization tandem MS detection (ESI+-MS). Two major enantiomers, i.e., sn-OOLb and sn-LLLb (O represents oleic acid; and L represents linoleic acid), were synthesized from the appropriate phosphatidylcholines. This allowed the identification of enantiomers after separation by chiral chromatography by tandem mass spectrometry. Similarly, TAGs from the seeds of Crepis foetida, and Leucas aspera were analyzed by reversed-phase chromatography and identified by mass spectrometry. Four enantiomers (sn-OOC, sn-LLC, sn-OOXi, and sn-LLXi) were synthesized. A total of six and three enantiomers of TAGs containing crepenynic and ximenynic acids, respectively, were identified by chiral column analysis. The retention times of TAGs containing allenic and acetylenic bonds were always greater on the reversed-phase column than TAGs with the same number of carbon atoms and the same unsaturation (e.g., LLL versus LLLb). From the chiral column, the regioisomers and enantiomers were eluted in the order of symmetric-asymmetric-asymmetric (i.e., sn-OCO, sn-COO, and sn-OOC). Through tandem mass spectrometry, we were able to identify and distinguish regioisomer [DAG]+-type ions, i.e., [MNH4NH3RCOOH]+, that can be considered diagnostic. Unfortunately, enantiomers and TAGs with the same numbers of carbon atoms and the same unsaturation levels have identical mass spectra, such as LLL and LLLb.
- MeSH
- Alkynes analysis chemistry MeSH
- Chromatography, Liquid MeSH
- Chromatography, Reverse-Phase MeSH
- Phosphatidylcholines chemistry MeSH
- Spectrometry, Mass, Electrospray Ionization MeSH
- Linoleic Acid analysis MeSH
- Oleic Acids analysis MeSH
- Fatty Acids analysis chemistry MeSH
- Seeds chemistry MeSH
- Stereoisomerism MeSH
- Tandem Mass Spectrometry * MeSH
- Triglycerides analysis chemistry isolation & purification MeSH
- Chromatography, High Pressure Liquid * MeSH
- Publication type
- Journal Article MeSH
Increased activity of the sympathetic nervous system (SNS) has been proposed as a risk factor for increased cardiovascular mortality in patients with chronic kidney disease (CKD). Information on the activity of cardiac sympathetic innervation is non-homogeneous and incomplete. The aim of our study was to evaluate the tonic effect of SNS on heart rate, norepinephrine turnover and direct and indirect effects of norepinephrine in left ventricles of subtotally nephrectomized rats (SNX) in comparison with sham-operated animals (SHAM). Renal failure was verified by measuring serum creatinine and urea levels. SNX rats developed increased heart rates and blood pressure (BP). The increase in heart rate was not caused by sympathetic overactivity as the negative chronotropic effect of metipranolol did not differ between the SNX and SHAM animals. The positive inotropic effects of norepinephrine and tyramine on papillary muscle were not significantly different. Norepinephrine turnover was measured after the administration of tyrosine hydroxylase inhibitor, pargyline, tyramine, desipramine, and KCl induced depolarization. The absolute amount of released norepinephrine was comparable in both groups despite a significantly decreased norepinephrine concentration in the cardiac tissue of the SNX rats. We conclude that CKD associated with renal denervation in rats led to adaptive changes characterized by an increased reuptake and intracellular norepinephrine turnover which maintained normal reactivity of the heart to sympathetic stimulation.
- MeSH
- Renal Insufficiency, Chronic complications metabolism physiopathology MeSH
- Cardiovascular Diseases etiology MeSH
- Kidney metabolism MeSH
- Disease Models, Animal MeSH
- Nephrectomy MeSH
- Neuropeptide Y metabolism MeSH
- Norepinephrine blood MeSH
- Calcitonin Gene-Related Peptide metabolism MeSH
- Rats, Wistar MeSH
- Heart Rate MeSH
- Heart Ventricles metabolism physiopathology MeSH
- Sympathetic Nervous System physiopathology MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
The diatom Phaeodactylum tricornutum was cultivated in a standard medium and under sulfur, silicon, nitrogen and phosphorus starvation and its triacylglycerols (TAGs) were analyzed by RP-HPLC/MS-APCI. Nearly 100 molecular species of polyunsaturated TAGs were identified. RP-HPLC was used to isolate positional isomers of TAGs, which were further separated by chiral HPLC. First eluted were those TAGs that have an eicosapentaenoic acid moiety in the sn-1 position. The ratios of symmetrical to asymmetrical TAGs in P. tricornutum were affected under sulfur-, nitrogen-, phosphorus- and silica-starvation, i.e. in cultivations involving cells in nutrient stress. The ratios of positional TAGs and also the proportions of enantiomers were changed. The ratios of symmetrical to asymmetrical TAGs in the control and under N- and P-starvation were very close. In the control, the ratio of 1,2-dipalmitoyl-3-eicosapentaenoyl-rac-glycerol to 1,3-dipalmitoyl-2-eicosapentaenoyl-rac-glycerol was 3:1 and the ratio of 1,2-dieicosapentaenoyl-3-palmitoyl-rac-glycerol to 1,3-dieicosapentaenoyl-2-palmitoyl-rac-glycerol was 9:1. Under N-starvation the ratios were reversed irrespective of the presence or absence of silicate in the medium. A similar pattern was found in P- and S-starvation.
Fatty acid (FA) and triacylglycerol (TG) composition of natural oils and fats intake in the diet has a strong influence on the human health and chronic diseases. In this work, non-aqueous reversed-phase (NARP) and silver-ion high-performance liquid chromatography with atmospheric pressure chemical ionization mass spectrometry detection and gas chromatography with flame-ionization detection (GC/FID) and mass spectrometry detection are used for the characterization of FA and TG composition in complex samples of animal fats from fallow deer, red deer, sheep, moufflon, wild boar, cock, duck and rabbit. The FA composition of samples is determined based on the GC/FID analysis of FA methyl esters. In total, 81 FAs of different acyl chain length, double bond (DB) number, branched/linear, cis-/trans- and DB positional isomers are identified. TGs in animal fats contain mainly monounsaturated and saturated FAs. High amounts of branched and trans-FAs are observed in the samples of ruminants. In NARP mode, individual TG species are separated including the separation of trans- and branched TGs. Silver-ion mode provides the separation of TG regioisomers, which enables the determination of their ratios. Great differences in the preference of unsaturated and saturated FAs in the sn-2 position on the glycerol skeleton are observed among individual animal fats. Unsaturated FAs are preferentially occupied in the sn-2 position in all animal samples except for wild boar with the strong preference of saturated FAs in the sn-2 position.
- MeSH
- Chromatography methods MeSH
- Dietary Fats MeSH
- Mass Spectrometry methods MeSH
- Ducks MeSH
- Rabbits MeSH
- Chickens MeSH
- Meat analysis MeSH
- Fatty Acids analysis chemistry MeSH
- Sheep MeSH
- Sus scrofa MeSH
- Triglycerides analysis chemistry MeSH
- Deer MeSH
- Animals MeSH
- Check Tag
- Rabbits MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Silver-ion high-performance liquid chromatography (HPLC) coupled to atmospheric pressure chemical ionization mass spectrometry (APCI-MS) is used for the regioisomeric analysis of triacylglycerols (TGs). Standard mixtures of TG regioisomers are prepared by the randomization reaction from 8 mono-acid TG standards (tripalmitin, tristearin, triarachidin, triolein, trielaidin, trilinolein, trilinolenin and tri-gamma-linolenin). In total, 32 different regioisomeric doublets and 11 triplets are synthesized, separated by silver-ion HPLC using three serial coupled chromatographic columns giving a total length of 75cm. The retention of TGs increases strongly with the double bond (DB) number and slightly for regioisomers having more DBs in sn-1/3 positions. DB positional isomers (linolenic vs. γ-linolenic acids) are also separated and their reverse retention order in two different mobile phases is demonstrated. APCI mass spectra of all separated regioisomers are measured on five different mass spectrometers: single quadrupole LC/MSD (Agilent Technologies), triple quadrupole API 3000 (AB SCIEX), ion trap Esquire 3000 (Bruker Daltonics), quadrupole time-of-flight micrOTOF-Q (Bruker Daltonics) and LTQ Orbitrap XL (Thermo Fisher Scientific). The effect of different types of mass analyzer on the ratio of [M+H-R(i)COOH](+) fragment ions in APCI mass spectra is lower compared to the effect of the number of DBs, their position on the acyl chain and the regiospecific distribution of acyl chains on the glycerol skeleton. Presented data on [M+H-R(i)COOH](+) ratios measured on five different mass analyzers can be used for the direct regioisomeric determination in natural and biological samples.
- MeSH
- Spectrometry, Mass, Electrospray Ionization methods MeSH
- Mass Spectrometry methods MeSH
- Isomerism MeSH
- Molecular Weight MeSH
- Triglycerides chemistry MeSH
- Chromatography, High Pressure Liquid methods MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Comparative Study MeSH
