Determination of fatty acid and triacylglycerol composition of human very-low-density lipoproteins

The fatty acid composition of human very-low-density lipoproteins (VLDL) was studied in a population from western Andalusia with a diet in which the fat content came mainly from olive oil. The lipid composition of VLDL, including the fatty acid composition of the phospholipids and triacylglycerols, was examined by capillary gas chromatography. Twenty-five peaks were resolved, ranging in chain length from 14 to 24 carbon atoms, including geometric and positional isomers. The major fatty acids present in phospholipids were 16:0, 18:0, 18:1(n − 9) and 18:2(n − 6), and in triacylglycerols were 18:1(n − 9), 16:0 and 18:2(n − 6). The major triacylglycerol was POO, followed by PLO and OOO. MLP, PPS and LLL were absent. The presence of a large amount of OOO in this fraction demonstrates that the triacylglycerol composition of the VLDL depends on the type of diet consumed.     

Short-term energy deficit causes net accumulation of linoleoyl-enriched triacylglycerols in rat liver

Energy depletion by reduced food intake over 4 days resulted in a 73% reduction in total rat liver triacylglycerols (TG). In liver TG of energy-depleted rats, dilinoleoyl oleoyl glycerol (OLL) and trilinoleoyl glycerol (LLL)) were quantitatively increased by 85% and 147%, respectively. The net increase in linoleoyl-enriched species could be quantitatively accounted for by the release of linoleate from monolinoleoyl species and its subsequent reacylation into dilinoleoyl species and trilinolein during energy depletion. Hence while palmitate, oleate and some linoleate are being hydrolyzed, presumably for oxidation some linoleate is retained and contributes to the remodelling of hepatic triacylglycerols during energy deficit.     

Differences in the Rates of Metabolism of Various Triacylglycerols during Seed Germination and the Subsequent Growth of Seedlings of Dioscorea tokoro, a Perennial Herb

Differences in the rates of metabolism of various triacylglycerols(TAGs) in the reserve oil of seeds during the course of germinationand seedling growth were studied in Dioscorea tokoro, an EastAsian perennial herb. Eleven molecular species of TAG were determinedin the reserve oil of seeds. These TAGs contained five fattyacids: palmitic acid (P); stearic acid (S); oleic acid (O);linoleic acid (L); and linolenic acid (Ln). At 20°C, levelsof TAGs with one or less than one L moiety, such as OLO, OOO,PLO, POO, OLS, and OLnO, decreased rapidly during germinationand the initial elongation of seedlings, whereas levels of otherTAGs decreased slowly during the subsequent development of seedlings.Even though seeds were unable to germinate at these temperatures,levels of some TAGs decreased during incubation at 5°C andat 30°C. Such changes might be related to the metabolismrequired for the survival of seeds at these temperatures. Itis also possible that they might be involved in the breakingof dormancy at low temperatures or in the induction of secondarydormancy at high temperatures. The present results provide anexample of the selective utilization of the TAGs in the reserveoil of seeds during germination and the subsequent growth ofseedlings. (Received October 3, 1995; Accepted January 31, 1996)     

Simultaneous determination of molecular species of monoacylglycerols,diacylglycerols and triacylglycerols in human very-low-density lipoproteins by reversed-phase liquid chromatography

The aim of the present study was to investigate the applicability of a previously developed method for the analysis of triacylglycerol molecular species to the simultaneous determination of triacylglycerols, diacylglycerols and monoacylglycerols of human very-low-density lipoproteins (VLDL). Ten elderly women were recruited for the study. Blood was obtained in fasting conditions and VLDL were isolated by ultracentrifugation. Neutral lipids were separated by solid-phase extraction and were subsequently injected on a reversed-phase HPLC system, with an elution system composed of acetone in acetonitrile. The method allowed the separation of four monoacylglycerols, 18 diacylglycerols and 24 triacylglycerols, including the resolution of positional isomers of diacylglycerols. Monoacylglycerols were composed of oleic, linoleic, palmitic and stearic acids. The major diacylglycerols were 1,2-dilinoleoyl-glycerol and 1,3-dilinoleoyl-glycerol (14.24+/-1.02 and 17.93+/-1.42%, respectively). The main triacylglycerols quantified were dioleoyl-stearoyl-glycerol (OOS), oleoyl-dipalmitoyl-glycerol (OPP), trilinoleoyl-glycerol (LLL) and linoleoyl-distearoyl-glycerol (LSS), accounting for 11.25+/-2.15, 10.14+/-2.05, 9.35+/-2.30 and 8.56+/-1.56%, respectively. An inverse relationship between polarity and fatty acid disappearance from triacylglycerols (r(2)=0.82, P<0.05) and from diacylglycerols (r(2)=0.93, P<0.01) was discovered. In conclusion, the method allowed, for the first time, the easy, rapid and simultaneous determination in a single chromatogram of triacylglycerol, diacylglycerol and monoacylglycerol molecular species of human VLDL by reversed-phase HPLC.     

Triacylglycerols composition, oxidation and oxidation compounds in camellia oil using liquid chromatography-mass spectrometry

Camellia seed oil is one of most important edible oil, rich in oleic acid and contains many natural antioxidants with various biological activities. During preparation of foods or storage camellia oil oxidizes by the auto-oxidation and produce oxidized compounds. Traditional analytical techniques like FFA, POV are used for the determination of oxidation and adulteration of oils and fats. These methods were rarely able to detect the oxidized compounds produced and extent of oxidation. This paper presents the uses of liquid chromatography coupled to electrospray ionization mass spectrometry (HPLC-ESI-MS) for the analysis of triacylglycerols (TAGs) composition and evaluation of auto-oxidation and oxidation products of camellia seed oil. The camellia oil was auto-oxidized for 12 months at room temperature. The TAGs were identified from their characteristics fragmentations such as protonated molecular ion, ammonium and sodium adducts, diacylglycerols, epoxy-diacylglycerols fragments and mono-acylglycerol fragments in ESI-MS mass spectra. HPLC-ESI-MS data revealed the separation and identification of 15 TAGs. The major TAGs separated and identified in camellia seed oil were POO, OOO, OLO, PLO/POL, OLL, SOO, ALO and OLLn. The auto-oxidation studies revealed a total loss of LnLLn, LnOLn, LLLn and OLLn amounting about 13.5% total oxidation. The auto-oxidation products were epoxy hydroperoxides, epoxy epidioxides, and mono-epoxides. It was observed that these were characteristic compounds produced in high oleic oils.     

Bile acids inhibit secretion of very low density lipoprotein by rat hepatocytes

The influence of taurocholate on very low density lipoprotein (VLDL) triacylglycerol synthesis and secretion was studied by isolated rat liver-parenchymal cells. The incorporation of [3H]glycerol into cell-associated and VLDL triacylglycerols were measured after incubation in medium containing 0.75 mM oleate. Taurocholate caused a maked decrease in VLDL [3H]triacylglycerol secretion from the hepatocytes: 50-150 microM taurocholate inhibited secretion of VLDL [3H]triacylglycerols by 70-90%. Similar results were obtained when the mass of secreted VLDL triacylglycerols was measured. Taurocholate caused a decreased secretion of VLDL [3H]triacylglycerols after 15-30 min incubation. A higher amount of cellular triacylglycerols was found in taurocholate-supplemented cells. Furthermore taurocholate did not change the intracellular lipolysis of triacylglycerols. These results suggest that bile acids interfere more probably with the assembly and/or secretion of VLDL-particles and not with earlier stages of VLDL formation, e.g. triacylglycerol synthesis.     

我国大豆油、花生油、芝麻油的甘油三酯组成研究

应用高效液相色谱技术,分离分析大豆油、花生油、芝麻油的甘油三酯组成,测试表明,大豆油的主要甘油三酯为LLL、LLO、LLP、OLO和PLO;花生油的主要甘油三酯为OLO、LLO、PLO;芝麻油的主要甘油三酯为LLO、OLO、POO和LLL。     

Triacylglycerol biosynthesis in everted sacs of rat intestinal mucosa.

The molecular specificity of the biosynthesis of triacylglycerols by rat intestinal mucosa was examined by means of radioactive and mass tracers, and thin-layer chromatography with silver nitrate and gas-liquid chromatography with radioactivity monitoring. Bile salt micelles of alternately labeled monoacylglycerols and free fatty acids were incubated with everted sacs of intestinal mucosa for various periods of time and the triacylglycerols isolated by solvent extraction and thin-layer chromatography. Analyses of the molecular species of the triacylglycerols labeled from monoacylglycerols showed that the 2-monoacylglycerol pathway was responsible for the biosynthesis of a maximum of 90% and the X-1-monoacylglycerol pathway for about 10% of the total radioactive triacylglycerols. Detailed analyses of the molecular species of triacylglycerols labeled fro free fatty acids showed that the phosphatidic acid pathway contributed a minimum of 20-30% of the total labeled triacylglycerol formed. There was a preferential utilization in triacylglycerol biosynthesis of the more unsaturated diacylglycerols arising from the monoacylglycerol pathway and of the more saturated diacylglycerols originating from the phosphatidic acid pathway. The above experiments do not allow a demonstration of the utilization of the sn-2,3-diacylglycerols in triacylglycerol biosynthesis but are not inconsistent with it.     

Characterization of seed oils from fresh Bokbunja (Rubus coreanus Miq.) and wine processing waste

The physicochemical characteristics, fatty acid (FA) profile, and triacylglyceride (TAG) composition of seed oils from fresh Bokbunja (Rubus coreanus Miq.) fruits and traditional Bokbunja wine processing waste were determined in this study. Oil contents of the fresh seeds and the seeds from wine processing waste were similar, accounting for about 18% of dry weight. The free fatty acid (FFA) content between the two seed oils was significantly different (0.50% for fresh seed oil and 73.14% for wine seed oil). Iodine, conjugated diene, saponification values, and unsaponifiable matter were very similar in the oil samples, but the specific extinction coefficients at 232 and 270 nm of wine seed oil were higher than those of fresh seed oil. Linoleic (C18:2, 50.45-53.18%, L) and linolenic (C18:3, 29.36-33.25%, Ln) acids were the dominant FAs in the two seed oils, whereas oleic (C18:1, 7.32-8.04%, O), palmitic (C16:0, 1.55-1.65%, P), and stearic (C18:0, 0.65-0.68%, S) acids were the minor FAs. LLL, OLL, LLLn, OOL, LLnLn, and OOO were the abundant TAGs, representing >90% of the oils.     

Salt tolerance of a cash crop halophyte Suaeda fruticosa: biochemical responses to salt and exogenous chemical treatments

Suaeda fruticosa Forssk is a leaf succulent obligate halophyte that produces numerous seeds under saline conditions. Seeds are a good source of high quality edible oil and leaves are capable of removing substantial amount of salt from the saline soil besides many other economic usages. Little is known about the biochemical basis of salt tolerance in this species. We studied some biochemical responses of S. fruticosa to different exogenous treatments under non-saline (0 mM), moderate (300 mM) or high (600 mM) NaCl levels. Eight-week-old seedlings were sprayed twice a week with distilled water, hydrogen peroxide (H₂O₂, 100 μM), glycine betaine (GB, 10 mM), or ascorbic acid (AsA, 20 mM) for 30 days. At moderate (300 mM) NaCl, leaf Na⁺, Ca²⁺ and osmolality increased, along with unchanged ROS and antioxidant enzyme activities, possibly causing a better plant growth. Plants grew slowly at 600 mM NaCl to avoid leaf Na⁺ buildup relative to those at 300 mM NaCl. Exogenous application of distilled water and H₂O₂ improved ROS scavenging mechanisms, although growth was unaffected. ASA and GB alleviated salt-induced growth inhibition at 600 mM NaCl through enhancing the antioxidant defense system and osmotic and ion homeostasis, respectively.     

Application of atmospheric pressure chemical ionization liquid chromatography–mass spectrometry in identification of lymph triacylglycerols

Atmospheric pressure chemical ionization liquid chromatography–mass spectrometry was used in the identification of triacylglycerol molecular species in lymph samples from rats given either a structured lipid or safflower oil. The structured lipid was MLM-type (M, medium-chain fatty acid; L, long-chain fatty acid) and manufactured from caprylic acid (8:0) and the oil (safflower oil or high-oleic sunflower oil). The triacylglycerol composition of lymph varied significantly between structured triacylglycerols and safflower oil. Diacylglycerol fragment ions were found for all triacylglycerols and we could also observe the ammonium adduct molecular ion [M+NH₄]⁺ for all the triacylglycerols at the selected conditions. Protonated molecular ions were formed from triacylglycerols containing unsaturated fatty acids, and fatty acid fragment ions were also observed in the case of strong fragmentation. The lymph triacylglycerols were identified from their ammonium adduct molecular ions and diacylglycerol fragment ions. In addition to the intact MLM-type structured triacylglycerols, the MLL- and LLL-type triacylglycerols were also identified. The absorption pathway of MLM-type structured triacylglycerols is most likely the same as that of conventional long-chain triacylglycerols, i.e. they were hydrolyzed into 2-monoacylglycerol and medium-chain fatty acids, which were then used for resynthesis of triacylglycerols. The present study thereby also demonstrates the possibility to study the absorption pathway of triacylglycerol via identification of triacylglycerol species in biological samples.     

Sodium chloride effects on lipase activity in germinating rape seeds

Seeds of rape (Brassica napus L.) were germinated at various NaCl concentrations up to 200 mM. Germinating levels, seedling growth, triacylglycerol mobilization and lipase activity were investigated. High salt concentrations resulted in retardation of seed germination. Seedling growth as measured by radicle length was severely reduced by NaCl doses higher than 50 mM. Moreover, the mobilization of storage oil in control rapeseed seedlings, started about 24 h after imbibition. As for germination and growth, elevated salt concentrations are found to delay triacylglycerol degradation. Experiments using triolein as substrate indicated clearly that lipase activity was inhibited by salt treatment.     

Stimulation of intermolecular ligation with E. coli DNA ligase by high concentrations of monovalent cations in polyethylene glycol solutions.

In the presence of high concentrations of the nonspecific polymer polyethylene glycol (PEG), intermolecular cohesive-end ligation with the DNA ligase from Escherichia coli was stimulated by high salt concentrations: 200 mM NaCl or 300 mM KCl in 10% (w/v) PEG 6000 solutions, and 100-200 mM NaCl or 150-300 mM KCl in 15% PEG 6000 solutions. Intermolecular blunt-end ligation with this ligase was also stimulated at 100-150 mM NaCl or 150-250 mM KCl in 15% PEG 6000 solutions. The extent of such intermolecular ligation increased and the salt concentrations at which ligation was stimulated extended to lower concentrations when we raised the temperature from 10 to 37 degrees C.     

Enhanced tolerance to salinity following cellular acclimation to increasing NaCl levels in Medicago truncatula

Salinity is a major abiotic stress that limits plant productivity. Plants respond to salinity by switching on a coordinated set of physiological and molecular responses that can result in acclimation. Medicago truncatula is an important model legume species, thus understanding salt stress responses and acclimation in this species is of both fundamental and applied interest. The aim of this work was to test whether acclimation could enhance NaCl tolerance in calli of M. truncatula. A new protocol is described incorporating multi-step up acclimation over 0–350 mM exogenous NaCl. By the end of the experiment, calli were tolerant to 150 mM and competent for embryogenesis at 100 mM NaCl. Positive and negative linear relationships between Na⁺ and K⁺ uptake and exogenous NaCl concentration intercepted at 160 mM suggesting a Na⁺/K⁺ homeostasis. Proline level peaked at 100/150 mM whilst highest osmolarity and lowest water content occurred at 250/350 mM NaCl. The concentration of water soluble sugars was positively related to 0–250 mM NaCl whilst callus growth and embryogenesis occurred regardless of endoreduplication. Expression of genes linked to growth (WEE1), in vitro embryogenesis (SERK), salt tolerance (SOS1), proline synthesis (P5CS) and ploidy level (CCS52 and WEE1) peaked at 100/150 mM NaCl. Hence, these genes and various physiological traits except sugar levels, served as useful markers of NaCl tolerance. To our knowledge, this is the first report of a multi-step acclimation conferring tolerance to 150 mM NaCl in leaf-derived calli of M. truncatula.     

Effects of nitric oxide on the germination of cucumber seeds and antioxidant enzymes under salinity stress

Effects of exogenous nitric oxide (NO) on the germination and antioxidant enzyme during cucumber seed germination were investigated under salt stress. Seeds of cucumber (Cucumis sativus L. cv. Jinyou 1) were treated with distilled water or NaCl in the presence or absence of NO donor sodium nitroprusside (SNP) during germination. Excess 50 mM NaCl reduced significantly the seed germination rate in a short term and speed of germination. When salt concentration increased, germination of cucumber seed was reduced and the time needed to complete germination lengthened. Addition of exogenous SNP in salt solution attenuated the salt stress effects in a dose-dependent manner, as indicated by accelerating the seed germination, as well as weight increase of budding seeds, and 50 μM SNP was optimal concentration. At 150 mM NaCl, the 50 μM exogenous SNP significantly increased the activities of superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6) and protein content, while decreased the contents of malondialdehyde (MDA). There were no obvious effects of exogenous NO on peroxidase (POD, EC 1.11.1.6) and ascorbate peroxidase (APX, EC 1.11.1.6) activities under salt stress. Exogenous NO also increased the SOD and CAT isozyme expression under salt stress, which was in accordance with the improved antioxidant activities in the germinating seeds. The NO-induced salt stress resistance was associated with activated enzymes, and enhanced protein content, thus decreasing MDA content. It is concluded that exogenous NO treatment on cucumber seeds may be a good option to improve seed germination under saline conditions.     

Triacylglycerol lipase activity in the rabbit renal medulla

Although the renal medulla is rich in triacylglycerols, the lipolysis of these intracellular triacylglycerols by a renomedullary triacylglycerol lipase has not been directly demonstrated. The present study demonstrates triacylglycerol lipase activity localized in the particulate subcellular fractions of rabbit renal medullae. Renomedullary triacylglycerol lipase activity, as determined by the hydrolysis of [14C]triolein to [14C]oleic acid, was observed to have a pH optimum of 5.8. Addition of cAMP/ATP/magnesium acetate resulted in an 80% activation of crude homogenate triacylglycerol lipase activity; addition of exogenous cAMP-dependent protein kinase resulted in a further activation of lipolysis. 3 mM CaCl2 had no effect on basal triacylglycerol lipase activity. 1 M NaCl did not inhibit lipolysis, suggesting that the lipase activity measured was not due to lipoprotein lipase. Endogenous renomedullary triacylglycerols were hydrolysed by a lipase in the 100,000 X g pellet of renomedullary homogenates, resulting in the release of free fatty acids including arachidonic and adrenic acids. Dispersed renomedullary cells were prepared to monitor hormone-sensitive triacylglycerol lipase activity in intact cells. Addition of 10 microM forskolin and 10 microM epinephrine resulted in 8-fold and 50-fold increases in triacylglycerol lipase activity, respectively, as defined by release of free glycerol from the cells. These studies demonstrate that a cAMP-dependent hormone-sensitive triacylglycerol lipase is present in the renal medulla, and is responsible for the hydrolysis of renomedullary triacylglycerols.     

Photosynthetic pigments,ionic and antioxidative behaviour of hulled tetraploid wheat in response to NaCl

Salt stress causes extensive losses to agricultural crops, including wheat, throughout the world and has been the focus of wide research. Though, information is scarce on the potential of ancient wheat relatives in tackling this major limiting factor. Thus, six hulled tetraploid wheat genotypes (HW) were compared to a free-threshing durum wheat genotype (FTW) under different NaCl concentrations, ranging from 0 to 150 mM, at early growth stages in a sand culture experiment. Salt stress induced significant declines in the leaf chlorophyll (Chl) a, Chl b, total Chl, and carotentoid contents; the extent of the declines was greater in FTW compared to HW. Mean leaf proline (3.6-fold) and Na⁺ (1.58-fold) concentrations and Na⁺/K⁺ (2.48-fold) drastically increased with 150 mM of NaCl; the magnitude of the increases was greater in HW compared to FTW. While the carotenoids concentration decreased with progressive salinity both in HW and FTW, the activities of antioxidant enzymes, i.e., catalase, ascorbate peroxidase, and peroxidase were reduced in FTW, but remained unchanged in HW. The above responses to 150 mM NaCl were associated with a significant decrease in shoot dry mass of FTW and lack of significant changes in that of HW. Findings of the present study could help pave the way for further studies on physiological and molecular mechanisms of salt tolerance in these durum wheat relatives.     

Low-affinity Na+ uptake in the halophyte Suaeda maritima

Na(+) uptake by plant roots has largely been explored using species that accumulate little Na(+) into their shoots. By way of contrast, the halophyte Suaeda maritima accumulates, without injury, concentrations of the order of 400 mM NaCl in its leaves. Here we report that cAMP and Ca(2+) (blockers of nonselective cation channels) and Li(+) (a competitive inhibitor of Na(+) uptake) did not have any significant effect on the uptake of Na(+) by the halophyte S. maritima when plants were in 25 or 150 mM NaCl (150 mM NaCl is near optimal for growth). However, the inhibitors of K(+) channels, TEA(+) (10 mM), Cs(+) (3 mM), and Ba(2+) (5 mM), significantly reduced the net uptake of Na(+) from 150 mM NaCl over 48 h, by 54%, 24%, and 29%, respectively. TEA(+) (10 mM), Cs(+) (3 mM), and Ba(2+) (1 mm) also significantly reduced (22)Na(+) influx (measured over 2 min in 150 mM external NaCl) by 47%, 30%, and 31%, respectively. In contrast to the situation in 150 mm NaCl, neither TEA(+) (1-10 mM) nor Cs(+) (0.5-10 mM) significantly reduced net Na(+) uptake or (22)Na(+) influx in 25 mM NaCl. Ba(2+) (at 5 mm) did significantly decrease net Na(+) uptake (by 47%) and (22)Na(+) influx (by 36% with 1 mM Ba(2+)) in 25 mM NaCl. K(+) (10 or 50 mM) had no effect on (22)Na(+) influx at concentrations below 75 mM NaCl, but the influx of (22)Na(+) was inhibited by 50 mM K(+) when the external concentration of NaCl was above 75 mM. The data suggest that neither nonselective cation channels nor a low-affinity cation transporter are major pathways for Na(+) entry into root cells. We propose that two distinct low-affinity Na(+) uptake pathways exist in S. maritima: Pathway 1 is insensitive to TEA(+) or Cs(+), but sensitive to Ba(2+) and mediates Na(+) uptake under low salinities (25 mM NaCl); pathway 2 is sensitive to TEA(+), Cs(+), and Ba(2+) and mediates Na(+) uptake under higher external salt concentrations (150 mM NaCl). Pathway 1 might be mediated by a high-affinity K transporter-type transporter and pathway 2 by an AKT1-type channel.     

Alleviation of Seed Dormancy in the Desert ForbZygophyllum simplexL. from Pakistan

Zygophyllum simplexL. is a succulent annual that grows on thecoastal and inland saline flats around Karachi, Pakistan. Theseeds are moderately salt tolerant during germination. GerminationofZygophyllum simplexseeds under various salinity, proline,betaine, GA and kinetin treatments was determined. Proline (0.1and 1 mM) and betaine (0.1 and 1 mM) alleviated the innate dormancyof seeds, and germination reached 60–70% compared to 12%in the control set. At low salinity compatible osmotica alleviatedsome effects of salinity, but at higher NaCl concentrationsboth proline and betaine were ineffective. Gibberellic acid(0.3 and 3 mM) and kinetin (0.05 and 0.5 mM) substantially alleviatedboth innate as well as salinity-induced seed dormancy. At highersalinity (125 mM), low concentrations of kinetin (0.05 mM) andhigh concentrations of GA (3 mM) were more effective. GA completelyalleviated the effect of salinity at all concentrations used. Betaine; desert; dormancy; forb; GA; germination; halophyte; kinetin; proline; seeds; Zygophyllum simplex     

Cyanophycin and glycogen synthesis in a cyanobacterial Scytonema species in response to salt stress

The response to salinity of a Scytonema species isolated from the central Australian desert was studied. Under nitrogen-fixing conditions the addition of increasing concentrations of salt (NaCl) caused progressive inhibition of growth, with growth ceasing at 150 mM NaCl. This correlated with a progressive loss of nitrogenase activity, a low level of activity being retained at 150 mM NaCl. The inhibition of growth was overcome when KNO₃ (10 mM) was added to the growth medium. In response to the salt stress, cells accumulated the reserve compounds cyanophycin and glycogen. Time course experiments showed that they were steadily synthesized over 48 h, after which the concentrations stabilized. Cyanophycin synthesis was enhanced in salt-stressed cells grown in nitrate. When cells were restored to their normal growth medium the content of these substances decreased towards control levels.