Tg Fraction Research Articles

Functional properties of cream and butter oil from milk of Holstein cows abomasally infused with increasing amounts of high-oleic sunflower fatty acids.

This research paper addresses the hypothesis that there is an optimal amount of intestinally available oleic acid (provided via abomasal infusion) to produce higher-oleic acid milk fat with satisfactory functional characteristics of cream and butter oil. A control and four increasing doses of free fatty acids from high oleic sunflower oil (HOSFA) were infused into the abomasum of four lactating dairy cows in a crossover experimental design with 7-d periods. Treatments were: (1) control (no HOSFA infused), (2) HOSFA (250 g/d), (3) HOSFA (500 g/d), (4) HOSFA (750 g/d), and (5) HOSFA (1000 g/d). All treatments included meat solubles and Tween 80 as emulsifiers. Viscosity, overrun and whipping time as well as foam firmness and stability were evaluated in whipping creams (33% fat). Solid fat content (from 0 to 40°C), melting point and firmness were determined in butter oil. Whipping time of cream increased linearly and viscosity decreased linearly as infusion of HOSFA increased. Overrun displayed a quadratic response, decreasing when 500 g/d or more was infused. Foam firmness and stability were not affected significantly by HOSFA. For butter oil, melting point, firmness, and solid fat content decreased as HOSFA infusion increased. Changes in 21 TG fractions were statistically correlated to functional properties, with 6-10 fractions showing the highest correlations consistently. Decisions on the optimal amount of HOSFA were dependent on the dairy product to which milk fat is applied. For products handled at commercial refrigeration temperatures, such as whipping cream and butter oil, the 250 g/d level was the limit to maintain satisfactory functional qualities. Palmitic acid needed to be present in at least 20% in milk fat to keep the functional properties for the products.

Are thyroid hormone levels and sarcopenia in elderly patients associated with type 2 diabetes mellitus and euthyroidism?

Aim to study the relationship between thyroid function parameters (free T3 (FT3), free T4 (FT4), FT3/FT4 ratio), thyroid-stimulating hormone (TSH) and parameters characterizing sarcopenia (muscle mass and strength, physical performance) in patients with type 2 diabetes mellitus (T2DM) and euthyroidism.
 Material and methods. The study included 312 elderly patients with T2DM in a state of euthyroidism (mean age 65.39 2.28 years). In all participants the following parameters were measured: anthropometric parameters, thyroid hormones (FT3, FT4), TSH, fasting blood glucose, glycated hemoglobin (HbA1c), fasting C-peptide and insulin, lipid profile, body composition, handgrip strength, walking speed.
 Results. Sarcopenia was prevalent in 26.9% of T2DM patients with euthyroidism. A binary logistic regression analysis adjusted for age, gender, smoking and alcohol consumption, T2DM duration, hypertension, HbA1c level, eGFR, TG, HDL and fat mass fraction showed that the high-normal FT3 level (OR = 0.522, 95% CI: 0.3040.895, p = 0.018), a low-normal FT4 level (OR = 1.126, 95% CI: 1.0091.258, p = 0.034) and an increased FT3/FT4 ratio (OR = 0.923, 95% CI: 0.8790.969, p = 0.001) were associated with a low prevalence of sarcopenia. FT3 concentration was positively associated with muscle strength (OR = 0.525, 95% CI: 0.3050.902, p = 0.020) and walking speed (OR = 0.443, 95% CI: 0.2590.758, p = 0.003), while FT4 concentration was negatively associated with muscle mass (OR = 1.114, 95% CI: 1.0091.232, p = 0.036). The FT3/FT4 ratio was positively associated with muscle mass (OR = 0.943, 95% CI: 0.9050.981, p = 0.006), muscle strength (OR = 0.945, 95% CI: 0.9010.992, p = 0.021), and walking speed (OR = 0.934, 95% CI: 0.8940.975, p = 0.002). TSH levels were not associated with sarcopenia.
 Conclusion. The high FT3/FT4 ratio is significantly associated with a low risk of sarcopenia in T2DM patients with euthyroidism.

Quercus based coffee-like beverage: effect of roasting process and functional characterization

Coffee is one of the world’s most widely consumed beverages but intake it is not encouraged in consumers with some health conditions. In this way, with the growing interest in developing healthier substitutes, a coffee-like beverage obtained from Quercus ilex and Quercus suber acorn´s species, was elaborates maintaining the flavor. Although, this beverage is a promising coffee alternative, little is known about effects of roasting process in its composition. To that end the antioxidant capacity and toxicity of the developed coffee were analyzed and phenolic compounds and fatty acids (esterified and free forms) were characterized through HPLC-DAD and GC-FID, respectively. The results showed that Quercus based beverages presented antioxidant capacity related to their phenolic content, mainly to ellagic acid as the primary phenolic compound identified. Due to this composition, the beverage also presented antimutagenic activity. The main fatty acids in the esterified lipids were mainly oleic, linoleic, palmitic, stearic and cis vaccenic. In the free fatty acids fraction (FFA) they were oleic, linoleic and palmitic acids. Heat processing produced a reduction in total fatty acid concentration in TG and FFA fraction of Q. ilex. Nevertheless, for Q. suber alterations were only found for FFA. Coffee production did not form genotoxic or cytotoxic compounds. Overall, these results show the feasibility of Quercus acorn-based foodstuffs and its potential to produce a functional coffee-like beverage.

Analysis of plasma n-terminal pro-brain natriuretic peptide levels and its influencing factors of patients with metabolic syndrome

Objective To analyze influence factors of the level of plasma n-terminal pro-brain natriuretic peptide(NT-proBNP) in patients with metabolic syndrome. Methods Two hundred cases patients with the metabolic syndrome in KaiLuan General Hospital from April 2012 to April 2015 were randomly selected as the observation group, and 200 cases non-metabolic syndrome patients were randomly selected as the control group.The plasma NT-proBNP levels of the two groups were compared.Pearson correlation analysis and multiple linear regression analysis test were used to analyze the influence factors of the level of plasma NT-proBNP in patients with metabolic syndrome. Results The level of plasma NT-proBNP of the observation group and the control group were (79.23±20.16) pg/L and (102.25±27.26) pg/L respectively, the observation group was significantly lower than the control group, and the difference had statistical significance(t=4.154, P<0.05). The pearson correlation analysis showed that in observation group the plasma NT-proBNP level were positively correlated with the age, Cr, UA, aortic root diameter(AOD), left ventricular mass(LVM), left atrium diameter(LAD), left ventricular end-diastolic diamete (LVIDd), end-diastolic wall thickness after(LVPWd)(r=0.376, 0.467, 0.167, 0.237, 0.392, 0.484, 0.374, 0.234; P<0.05), and negatively correlated with BMI, TG, TC, LDL-C, Hb and left ventricular ejection fraction(LVEF)(r=-0.167, -0.253, -0.187, -0.342, -0.354, -0.563; P<0.05). The multivariate Logistic regression analysis showed that in observation group, age(OR=1.236, 95%CI 1.072-2.726, P<0.05), BMI(OR=2.256, 95%CI 1.346-2.983, P<0.05), SBP(OR=1.687, 95%CI 1.183-3.209, P<0.05), DBP(OR=1.125, 95%CI 1.052-2.037, P<0.05), FPG(OR=1.887, 95%CI 1.495-3.494, P<0.05), TG(OR=1.504, 95%CI 1.256-2.609, P<0.05), LVMI(OR=1.983, 95%CI 1.363-2.574, P<0.05) and LVEF(OR=1.485, 95%CI 1.147-2.485, P<0.05) were independent factors affecting the level of plasma NT-proBNP. Conclusion The level of plasma NT-proBNP in patients with metabolic syndrome is significantly lower than the patients without metabolic syndrome, the reason may be related to the increase in body mass index. Key words: Metabolic syndrome; N-terminal pro-brain natriuretic peptide; Factors

Ursolic acid from Trailliaedoxa gracilis induces apoptosis in medullary thyroid carcinoma cells.

Medullary thyroid carcinoma (MTC) originates from the C-cells of the thyroid and is not sensitive to radiation or chemotherapy. Therefore, surgical removal of the tumor tissue in its entirety is the only curative treatment for MTC. The present study aimed to examine the potential mechanisms of action of extracts of Trailliaedoxa gracilis (TG; WW Smith & Forrest), a plant from the province of Sichuan, China, and of ursolic acid (UA), a pentacyclic triterpen present in TG, on the MTC-SK MTC cell line. A total of 13 TG fractions and UA were examined in vitro for their effects on cell morphology, cell number, proliferation and rates of apoptosis. Reverse transcription-quantitative polymerase chain reaction of nuclear factor-κB essential modifier (NEMO) was performed to delineate the role of the apoptotic pathway following treatment with UA. TG and UA were examined in vivo in xenotransplanted MTC-bearing severe combined immunodeficient mice. The TG fractions exhibited antiproliferative effects, with inhibition of mitochondrial activity in the tumor cells at concentrations, which caused no impairment of the normal control cells. The apoptotic rates of the MTC-SK cells treated with the TG fractions and UA were determined, in which no marked tumor inhibition was observed in the treated MTC-mice, and no change in the expression of NEMO was detected in the treated MTC-SK cells. The observation of early-onset activation of caspase 8 suggested that the responsible factor was linked to NEMO, an anti-apoptotic protein. However, no differences in the mRNA transcription levels of NEMO were detected in MTC-SK cells treated with UA, suggesting that this protein was not associated with the signal transducer and activator of transcription 3 pathway.

Blood Immunological and Biochemical Indicators in Turkey Hens Fed Diets With a Different Content of the Yeast Yarrowia Lipolytica

Abstract The aim of this study was to determine immunological and biochemical blood indicators of turkey hens administered feed mixtures with 3 or 6% of Yarrowia lipolytica strain yeast as a dietary nutrient. The experiment was carried out on 240 turkey hens, aged from 1 to 16 weeks. The hens were randomly assigned to 3 experimental groups of 80 birds. Group I served as a control (K) and did not receive any experimental compounds. The turkey hens from experimental groups (YL3, YL6) were administered dried Yarrowia lipolytica yeast in two doses: 3% (YL3) and 6% (YL6) in feed mixtures. The study showed that the addition of Yarrowia lipolytica yeast in a dose of 3% but mainly in a dose of 6% stimulated the body’s immune defence mechanisms, which was evidenced by the increase in plasma lysozyme, % KF, IF, and reduction of monocyte ratio H/L in turkey hens. The advantage of using Yarrowia lipolytica in the nutrition of turkey hens was also a decrease in the content of blood indicators of lipid peroxidation such as CHOL, TG and LDL-cholesterol fraction, and an increase in the percentage of HDL-cholesterol fraction. The use of yeast component in the feeding of turkey hens affected the health status of birds and contributed to proper (not deviating from the reference values) biochemical indicators of metabolism.

Fractionation and characterization of industrial lignins

Three industrial lignins (Indulin AT Kraft softwood, Protobind 1000, and corn stover) were fractionated in methanol to obtain soluble and insoluble fractions. The original lignin and obtained lignin fractions were characterized chemically by a combination of FTIR spectroscopy, pyrolysis-gas chromatography–mass spectrum (py-GC–MS), and derivatization followed by reductive cleavage (DFRC). The methanol soluble lignin fractions contained fewer condensed structures than the original lignin and methanol insoluble fractions. The thermal (e.g. glass transition temperature (Tg) and thermal stability) and rheological properties of the lignins were characterized by a combination of DSC, TMA, TGA and parallel plate rheology. TMA and modulated temperature DSC (MTDSC) proved to be sensitive techniques in determining Tg. The methanol soluble lignin fractions had the lowest Tg values relative to the original lignin and methanol insoluble fractions. The original lignin and methanol insoluble fractions had a higher thermal degradation temperature relative to the methanol soluble lignin. Solvent partitioning offers a practical approach to fractionate lignin into low Mw and Tg fractions which show promise as starting materials for making lignin-copolymers.

Oxidoreductase Interactions Include a Role for ERp72 Engagement with Mutant Thyroglobulin from the rdw/rdw Rat Dwarf

Newly synthesized thyroglobulin (Tg), the secretory glycoprotein that serves as precursor in thyroid hormone synthesis, normally forms transient covalent protein complexes with oxidoreductases of the endoplasmic reticulum (ER). The Tg-G2320R mutation is responsible for congenital hypothyroidism in rdw/rdw rats, in which a lack of secondary thyroid enlargement (goiter) implicates death of thyrocytes as part of disease pathogenesis. We found that mutant Tg-G2320R was retained within the ER with no detectable synthesis of thyroxine, had persistent exposure of free cysteine thiols, and was associated with activated ER stress response but incomplete ER-associated degradation (ERAD). Tg-G2320R associated with multiple ER resident proteins, most notably ERp72, including covalent Tg-ERp72 interactions. In PC Cl3 thyrocytes, inducible overexpression of ERp72 increased the ability of cells to maintain Tg cysteines in a reduced state. Noncovalent interactions of several ER chaperones with newly synthesized Tg-G2320R diminished over time in parallel with ERAD of the mutant protein, yet a small ERAD-resistant Tg fraction remained engaged in covalent association with ERp72 even 2 days post-synthesis. Such covalent protein aggregates may set the stage for apoptotic thyrocyte cell death, preventing thyroid goiter formation in rdw/rdw rats.

Contrasting effects of t10,c12- and c9,t11-conjugated linoleic acid isomers on the fatty acid profiles of mouse liver lipids.

The purpose of this study was to examine the effects of two purified isomers of CLA (c9,t11-CLA and t10,c12-CLA) on the weights and FA compositions of hepatic TG, phospholipids, cholesterol esters, and FFA. Eight-week-old female mice (n = 6/group) were fed either a control diet or diets supplemented with 0.5% c9,t11-CLA or t10,c12-CLA isomers for 8 wk. Weights of liver total lipids and those of individual lipid fractions did not differ between the control and the c9,t11-CLA groups. Livers from animals fed the t10, c12-CLA diet contained four times more lipids than those of the control group; this was mainly due to an increase in the TG fractions (fivefold), but cholesterol (threefold), cholesterol esters (threefold), and FFA (twofold) were also significantly increased. Although c9,t11-CLA did not significantly alter the weights of liver lipids when compared with the control group, its intake was associated with significant reductions in the weight percentage (wt% of total FAME) of 18:1n-9 and 18:1n-7 in the TG fraction and with significant increases in the weight percentage of 18:2n-6 in the TG, cholesterol ester, and phospholipid fractions. On the other hand, t10,c12-CLA intake was linked with a significant increase in the weight percentage of 18:1n-9 and a decrease in that of 18:2n-6 in all lipid fractions. These changes may be the result of alterations in the activity of delta9-desaturase (stearoyl CoA desaturase) and the enzymes involved in the metabolism of 18:2n-6. Thus, the two isomers differed not only in their effects on the weights of total liver lipids and lipid fractions but also on the FA profile of the lipid fractions.

Dietary n-3 FA modulate long and very long chain FA content, rhodopsin content, and rhodopsin phosphorylation in rat rod outer segment after light exposure.

A previous study has shown that the long and very long chain FA (VLCFA) content of the rat retina responds to changes in dietary n-6/n-3 ratio of the fat fed (1). The present study tested whether similar changes in these FA are associated with alterations in rhodopsin content and rhodopsin phosphorylation after light treatment. Weanling rats were fed diets containing 20% (w/w, 40% energy) fat with either high (4.8%, w/w) or low (1.2%, w/w) n-3 FA. After 6 wk of feeding, half of the animals in each group were exposed to light for 48 h at 350 lx or were kept in complete darkness. In the rod outer segment, the high n-3 diet treatment increased the level of 20:5n-3 and 22:6n-3 and reduced the levels of 20:4n-6 and 24:4n-6 in PC, PE, and PS. After the feeding of a high n-3 FA diet, total n-3 pentaenoic VLCFA from C24 to C34 increased in PC, whereas the n-6 tetra- and pentaenoic VLCFA decreased. No changes occurred in n-3 hexaenoic VLCFA regardless of the level of 22:6n-3 in the diet. After light exposure, animals fed a high n-3 FA diet showed reduction in 22:6n-3 as well as in n-6 and n-3 VLCFA in PC. FFA and TG fractions contained increased levels of both 20:4n-6 and 22:6n-3 after light exposure. Dark-adapted rhodopsin content and rhodopsin phosphorylation in the rod outer segment of rats fed the low n-3 FA diet were higher than in animals fed a high n-3 FA diet. After light exposure, animals fed the low n-3 FA diet lost more rhodopsin compared to animals fed the high n-3 FA diet, resulting in less phosphorylation of rhodopsin. Results indicate that the FA composition, rhodopsin content, and phosphorylation in visual cells is influenced by the dietary n-3 FA fed as well as by light exposure. The results also imply that 22:6n-3 may not be the precursor for synthesis of hexaenoic VLCFA.

Erratum

Objective: The aim of this study was to examine the distribution of trans fatty acids (TFA) in plasma lipid classes and the relationship with dietary intake of TFA. Design: After a 2 week baseline (habitual) diet, all subjects consumed a moderate fat (MF) diet for 3 weeks with the fat being derived mainly from margarine and the rest from lean beef and then a very low fat (VLF) diet for 3 weeks with the TFA being derived only from the lean beef. Blood samples were collected 2 days prior to the end and also on the last day of each dietary period. Setting: Deakin Institute of Human Nutrition, Deakin University, Geelong, Australia. Subjects: Ten free-living mildly hypercholesterolaemic subjects aged 22–66 were recruited in Geelong. Outcome measures: TFA intake was calculated from analyses of Australian margarines, butter, lean meat and animal fat. The TFA in plasma lipid fractions were separated by AgNO3 thin-layer chromatography and quantitated by capillary gas–liquid chromatography using internal standards. Results: The phospholipid (PL) fraction contained more than 60% of the trans-18:1 isomers in the plasma lipids in all subjects. On the baseline diet, the predominant positional isomer of trans-18:1 in PL was Δ11, whereas in the other lipid classes it was the Δ9 isomer. The concentration of the Δ9 isomer increased on the MF diet, particularly in the PL fraction, while the concentration of the Δ11 isomer decreased in all fractions. On the VLF diet, the total TFA level decreased by approximately 50%, mainly due to decreases in the TFA isomers in the PL and TG fractions. Changes in plasma total and PL TFA, PL Δ9, Δ10 and Δ11 were strongly correlated with dietary TFA intake (P<0.0001). There were also significant association between dietary TFA intake and PL Δ12 (P=0.003), triacylglycerol Δ9 (P=0.009), Δ11 (P=0.0005), total triacylglycerol (P=0.023) and free fatty acid TFA (P=0.042). Conclusions: The results suggest that the measurement of trans-18:1 in plasma PL and TAG and plasma total TFA could be used to estimate the intake of TFA. Sponsorship: Meat Research Corporation of Australia. European Journal of Clinical Nutrition (2001) 55, 59–64

The occurrence of trans-18:1 isomers in plasma lipids classes in humans.

The aim of this study was to examine the distribution of trans fatty acids (TFA) in plasma lipid classes and the relationship with dietary intake of TFA. After a 2 week baseline (habitual) diet, all subjects consumed a moderate fat (MF) diet for 3 weeks with the fat being derived mainly from margarine and the rest from lean beef, and then a very low fat (VLF) diet for 3 weeks with the TFA being derived only from the lean beef. Blood samples were collected 2 days prior to the end and also on the last day of each dietary period. Deakin Institute of Human Nutrition, Deakin University, Geelong, Australia. Ten free-living mildly hypercholesterolaemic subjects aged 22-66 were recruited in Geelong. TFA intake was calculated from analyses of Australian margarines, butter, lean meat and animal fat. The TFA in plasma lipid fractions were separated by AgNO3 thin-layer chromatography and quantitated by capillary gas-liquid chromatography using internal standards. The phospholipid (PL) fraction contained more than 60% of the trans-18:1 isomers in the plasma lipids in all subjects. On the baseline diet, the predominant positional isomer of trans-18:1 in PL was delta11, whereas in the other lipid classes it was the delta9 isomer. The concentration of the delta9 isomer increased on the MF diet, particularly in the PL fraction, while the concentration of the delta11 isomer decreased in all fractions. On the VLF diet, the total TFA level decreased by approximately 50%, mainly due to decreases in the TFA isomers in the PL and TG fractions. Changes in plasma total and PL TFA, PL delta9, delta10 and delta11 were strongly correlated with dietary TFA intake (P < 0.0001). There were also significant association between dietary TFA intake and PL delta12 (P = 0.003), triacylglycerol delta9 (P=0.009), delta11 (P=0.0005), total triacylglycerol (P=0.023) and free fatty acid TFA (P = 0.042). The results suggest that the measurement of trans-18:1 in plasma PL and TAG, and plasma total TFA could be used to estimate the intake of TFA.

T cell clones raised from chronically infected healthy humans by stimulation with Toxoplasma gondii excretory-secretory antigens cross-react with live tachyzoites: characterization of the fine antigenic specificity of the clones and implications for vaccine development.

Excreted-secreted Ags (ESA) of Toxoplasma gondii (Tg) play an important role in the stimulation of the host immune system in both acute and chronic infections. To identify the parasite Ag(s) involved in the maintenance of T cell-mediated long term immunity, 40 ESA-specific T cell clones were derived from three chronically infected healthy subjects. All the clones were CD4+ and recognized both ESA and live tachyzoites in a HLA-DR-restricted manner. Conversely, CD4+ tachyzoite-specific T cell clones from the same subjects proliferated in response to ESA, pointing to shared immunodominant Ags between ESA and Tg tachyzoites. By T cell blot analysis using SDS-PAGE-fractionated parasite extracts, the following patterns of reactivity were detected. Of 25 clones, 6 recognized Tg fractions in the 24- to 28-kDa range and proliferated to purified GRA2, 5 reacted with Tg fractions in the 30- to 33-kDa range; and 4 of them proved to be specific for rSAg1. Although surface Ag (SAg1) is not a member of ESA, small amounts of this protein were present in ESA preparation by Western blot. Of 25 clones, 8 responded to Tg fractions in the 50- to 60-kDa range but not to the 55-kDa recombinant rhoptries-2 parasite Ag, and 6 did not react with any Tg fraction but proliferated in response to either ESA or total parasite extracts. In conclusion, CD4+ T cells specific for either ESA (GRA2) or SAg1 may be involved in the maintenance of long term immunity to Tg in healthy chronically infected individuals.

Carbohydrate analysis of porcine thyroglobulin isoforms with different iodine contents

To further validate the relationship between thyroid hormone formation and the carbohydrate structure of thyroglobulin (Tg), we reinvestigated the relationship between the iodine content and the asparagine-linked oligosaccharide structures of porcine Tg. Purified porcine Tg was further separated into isoforms (Tg-F1, -F2 and -F3) with a DEAE–cellulose ion-exchange chromatography column. The iodine residues, neutral sugar and sialic acid were analyzed for the separated Tg isoforms and their asparagine-linked oligosaccharide structures were analyzed. The asparagine-linked oligosaccharides were released from Tg-F1, -F2 and -F3 by hydrazinolysis and each oligosaccharide was labeled with p-aminobenzoic acid octyl ester (ABOE). The ABOE-labeled oligosaccharides from Tg-F1, -F2 and -F3 were analyzed for their relative content in oligosaccharides of each structure type by chemical methods and DEAE- and ConA high-performance liquid chromatography (HPLC) columns. As a result, it was revealed that the Tg fraction eluted at higher ionic strength from a DEAE–cellulose column is apt to contain more of each iodoamino acid, as well as total content of iodine, larger negative zeta-potential, conforming to sialic acid content in the Tg molecule and to a higher content of di-sialo-bi-antennary complex and to high mannose-type oligosaccharides. These results support the conclusion that iodine organification of the Tg molecule is correlated with asparagine-linked oligosaccharide completion.

Determination of free fatty acids in cheese: comparison of two analytical methods

Two methods were compared for the determination of free fatty acids (FFA) from acetic to long-chain acids in samples with a large excess of triacylglycerols (TG) (1[ratio ]200, w/w), such as cheese and other dairy products. In method 1, after fat extraction, FFA were separated from TG by aminopropyl-bonded phase chromatography, injecting the fraction containing FFA directly into the gas chromatograph. In method 2, extracted fat was treated with tetramethylammonium hydroxide, the methyl ester derivatives being formed in the injector. Cheese samples and standard mixtures of FFA and TG in different proportions were analysed by both methods. The cheese sample contained 2·4 times more FFA when analysed by method 2 as compared with the result obtained with method 1. The composition of the standard mixtures analysed by method 1 closely reflected that of the original mixture and gave 90–100% recovery of FFA, regardless of their chain length and the ratio of FFA[ratio ]TG (1[ratio ]1 or 1[ratio ]200, w/w). The composition of samples with a FFA[ratio ]TG ratio of 1[ratio ]200 (w/v) was severely distorted (as compared with the original composition of the sample) when analysed by method 2. Varying recoveries of FFA were also obtained, the largest differences being found for the shorter-chain components. We conclude that the FFA fraction should be separated from the TG fraction before derivatization and chromatographic analysis, particularly for samples in which the FFA represent a minor fraction of the TG.

Docosahexaenoic acid causes accumulation of free arachidonic acid in rat pineal gland and hippocampus to form hepoxilins from both substrates

Hepoxilins (Hx) are biologically active metabolites of arachidonic acid (AA) formed regioselectively from 12(S)-HPETE by `hepoxilin synthase'. Hx modulate synaptic neurotransmission in hippocampal CA1 neurons, and inhibit norepinephrine release in hippocampal slices. During the course of our studies we investigated whether docosahexaenoic acid (DHA) was a substrate for hepoxilin formation. We used two tissues, the pineal gland and hippocampal slices. Tissues were incubated alone or with AA (20 μg/ml) or DHA (20 μg/ml). After 60 min at 37°C, samples were acid-extracted to convert Hx into their stable trioxilin (TrX) form and analyzed as the Me-TMSi derivatives by EI-GC/MS to determine the structures of the DHA metabolites, and as PFB-TMSi derivatives by GC/MS in the NICI mode using SIM to simultaneously quantify TrX products of the 3-series (derived from AA) monitored at m/z 569, while those of the 5-series (derived from DHA) were monitored at m/z 593. Results show good conversion of both substrate fatty acids by the rat pineal gland and hippocampal slices, into the 3-series (21.3±5.8 and 12.5±2.2 ng/ μg protein, respectively) and 5-series TrX (12.3±2.7 and 2.9±0.4 ng/ μg protein, respectively). Surprisingly though, experiments with DHA, in both tissues, also showed formation of TrX derived from endogenous AA (3-series) (10.4±8.3 and 3.1±2.1 ng/ μg protein, respectively). These experiments demonstrate previously unreported actions of DHA causing the accumulation of AA, which is converted into hepoxilins. In order to prove that AA is accumulated during DHA stimulation of the tissue, we carried out separate experiments with hippocampal slices in which the neutral lipids and phospholipids were labeled with [ 14C]AA. DHA caused a time-dependent appearance of free [ 14C]AA which was released mostly from the TG pool. Measurement of the AA/DHA ratio in the TG pool by GC/MS further indicated that DHA is incorporated into the TG at the expense of AA. These results demonstrate that DHA competes with AA for acylation into the metabolically active TG fraction, and both fatty acids are converted into hepoxilins of the corresponding series.

The influence of dietary fat source (sunflower oil or olive oil) on LDL composition and serum lipid levels in miniature swine ( Sus scrofa)

A total of 24 miniature swine ( Sus scrofa) were fed with two diets of 9% fat content, differing only in the quality of the fat source (sunflower oil and olive oil). Two groups of animals were fed for a 12-week period, and the other two groups were fed for a 50-week period. After the two experimental periods, the influence of the dietary fat on serum lipids and protein and fatty acid composition of isolated LDLs was studied. In the short term, the serum cholesterol level was slightly higher in the olive oil group but, with the time of adaptation to the diet, serum levels of TC, FC and PL increased significantly in the sunflower group. In the long term, LDL and HDL were also significantly higher in the sunflower group when compared to the monounsaturated diet. In the sunflower group, PROT/TC and PROT/LIP ratios decreased significantly with the experimental period, while in the olive oil group they increased, due to the decrease in EC and TG fractions. The LDL particle in the olive group contained fewer saturated fatty acids and more monounsaturated fatty acids, specially oleic acid, than the LDL in the sunflower group. The changes found in chemical and fatty acid compositions of LDL, according to the saturation degree of the predominant fat of the diet, could alter its cellular metabolism.

Separation and quantification of the triacylglycerols in evening primrose and borage oils by reversed-phase high-performance liquid chromatography

Evening primrose and borage oil are used frequently in nutritional and clinical studies where an impaired Δ 6-desaturase enzyme activity may be bypassed by supplementation with γ-linolenic acid (GLA, 18:3 n − 6). The separation and quantification of the triglycerides of borage oil and evening primrose oil has been carried out using reversed-phase HPLC with UV detection. Borage oil was found to have 34 UV-detectable fractions and evening primrose 22. The TG fractions were collected manually, their fatty acid composition determined and quantified with an internal standard. The probable identify of the individual TG fractions was deduced using the fatty acid composition of the TG fractions, calculated theoretical carbon numbers (TCN) for the various TG species and the predicted probability of occurrence. Correction factors, U i, for GLA (18:3 n − 6), gadoleic acid (20:1 n − 9), erucic acid (22:1 n − 9) and nervonic acid (24:1 n − 9) were estimated to be 0.3–0.4, 0.6, 0.4 and 0.3, respectively, and are used along with other known U i correction factors for unsaturated fatty acids to calculate TCN values for all the TG species. These U i values represent the loss in affinity of the unsaturated fatty acid for the reversed-phase C-18 stationary phase. The reversed-phase HPLC trace of borage oil is much more complex compared to evening primrose oil. Apart from differences in the total fatty acid composition there are substantial differences in the quantity of individual TG species present in the two oils. The clinically important fatty acid, γ-linolenic acid, is distributed much more widely throughout the TG species of borage oil compared to evening primrose oil. Over 90% of the GLA present in evening primrose occurs in the first 9 eluting TG species whereas only about 65% is found in these TG species of borage oil.

Lipid esterification and synthesis by the IEC-6 intestinal epithelial crypt cell line: effect of transforming growth factor β

The capacity of immature intestinal epithelial crypt cells to synthesize lipids and the factors that promote their differentiation remain largely unknown. We examined the profile of lipids synthesized by a normal rat intestinal epithelial crypt cell line (IEC-6) and determined the effects of transforming growth factor beta (TGF beta), a putative crypt cell differentiating factor, on their lipid handling. Incubation of IEC-6 cells with [14C]oleic acid (20 h) resulted in lipid esterification and synthesis, mainly as triglycerides (TGs, 57 +/- 0.6%) and phospholipids (PLs, 30 +/- 0.6%), with a PL/TG ratio of 0.53. When cells were pulsed (2.5 h) with [14C]oleic acid and then maintained 20 h in medium alone, a significant elevation of the PL/TG ratio (10.2 +/- 1.3, p < 0.01) was observed, primarily accounted for by a significant decrease of the TG fraction (p < 0.01). IEC-6 cells secreted only trace amounts of lipids under the latter experimental condition. Incubation with TGF beta (20 h) significantly inhibited IEC-6 cell proliferation but did not promote the expression of cell sucrase activity. TGF beta induced a significant increase in the cellular composition of PL (p < 0.05) and a decrease in the TG fraction (p < 0.02), after a 2.5-h pulse of [14C]oleic acid. Lipid production was unaffected by TGF beta during the 20-h incubation with [14C]oleic acid. Lipid secretion into the medium remained negligible in the presence of TGF beta, after 2.5 h of incubation with substrate as above. Our findings suggest that immature crypt IEC-6 cells are capable of lipid esterification and synthesis but secrete minute amounts of lipoproteins.(ABSTRACT TRUNCATED AT 250 WORDS)

Fractionation of the triacylglycerols of lipase‐modified butter oil

AbstractTriacylglycerols (TGs) of lipase‐modified butter oil were separated into saturated, monoene, diene and triene fractions on ap‐propylbenzene sulfonic acid solid‐phase extraction column loaded with silver ions. Fatty acid analysis of the fractions showed that the amounts of saturated TGs (98.4 mol%) and monoene TGs (26.0 mol%) in the saturated and monoene fractions, respectively, were close to the theoretical amounts of TGs in pure fractions. The column method provides a useful alternative to AgNO3‐thin‐layer chromatography as a means of separating the TGs of butterfat and producing relatively pure TG fractions for further analysis by gas chromatography (GC) or GC‐mass spectrometry.