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Improvement of oxidative stability of conjugated linoleic acid (CLA) by microencapsulation in cyclodextrins.

Oxidative stability of conjugated linoleic acid (CLA) encapsulated in alpha-, beta-, and gamma-cyclodextrins (designated CLA/CDs microencapsules) was studied by measuring the headspace-oxygen depletion in airtight serum bottles and by measuring the peroxide values (POV). The rate of oxygen depletion was reduced from 41.0 (control) to 21.5, 2.1, 1.2, and 1.1 micromol/L.h(-)(1) by CLA/alpha-CD microencapsules at 1:1, 1:2, 1:4, and 1:6 mole ratios, respectively, indicating that CLA oxidation was completely protected by a 1:4 mole ratio of CLA/alpha-CD. Such a protective effect by CLA/beta-CD or CLA/gamma-CD microencapsules was achieved at a 1:6 mole ratio, but the effect by CLA/beta-CD was slightly greater than that by CLA/gamma-CD. The protective effect of alpha-, beta-, and gamma-CDs for CLA oxidation was confirmed by their POV-reducing abilities in CLA/CDs. These results suggest that alpha-CD was the most effective for the protection of CLA oxidation by microencapsulation, followed by beta-CD and gamma-CD.

Cyclodextrins↗

Conjugated linoleic acid induces lipid peroxidation in men with abdominal obesity.

Conjugated linoleic acid (CLA) has been shown in experimental studies to have chemoprotective properties, and may decrease the deposition of body fat. CLA is prone to oxidation, and it has been suggested that increased lipid oxidation may contribute to the anti-tumorigenic effects of this agent. The present study investigates the urinary levels of 8-iso-prostaglandin F(2alpha) (8-iso-PGF(2alpha)), a major isoprostane, and of 15-oxo-dihydro-PGF(2alpha), a major metabolite of PGF(2alpha), as indicators of non-enzymic and enzymic arachidonic acid oxidation respectively after dietary supplementation with CLA in middle-aged men (mean age 53 years) with abdominal obesity for 1 month in a randomized controlled trial. Significant increases in the levels of both 8-iso-PGF(2alpha) and 15-oxo-dihydro-PGF(2alpha) in urine (P<0. 0001 and P=0.0013 respectively) were observed after 1 month of daily CLA intake (4.2 g/day) as compared with the control group. The lipid peroxidation parameters had returned to their basal levels at 2 weeks after the cessation of CLA intake, and remained at the same levels for a further 2 weeks until the end of the study. CLA had no effect on serum alpha-tocopherol and gamma-tocopherol levels, or on the urinary levels of 2,3-dinor-thromboxane B(2). Thus CLA may induce both non-enzymic and enzymic lipid peroxidation in vivo in middle-aged men with abdominal obesity, without any side effects. The consequences of the increased lipid peroxidation after CLA supplementation are unknown.

Adult↗

Cis 9, trans 11- and trans 10, cis 12-conjugated linoleic acid isomers induce apoptosis in cultured SW480 cells.

Dietary conjugated linoleic acid (CLA) has been shown to reduce colon tumor incidence in rodents by mechanisms probably involving apoptosis. The aim of this study was to evaluate the effects of three commercial CLA preparations (pure c9, t11-CLA, pure t10, c12-CLA and a CLA mixture, containing 29.5% c9, t11 and 29% t10, c12-CLA) on caspase-dependent apoptosis in colon SW480 tumor cells. After 4 days incubation, all CLA-treated cells displayed an increase in caspase 3 (27-34%) and caspase 9 activities (37-47%), cleavage of pro-caspase 3 (32 kDa) to 17 and 12 kDa subunits, increased membrane annexin V levels and reduced expression of bcl-2 compared with untreated controls. Cytosolic cytochrome c was increased (p < 0.05) by all CLA preparations, with the t10, c12-CLA isomer being the most potent. The data indicate that t10, c12-CLA may be the more biologically active isomer for inhibition of colon tumor cell proliferation in vitro.

Annexin A5↗

Effect of conjugated linoleic acid immobilization on the hemocompatibility of cellulose acetate membrane.

Conjugated linoleic acid (CLA) was covalently immobilized onto cellulose acetate (CA) membranes. The effects of CLA immobilization on the blood coagulation, platelet aggregation, and oxidative stress were evaluated using human blood. The resulting CLA grafting CA membranes were characterized with X-ray photoelectronic spectroscopy (XPS). The complete blood count (CBC) and coagulation time (CT) was evaluated in vitro for the hemocompatibility. Human serum albumin (HSA) and human plasma fibrinogen (HPF) was evaluated for the protein affinity. The production of reactive oxygen species (ROS) was measured by chemiluminescence (CL) method to evaluate the oxidative stress. The results showed that the CLA-immobilizing CA membrane could keep the CBC values more stable than unmodified CA membrane. The CLA-immobilized CA membranes also showed longer CT and less adsorption of plasma proteins. CLA-immobilized CA membrane could keep the CL counts of hydrogen peroxide and superoxide values more stable than unmodified CA membrane. These results suggest that a CLA-immobilized CA membrane could offer protection for patients against oxidative stress and would be helpful for reducing the dosage of anticoagulant during hemodialysis.

Blood Coagulation↗

Dietary, but not topical, alpha-linolenic acid suppresses UVB-induced skin injury in hairless mice when compared with linoleic acids.

Peroxidizability of fatty acids in the air is roughly proportional to the number of double bonds, but in vivo peroxidation proceeds in a more complex manner. Here, we compared the effects of dietary and topically applied oils enriched with linoleic acid (LA, 18:2n-6) or alpha-linolenic acid (ALA, 18:3n-3) on UV-induced skin injury in a strain of hairless mice. The UVB-induced erythema score was significantly lower in mice with topically applied creams containing LA and ALA than in mice with the basal cream; no significant increase in the score was detected in the ALA group compared with the LA group. However, dietary ALA inhibited the increase in erythema score after UVB irradiation compared with LA. The peroxidizability index of the skin total lipids was significantly higher, but UVB-induced prostaglandin E2 (PGE2) production was significantly lower in the group fed an ALA-rich diet compared with the group fed an LA-rich diet. The levels of thiobarbituric acid-reactive substances, estimated in the presence of butylated hydroxytoluene in the assay mixture, were not affected by UVB treatment or by the dietary fatty acids, but the severity of the skin lesion was associated with PGE2 levels. These results indicate that the type of fatty acids, n-6 or n-3, is critical for the suppression of UVB-induced skin lesion when the skin fatty acids are modified by dietary manipulation. Anti-inflammatory activity of dietary flaxseed oil with relatively high ALA and low LA contents was demonstrated in UVB-irradiated hairless mice.

Administration, Cutaneous↗

Regulation of endothelial nitric oxide synthase gene expression by oxidized linoleic acid.

Hypercholesterolemia is associated with impairments in endothelium-dependent vascular relaxations. Paradoxically, endothelial production of nitrogen oxides is increased in early stages of hypercholesterolemia. Prior work has shown that oxidized low density lipoprotein (LDL) has both stimulatory and inhibitory effects on endothelial nitric oxide synthase expression (eNOS) and has focused on lysophosphatidyl choline (LPC) as a component of oxidized LDL which may modulate this effect. Another biologically active component of oxidized LDL is 13-hydroperoxyoctadecadienoic acid (13-HPODE), an oxidized form of linoleic acid. The purpose of this study was to determine the effect of HPODE on the expression of eNOS in bovine aortic endothelial cells (BAECs). Twenty four hour treatment of endothelial cells with HPODE caused a dose-dependent increase in eNOS mRNA levels as assessed by Northern analysis. The time response studies show that HPODE treatment significantly increased eNOS mRNA levels at 12 and 24 h. Concomitant with the increase in eNOS mRNA levels, 20 microM HPODE treatment significantly increased eNOS protein content and enzyme activity. Nuclear run-on studies indicated that the rate of transcription of eNOS gene was significantly elevated 4 h after HPODE treatment when compared to control cultures. Also, actinomycin D studies demonstrated that the half-life of eNOS mRNA was increased from 6 h to 12 h by HPODE treatment. Thus, HPODE-induced up-regulation of eNOS expression is mediated by both transcriptional and posttranscriptional mechanisms. These observations suggest that endothelial cells may attempt to compensate for oxidative injury by increasing expression of eNOS in early stages of hypercholesterolemia.

Animals↗

Inhibition of the autoxidation of linoleic acid by phenylpropanoid glycosides from Pedicularis in micelles.

The activities of six phenylpropanoid glycosides as chain-breaking antioxidants have been studied for the autoxidation of linoleic acid in cetyl trimethylammonium bromide (CTAB) micelles at 37 degrees C. Verbascoside, isoverbascoside, echinacoside and pedicularioside A, which possess four phenolic hydroxyl groups, exhibited antioxidant activities, while cistanoside D possessing only two phenolic hydroxyl groups and permethyl verbascoside without phenolic hydroxyl group did not suppress the oxidation appreciably. The ratio of rate constants for inhibition and propagation kinh/kp and stoichiometric factor n were determined.

Amidines↗

Conjugated linoleic acid isomers differ in their free radical scavenging properties.

Conjugated linoleic acid (CLA) isomers were investigated for free radical scavenging properties against the stable 2,2-diphenyl-1-picryhydrazyl radical (DPPH(*)) by electron spin resonance (ESR) spectrometry and spectrophotometric methods. ESR measurements confirmed that both c9,t11-CLA and t10,c12-CLA directly reacted with and quenched DPPH radicals, whereas spectrophotometric analysis demonstrated that c9,t11-CLA and t10,c12-CLA differed in their kinetic and thermodynamic properties in reacting with DPPH radicals. t10,c12-CLA was shown to exhibit a greater initial velocity in CLA-DPPH radical reactions at levels of 2.5-80 mg/mL, and c9,t11-CLA scavenged more DPPH radicals at steady state. Similar dose and time relationships were observed for both isomers. In addition, a mixture of c9,t11- and t10,c12-CLA isomers demonstrated a greater initial velocity in quenching DPPH radicals than either isomer alone on the same concentration basis, suggesting that a synergistic effect between CLA isomers existed in their reactions with DPPH radicals. These results support the conclusion that individual CLA isomers differ in their biological actions and indicate that interaction(s) between isomers may contribute to their beneficial effects.

Antioxidants↗

Safety profile of conjugated linoleic acid in a 12-month trial in obese humans.

Conjugated linoleic acid (CLA) is marketed in numerous commercially available dietary supplements, but few studies have looked at the long-term safety of this product. The current study evaluated the safety of one CLA product (Clarinol) over a one-year period in obese humans who were generally healthy. This was a randomized, double-blind study consisting of three phases in which subjects were given 6 g/day of CLA or placebo. Phase 1 was a low calorie diet (13 kcal/kg desirable weight) for 12 weeks or until 10-20% of initial body weight was lost. In phase 2, from weeks 12 to 28, subjects were re-fed a diet providing 25-30 kcal/kg of desirable body weight. Phase 3 was open label, with subjects from both groups taking CLA from weeks 28 to 52. At biweekly visits, subjects completed a questionnaire evaluating side effects and adverse events. Blood was taken for assay of liver function, glucose, insulin, serum lipids, blood counts, and general chemistry. Overall, body composition did not differ between groups. Laboratory tests showed no adverse effects of CLA. Adverse events and side effects were less in the CLA group compared to placebo. We conclude that CLA as Clarinol is safe for use in obese humans for at least one year.

Adult↗

Effect of triacylglycerol structure on absorption and metabolism of isotope-labeled palmitic and linoleic acids by humans.

The effect of dietary TAG structure and fatty acid acyl TAG position on palmitic and linoleic acid metabolism was investigated in four middle-aged male subjects. The study design consisted of feeding diets containing 61 g/d of native lard (NL) or randomized lard (RL) for 28 d. Subjects then received an oral dose of either 1,3-tetradeuteriopalmitoyl-2-dideuteriolinoleoyl-rac-glycerol or a mixture of 1,3-dideuteriolinoleoyl-2-tetradeuteriopalmitoyl-rac-glycerol and 1,3-hexadeuteriopalmitoyl-2-tetradeuteriolinoleoyl-rac-glycerol. Methyl esters of plasma lipids isolated from blood samples drawn over a 2-d period were analyzed by GC-MS. Results showed that absorption of the 2H-fatty acids (2H-FA) was not influenced by TAG position. The 2H-FA at the 2-acyl TAG position were 85+/-4.6% retained after absorption. Substantial migration of 2H-16:0 (31.2+/-8.6%) from the sn-2 TAG position to the sn-1,3 position and 2H-18:2n-6 (52.8+/-6.4%) from the sn-1,3 position to the sn-2 position of chylomicron TAG occurred after initial absorption and indicates the presence of a previously unrecognized isomerization mechanism. Incorporation and turnover of the 2H-FA in chylomicron TAG, plasma TAG, and plasma cholesterol esters were not influenced by TAG acyl position. Accretion of 2H-16:0 from the sn-2 TAG position in 1-acylphosphatidylcholine was 1.7 times higher than 2H-16:0 from the sn-1,3 TAG positions. Acyl TAG position did not influence 2H-18:2n-6 incorporation in PC. The concentration of 2H-18:2n-6-derived 2H-20:4n-6 in plasma PC from subjects fed the RL diet was 1.5 times higher than for subjects fed the NL diet, and this result suggests that diets containing 16:0 located at the sn-2 TAG position may inhibit 20:4n-6 synthesis. The overall conclusion is that selective rearrangement of chylomicron TAG structures diminishes but does not totally eliminate the metabolic and physiological effects of dietary TAG structure.

Absorption↗

The linoleic acid derivative DCP-LA selectively activates PKC-epsilon, possibly binding to the phosphatidylserine binding site.

This study examined the effect of 8-[2-(2-pentyl-cyclopropylmethyl)-cyclopropyl]-octanoic acid (DCP-LA), a newly synthesized linoleic acid derivative with cyclopropane rings instead of cis-double bonds, on protein kinase C (PKC) activity. In the in situ PKC assay with reverse-phase high-performance liquid chromatography, DCP-LA significantly activated PKC in PC-12 cells in a concentration-dependent (10 nM-100 microM) manner, with the maximal effect at 100 nM, and the DCP-LA effect was blocked by GF109203X, a PKC inhibitor, or a selective inhibitor peptide of the novel PKC isozyme PKC-epsilon. Furthermore, DCP-LA activated PKC in HEK-293 cells that was inhibited by the small, interfering RNA against PKC-epsilon. In the cell-free PKC assay, of the nine isozymes examined here, DCP-LA most strongly activated PKC-epsilon, with >7-fold potency over other PKC isozymes, in the absence of dioleoyl-phosphatidylserine and 1,2-dioleoyl-sn-glycerol; instead, the DCP-LA action was inhibited by dioleoyl-phosphatidylserine. DCP-LA also activated PKC-gamma, a conventional PKC, but to a much lesser extent compared with that for PKC-epsilon, by a mechanism distinct from PKC-epsilon activation. Thus, DCP-LA serves as a selective activator of PKC-epsilon, possibly by binding to the phosphatidylserine binding site on PKC-epsilon. These results may provide fresh insight into lipid signaling in PKC activation.

Animals↗

Increasing amounts of conjugated linoleic acid progressively reduces milk fat synthesis immediately postpartum.

Mixed conjugated linoleic acid (CLA) isomers decrease milk fat synthesis during established lactation, but their ability to cause milk fat depression (MFD) immediately postpartum remains unclear. Multiparous Holstein cows (n = 19) were randomly assigned to 1 of 4 doses of rumen-protected (RP) CLA supplements (0, 200, 400, and 600 g/d); each dose provided equal amounts of fatty acids by replacing and balancing treatments with an RP supplement of palm fatty acid distallate. Doses provided a total of 468 g fatty acids/d and 0, 62, 125, or 187 g of mixed CLA isomers/d, respectively. The CLA supplement contained a variety of CLA isomers: 5.4% trans-8, cis-10; 6.3% cis-9, trans-11; 7.9% trans-10, cis-12; and 8.2% cis-11, trans-13 CLA. Each group received treatments from approximately -10 to 21 d relative to calving. To improve palatability and ensure complete consumption, doses were mixed with equal amounts of steam-flaked corn and dried molasses; one-half the supplement was fed at 0600 h, and the remaining supplement was fed at 1800 h. Milk yield and individual feed intake were recorded daily, and milk samples were obtained from each cow every 2nd day (at both milkings) starting on d 1 postpartum. There were no differences in dry matter intake (17.1 kg/d), milk yield (34.2 kg/d), protein content (3.74%), lactose content (4.61%), or yield of milk protein or lactose. The CLA supplementation decreased overall milk fat content in a dose-responsive manner (4.57, 3.97, 3.32, and 3.10, respectively), and milk fat yield displayed the same progressive decline. The dose-dependent decrease in milk fat content was evident during wk 1 and became highly significant during wk 2 and 3. The milk fat yield response pattern was similar, and by d 21, the highest RP-CLA supplement decreased milk fat content and yield by 49 and 56%, respectively. These data clearly indicate RP-CLA can markedly (40 to 50%) induce MFD immediately postpartum without negatively affecting other production parameters.

Animal Feed↗

Effect of diets containing different levels of linoleic acid on human breast cancer growth and lung metastasis in nude mice.

The purpose of the study was to determine the effect of three different levels of dietary linoleic acid (LA) intake on the growth of MDA-MB-435 human breast cancer cells in the mammary fat pads of nude mice, and their metastasis to the lungs. These diets were isocaloric, and contained different mixtures of safflower (LA-rich) and coconut (saturated fatty acid-rich) oils to provide 23% (w/w) total fat, with 2, 8, and 12% (w/w) LA. A fourth group was fed a low-fat, 5% (w/w) corn oil diet. There were 25 mice in each dietary group. A necropsy, 12 weeks after the tumor cell injections, the primary tumor weights in the 12% LA (4.1 +/- 2.7 g)- and 8% LA (3.5 +/- 1.7 g)-fed groups were significantly greater (P < 0.05) than those fed the 2% LA diet (2.5 +/- 1.5 g); they did not differ significantly from the weights of mammary fat pad tumors in the 5% corn oil-fed mice. The incidence of grossly visible pulmonary metastatic nodules was not significantly different between the 8 and 12% LA-fed mice, but was higher for both groups compared with the 2% LA-fed group (P < 0.05), with a similar trend in comparison with the 5% corn oil group. The mean total calculated volumes of the macroscopic metastases per tumor-bearing mouse were significantly greater in the 8 and 12% LA (157 +/- 250.7 and 99.1 +/- 140.0 mm3, respectively), compared with the 2% LA (23.3 +/- 51.8 mm3)- and 5% corn oil (24.5 +/- 35.1 mm3)-fed mice; all P < 0.05. Micrometastases were observed most frequently in the 5% corn oil and 2% LA dietary groups, but none of the differences were statistically significant. No differences were detected in the concentrations of prostaglandin E, leukotriene B4, or 5-hydroxyeicosatetraenoic acid in tumors from mice fed the four different diets.

Adipose Tissue↗

Flavor and stability of pasteurized milk with elevated levels of conjugated linoleic acid and vaccenic acid.

The objectives of this study were to determine if flavor differences between 2% fat pasteurized milks with and without naturally enhanced vaccenic acid (VA) and cis-9, trans-11 conjugated linoleic acids (CLA) levels could be detected over the commercial shelf life of the product and to determine if milk with elevated VA and cis-9, trans-11 CLA levels was more susceptible to development of light-induced oxidative flavor defects. Cows were fed a control diet or the same ration supplemented with 2% soybean oil and 1% fish oil (CLA diet). The milk, standardized to 2% fat, was pasteurized, homogenized, and stored in plastic containers at 4 degrees C. Oxidation was induced by exposing half of the containers to light. Testing was conducted at 1, 7, and 14 d postpasteurization. Average cis-9, trans-11 CLA content of the milks from the control and CLA diet groups was 0.52 and 4.74 g/100 g of fatty acids, respectively (8-fold increase). Average VA content of the milk from the control and CLA diet groups was 1.43 and 12.06 g/100 g of fatty acids, respectively (7.5-fold increase). Together, VA plus CLA represented almost 17% of the total milk fatty acids. There was no effect of light exposure on fatty acid composition initially or over the 14-d storage period. Although VA, cis-9, trans-11 CLA, and degree of unsaturation were significantly elevated in the milk from the CLA diet group, untrained panelists were unable to detect flavor differences initially or over time in 15 of 16 triangle test evaluations. Similarly, sensory results indicated no difference in susceptibility to the development of oxidized off-flavors between the milk from the control and CLA diet groups, even when oxidation was induced by light exposure.

Animals↗

Effects of sodium saccharin and linoleic acid on mRNA levels of Her2/neu and p53 in a human breast epithelial cell line.

The effects of two food-related chemicals (sodium saccharin and linoleic acid) on the levels of Her2/neu and p53 mRNA in a non-cancerous human breast epithelial cell line (HBL-100) were tested in comparison with the effects of the known tumor promoter phorbol 12-myristate 13-acetate (TPA). Treatments were made both with and without prior treatment with two well-known tumor initiators, N-nitroso-N-methylurea (NMU) or 7,12-dimethylbenz[a]anthracene (DMBA). The effects in general were small, the greatest being increases of 46-67% in Her2/neu mRNA levels in response to treatments with TPA or sodium saccharin following NMU treatments. These results demonstrate that sodium saccharin following NMU treatments might be involved in transcriptional regulation of Her2/neu in HBL-100 cells and suggest that its effects may not be limited to urinary bladder.

9,10-Dimethyl-1,2-benzanthracene↗

Linoleic acid in the treatment of lithium toxicity and familial tremor.

Lithium inhibits the synthesis of prostaglandin (PG) EI by blocking the mobilisation of dihomogammalinolenic acid (DGLA). Toxicity due to lithium might re related to reduced PGEI formation. In five patients who developed toxic effects on low doses of lithium, linoleic acid in the forms of the safflower oil was given in an attempt to raise levels of the linoleic acid metabolite, DGLA. In al five patients the safflower oil was effective in remitting the symptoms of neurotoxicity. Safflower oil was also effective in a patient with familial tremor.

Adult↗

Effects of a low dietary linoleic acid level on intestinal morphology and enterocyte brush border membrane lipid composition.

The influence of low dietary linoleic acid level (an essential fatty acid deficiency) on the intestine mucosal morphology and the purified brush border membrane (BBM) lipid composition was investigated in the rat. Electron micrographs and morphometric measurements showed that villi and crypt sizes as well as the ultrastructure of epithelial cells were altered. Cholesterol (CHOL) and phospholipid (PL) levels, CHOL/PL ratio and PL class distribution were not changed by the low linoleate diet. However, the fatty acid composition of phospholipids was markedly modified in the enterocyte BBM, showing elevated amounts of palmitoleic (16:1n-7), oleic (18:1n-9) and 5,8,11-eicosatrienoic (20:3n-9) acids and, by contrast, depressed linoleic (18:2n-6) and arachidonic (20:4n-6) acid levels. Although the underlying mechanisms remain unknown the results obtained suggest that essential fatty acids (EFA) could be directly involved in the trigger action of the observed alterations, as regards both their dynamic (metabolic) and structural roles.

Animals↗

[Chronologic studies of the effects of a hypoproteinic diet followed by an equilibrated diet on delta-6 and delta-5 desaturations of linoleic acid in liver microsomes in the rat].

A low protein diet affects amounts of linoleic and arachidonic acids in hepatic microsomal phospholipids of growing rats. Are the changes related to modifications in microsomal delta 6- and delta 5- linoleic acid desaturase activities? Two groups of Wistar rats weighing 80 +/- 5 g at the beginning of the experiment were used: Control group (T) was fed on a 16% gluten + 4% casein diet for 53 days; Experimental group (E) was fed on a 4% gluten + 1% casein diet for 26 days (MP) then Control diet for 27 days (RE). After 2, 14 and 26 days of MP and 2, 15 and 27 days of RE, rats of each group were sacrificed. Protein and water contents of liver, quantitative fatty acid, composition of total lipids in liver and hepatic microsomes were determined. delta 6- and delta 5- linoleic acid desaturase activities were estimated from incubation of liver microsomes with [1-14C] C 18: 2 n-6 or [2(14)C] C 20: 3 n-6 respectively. The low protein diet stops practically ponderal growth. The fatty-acid compositions of microsomal total lipids of E rats were affected in comparison with values of T rats. These modifications persist after 27 days of RE. The C 20: 4 n-6/C 18: 2 n-6 ratio in microsomal total lipids was slightly different between T and E rats but increased strongly during refeeding. Same modifications take place in the fatty-acid composition of hepatic total lipids. After two days of MP, delta 6- and delta 5- desaturase activities were depressed, phenomenon that not persist in the course of MP. These enzyme activities increase to higher values than those of the T after two days of RE.

Animals↗