bluuebunny/crossref_metadata_2025 huggingface dataset:
Studies of lipase‐catalyzed esterification reactions of some acetylenic fatty acids
Authors: Lie Ken Jie Marcel S. F. , Xun Fu
Published: November 2006

Esterification of five positional isomers of acetylenic fatty acids [viz. 9:1(2a), 11:1(10a), 18:1(6a), 18:1(9a) and 22:1(13a)] of different chain lengths with n‐butanol in n‐hexane in the presence of eight different lipases [Lipozyme IM 20 (Rhizomucor miehei), Lipolase 100T (R. miehei), Novozyme 435 (Candida antarctica), PPL (porcine pancreatic lipase), CCL (C. cylindracea), PS‐D (Pseudomonas cepacia), Lipase A‐12 (Aspergillus niger) and Lipase AY‐30 (C. rugosa)] was studied. 2‐Nonynoic acid was not esterified except when catalyzed by the lipase from C. antarctica (Novozyme 435) to give 42% butyl ester after 48 h. The lipases from A. niger (Lipase A‐12) and C. rugosa (Lipase AY‐30) showed poor biocatalytic behavior in the esterification of the acetylenic fatty acids studied. 10‐Undecynoic acid gave the highest conversion rate of esterification with each kind of lipase used. 6‐Octadecynoic acid showed a marked degree of resistance to esterification carried out in the presence of C. cylindracea (CCL), P. cepacia (PS‐D), or porcine pancreatic (PPL) lipase but not significantly in the presence of the lipases of R. miehei (Lipozyme IM 20), R. miehei (Lipolase 100T), or Novozyme 435. 9‐Octadecynoic acid and 13‐docosynoic acid were not discriminated and were readily esterified by the remaining six lipases, but when compared to oleic acid the acetylenic fatty acids were comparatively much slower in conversion to the esters.
Bile acid metabolism in analbuminemic rats
Authors: Kambe Akira , Uchida Kiyohisa , Takase Haruto , Nomura Yasuharu , Adachi Yukihiko
Published: November 2006

The bile acid concentrations in the serum, liver, bile, intestines, and feces of 3‐ and 19‐mon‐old male and female Nagase analbuminemic (NA) rats were compared with those in Sprague‐Dawley (SD) rats. There was no significant difference in the bile acid levels between NA and SD rats. However, increased biosynthesis and pool size of cholic acid (CD) derivatives and decreased levels of chenodeoxycholic acid (CDCA) derivatives (increased CA/CDCA ratio) were detected in male NA rats as compared to SD rats. The CA/CDCA ratio in female NA rats was not different from that in their SD rats in the biliary bile flow, bile acid levels in the small and large intestines, fecal bile acid excretion, bile acid concentration in the portal and systemic circulation, and in the pool size of bile acids. The blood lipid concentrations were significantly higher in the NA rats than in the SD rats. The hepatic levels of lipids were not significantly different between the two rat strains. In conclusion, this study showed that metabolism of bile acids in NA rats is not significantly affected, and that the hypercholesterolemia observed in these strains is not related to abnormalities of bile acid metabolism.
Arachidonic acid supplementation enhances synthesis of eicosanoids without suppressing immune functions in young healthy men
Authors: Kelley Darshan S. , Taylor Peter C. , Nelson Gary J. , Mackey Bruce E.
Published: November 2006

This study was conducted to determine the effects of arachidonic acid (AA) supplementation on human immune response (IR) and on the secretion of prostaglandin E2 (PGE2) and leukotriene B4 (LTB4). Ten healthy men (20–38 yr) participated in the study and lived at the Metabolic Suite of the Western Human Nutrition Research Center. They were fed a basal diet (57, 27, and 16 energy percentage from carbohydrate, fat, and protein, respectively, and AA 200 mg/d) for the first 15 d of the study. Additional AA (1.5 g/d) was added to the diet of six men from day 16 to 65, while the remaining four subjects remained on the basal diet. The diets of the two groups were crossed‐over from day 66 to 115. In vitro indices of IR were examined using blood drawn on days 15, 58, 65, 108, and 115. Influenza antibody titers were determined in the sera prepared from blood drawn on days 92 and 115 (23 d postimmunization). AA supplementation caused significant increases in the in vitro secretion of LTB4, and PGE2, but it did not alter the in vitro secretion of tumor necrosis factor α; interleukins 1β, 2, 6; and the receptor for interleukin 2. Nor did it change the number of circulting lymphocytes bearing markers for specific subsets (B, T, helper, suppressor, natural killer) and the serum antibody titers against influenza vaccine. The opposing effects of PGE2 and LTB4 may have led to the lack of change in immune functions tested.
Effects of highly purified eicosapentaenoic acid and docosahexaenoic acid on fatty acid absorption, incorporation into serum phospholipids and postprandial triglyceridemia
Authors: Hansen John‐Bjarne , Grimsgaard Sameline , Nilsen Hugo , Nordøy Arne , Bønaa Kaare H.
Published: November 2006

Fourteen healthy volunteers were randomly allocated to receive 4 g highly purified ethyl esters of eicosapentaenoic acid (EPA) (95% pure, n=7) or docosahexaenoic acid (DHA) (90% pure, n=7) daily for 5 wk in supplement to their ordinary diet. The n−3 fatty acids were given with a standard high‐fat meal at the beginning and the end of the supplementation period. EPA and DHA induced a similar incorporation into chylomicrons which peaked 6 h after the meal. The relative uptake of EPA and DHA from the meal was >90% compared with the uptake of oleic acid. During absorption, there was no significant elongation or retroconversion of EPA or DHA in total chylomicron fatty acids. The concentration of EPA decreased by 13% and DHA by 62% (P<0.001) between 6 and 8 h after the meal. During the 5‐wk supplementation period, EPA showed a more rapid and comprehensive increase in serum phospholipids than did DHA. DHA was retroconverted to EPA, whereas EPA was elongated to docosapentaenoic acid (DPA). The postprandial triglyceridemia was suppressed by 19 and 49% after prolonged intake of EPA and DHA, respectively, indicating that prolonged intake of DHA is equivalent to or even more efficient than that of EPA in lowering postprandial triglyceridemia. This study indicates that there are metabolic differences between EPA and DHA which may have implications for the use of n−3 fatty acids in preventive and clinical medicine.
Effect of apolipoprotein E polymorphism on serum lipoprotein response to saturated fatty acids
Authors: Tso Tim K. , Park Sunmin , Tsai Yu‐Hwai , Williams Glenn , Snook Jean T.
Published: November 2006

This report summarizes two studies which investigated the effects of apolipoprotein E (apoE) polymorphism on the serum total cholesterol (TC) and lipoprotein cholesterol responses to 8∶0+10∶0 and 12∶0 diets (Study I) and 14∶0, 16∶0, and 18∶0 diets (Study II). Eighteen healthy premenopausal women (3 apoE 3/2, 12 apoE 3/3, 3 apoE 4/3) in study I and another 18 healthy premenopausal women (4 apoE 3/2, 10 apoE 3/3, 3 apoE 4/3, 1 apoE 4/2) in study II consumed a baseline diet providing 40 en% total fat, 11 en % 18∶2, 15 en% 18∶1, 11.5 en% saturated fat for the first week of each 5‐wk period. The experimental diets for both studies provided 40 3n% total fat, 13–14 en% as one of five test saturated fatty acids (SFA), 14–16 en% 18∶1, and 3–4 en% 18∶2. Analysis by apoE phenotypes showed that both the 8∶0+10∶0 diet and the 12∶0 diet in Study Linduced significant increases in serum TC in subjects with different apoE phenotypes with the exception of apoE 3/2 in the medium‐chain triglyceride group. In contrast, in Study II, individuals with apoE 4/3 consuming the 14∶0 diet showed significant increases in serum TC, high density lipoprotein‐cholesterol (HDL‐C), and HDL2‐C, but the same subjects consuming the 16∶0 diet showed significant increases in serum TC and low density lipoprotein‐cholesterol. The findings from both studies indicated serum lipoprotein responses to SFA were different and the variation of responsiveness may be regulated, at least in part, by apoE polymorphism, especially when 14∶0, 16∶0, or 18∶0 was consumed.
Changes in cultured arterial smooth muscle cells isolated from chicks upon cholesterol feeding
Authors: Carazo Angel , Alejandre M. José , Diaz Ramón , Ríos Antonio , Castillo Mercedes , Linares Ana
Published: November 2006

We have developed cultures of smooth muscle cells (SMC) isolated from arterial hypercholesterolemic chicks (cholesterol‐SMC). These cultures are suitable for the study at the molecular level of the changes in arterial SMC induced by a cholesterol diet. By using a strong dose of cholesterol (5%) for 10 d, we obtained very proliferative SMC which became foam cells after 30 d in culture. On the other hand, SMC cultures isolated from control‐fed chicks has a lower growth rate than the SMC ones under the same culture conditions. DNA synthesis was fourfold greater in cholesterol‐SMC than in control‐SMC cultures. Intracellular cholesterol concentrations were the same in both cholesterol and control SMC during the first 14 d of culture but afterward increased in differing ways: after 20 d of culture the cholesterol‐SMC increased their cholesterol content to double the control. We give here the results obtained from transmission electron microscopy, lipid analysis, proliferation studies, DNA, RNA and protein synthesis, and then discuss their implications.
Silver‐ion high‐performance liquid chromatographic separation and identification of conjugated linoleic acid isomers
Authors: Sehat Najibullah , Yurawecz Martin P. , Roach John A. G. , Mossoba Magdi M. , Kramer John K. G. , Ku Youh
Published: November 2006

This is the first report of the application of silverion impregnated high‐performance liquid chromatography (Ag+‐HPLC) to the separation of complex mixtures of conjugated linolenic acid (CLA) isomers present in commercial CLA sources and foods and in biological specimens. This method showed a clear separation of CLA isomers into three groups related to their trans,trans, cis,trans or trans,cis, and cis,cis configuration of the conjugated double‐bound system. In addition, this method separated within each geometrical isomeric group. Following Ag+‐HPLC isolation, gas chromatography (GC)‐electron impact mass spectrometry, and GC‐direct deposition‐Fourier transformed infrared spectroscopy were used to confirm the identity of two major positional isomers in the cis/trans region, i.e., Δ8,10‐ and Δ11,13‐octadecadienoic acid, which had not been chromatographically resolved previously, Furthermore, the potential of this method was demonstrated by showing different Ag+‐HPLC profiles exhibiting patterns of isomeric distributions for biological specimens from animals fed a diet containing a commerical CLA preparation, as well as for a commerical cheese product.
Diabetes‐induced and age‐related changes in fatty acid proportions of plasma lipids in rats
Authors: Nishida Shigeru , Kanno Tatsuhiko , Nakagawa Shigeki
Published: November 2006

Diabetes‐induced and age‐related proportional changes in plasma fatty acids of triglycerides (TG), phospholipids (PL), and cholesteryl esters (CE) were investigated using streptozotocin‐induced diabetic and control rats. Among n‐6 fatty acids from diabetic rat plasma, increased proportions of 18∶2n‐6 and 20∶3n‐6 in all three lipid classes and of 18∶3n‐6 in PL at 1–3 months old and in TG at 3–5 months old were observed. The proportions of 20∶4n‐6 decreased in both PL and CE, but were unchanged in diabetic TG. Among the n‐3 fatty acids, in the early stage, diabetes caused increases in the proportions of 18∶3n‐3 in PL and CE and of 20∶5n‐3 and 22∶6n‐3 in TG, while 22∶5n‐3 was decreased later in the disease course. These results suggest reduced Δ5‐desaturase activities on 20∶3n‐6 but not on 20∶4n‐3, while Δ6‐desaturase activity on 18∶2n‐6 was essentially unaffected. Furthermore, the reduction in Δ9‐desaturase activity in diabetic rats may well explain the decreases in the proportions of 16∶1n‐7 and 18∶1n‐7. However, the proportion of 18∶1n‐9, another product of Δ9‐desaturase, was significantly increased in CE and PL as compared to the controls. Thus, there was a discrepancy between our results and those of earlier studies with respect to the n‐9, n‐6, and n‐3 fatty acid proportions of plasma lipids in diabetic rats. We also investigated age‐related changes in the proportions of plasma fatty acids. Although rather small, age‐related changes were evident in both diabetic and control rats.
Effect of continuous enteral medium‐chain fatty acid infusion on lipid metabolism in rats
Authors: You Yi Qian , Ling Pei Ra , Qu Zhensheng , Bistrian Bruce R.
Published: November 2006

This study compared (i) the relative effects of long‐chain triglycerides (LCT) and medium‐chain triglycerides (MCT), (ii) the influence of amount of MCT, and (iii) the impact of medium‐chain fatty acid position, on plasma and lymphatic triglycerides and portal vein free fatty acids. The animals were fed approximately at 250 kcal/kg · day for 20h. The lymph from lymphatic duct and blood from portal vein and systemic circulation were collected. The results showed that feeding 100% MCT for 20h was sufficiently long to reduce significantly the level of linoleic acid in portal vein fatty acids and plasma and lymph triglycerides. However, this alteration induced by MCT feeding was partially prevented by adding LCT to the diet. The level of arachidonic acid was significantly reduced in plasma triglycerides by any of the diets containing medium‐chain fatty acids compared to 100% LCT. When feeding MCT only, palmitoleic acid, presumably reflecting de novo lipogenesis, was increased in lymphatic triglycerides and portal vein fatty acids. Total saturated fatty acids as a total percentage of total fatty acids were also significantly increased in plasma and lymphatic triglycerides and portal vein fatty acids. Thus, when linoleic acid is limiting, the conversion of MCT into long‐chain fatty acids by de novo lipogenesis is likely to be an important metabolic route. Providing LCT with MCT or 2‐monodecanoin appears to limit this pathway.
Cytotoxicity of tocopherols and their quinones in drug‐sensitive and multidrug‐resistant leukemia cells
Authors: Cornwell David G. , Jones Kenneth H. , Jiang Zongcheng , Lantry Laura E. , Southwell‐Keely Peter , Kohar Indrajati , Thornton Donald E.
Published: November 2006

Cytotoxicities of tocopherols (α‐T, γ‐T, δ‐T), their para (α‐TQ, γ‐TQ, δ‐TQ)‐and ortho (Tocored)‐quinone oxidation products, the synthetic quinone analog of γ‐TQ containing a methyl group substituted for the phytyl side‐chain (TMCQ) and the synthetic quinone analog of Tocored containing a methyl group substituted for the phytyl side‐chain (PR) were measured in acute lymphoblastic leukemia cell lines that are drug‐sensitive (CEM) and multidrug‐resistant (CEM/VLB100). Among tocopherols, only δ‐T exhibited cytotoxicity. Among para quinones, α‐TQ showed no cytotoxicity, while γ‐TQ and δ‐TQ were highly cytotoxic in both CEM and CEM/VLB100 cell lines (LD50<10 μM). δ‐TQ and γ‐TQ were more cytotoxic than the widely studied chemotherapeutic agent doxorubicin, which also showed selective cytotoxicity to CEM cells. The orthoquinone Tocored was less cytotoxic than doxorubicin in drug‐sensitive cells but more cytotoxic than doxorubicin in multidrug‐resistant cells. Cytotoxicity was not a function of the phytyl side‐chain since both TMCQ and PR were cytotoxic in leukemia cells. Cytotoxic para and ortho quinones were electrophiles that formed adducts with nucleophilic thiol groups in glutathione and 2‐mercaptoethanol. Cytotoxicity was enhanced when the glutathione pool was depleted by preincubation with buthionine‐[S,R]‐sulfoximine, but cytotoxicity was diminished by the addition of N‐acetylcysteine to cultures. α‐T also diminished the cytotoxicity of para‐ and or‐thoquinones. Buthionine‐[S,R]‐sulfoximine did not block the inhibitory effect of either N‐acetylcysteine or α‐T, showing that these agents did not act solely by maintaining the glutathione pool as an essential antioxidant system. In conclusion, tocopherylquinones represent a new class of alkylating electrophilic quinones that function as highly cytotoxic agents and escape multidrug resistance in acute lymphoblastic leukemia cell lines.
Conversion of palmitate to unsaturated fatty acids differs in a Neurospora crassa mutant with impaired fatty acid synthase activity
Authors: Stafford Allan E. , McKeon Thomas A. , Goodrich‐Tanrikulu Marta
Published: November 2006

The Neurospora crassa cel (fatty acid chain elongation) mutant has impaired fatty acid synthase activity. The cel mutant requires exogenous 16:0 for growth and converts 16:0 to other fatty acids. In contrast to wild‐type N. crassa, which converted only 42% of the exogenous [7,7,8,8‐2H4]16:0 that was incorporated into cell lipids to unsaturated fatty acids, cel converted 72%. In addition, cel contains higher levels of 18:3δ9,12,15 than wild‐type, and synthesizes two fatty acids, 20:2δ11,14 and 20:3δ11,14,17, found at only trace levels in wild‐type. Thus, the Δ15‐desaturase activity and elongation activity on 18‐carbon polyunsaturated fatty acids are higher for cel than wild‐type. This altered metabolism of exogenous 16:0 may be directly due to impaired flux through the endogenous fatty acid biosynthetic pathway, or may result from altered regulation of the synthesis of unsaturated fatty acids in the mutant.
Identification of ceramide‐phosphorylethanolamine in Oomycete plant pathogens: Pythium ultimum, phytophthora infestans, and Phytophthora capsici
Authors: Moreau Robert A. , Young David H. , Danis Paul O. , Powell Michael J. , Quinn Chad J. , Beshah Kebede , Slawecki Richard A. , Dilliplane Robert L.
Published: November 2006

Cellular lipids were extracted from three species of Oomycete plant pathogens (Pythium ultimum, Phytophthora infestans, and Ph. capsici) and analyzed via normal‐phase high‐performance liquid chromatography with flame‐ionization detection. The most abundant polar lipids in each of the three species were the polar membrane lipids, phosphatidylethanolamine (PE), phosphatidylcholine, and a phosphosphingolipid that eluted soon after PE. Structural analysis via mass spectrometry and nuclear magnetic resonance spectrometry revealed that the phosphosphingolipid was ceramide phosphorylethanolamine (Cer‐PE). The most abundant molecular species of Cer‐PE in P. ultimum had a molecular weight of 670.5, contained an unusual 19‐carbon branched triunsaturated sphingoid (C19‐Δ4, 8, 10, 9‐methyl long‐chain base) and palmitic acid as the amidelinked fatty acid. The most abundant molecular species of Cer‐PE in Ph. infestans has a molecular weight of 714.5, contained a common 16‐carbon 1,3 di‐OH sphingoid, and erucic (cis 13‐docosenoic, C22‐Δ13) acid as the amide‐linked fatty acid. The Cer‐PE in Ph. capsici comprised a mixture of each of the two molecular species found in P. ultimum and Ph. infestans.
Lipids of Thermococcus hydrothermalis, an archaea isolated from a deep‐sea hydrothermal vent
Authors: Lattuati Agnès , Guezennec Jean , Metzger Pierre , Largeau Claude
Published: November 2006

The membrane lipids of a deep‐sea hydrothermal vent archaea, Thermococcus hydrothermalis, were isolated, purified, and structurally characterized. On the basis of acid methanolysis and spectroscopic studies, the polar lipids, amounting to 4.5% (w/w) of the dry cells, comprised diphytanyl glycerol diethers and dibiphytanyldiglycerol tetraethers, in a 45∶55 ratio. No cyclopentane ring was present in the tetraethers. From the neutral lipids, accounting for 0.4% (w/w) of the dry cells, besides low amounts of di‐ and tetraethers occurring in a free form, four acyclic tetraterpenoid hydrocarbons, di‐ and triunsaturated were identified. All were structurally related to lycopane. The presence of these hydrocarbons provides some evidence that lycopane, widely distributed in oceans, could be derived, at least partially, from the hydrocarbons synthesized by some thermophilic Archaea. Finally, analysis of the uninoculated culture medium indicates that fatty acid derivatives and some steroid and triterpenoid compounds identified in the lipidic extract of the archaea originated from the culture medium.
Fatty acid composition of the cellular slime mold Polysphondylium pallidum
Authors: Saito Tamao , Ochiai Hiroshi
Published: November 2006

The cellular slime mold Polysphondylium pallidum was grown upon Escherichia coli B/r, and the fatty acid compositions of total lipids obtained from vegetative amebae and aggregation‐competent cells were compared. Fatty acids isolated from vegetative cells included C‐17 and C‐19 cyclopropane fatty acids and also straight‐chain, saturated fatty acids. The cyclopropane fatty acids were derived from the ingested bacteria. Development of amebae to aggregation‐competent cells was accompanied by a substantial decrease in saturated cyclopropane fatty acids and a concomitant increase in unsaturated fatty acids and unsaturated cyclopropane fatty acids, mostly as 18∶3 (5,9,12). We report here the fatty acid composition and identify the occurrence of Δ5 desaturation of cyclopropane fatty acids, namely, 9,10‐methylene 5‐hexadecenoic acid and 11,12‐methylene 5‐octadecenoic acid. These fatty acids have not been reported previously in the related species Dictyostelium discoideum, which also feeds on E. coli B/r and has Δ5‐desaturation activity.
Isolation and structure of a new galactolipid from oat seeds
Authors: Hamberg Mats , Liepinsh Edvards , Otting Gottfried , Griffiths William
Published: November 2006

Seeds of oat (Avena sativa L.) were recently shown to contain significant quantities of a new hydroxy acid, (15R)‐hydroxy‐(9Z)‐(12Z)‐octadecadienoic acid (trivial name, avenoleic acid). In the present work, avenoleate was found to be mainly (63%) localized in the glycolipid fraction of oat seed lipids. Fractionation of the glycolipids by thin‐layer chromatography and reversed‐phase high‐performance liquid chromatography revealed the presence of a main molecular species which accounted for 20% of the total avenoleate content of oat seeds. Structural studies by chemical methods and mass spectrometry demonstrated that the avenoleate‐containing glycolipid was a galactolipid assembled of one molecule of avenoleic acid, two molecules of linoleic acid, two molecules of D‐galactose, and one molecule of glycerol. Degradation of the new galactolipid by chemical and enzymatic methods demonstrated the localization of acyl chains, i.e., linoleate at sn‐1 and linoleoylavenoleate at sn‐2. Nuclear magnetic resonance spectroscopy gave independent support for this structure and also demonstrated that the two galactoses formed an α‐d‐galactopyranosyl‐1‐6‐β‐d‐galactopyranosyl moiety which was bound to the sn‐3 position. Based on these experiments, the new galactolipid could be formulated as 1‐[(9′Z),(12′Z)‐octadecadienoyl]‐2‐[(15″R)‐{9″′Z), (12″′Z)‐octadecadienoyloxy}‐(9″Z),(12″Z)‐octadecadienoyl]‐3‐(α‐d‐galactopyranosyl‐1‐6‐β‐d‐galactopyranosyl)‐glycerol. Quantitatively, the amount of the avenoleate‐containing galactolipid was of the same order of magnitude as those of individual molecular species of digalactosyldiacylglycerol containing nonoxygenated acyl chains. The content of the new galactolipid in oat seeds was 0.5–0.6 mg per g of seed.
Stoichiometric and kinetic studies on Ginkgo biloba extract and related antioxidants
Authors: Shi Honglian , Niki Etsuo
Published: November 2006

Owing to increasing evidence showing the importance of lipid peroxidation in oxidative stress in vivo, the role and evaluation of antioxidants have received much attention. Ginkgo biloba extract (GBE), well‐known as an efficient drug against diseases induced by free radicals, has been suggested to exert its effect by antioxidant action. A method was established to determine the activity of GBE as a hydrogen donor by stoichiometric and kinetic studies, and GBE was compared with several other antioxidants such as α‐tocopherol, propyl gallate, and two kinds of flavonoids which are found in GBE, quercetin, and kaempferol. It was found that there were 6.62×1019 active hydrogens in 1g of GBE. Stoichiometric studies showed that one molecule of α‐tocopherol reacted with one molecule of galvinoxyl radical. For quercetin, kaempferol and propyl gallate, the experimental stoichiometric numbers were 4.0, 1.9, and 3.1, respectively. The rates of reaction of antioxidants with galvinoxyl in ethanol were determined spectrophotometrically, using a stopped‐flow technique. The second‐order rate constant, k2, obtained at 25°C was 0.13 (g/l)−1s−1 for GBE and 5.9×103, 2.1×103, 1.2×104, and 2.4×103 M−1s−1 for quercetin, kaempferol, propyl gallate, and α‐tocopherol, respectively. The second‐order rate constant, k2′, on the molar basis of active hydroxyl groups in the tested substances obtained at 25°C decreased in the order of propyl gallate > α‐tocopherol> quercetin>GBF∼kaempferol. This is the first study on GBE as an antioxidant which reports both stoichiometric and kinetic results.
Lipid molecular order in liver mitochondrial outer membranes, and sensitivity of carnitine palmitoyltransferase I to malonyl‐CoA
Authors: Zammit Victor A. , Corstorphine Clark G. , Kolodziej Marys P. , Fraser Fiona
Published: November 2006

Mitochondrial outer membranes were prepared from livers of rats that were in the normal fed state, starved for 48 h, or made diabetic by injection of streptozotocin. Membranes were also prepared from starved late‐pregnant rats. The latter three conditions have previously been shown to induce varying degrees of desensitization of mitochondrial overt carnitine palmitoyltransferase (CPT I) to malonyl‐CoA inhibition. We measured the fluorescence polarization anisotropy of two probes, 1,6‐diphenyl‐1,3,5‐hexatriene (DPH) and 1‐(4‐trimethylammoniumphenyl)‐6‐phenyl‐1,3,5‐hexatriene‐p‐toluenesulfonate (TMA‐DPH) which, when incorporated into membranes, report on the hydrophobic core and on the peripheral regions of the bilayer, respectively. The corresponding polarization indices (rDPH and rTMA‐DPH) were calculated. In membranes of all three conditions characterized by CPT I desensitization to malonyl‐CoA, rDPH was decreased, whereas there was no change in rTMA‐DPH, indicating that CPT I is sensitive to changes in membrane core, rather than peripheral, lipid order. The major lipid components of the membranes were analyzed. Although significant changes with physiological state were observed, there was no consistent pattern of changes in gross lipid composition accompanying the changes to membrane fluidity and CPT I sensitivity to malonyl‐CoA. We conclude that CPT I kinetic characteristics are sensitive to changes in lipid composition that are localized to specific membrane microdomains.
Biochemical and structural characterization of triacylglycerol lipase from Penicillium cyclopium
Authors: Ibrik Aida , Chahinian Henri , Rugani Nathalie , Sarda Louis , Comeau Louis‐Claude
Published: November 2006

An extracellular lipase, active on water‐insoluble triacylglycerols, has been isolated from Penicillium cyclopium. The purified enzyme has a molecular mass of 29 kDa by gel filtration and SDS‐polyacrylamide gel electrophoresis. It hydrolyzes emulsions of tributyrin, trioctanoin, and olive oil at the same rate as pancreatic lipase and shows very low activity against partial acylglycerols (monooctanoin and dioctanoin) and methyl esters. It is stable at 35°C for 60 min and has maximal activity in a pH range of 8–10. Hydrolysis of triacylglycerols by P. cyclopium lipase is inhibited by detergents such as Triton X‐100. Comparison of the sequence of the 20 first amino acid residues of P. cyclopium triacylglycerol lipase with other Penicillium lipases indicates a high homology with previously characterized lipases produced by P. expansum and P. solitum which are enzymes of comparable size and substrate specificity. Conversely, homology between P. cyclopium lipase and P. simplicissimum lipase, a nonspecific lipolytic enzyme, is low. Penicillium cyclopium triacylglycerol lipase shows no homology with P. camembertii lipase which is specific to monoacylglycerol and diacylglycerol.
Enhanced macrophage uptake of lipoprotein(a) after Ca2+‐induced aggregate‐formation
Authors: Tanaka Seiya , Yashiro Akira , Tasaki Hiromi , Nakashima Yasuhide
Published: November 2006

We tested the hypothesis that aggregated lipoprotein(a) [Lp(a)] is avidly taken up by macrophages. Lp(a) was isolated by sequential centrifugations and gel chromatography from a patient with high plasma levels of Lp(a) who was being treated with low density lipoprotein (LDL)‐apheresis. Aggregated Lp(a) was prepared by mixing native Lp(a) with 2.5 mmol/L CaCl2, and 54% of the 125I‐Lp(a) aggregated after interacting with CaCl2. The binding and degradation of aggregated Lp(a) in macrophages were 4.6‐ and 4.7‐fold higher than those of native Lp(a), respectively. An excess amount of LDL did not inhibit either increase. Cholesterol esterification in macrophages was markedly stimulated by aggregated Lp(a), and macrophages were transformed into foam cells. Cytochalasin B, a phagocytosis inhibitor, strongly inhibited the degradation and cholesterol esterification (78 and 83%, respectively). These findings suggested that aggregation may be partially involved in Lp(a) accumulation, thereby contributing to the acceleration of atherosclerosis.
Conjugated linoleic acids alter bone fatty acid composition and reduce ex vivo prostaglandin E2 biosynthesis in rats fed n‐6 or n‐3 fatty acids
Authors: Li Yong , Watkins Bruce A.
Published: November 2006

This study evaluated the effects of conjugated linoleic acids (CLA) on tissue fatty acid composition and ex vivo prostaglandin E2 (PGE2) production in rats given diets varying in n‐6 and n‐3 fatty acids. Four groups of rats were given a basal semipurified diet (AIN‐93G) containing 70 g/kg of added fat for 42 d. The fat treatments were formulated to contain CLA (0 vs. 10 g/kg of diet) and n‐6 (soybean oil having an n‐6/n‐3 ratio of 7.3) and n‐3 fatty acids (menhaden oil+safflower oil having an n‐6/n‐3 ratio of 1.8) in different ratios in a 2×2 factorial design. Fatty acids in liver, serum, muscle, heart, brain, spleen, and bone (cortical, marrow, and periosteum) were analyzed by capillary gas‐liquid chromatography. The various dietary lipid treatments did not affect growth; however, CLA improved feed efficiency. The CLA isomers were found in all rat tissues analyzed although their concentrations varied. Dietary CLA decreased the concentrations of 16∶1n−7, 18∶1, total monounsaturates and n−6 fatty acids, but increased the concentrations of n−3 fatty acids (22∶5n−3 and 22∶6n−3), and saturates in the tissues analyzed. Ex vivo PGE2 production in bone organ culture was decreased by n−3 fatty acids and CLA. We speculate that CLA reduced the concentration of 18∶1 fatty acids by inhibiting liver Δ9‐desaturase activity. The fact that CLA lowered ex vivo PGE2 production in bone organ culture suggests that these conjugated fatty acids have the potential to influence bone formation and resorption.
Cycloserine‐induced decrease of cerebroside in myelin
Authors: Miller Sheldon L. , Denisova Lyudmila
Published: November 2006

l‐Cycloserine has been shown specifically to lower brain cerebroside levels in vivo, but the extent to which the decrease in whole brain cerebroside content reflects lower myelin cerebroside levels is not known since a substantial portion of cerebroside is found in nonmyelin membranes. The present report demonstrates that chronically administered cycloserine lowers the proportion of cerebroside in rat brain myelin. Cycloserine‐induced decrease of myelin cerebroside should provide a useful tool in investigating the role of cerebroside in maintaining myelin stability.
A 13C nuclear magnetic resonance study of free fatty acid incorporation in acylated lipids in differentiating predipocytes
Authors: Guo Wen , Kirkland James L. , Corkey Barbara , Hamilton James A.
Published: November 2006

To understand the role of free fatty acid (FFA) incorporation in the accumulation of lipids in the adipocyte and ultimately in the development of obesity, 13C nuclear magnetic resonance was used to study lipid metabolism in differentiating preadipocytes. The incorporation of 13C=O‐labeled FFA into cellular lipids in primary cultured rat preadipocytes and 3T3L1 preadipocytes at different stages of differentiation was monitored by the 13C carbonyl chemical shift. Significant incorporation of palmitic acid into phosphatidylcholine in both the α and β acyl chain positions was found in cells at early stages of differentiation. At later differentiation stages or after extended incubation periods, most of the 13C=O signals were found in the triacylglycerol (TG) molecules. Unsaturated 13C=O‐labeled acyl chains were detected in the TG molecules when cells were incubated with saturated 13C=O‐labeled FFA, indicating that intracellular dehydrogenation had occurred in the 13C=O‐labeled palmitoyl chain. By using 13C‐labeled methyl myrisfate as an internal intensity reference, incorporation of 13C FFA into each acyl chain position of the major intracellular lipids was determined quantitatively.
Tocotrienols inhibit the growth of human breast cancer cells irrespective of estrogen receptor status
Authors: Nesaretnam Kalanithi , Stephen Ruth , Dils Ray , Darbre Philippa
Published: November 2006

Potential antiproliferative effects of tocotrienols, the major vitamin E component in palm oil, were investigated on the growth of both estrogen‐responsive (ER+) MCF7 human breast cancer cells and estrogen‐unresponsive (ER‐) MDA‐MD‐231 human breast cancer cells, and effects were compared with those of α‐tocopherol (αT). The tocotrienol‐rich fraction (TRF) of palm oil inhibited growth of MCF7 cells in both the presence and absence of estradiol with a nonlinear dose‐response but such that complete suppression of growth was achieved at 8 μg/mL. MDA‐MB‐231 cells were also inhibited by TRF but with a linear dose‐response such that 20 μg/mL TRF was needed for complete growth suppression. Separation of the TRF into individual tocotrienols revealed that all fractions could inhibit growth of both ER+ and ER‐ cells and of ER+ cells in both the presence and absence of estradiol. However, the γ‐ and δ‐fractions were the most inhibitory. Complete inhibition of MCF7 cell growth was achieved at 6 μg/mL of γ‐tocotrienol/δ‐tocotrienol (γT3/δT3) in the absence of estradiol and 10μm/mL of δT3in the presence of estradiol, whereas complete suppression of MDA‐MB‐231 cell growth was not achieved even at concentrations of 10μg/mL of δT3. By contrast to these inhibitory effects of tocotrienols, αT had no inhibitory effect on MCF7 cell growth in either the presence or the absence of estradiol, nor on MDA‐MB‐231 cell growth. These results confirm studies using other sublines of human breast cancer cells and demonstrate that tocotrienols can exert direct inhibitory effects on the growth of breast cancer cells. In searching for the mechanism of inhibition, studies of the effects of TRF on estrogen‐regulated pS2 gene expression in MCF7 cells showed that tocotrienols do not actviaan estrogen receptor‐mediated pathway and must therefore act differently from estrogen antagonists. Furthermore, tocotrienols did not increase levels of growth‐inhibitory insulin‐like growth factor binding proteins (IGFBP) in MCF7 cells, implying also a different mechanism from that proposed for retinoic acid inhibition of estrogen‐responsive breast cancer cell growth. Inhibition of the growth of breast cancer cells by tocotrienols could have important clinical implications not only because tocotrienols are able to inhibit the growth of both ER+ and ER‐ phenotypes but also because ER+ cells could be growth‐inhibited in the presence as well as in the absence of estradiol. Future clinical applications of TRF could come from potential growth suppression of ER+ breast cancer cells otherwise resistant to growth inhibition by antiestrogens and retinoic acid.
Effects of the ratio of polyunsaturated and monounsaturated fatty acid to saturated fatty acid on rat plasma and liver lipid concentrations
Authors: Chang Nai Wen , Huang Po Chao
Published: November 2006

The effects of dietary monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid+MUFA/saturated fatty acid (PUFA+MUFA/SFA) ratio on plasma and liver lipid concentrations were studied. In experiment I, when rats were fed with 40% fat (energy%, PUFA/SFA ratio 1.0) and 1% (w/w) cholesterol (C) diets for 21 d, a large amount of MUFA (28.1 energy%, PUFA+MUFA/SFA=5.7) in the diet was found to increase the plasma total C, triacylglycerol (TAG), and phospholipid (PL) as compared with the low‐MUFA diet (7.0 energy%, PUFA+MUFA/SFA=1.4). The plasma very low density lipoprotein (VLDL)‐C, VLDL‐TAG, VLDL‐PL, and low density lipoprotein (LDL)‐C increased significantly in the high‐MUFA diet group, but high density lipoprotein (HDL)‐C did not change significantly. The high‐MUFA diet resulted in greater accumulation of liver C but lesser accumulation of TAG. In experiment II, when dietary SFA was fixed at a certain level (13.2 energy%; PUFA+MUFA/SFA=2.0), rats given a larger amount of MUFA (23.1 energy%; PUFA/MUFA=0.2; MUFA/SFA=1.8) showed higher plasma and liver C levels than did the low‐MUFA diet (7.7 energy%; PUFA/MUFA=2.5; MUFA/SFA=0.6). When PUFA was fixed at a certain level (24.4 energy%), there was not a significant difference in the plasma C level between the high‐and low‐MUFA dietary groups (PUFA+MUFA/SFA=4.8 and 8.4), but the higher PUFA+MUFA/SFA diet, which was high in MUFA/SFA ratio, significantly decreased the plasma HDL‐C and TAG levels. However, when MUFA content was fixed at a certain level (16.4 energy%), no significant difference was observed between the two groups with different PUFA/SFA ratios of 0.2 and 4.1, but liver C level was raised in the higher PUFA/SFA diet. It appears that the PUFA/SFA ratio alone is unsuitable to predict the change of plasma C level, because a large amount of dietary MUFA may lead to an increase of plasma and liver lipids in rats. It seems that the prerequisites for keeping low plasma and liver C are (i) low MUFA/SFA ratio, (ii) high PUFA/MUFA ratio, and (iii) PUFA+MUFA/SFA ratio not to exceed 2.
Lipid‐lowering effects of WAY‐121,898, an inhibitor of pancreatic cholesteryl ester hydrolase
Authors: Krause Brian R. , Sliskovic Drago R. , Anderson Maureen , Homan Reynold
Published: November 2006

WAY‐121,898 is an inhibitor of pancreatic cholesteryl ester hydrolase (pCEH). After confirming its in vitro potency and relative lack of a major effect on acyl‐CoA:cholesterol acyltransferase (ACAT), it was found that this compound lowers plasma cholesterol in cholesterol‐fed, but not chow‐fed, rats. Measures of liver cholesteryl ester content and the direct determination of cholesterol absorption (lymph‐fistula model) show that inhibition of cholesterol absorption is at least one mechanism for the observed cholesterol lowering. However, WAY‐121,898 was also active when administered parenterally to cholesterol‐fed rats, and in cholesterol‐fed hamsters cholesterol‐lowering occurred with oral dosing despite no change in cholesterol absorption, suggesting other modes of action possibly relating to inhibition of liver CEH. Combination treatment in cholesterol‐fed rats with the ACAT inhibitor CI‐976 resulted in a greater‐than‐additive reduction in plasma cholesterol, imlying that both pCEH and ACAT may play a role in cholesterol absorption in this species. In rabbits, WAY‐121,898 prevented the rise in plasma cholesterol due to the feeding of cholesteryl ester but not in rabbits fed (free) cholesterol. In guinea pigs, the compound induced an increase in adrenal cholesteryl ester mass. Taken together, the overall profile in these animal models suggests that WAY‐121,898 inhibits more than just the intestinal (lumenal) pCEH, and that the role of this enzyme in cholesterol metabolism may be different within and across species, the former depending upon the dietary cholesterol load.
Effect of n−3 fatty acid supplementation on lipid peroxidation and protein aggregation in rat erythrocyte membranes
Authors: Ando Ken , Nagata Kunihide , Beppu Masatoshi , Kikugawa Kiyomi , Kawabata Terue , Hasegawa Kyoko , Suzuki Masao
Published: November 2006

Human erythrocytes in the circulation undergo dynamic oxidative damage involving membrane lipid peroxidation and protein aggregation during aging. The present study was undertaken to determine the effect of n−3 fatty acid supplementation on lipid peroxidation and protein aggregation in the circulation and also the in vitro susceptibility of rat erythrocyte membranes to oxidative damage. Wistar male rats were fed a diet containing n−6 fatty acid‐rich safflower oil or n−3 fatty acid‐rich fish oil with an equal amount of vitamin E for 6 wk. n−3 Fatty acid content in erythrocyte membranes of rats fed fish oil was significantly higher than that of rats fed safflower oil. The degree of membrane lipid peroxidation and protein aggregation of rats fed fish oil was not significantly higher than that of rats fed safflower oil when the amounts of phospholipid hydroper‐oxides, thiobarbituric acid‐reactive substances, and detergent‐insoluble protein aggregates were measured. When isolated erythrocytes were oxidized under aerobic conditions in the presence of Fe(III), the degree of membrane lipid peroxidation of erythrocytes from rats fed fish oil was increased to a greater extent than that of rats fed safflower oil, whereas the degree of membrane protein aggregation of both groups was increased in a similar extent. Hence, n−3 fatty acid supplementation did not affect lipid peroxidation and protein aggregation in membranes of circulating rat erythrocytes, and the supplementation increased the susceptibility of isolated erythrocytes to lipid peroxidation, but not to protein aggregation, under the aerobic conditions. If a sufficient amount of vitamin E is supplied, n−3 fatty acid supplementation may give no undesirable oxidative effects on rat erythrocytes in the circulation.
Modification of milk formula to enhance accretion of long‐chain n−6 and n−3 polyunsaturated fatty acids in artificially reared infant rats
Authors: Yeh Yu‐Yan , Yeh Shaw‐Mei , Lien Eric L.
Published: November 2006

Artificially reared infant rats were used to determine the effects of long‐chain polyunsaturated fatty acid (LCP‐UFA) supplementation on blood and tissue concentrations of arachidonic acid (AA) and docosahexaenoic acid (DHA). Beginning at 7 d of age, infant rats were fed for 10 d with rat milk formulas supplemented with AA at 0,0.5 and 1.0%, or supplemented with DHA at 0,0.5 and 1.0% of total fatty acid. The supplementation of AA increased accretion of the fatty acid in tissue and blood phospholipids with a maximum increase of 9% in brain, 15% in liver, 25% in erythrocytes, and 43% in plasma above the values of unsupplemented infant rats. Rat milk formula containing 1.0% of AA had no added benefits over that containing 0.5% of AA. The supplementation of DHA increased phospholipid DHA by a maximum of 24% in brain, 87% in liver, 54% in erythrocytes, and 360% in plasma above the unsupplemented control. The increase in tissue and blood DHA was concentration‐dependent on formula fatty acid. Brain phosphatidylcholine and phosphatidylethanolamine were similarly enriched with AA and DHA by supplementation of the corresponding fatty acids. In general the observed increase of AA was accompanied by a decrease in 16:0, 18:1n−9, and/or 18:2n−6, whereas the increased DHA was associated with a reduction of 18:1n−9, 18:2n−6, and/or 20:4n−6. Clearly, infant rats were more responsive to DHA than AA supplementation, suggesting a great potential of dietary manipulation to alter tissue DHA concentrations. However, the supplementation of DHA significantly decreased tissue and blood AA/DHA ratios (wt%/wt%), whereas there was little or no change in the ratio by AA supplementation. Although the physiological implications of the levels of AA and DHA, and AA/DHA ratios achieved under the present experimental conditions are not readily known, the findings suggest that artificial rearing could provide a suitable model to investigate LCPUFA requirements using various sources of AA and DHA in rats.
Simultaneous quantitation of ceramides and 1,2‐diacylglycerol in tissues by latroscan thin‐layer chromatography‐flame‐ionization detection
Authors: Okumura Kenji , Hayashi Kazunori , Morishima Itsuro , Murase Kichiro , Matsui Hideo , Toki Yukio , Ito Takayuki
Published: November 2006

Ceramides and 1,2‐diacylglycerol have been demonstrated in intracellular signaling pathways. A method of simultaneous mass determination of ceramides and 1,2‐diacylglycerol in tissues was developed using the latroscan which combines thin layer chromatography and flame ionization detection (TLC/FID) techniques. Because of relatively low amounts of these components in tissues, the fraction of nonpolar lipids, which included ceramides and glycerides, was eluted with chloroform/acetone mixture (3∶1, vol/vol) through a silicic acid column to eliminate the polar phospholipids. Development of Chromarods was carried out using three solvent systems in a four‐step development technique. The relationship of the peak area ratio to weight ratio compared with cholesteryl acetate added as an internal standard was linear. The amount of ceramides increased with incubation of rat heart homogenate and human erythrocyte membranes in the presence of sphingomyelinase (E.C. 3.1.4.12). The latroscan TLC/FID system provided a quick and reliable assessment of ceramides and 1,2‐diacylglycerol.
A gas chromatographic‐mass spectrometric method using a PoralPLOT column for the detection of hydroperoxide lyase in Chlorella pyrenoidosa
Authors: Nuñez Alberto , Foglia Thomas A. , Piazza George J.
Published: November 2006

A gas chromatographic‐mass spectrometric (GC‐MS) method using a PoraPLOT Q column was developed for the analysis and identification of the volatile products produced by the action of hydroperoxide lyase (HPLS) upon 13‐hydroperoxylinoleic or 13‐hydroperoxylinolenic acids. The developed procedure required no derivatization, was not affected by the presence of water, did not require cryogenic conditions to be maintained during injection, and allowed for the quantiation of most products. An acetone powder preparation of Chlorella pyrenoidosa cells was triturated with borate buffer pH=8.0, and the mixture centrifuged at 12,000×g. The supernatant and pellet were assayed for HPLS activity by GC‐MS analysis of the volatile products given by linoleic acid hydroperoxide. The data showed that the majority of HPLS activity resides in the pellet fraction, and that the primary volatile component was pentane, with smaller amounts of 2‐(Z)‐pentene and 1‐pentene being produced. The fact that HPLS activity resides in the water‐insoluble fraction of the acetone powder suggests that HPLS from Chlorella is a membrane‐associated enzyme. This investigation also determined that a spectrophotometric assay using alcohol dehydrogenase for measuring HPLS activity was not specific, but measured enzymatic activity other than HPLS.