bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of 25‐hydroxycholesterol on cholesteryl ester formation in Caco‐2 cells
Authors: Kusuhara Hidenobu , Shimada Osafumi , Inui Jun
Published: November 2006

Incubation of Caco‐2 cells, a human intestinal cell line, with 25‐hydroxycholesterol (25‐HOC) markedly enhanced cellular cholesteryl ester formation determined by incorporation of [14C]oleic acid into intracellular cholesteryl [14C]oleate. The stimulation by 25‐HOC of cholesteryl ester formation was suppressed by staurosporine, a kinase inhibitor, but not by cycloheximide or actinomycin D. The specific activity of microsomal acyl‐coenzyme A:cholesterol acyltransferase (ACAT) increased two‐fold in cells treated with 10 μM 25‐HOC for 5 h. ACAT activity decreased when microsomes were incubated without sodium fluoride, a phosphatase inhibitor, but the decrease in ACAT activity in cells stimulated with 25‐HOC was more pronounced. The results suggest that protein phosphorylation may be involved in the stimulation of cholesteryl ester formation by 25‐HOC in Caco‐2 cells.
Analysis of exoskeleton triglycerides ofLeptinotarsa decemlineata say
Authors: Dubis Eligiusz , Poplawski Janusz , Wróbel Jerzy T. , Kúsmierz Józef , Maliński Edmund , Szafranek Janusz
Published: November 2006

The exoskeleton triglyceride fraction isolated from the Colorado beetle,Leptinotarsa decemlineata Say, was studied. Analysis of the hydrolysis products and direct capillary column gas chromatography showed that, apart from triglycerides, the fraction contained no diol lipids. The composition of the natural triacylglccerol mixture was assessed by gas chromatography and by field desorption and electron impact mass spectrometry. Triglycerides isolated from the Colorado beetle are typical glycerol triesters, with long chain (mainly oleic or palmitic) fatty acids. The structures of two major components (91%), i.e., trioleoyl glycerol and 1,2‐dioleoyl‐3‐palmitoyl glycerol, were determined.
A liquid crystalline phase in human intestinal contents during fat digestion
Authors: Holt Peter R. , Fairchild Beatrice M. , Weiss Jordan
Published: November 2006

A viscous, gel‐like phase was found in ultracentrifuged human upper intestinal contents collected during rapid fat lipolysis. This “gel” phase was layered in the ultracentrifuge tube between the micellar and oil phase. The gel contained lipids typical of the micellar and not the oil phase. The concentration of these lipids was higher and the ratio of total bile salts to lipids was lower in the gel than in the micellar phase. The gel, unlike the micellar phase, was birefringent between crossed polarizers. These data demonstrate that lipids in this gel phase can form liquid crystals in the gut during fat digestion.
The peroxidizing effect of α‐tocopherol on autoxidation of methyl linoleate in bulk phase
Authors: Terao J. , Matsushita S.
Published: November 2006

In order to understand the effect of α‐tocopherol on the autoxidation mechanism of edible oil under storage conditions, methyl linoleate was allowed to autoxidize at 50 C in bulk phase without any radical initiator. The reaction was monitored by determining the production of four isomeric hydroperoxides (13‐cis,trans; 13‐trans,trans; 9‐cis,trans; 9‐trans,trans) by high performance liquid chromatographic analysis after reduction. In the absence of α‐tocopherol, the rate of autoxidation depended on the sample size, and the duration of the induction period was affected by the initial level of hydroperoxides. However, the distribution of c‐t and t‐t hydroperoxide isomers remained constant during the propagation period regardless of the sample size. The addition of α‐tocopherol at 0.1 and 1.0% caused a linear increase in the amount of hydroperoxides and elevated the distribution of the c‐t isomers. The rate of hydroperoxidation appeared to be governed by the initial concentration of α‐tocopherol rather than the sample size or the initial hydroperoxide level. This peroxidizing effect of α‐tocopherol was suppressed by the presence of ascorbyl palmitate. A mechanism in which chromanoxy radical participates is proposed for the effect of α‐tocopherol on lipid autoxidation in bulk phase. It is therefore suggested that α‐tocopherol at high concentrations influences the mechanism of autoxidation of edible oil.
Enzymic oxidation of linolenic acid to 1,Z‐5‐octadien‐3‐ol, Z‐2,Z‐5‐octadien‐1‐ol and 10‐oxo‐E‐8‐decenoic acid by a protein fraction from mushrooms (Psalliota bispora)
Authors: Wurzenberger Margot , Grosch Werner
Published: November 2006

The pathway for the oxidative cleavage of linolenic acid was investigated using a protein fraction from mushrooms (Psalliota bispora) as the enzyme source. Incubation of the protein fraction with linolenic acid resulted in 1,Z‐5‐octadien‐3(R)‐ol and Z‐2,Z‐5‐octadien‐1‐ol (in a 3∶2 ratio) and 10‐oxo‐E‐8‐decenoic acid (10‐ODA). Experiments with molecular oxygen‐18 indicated that the oxygen in the hydroxy group of both octadienols originates from the gaseous phase and not from water. The protein fraction was incubated with the individual 9‐, 10‐, 12‐, 13‐, 15‐ and 16‐hydroperoxide isomers of linolenic acid. Only the 10‐hydroperoxy‐E‐8,Z‐12,Z‐15‐octadecatrienoic acid (10‐HPOT) served as substrate, and was cleaved into the two octadienols and the 10‐ODA. This result suggests that in the oxidation of linolenic acid by the mushroom fraction the 10‐HPOT is an intermediate cleaved by a hydroperoxide lyase into the octadienols and the 10‐ODA. Model experiments in which the methyl esters of both 10‐hydroperoxy‐E‐8,Z‐12‐octadecadienoic acid (10‐HPOD) and 10‐HPOT were treated with the Lewis acid BF3 yielded only various C9 compounds. It was therefore concluded that the reactions of the mushroom hydroperoxide lyase cannot be explained by the heterolytic rearrangement mechanism, which was proposed for the corresponding plant enzyme. β‐Scission of the 10‐HPOD and the 10‐HPOT explains the reactions of the mushroom hydroperoxide lyase and is discussed in detail.
Calories, fat and cancer
Authors: Kritchevsky David , Weber Maxine M. , Buck Carrie L. , Klurfeld David M.
Published: November 2006

The experiments reported are part of our effort to dissociate the tumor‐enhancing effects of dietary fat and high caloric intake. Rats either were fed ad libitum diets containing 4% corn oil or their calories were restricted by 40% and their diets contained 13.1% corn oil. Incidence of 7,12‐dimethylbenz(a)anthracene (DMBA)‐induced mammary tumors was 80% in rats fed ad libitum and 20% in those fed the calorie‐restricted diets. Incidence of 1,2‐dimethylhydrazine (DMH)‐induced colon tumors was 100% in rats fed ad libitum and 53% in those whose caloric intake was restricted by 40%. The tumor yield (tumors per tumor‐bearing rat) was significantly lower in rats on caloric restriction. In another series, rats were fed diets containing 5, 15 or 20% corn oil ad libitum or were fed calorie‐restricted (by 25%) diets which provided 20 or 26.6% corn oil (therefore, the same absolute amount of fat was consumed in each of the pair‐fed groups). Tumor incidence and tumor yield in the two calorie‐restricted groups were similar to those seen in the rats fed 5% fat ad lititum; tumor burden (total g of tumor) was 45–65% lower in the calorie‐restricted rats. The data suggest that caloric intake is a more stringent determinant of tumor growth than fat intake.
Effects of high levels of dietary fats on the growth of azaserine‐induced foci in the rat pancreas
Authors: Roebuck B. D.
Published: November 2006

Azaserine induced two phenotypically different populations of foci, namely, acidophilic and basophilic foci. The effects of dietary modification during the post‐initiation phase of carcinogesis were examined. A diet of 20% (w/w) unsaturated fat (unsat) compared to a 20% saturated fat (sat) diet or a control diet (5% unsaturated fat) increased the number of acidophilic foci, as well as the thymidine labeling index (LI) of their nuclei. While the basophilic foci are carcinogen‐induced and at 2.5 mo post‐initiation have a similarly high growth rate to the acidophilic foci, this rate is not sustained as indicated by examination of both the LI and mean size of foci at 4 mo post‐initiation.
Dietary polyunsaturated fat in relation to mammary carcinogenesis in rats
Authors: Braden Laura M. , Carroll Kenneth K.
Published: November 2006

High fat diets promote the development of mammary tumors induced in rats by 7,12‐dimethylbenz(a)anthracene (DMBA), and polyunsaturated fats are more effective than saturated fats. This difference is related to the linoleic acid content of polyunsaturated vegetable oils, but the amount of linolealte required for maximum tumor promotion appears to be higher than indicated by earlier experiments. Comparison of the effects of a polyunsaturated vegetable oil (corn oil) containing linoleate with a fish oilo (menhaden oil) containing polyunsaturated fatty acids derived from linolenic acid showed that higher dietary mammary tumors, while corresponding levels of menhaden oil had an inhibitory effect. This is further evidence that promotion of mammary tumorigenesis by polyunsaturated vegetable oils may be mediated by prostaglandins or other biologically active eicosanoids derived from n−6 fatty acids.
Ether lipid derivatives: Antineoplastic activity in vitro and the structure‐activity relationship
Authors: Berdel Wolfgang E. , Von Hoff Daniel D. , Unger Clemens , Schick Hans D. , Fink Ulrich , Reichert Anneliese , Eibl Hansjorg , Rastetter Johann
Published: November 2006

The antineoplastic activity of two ether lipid derivatives, the alkyl‐lysophospholipid derivative (ALP) ET‐18‐OCH3 and the ether‐linked lipoidal amine CP‐46,665 was tested in a human tumor clonogenic assay (HTCA) in vitro. CP‐46,665 suppresed the colony formation of various human tumors with a slight dose response relation after 1 hr incubation and with a clear optimum (85% response rate) after continuous exposure in the higher dose range tested (10 μg/ml). ET‐18‐0CH3 did not have substantial activity after 1 hr of incubation. However, when continuous exposure to the compound was used, ET‐18‐OCH3 seemed to have a modest dose response effect and yielded a response in about 60% of the tumor cell samples tested in the higher dose range (10 μg/ml). Thus, both compounds have in vitro antitumor activity in the HTCA within a dose range of 1–10 μg/ml, especially during continuous exposure. The tumor specific type activity was found in breast cancer, ovarian cancer, lung cancer and mesothelioma. Both compounds caused decreases in colony formation down to the 0%, 2% and 4% levels. In a comparison of specimens in which both compounds were used, only one of five times showed a discordance in sensitivity or resistance; therefore the compounds appear similar in their in vitro activity.

In a second set of experiments we tested the structure‐activity relationship among a variety of ALP in the [3H]thymidine incorporation assay after incubation with HL‐60 leukemic blasts and other neoplastic cells from human origin. From these studies it can be concluded that in the ALP the alkyl linkage in the sn‐1 position is a necessary prerequisite for cytotoxicity; furthermore, in the majority of tumors tested the substitution of the sn‐2 position to prevent reacylation of the molecule is necessary for cytotoxicity.
The metabolism of malondialdehyde
Authors: Draper H. H. , McGirr L. G. , Hadley M.
Published: November 2006

Interest in malondialdehyde (MDA) metabolism stems from its formation as a product of lipid peroxidation in the diet and in the tissues; its reactivity with functional groups of nucleic acid bases, proteins and phospholipids; its mutagenicity in bacteria, and its reported skin and liver carcinogenicity in animals. Administration of the Na enol salt of MDA in the drinking water of mice over a range of 0.1–10.0 μg/g/day for 12 mo produced dose‐dependent hyperplastic and neoplastic changes in liver nuclei and increased mortality at the highest level but produced no gross hepatic tumors. Addition of MDA to the medium of rat skin fibroblasts grown in culture caused nuclear abnormalities at concentrations as low as 10−6M despite an uptake of only 4%. [1,3‐14C]MDA was rapidly oxidized to [14C]acetate in rat liver mitochondria and to14CO2 in vivo: however, ∼10% of the radioactivity was recovered in the urine. Chromatographic analysis of rat urine revealed the presence of several compounds which yield MDA on acid hydrolysis. Total MDA excretion increased in response to conditions which stimulate lipid peroxidation in vivo, including vitamin E deficiency, Fe or CCl4 administration, and enrichment of the tissues with PUFA. N‐acetyl‐e‐(2‐propenal)lysine was identified as a major urinary metabolite of MDA in rat and human urine. This compound is derived primarily from N‐α‐(2‐propenal)lysine released in digestion as a product of reactions between MDA and the ε‐amino groups of N‐terminal lysine residues in food proteins. However, its presence in the urine of animals fasted or fed MDA‐free diets indicates that it is also formed in vivo. Identification of the metabolites of MDA excreted in the urine may provide clues to the mechanisms of cellular damage caused by this compound in the tissues.
Ion‐pair high‐performance liquid chromatography of bile salt conjugates: Application to pig bile
Authors: Legrand‐Defretin Véronique , Juste Catherine , Henry Robert , Corring Tristan
Published: November 2006

The high‐performance liquid chromatographic separation and quantitation of conjugated bile salts from pig bile is reported. Synthetic standards and bile samples were chromatographed on a C18 reversed phase column using acetonitrile/water/tetrabutyl ammonium phosphate as an isocratic mobile phase at a flow rate of 1 mL/min. Detection of the ion‐pairs was at 214 nm. The method permits efficient separation of all conjugated pig biliary bile salts without prior modification or treatment of the samples. Analysis of 12 pig biles showed that 85% of the bile salts are conjugated to glycine. The three main conjugated bile salts were glyco‐3α,6α,7α‐trihydroxy‐5β‐cholanoic acid (GHC), glyco‐3α,7α‐dihydroxy‐5β‐cholanoic acid (GCDC), and glyco‐3α,6α‐dihydroxy‐5β‐cholanoic acid (GHDC). Glyco‐3α‐hydroxy‐6‐oxo‐5β‐cholanoic acid (G3α6oxo), tauro‐3α,7α‐dihydroxy‐5β‐cholanoic acid (TCDC), tauro‐3α,6α,7α‐trihydroxy‐5β‐cholanoic acid (THC), and tauro‐3α,6α‐dihydroxy‐5β‐cholanoic acid (THDC) were found to contribute each for 4 to 5% ot the total. An excellent correlation was found between the sum of conjugated bile salts quantitated by high‐performance liquid chromatography (HPLC) and values obtained by conventional enzymatic assay. Simplicity, efficiency and relative rapidity of the method render it suitable for routine analyses.
Diversity of sterol biosynthetic capacity in the caryophyllidae
Authors: Salt Thomas A. , Xu Sihua , Patterson Glenn W. , Adler John H.
Published: November 2006

The order Caryophyllales, along with its two associated orders, the Polygonales and Plumbaginales, comprise the angiosperm subclass Caryophyllidae. We have now characterized the sterol compositon of 231 members of this subclass. This includes 210 species and 21 cultivars in 108 genera within the 14 families of these three orders. From these data, clear differences in biosynthetic capability and putative relationships between taxa have been established. Members of the two monofamilial orders (Polygonales and Plumbaginales) contain Δ5‐sterols in ratios typical of “main line” angiosperms. Members of families in the Caryophyllales contain Δ5‐sterols, or Δ7‐sterols or mixtures of Δ5‐ and Δ7‐sterols. In the majority of species where Δ7‐sterols are the dominant sterols produced, trace amounts to almost equal amounts of Δ5‐sterols are also present. Replicate samples of many of these species have shown that the ratio of Δ5‐sterols to Δ7‐sterols in these species is stable over time and/or location. From these data, it appears that the conversion of Δ7‐sterols to Δ5‐sterols is highly regulated in the majority of species within this order. In these families, similarities in sterol composition correlate well with taxonomic relatedness. Relationships between these taxa with respect to biosynthetic capability can now be postulated.
Neutral sterol metabolism in insects
Authors: Svoboda James A. , Feldlaufer Mark F.
Published: November 2006

Since they are unable to biosynthesize sterols, many phytophagous and omnivorous insects satisfy their cholesterol requirement by side chain dealkylation of the C‐24 alkyl group of dietary C28 and C29 phytosterols. However, not all insects that can dealkylate the phytosterol side chain produce cholesterol. In addition, certain insects,e.g., some Hymenoptera, Hemiptera, and Diptera, are unable to dealkylate the sterol side chain. Although C27 ecdysteroids (molting hormones), which are biosynthesized from cholesterol, are the major ecdysteroids in most insects, many of those species that are unable to dealkylate phytosterols utilize campesterol as a precursor for the C28 ecdysteroid makisterone A. The considerable diversity of steroid utilization between certain insect species makes it difficult to generalize about insect steroid biochemistry. The ability to disrupt certain unique aspects of steroid utilization and metabolism in insects might be exploited for developing new insect control technology.
Metabolism of plant sterols by nematodes
Authors: Chitwood David J. , Lusby William R.
Published: November 2006

Parasitic nematodes do not biosynthesize sterolsde novo and therefore possess a nutritional requirement for sterol, which must be obtained from their hosts. Consequently, the metabolism of phytosterols by plant‐parasitic nematodes is an important process with potential for selective exploitation. The sterol compositions of several species of plant‐parasitic nematodes were determined by capillary gas chromatography‐mass spectrometry and compared with the sterol compositions of their hosts. Saturation of the phytosterol nucleus was the major metabolic transformation performed by the root‐knot nematodesMeloidogyne arenaria andM. incognita and the corn root lesion nematode,Prytalenchus agilis. In addition to saturation, the corn cyst nematode,Heterodera zeae, dealkylated its host sterols at C‐24. Because free‐living nematodes can be cultured in sterol‐defined artificial medium, they have been successfully used as model organisms for investigation of sterol metabolism in plant‐parasitic nematodes. Major pathways of phytosterol metabolism inCaenorhabditis elegans, Turbatrix aceti andPanagrellus redivivus incleded C‐24 dealkylation and 4α‐methylation (a pathway unique to nematodes).C. elegans andT. aceti introduced double bonds at C‐7, andT. aceti andP. redivivus saturated the sterol nucleus similarly to the plant‐parasitic species examined. Several azasteroids and long‐chain dimethylalkylamines inhibited growth and development ofC. elegans and also the Δ24‐sterol reductase enzyme system involved in the nematode C‐24 dealkylation pathway.
Cloning and disruption of the yeast C‐8 sterol isomerase gene
Authors: Ashman William H. , Barbuch Robert J. , Ulbright Corinne E. , Jarrett Harry W. , Bard Martin
Published: November 2006

The yeastERG2 gene codes for the C‐8 sterol isomerase, an enzyme required for the isomerization of the δ8 double bond to the δ7 position in ergosterol biosynthesis. TheERG2 gene was cloned by complementation of a C‐8 sterol isomerase mutant strain (erg2). The complementing region of DNA required to restore ergosterol synthesis toerg2 was limited to a 1.0 kbStuI‐BglII fragment. In order to determine whether theERG2 gene was essential for yeast viability, aLEU2 gene was inserted into theNdeI site (made blunt) of this 1.0 kb fragment. Transformation of a wild type diploid strain with theERG2 substituted DNA resulted in the generation of viable haploids containing theerg2 null allele (erg2–4∶∶Leu2). These results suggest that the C‐8 sterol isomerase activity is not essential for yeast cell viability. This disruption represents the second ergosterol biosynthetic gene in the distal portion of the pathway to be disrupted without adversely affecting cell viability.
Aspects related to mevalonate biosynthesis in plants
Authors: Bach Thomas J. , Boronat Albert , Caelles Carmé , Ferrer Albert , Weber Thomas , Wettstein Annette
Published: November 2006

We purified and characterized a membrane‐associated enzyme system from radish (Raphanus sativus L.) that is capable of converting acetyl‐CoA into 3‐hydroxy‐3‐methylglutaryl‐coenzyme A (HMG‐CoA). The enzyme system apparently comprises acetoacetyl‐CoA thiolase (EC 2.3.1.9) and HMG‐CoA synthase (EC 4.1.3.5). Its activityin vitro can be strongly stimulated by FeII. When ferrous ions are applied chelated with ethylenediaminetetraacetate, citrate or adenosine 5′‐triphosphate (ATP), the stimulation is further increased. Stimulation is due to a higher catalytic efficiency as indicated by an increase in Vmax, whereas the affinity of the enzyme towards acetyl‐CoA remains constant (Km=6 μM). A considerable portion of HMG‐CoA lyase activity is associated with the same membranes. HMG‐CoA lyase (EC 4.1.3.4) is also solubilized and partially co‐purified. Its activity requires comparatively high concentrations of Mg2+. The conversion of HMG‐CoA to mevalonic acid is catalyzed by HMG‐CoA reductase (EC 1.1.1.34) that is associated with the same membranes. By cDNA encoding theArabidopsis HMG‐CoA reductase, we isolated a corresponding gene from a cDNA library newly established from etiolated radish seedlings. This full‐length cDNA, referred to as λcRS3, encodes a polypeptide of 583 amino acids with a molecular mass of about 63 kDa. The hydropathy profile suggests the presence of two hydrophobic membrane‐spanning domains within the N‐terminal 165 amino acids. The carboxy‐terminal part, where the catalytic site resides, is highly conserved in all eukaryotic HMG‐CoA reductase genes sequenced so far.
On the structure, biosynthesis, function and phylogeny of isoarborinol and motiol
Authors: Nes W. David , Wong Rosalind Y. , Griffin Jane F. , Duax William L.
Published: November 2006

The solid‐state conformations of the C‐3 acetates of two isomeric hopanoids—1, isoarborinol (D∶C‐friedo‐B1∶A1‐3β,5α,8α,10β,13β,14α,17β,18α,21β) and 2, motion (D∶C‐friedo‐B1∶A1‐neogammacer‐7(8)‐en‐3β‐ol[3β,5α,9α,10β,13α,14β,17α,18β,21α])—have been determined by X‐ray crystallography. The data show that whereas both molecules are planar, 1 orients into a chair‐halfchair‐chair‐chair‐halfchair conformation while 2 orients into a chair‐sofa‐twist‐halfchair‐halfchair conformation. To explain the biogenesis of 1 and 2 from squalene oxide, a step‐wise mechanism is proposed which proceeds through the protosteroid cation (for 1) and dammarenyl cation (for 2). After ring enlargement from the corresponding 13(17)bond followed by concerted 1,2‐migrations and loss of the 11β‐H and 7β‐H as protons, respectively, a 9,11‐double bond (in 1) and a 7,8‐double bond (in 2) is introduced into the nucleus. The mechanism is discussed in relation to the classical view of a non‐stop cyclization process where, for example, squalene oxide folds in a chair‐chair‐chair‐chair‐boat conformation to give a cyclized product (motiol) presumably with the same conformational disposition as the cyclizing material. The three‐dimensional geometry of 1 and 2 was found to be structurally dissimilar from sterols. For instance, 1 and 2 are shorter and volumetrically smaller molecules than cholesterol, and this may explain their diminished importance as membrane inserts compared with sterols in eukaryote evolution.
Fatty acid metabolism inTaphrina deformans treated with sterol biosynthesis inhibitors
Authors: Weete J. D. , Sancholle M. , Patterson K. A. , Miller K. S. , Huang M. Q. , Campbell F. , Van den Reek M.
Published: November 2006

Changes in the unsaturated fatty acid content of the fungusTaphrina deformans as a function of growth, temperature, and sterol content were investigated. It was found that the highest growth rate was accompanied by a relatively high degree of fatty acid unsaturation (18∶1<18∶2+18∶3) and low sterol (brassicasterol) content. Also, a substantial shift in the degree of unsaturation from mainly 18∶2+18∶3 to 18∶1 occurred in the later stages (mid‐linear) of culture development. Cells readily adapted from 18°C to 13°C, and exhibited a higher growth rate at the lower temperature after a period of acclimation. Growth was readily inhibited by the sterol biosynthesis inhibitors propiconazole and naftifine, which blocked brassicasterol biosynthesis at C‐14 demethylation and squalene epoxidation, respectively. Growth was also inhibited by tunicamycin which did so without affecting sterol content. The shift in degree of fatty acid unsaturation did not occur in cells from cultures at reduced temperature or treated with any of the inhibitors. Since tunicamycin did not affect the sterol content, delay in the shift in the degree of fatty acid unsaturation was attributed to factors other than a reduction in sterol content.
A facile chemical synthesis of cholest‐4‐en‐3‐one. Carbon‐13 nuclear magnetic resonance spectral properties of cholest‐4‐en‐3‐one and cholest‐5‐en‐3‐one
Authors: Parish Edward J. , Honda Hiroshi , Chitrakorn Sarawanee , Livant Peter
Published: November 2006

This paper presents a simplified method for the chemical synthesis of cholest‐4‐en‐3‐one, which is a naturally occurring steroid and a salient intermediate in steroid chemistry. Pyridinium chlorochromate (PCC) in refluxing benzene has been found to be an effective and convenient reagent for the oxidation and concomitant isomerization of cholest‐5‐en‐3β‐ol (cholesterol) to cholest‐4‐en‐3‐one in high yield. Also described are the carbon‐13 nuclear magnetic resonance spectral properties of cholest‐4‐en‐3‐one and of the isolated reaction intermediate cholest‐5‐en‐3‐one.
Hydroxylation of fatty acids and alcohols by hepatic microsomal cytochrome P‐450 system from the Mongolian gerbil
Authors: Miura Yoshiro , Hisaki Harumi , Siems Werner , Oda Sen‐ichi
Published: November 2006

The liver microsomes of the Mongolian gerbilMeriones unguiculatus catalyzed the hydroxylation of various saturated fatty acids (C8−C18), alcohols (C12 and C16) and hydrocarbon (C12) to the corresponding ω‐ and (ω‐1)‐hydroxy derivatives. Lauric acid was hydroxylated most effectively among saturated fatty acids and the order of activity as hydroxylation substrates was C12>C14>C13>C16>C10>C18>C8. The specific activity of laurate hydroxylation (5.99 nmol/mg microsomal protein/min) in gerbil liver microsomes was higher than that observed in other species. 1‐Dodecanol was also hydroxylated very effectively (4.58 nmol/mg microsomal protein/min) by gerbil liver microsomes, but in general the hydroxylation rates for fatty alcohols were much lower than those for the corresponding acids. It was found from both inhibitor and cofactor studies that the enzyme catalyzing the hydroxylation of fatty acids and alcohols in the liver microsomes of the Mongolian gerbil was a typical cytochrome P‐450‐linked monooxygenase, and at least two different cytochrome P‐450 species were involved in the hydroxylation.
Properties of monoacylglycerol lipase in rabbit aorta
Authors: Hee‐Cheong Mariette , Severson David L.
Published: November 2006

Monoacylglycerol lipase activity was characterized in a soluble preparation from rabbit aorta (intima‐media) obtained by combining a 100,000×g supernatant fraction with activity solubilized from the 100,000×g precipitate fraction by treatment with Triton X‐100. Rates of hydrolysis with 1‐monoolein and 2‐monoolein substrates were nearly identical. 1‐Monoolein was a competitive inhibitor (Ki 65 μM) of 2‐monoolein hydrolysis. 2‐Monoolein and 2‐monopalmitin were both hydrolyzed more rapidly than 2‐monoarachidonin. Lipase activity measured with a 2‐monoolein substrate was inhibited by the addition of oleate, NaF and CaCl2 to the assay. Preincubation of the lipase preparation with p‐bromophenacyl bromide resulted in a potent inhibition of lipase activity; this inhibition could be prevented by dithiothreitol.
The result of feeding palmitoyl glycerol on lymph and plasma lipids
Authors: Trumbo Paula , Sutherland‐Smith Margaret , Kitchell Barbara , Tove Samuel B.
Published: November 2006

Palmitoyl glycerol is toxic when fed to mice, but the toxicity is alleviated by supplementing the toxic diet with 2–4% oleate or linoleate at the expense of sucrose. Lipid and fatty acid composition of lymph and plasma were studied in mice fed chow and palmitoyl glycerol diets to help explain the toxicity mechanism. When mice were fed chow, intestinal lymph contained a high proportion of saturated fatty acids; when they were given palmitoyl glycerol, the proportion approached 90% saturated fatty acids. The cholesteryl ester fraction was higher in lymph from mice fed a toxic diet than when the diet was fortified with supplemental safflower oil. However, there were no differences between diets in lipid composition of blood plasma. Similarly, except for plasma cholesterol esters, there were no differences in fatty acid composition between mice fed palmitoyl glycerol as the only fat or supplemented with a protective unsaturated fat. In the plasma, cholesteryl palmitate was elevated and cholesteryl oleate and cholesteryl linoleate were depressed when mice were given a toxic diet. Although a monoacylglycerol was toxic when fed, the percentages of monoacylglycerols in lymph or plasma were not materially elevated. The findings indicate that neither the total proportion of saturated fatty acids nor the amount of circulating monoacylglycerols was directly involved in the toxicity of palmitoyl glycerol.
Effect of cupric ions on serum and liver cholesterol metabolism
Authors: Tanaka Mitsuo , Iio Toshihiro , Tabata Toshikazu
Published: November 2006

Cupric ions were administered subcutaneously to male Sprague‐Dawley rats at a single dose of 200 μmol/kg. At 24 hr after administration, a remarkable increase of total and free cholesterol was seen in the rat serum. Also, when lecithin‐cholesterol acyltransferase (LCAT) (E.C. 2.3.1.43) activity was expressed as the percentage of the total serum that free cholesterol esterified, the acyltransferase activity in rats treated with cupric ions showed a slight decrease while the triglyceride content in rat serum and liver decreased by 54% and 61%, respectively. However, the content of hepatic cholesterol in rats treated with cupric ions did not show such a marked change.

On the other hand, acid cholesteryl ester hydrolase activity (Acid CEH) (E.C. 3.1.1.14) in liver lysosomes of rats treated with cupric ions showed a marked decrease with increasing cupric ion concentration both in vivo and in vitro. Furthermore, cupric ions caused a marked release of the lysosomal enzymes cathepsin D and β‐glucuronidase into the cytosolic fraction. The changes in acid cholesteryl ester hydrolase activity induced by cupric ions appear to be a direct effect of cupric ions on the enzyme. These results suggest that excessive cupric ion concentrations could cause various disorders in lipid metabolism.
Effects of ketoconazole on cholesterol synthesis and precursor concentrations in the rat liver
Authors: Strandberg Timo E. , Tilvis Reijo S. , Miettinen Tatu A.
Published: November 2006

Ketoconazole, an antimycotic agent, given to rats for a week as 0.05% food addition had no effect on the hepatic concentrations of free and esterified cholesterol or on the activity of acyl coenzyme A: cholesterol‐acyltransferase (ACAT). However, the levels of free methylated cholesterol precursors, especially lanosterols, less markedly Δ8,24 and Δ8‐dimethyl sterols and monomethyl sterols, were increased after only one day's treatment, while those of esterified methyl sterols were increased inconsistently, and those of free and esterified Δ8‐lathosterol, lathosterol and desmosterol were not affected at all. Cholestyramine treatment had no significant effect on ACAT in spite of a decrease in the hepatic content of esterified cholesterol and caused a marked increase in the free cholesterol precursor levels, especially in those of lathosterols. Cholestyramine given to ketoconazole‐treated rats increased the hepatic levels of Δ8 and Δ7‐lathosterols but not desmosterol or methylated cholesterol precursors. Ketoconazole increased and cholestyramine markedly decreased plantssterols, sitosterol and campesterol in the liver. In serum, the contents of both lanosterols and lathosterol were increased but that of cholesterol tended to be decreased by ketoconazole (−19%). The results indicate that ketoconazole impairs demethylation processes at C‐14 and to some extent at C‐4 in the rat liver, resulting in lowered serum cholesterol level.
Injection of tridocosahexaenoyl‐glycerol emulsion and fatty acid composition of blood cells
Authors: Hamazaki Tomohito , Urakaze Masaharu , Yano Saburo , Soda Yasuji , Miyamoto Akira , Kubota Kohji , Ibuki Fumio
Published: November 2006

An injectable emulsion of docosahexaenoic acid (DHA) was prepared. One hundred ml of the emulsion contained 3 g of 93%‐pure 1,2,3‐tridocosahexaenoylglycerol (DHA‐TG), 1.2 g of 93%‐pure 2‐docosahexaenoyl‐phosphatidylcholine as an emulsifier and 2.5 g of glycerol. Thirty ml of the emulsion of DHA‐TG was injected into three rabbits on days 1 and 4 of the study. Blood was taken on day 0, on day 4 just before the second injection and on day 7. The percentage of DHA in the total phospholipid fraction of platelets increased from 0.46% (day 0) to 1.88% (day 4, p<0.05) and 3.66% (day 7; p<0.02 vs day 0). The percentage of eicosapentaenoic acid (EPA) increased from 0.46% (day 0) to 1.03% (day 4, p<0.02) and 1.63% (day 7: p<0.05 vs day 0). The percentage of arachidonic acid (AA) decreased from 9.45% (day 0) to 4.31% (day 4, p<0.05) and 6.68% (day 7; p<0.02 vs day 0). The percentage of DHA in the total phospholipid fraction of erythrocyte membranes increased from 0.23% (day 0) to 0.91% (day 4, p<0.05) and 1.52% (day 7; p<0.005 vs day 0); that of EPA increased from 0.21% (day 0) to 0.34% (day 4, p<0.005%) and 0.52% (day 7, p<0.01 vs day 0); that of AA was unchanged. Blood lipids were the same before and after the two injections of the emulsion, except that free fatty acids decreased markedly from 0.32 to 0.06 mEq/1 (p<0.02). On day 8, free AA (2 mg/kg) was injected into ear veins of the three treated rabbits and of four control rabbits (not treated with DHA‐TG). All the control rabbits died a few minutes after the AA injection, but none of the DHA‐treated rabbits died after AA injection (p<0.01). An emulsion of DHA‐TG may be useful for patients having immediate risk of thrombosis or for those who need DHA but cannot take it orally.
Effects of dietary primrose oil on mammary tumorigenesis induced by 7,12‐dimethylbenz(a) anthracene
Authors: Abou El‐Ela Soad H. , Prasse Keith W. , Carroll Richard , Bunce Opal R.
Published: November 2006

The mammary tumor‐promoting effect of a high‐fat diet containing 20% evening primrose oil (PO) was compared to that of a 20% corn oil (CO) diet. Mammary tumors were induced in female Sprague‐Dawley rats using 10 mg (Study 1) and 5 mg (Study 2) 7,12‐dimethylbenz(a)anthracene (DMBA). The 10 mg dose of DMBA gave a total mammary tumor incidence of 47% in rats fed the PO diet and 80% for those fed the CO diet. When only adenocarcinomas were counted, the malignant mammary tumor incidences were 41% in rats fed the PO diet and 73% in rats fed the CO diet. In a second study using 5 mg DMBA to induce mammary tumors, total tumor incidences were 50% for PO‐fed rats and 63% for those receiving a CO diet. Again, when only adenocarcinomas were counted, tumor incidences were 27% for PO‐ and 63% for CO‐dieted rats. Analysis of plasma fatty acid profiles indicated that animals fed a 20% PO diet showed significant increases in 18∶3 and 20∶4 fatty acids and significant decreases in 16∶0 and 18∶1 compared to animals fed a 20% CO diet. These results indicate that the mammary tumor promoting effect of a diet containing 20% fat can be diminished by substituting PO for CO. Moreover, the promoting effect on mammary cancer by a high‐fat diet could be depressed by feeding a source of γ‐linolenic acid (GLA).
Short‐Term changes in hepatic HMG‐CoA reductase in rats fed diets containing cholesterol or oat bran
Authors: Kelley Micheal J. , Story Jon A.
Published: November 2006

In vivo regulation of hepatic HMG‐CoA reductase (HMGR) (mevalonate: NADP+ oxidoreductase [acylating CoA]; EC 1.1.1.34] by phosphorylation/dephosphorylation has not been demonstrated. Rats were meal‐fed semipurified diets; effects of inclusion of cholesterol (2%) or oat bran (15%) in a single meal on expressed (phosphorylated) and total (dephosphorylated) activities of HMGR were measured from 15 min to 4 hr after presentation of the meal. Expressed activity was not significantly altered in response to the control diet during the time periods examined, while total HMGR activity declined by 15 min and increased through 4 hr to an activity about 1.5 times control levels. Addition of cholesterol resulted in little change in expressed activity but a greater and more sustained reduction in total activity. Oat bran caused reductions in both total and expressed activities, which were maintained through 4 hr. Total HMGR activity was best correlated with apparent demand for cholesterol synthesis.
Relationship between translocation of long‐chain acyl‐CoA hydrolase, phosphatidate phosphohydrolase and CTP:Phosphocholine cytidylyltransferase and the synthesis of triglycerides and phosphatidylcholine in rat liver
Authors: Asiedu Daniel , Skorve Jon , Demoz Abraham , Willumsen Nina , Berge Rolf K.
Published: November 2006

Translocation of long‐chain acyl‐coenzyme A hydrolase from the microsomal fraction to the cytosolic fraction was promoted in cell‐free extracts of rat liver by palmitic acid, oleic acid, tetradecylthioacetic acid, and tetradecylthiopropionic acid, and by their CoA esters. The CoA esters were more effective than the non‐esterified acids in the translocation of the enzyme. Treatment of normolipidemic rats with sulfur‐substituted non‐β‐oxidizable fatty acid analogues resulted in a transitory increase in hepatic concentration of long‐chain acyl‐CoA. Longer feeding times almost normalized the hepatic long‐chain acyl‐CoA content. Microsomal long‐chain acyl‐CoA hydrolase activity was inhibited, whereas the activity of the cytosolic form was stimulated. The rise in enzyme activity coincided with a reduction in liver content of triglyceride and an increase in hepatic phospholipid content. The results suggest that the activity of long‐chain acyl‐CoA hydrolase in the cytosol may control the amount of acyl‐CoA thioesters in the liver. Esterified and non‐esterified fatty acids causedin vitro translocation of phosphatidate phosphohydrolase and cytidine 5′‐triphosphate (CTP):phosphocholine cytidylyltransferase from the cytosolic fraction to the microsomal fraction. However, the translocation of these two enzyme systems was not obtainedin vivo. The activity of phosphatidate phosphohydrolase decreased in microsomal and cytosolic fractions while the activity of cytidylyltransferase in these fractions increased. The activities of soluble phosphatidate phosphohydrolase and long‐chain acyl‐CoA hydrolase appeared to be inversely correlated. The results imply that in cytoplasm, long‐chain acyl‐CoA hydrolase may compete with the biosynthetic enzymes for the acyl‐CoA substrate, thus influencing the rate of lipid synthesis. The reduced hepatic triglyceride content observed in tetradecylthioacetic acid‐treated rats is probably due to reduced triglyceride synthesis, which is mediated by an inhibition of phosphatidate phosphohydrolase accompanied with translocation and stimulation of long‐chain acyl‐CoA hydrolase. Development of fatty liver as an effect of tetradecylthiopropionic acid is probably due to accelerated triglyceride biosynthesis, which is mediated by a stimulation of phosphatidate phosphohydrolase and a decrease in cytosolic palmitoyl‐CoA hydrolase activity.
Transfer of liposomes containing dolichol into isolated hepatocytes
Authors: Tollbom Örjan , Chojnacki Tadeusz , Dallner Gustav
Published: November 2006

Isolated rat hepatocytes were preincubated with egg lecithin liposomes containing [3H]dolichol and [3H]‐dolichyl ester, and the intracellular levels and distributions of these lipids were subsequently determined after incubation in a liposome‐free medium. [3H]Dolichol was recovered initially mainly in microsomes, and no increase with time in the low level of this compound in the mitochondrial/lysosomal fraction could be observed. A small portion of the labeled dolichol was esterified in the endoplasmic reticulum and transferred to the lysosomecontaining fraction. [3H]Dolichyl linoleate was initially localized in microsomes and supernatant, but later accumulated in the mitochondria/lysosomes. Dolichyl linoleate was found in the membrane of microsomes, in the membrane and lumen of lysosomes, and in the soluble cytoplasm. Exogenous dolichol recovered in microsomes was not phosphorylated to any significant extent. Liposomal phosphatidylcholine also showed preferential accumulation in microsomes after incubation with hepatocytes. These results indicate that exogenous or endogenously formed dolichyl esters are transferred from the endoplasmic reticulum to lysosomes, probably through the cytoplasm. It appears that fatty acids play a role in targeting these lipids to their intracellular locations.
The dual effect of oxidation on lipid bilayer structure
Authors: Borchman Douglas , Lamba Om P , Salmassi Samira , Lou Marjorie , Cecilia Yappert M.
Published: November 2006

Sphingomyelin membranes were prepared with different levels of oxidative damage caused bytert‐butyl hydroperoxide (TBH). Temperature‐induced changes in membrane hydrocarbon chain packing (phase transitions) were monitored using infrared spectroscopy. Lipid phase transition characteristics were evaluated from thermodynamic parameters fitted to the experimental transition curve data. At temperatures below the lipid phase transition Tc, hydrocarbon chains pack in an ordered state whereas above the Tcthe hydrocarbonchains pack in a disordered state. Compared to the non‐oxidized control, the packing of the hydrocarbon chains of mildly oxidized sphingomyelin (<10 nmol TBH/mg lipid) was no different at all temperatures below the Tc, and was more ordered above the Tc. The hydrocarbon chains of strongly oxidized sphingomyelin (>10nmol TBH/mg lipid) were more disordered at temperatures above and below the Tccompared to the control samples. These results suggest that lipid oxidation has a dual effect on lipid order. A more ordered or disordered state may result depending on the degree of oxidation and the state of lipid order prior to oxidation. These results could be important for explaining the structural changes in oxidized membranes high in sphingomyelin such as those found in the ocular lens and liver plasma membranes.
Decreased lecithin: Cholesterol acyltransferase activity in the plasma of hypercholesterolemic pigs
Authors: Lacko Andras G. , Lee S‐M. , Mirshahi Iraj , Hasler‐Rapacz Judith , Kudchodkar Bhalchandra J.
Published: November 2006

Lecithincholesterol acyltransferase (LCAT) activity levels were determined, as function of plasma total cholesterol (TC) in 13 normocholesterolemic (TC<85 mg/dL) and in 28 hypercholesterolemic (TC>98 mg/dL) pigs. The normocholesterolemic group consisted of pigs that carried apo‐B allelic genes other thanLpb5 and orLpb8. The hypercholesterolemic group consisted ofLpb5/x andLpb5/8 heterozygous andLpb5/5 homozygous animals. The data reported in this study show that the LCAT activity in the plasma of hypercholesterolemic (HC) pigs (79±43 units) was significantly lower (p<0.0005) compared to the normocholesterolemic controls (175±45 units). Furthermore, LCAT activity was positively correlated with TC in the normocholesterolemic group (r=+0.54; p<0.05), whereas it was negatively correlated with TC in the hypercholesterolemic group (r=−0.73; p<0.001). Additional data obtained from incubation experiments suggest that the lower LCAT activity in hypercholesterolemic pigs may be due, at least in part, to inhibition of LCAT activity by components found in the lipoprotein‐deficient fractions of the plasma of hypercholesterolemic pigs.
Oxysterols in cultured bovine aortic smoth muscle cells and in the monocyte‐like cell line U937
Authors: Pie Jae Eun , Seillan Christine
Published: November 2006

Oxysterols in cultured bovine aortic smooth muscle cells and in the monocyte‐like cell line U937 were analyzed by gas‐liquid chromatography. The following products were detected: 7α‐ and 7β‐hydroxycholesterol, 7‐ketocholesterol, cholesterol‐α‐epoxide, cholesterol‐β‐epoxide, cholestanetriol, and 20‐ and 25‐hydroxycholesterol. The proportions of oxidized cholesterol varied from 1 to 3% of total cholesterol. The concentrations of oxysterols were lower when the smooth muscle cells were maintained in a vitamin E‐enriched medium, and were higher in U937 cells when the cells were activated with phorbol myristate acetate. The cell oxysterol content appears to be regulated by factors that inhibit or enhanced free radical generation. The concentrations of oxysterols found in cells may serve as an indicator of the extent of lipid peroxidation.
Phospholipids inTrypanosoma cruzi: Phosphoinositide composition and turnover
Authors: Racagni G. , Garcia de Lema M. , Domenech C. E. , Machado de Domenech E. E.
Published: November 2006

The polyphosphoinositides fromTrypanosoma cruzi were isolated by preparative thin‐layer chromatography (TLC) and identified. When myo‐[3H]inositol was present in the culture medium for five days, analyses showed the presence of phosphatidylinositol (PI), lysophosphatidylinositol (lysoPI), phosphatidylinositol 4‐phosphate (PIP) and phosphatidylinositol 4,5‐bisphosphate (PIP2). Short‐term incubation with32Pi led to higher percentages of incorporation into phosphatidylethanolamine (PE), lysophosphatidylethanolamine (lysoPE) and PI compared to the other glycerophospholipids. The phosphoinositides (PI, PIP and PIP2) contained a larger proportion of unsaturated than saturated fatty acids. High proportions of 18∶2 were found in the three phosphoinositides analyzed, whereas the major saturated fatty acid was 18∶0. Watersoluble inositol phosphates (IP, IP2 and IP3) were also identified.
Chemiluminescence detection of 8a‐hydroperoxy‐tocopherone in photooxidized α‐tocopherol
Authors: Miyazawa Teruo , Yamashita Toyonobu , Fujimoto Kenshiro
Published: November 2006

Photosensitized oxidation products of α‐tocopherol were analyzed by reversed phase high‐performance liquid chromatography (CL‐HPLC) in combination with chemiluminescence detection, to measure hydroperoxide derivatives. An intense peak was observed on the chemiluminescence chromatogram, corresponding to 8a‐hydroperoxy‐α‐tocopherone (α‐tocopherol hydroperoxide), the identity of which was confirmed by1H nuclear magnetic resonance and mass spectrometry. Observed peak intensities correlated with the peroxide value of oxidized α‐tocopherol. As little as 50 pmol of 8a‐hydroperoxy‐α‐tocopherone was detectable. The CL‐HPLC method, which is specific for the detection of hydroperoxides, should prove useful in studies on tocopherol oxidation in foods and biological systems.
α‐Helical requirements for free apolipoproteins to generate HDL and to induce cellular lipid efflux
Authors: Hara Hitoshi , Hara Hiroko , Komaba Akira , Yokoyama Shinji
Published: November 2006

The structural requirement has been studied for apolipoproteins in their free form to interact with cells, to generate high density lipoprotein (HDL), and to cause cellular lipid efflux (J. Biol. Chem. 266, 3080–3086, 1991). It is shown that human apolipoprotein (apo) A‐IV and apolipophorin III ofManduca sexta cause cholesterol efflux from cholesterol‐loaded mouse peritoneal macrophages and reduce intracellularly accumulated cholesteryl ester as a results of forming HDL‐like particles with cellular lipids, as do apoA‐I, A‐II and E. On the other hand, similar to apoC‐III, reduced‐and‐carboxymethylated human apoA‐II had no such effect. Thus, apolipoproteins seem to require at least four amphiphilic helical segments per molecule to express this function.