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PubMed1998

Digestion and absorption rates of [3H]-oleic acid and [14C]-triolein do not differ in rats fed heated (-) and (+) gossypol cottonseed and soybean flours.

This study was conducted to compare in vivo the acute effects of heated (+) and (-) gossypol cottonseed flours with those of soybean flour on lipid digestion and absorption in growing rats. Rats were fed by gastric intubation mixed [3H]-oleic acid and [14C]-triolein with heated flours or without flour (control). Lipid digestion and absorption were determined for 6 h after meal intubation. Both radioactivities recovered in gastrointestinal tract were significantly higher in rats fed (+) gossypol cottonseed flour than in all other groups. The majority of both recovered radioactivities was found in stomach contents, then in stomach wall and finally in intestinal wall. The distribution of both radioactivities at different gastrointestinal levels was similar. In stomach contents and wall, [14C]-radioactivity was primarily in triacylglycerols, but was also recovered in free fatty acids and diacylglycerols. In intestinal wall (mucosa + tunica) [3H]-radioactivity was found at greatest levels in free fatty acids, then in triacylglycerols and diacylglycerols. Greatest [14C]-radioactivity was found in triacylglycerols, then in free fatty acids, in diacylglycerols and last in phospholipids in rats fed the three flours. Therefore no quantitative differences in lipid digestion and absorption were observed among the rats fed the three flours. In conclusion, both cottonseed flours slowed lipid digestion and absorption compared with soybean flour and this delay was greater in the rats fed (+) gossypol cottonseed flour than in those fed (-) gossypol cottonseed flour. However, this inhibiting effect was probably too low to induce physiologically important effects on lipid digestion or absorption.

Animals↗
PubMed2003

Differential incorporation of docosahexaenoic and arachidonic acids by the yolk sac membrane of the avian embryo.

During avian development, lipoproteins derived from yolk lipid are assembled in the yolk sac membrane (YSM) for secretion into the embryonic circulation. To investigate how yolk polyunsaturated fatty acids, essential for the development of certain tissues, are distributed among the lipid classes of the lipoproteins, pieces of YSM were incubated in vitro with [14C]arachidonic and [14C]docosahexaenoic acids (DHA). There was a marked difference in the partitioning of these two precursors among the lipid classes of the tissue. Of the radioactivity incorporated into total lipid from [14C]-arachidonic acid during 1 h of incubation, 67.3% was esterified as phospholipid and 29.5% as triacylglycerol. In contrast, only 14.6% of the label incorporated from [14C]-DHA was esterified as phospholipid, whereas 73.2% was recovered in triacylglycerol. This pattern of differential partitioning was observed at all time points and across a 20-fold range of fatty acid concentrations. There was no evidence for conversion of the radioactive arachidonic and DHAs to other fatty acids prior to incorporation into tissue lipids. It is suggested that the selective incorporation of yolk-derived DHA into the triacylglycerol of secreted lipoproteins represents part of a mechanism for directing this polyunsaturate to particular embryonic tissues.

Animals↗
PubMed1976

Utilisation of exogenous fatty acids by rat epididymal fat cells in vitro.

When isolated rat fat cells are incubated with radioactively labelled palmitate the extracellular fatty acid pool is diluted by fatty acids of endogenous origin. Using fat cell concentration as an experimental variable, a simple graphical procedure has been devised to correct for the difficulty such precursor dilution causes in measurement of oxidation and esterification of exogenous fatty acids. After application of such corrections it was calculated that extracellular palmitate was utilised for oxidation with a composite Km of approx. 7 mu M. The composite Km for palmitate esterification was 29 mu M (no other substrates present, albumin included 1t 10 mg/ml).

Adipose Tissue↗
PubMed1983

Separation and quantitation of free fatty acids and fatty acid methyl esters by reverse phase high pressure liquid chromatography.

Reverse phase high pressure liquid chromatography (HPLC) on octadecylsilyl columns separates mixtures of either free fatty acids or fatty acid methyl esters prepared from mammalian tissue phospholipids. Acetonitrile-water mixtures are used for the elution of esters. Aqueous phosphoric acid is substituted for water for the separation of the free acids. Unsaturated compounds are detected and quantitated by their absorption at 192 nm. Saturates are detected better at 205 nm. The order of elution of fatty acids in complex mixtures varies as a function of acetonitrile concentration. At any given concentration, some compounds overlap. However, by varying the solvent strength, any fatty acid of interest can be resolved including many geometrical and positional isomers. Methyl esters prefractionated according to unsaturation by argentation thin-layer chromatography (TLC) are rapidly and completely separated by elution with CH3CN alone. Argentation TLC-reverse phase HPLC can be used as an analytical as well as a preparative procedure. Octylsilyl columns are used for rapid resolution and improved detection of minor or low ultraviolet-absorbing components in the fractions. For example, monoenoic fatty acids with up to 32 carbons have been detected in bovine brain glycerophospholipids. Specific radioactivities of 3H- and 14C-labeled fatty acids and the distribution of radioactivity among acyl groups from complex lipids are measured. The method is not recommended for complete compositional analysis, but is useful for determinations of specific radioactivities during studies on turnover and metabolic conversions of labeled fatty acids.

Animals↗
PubMed2003

Synthesis of lipids in isolated nuclei from rat thymus and liver cells.

Synthesis of lipids was studied in isolated nuclei from rat thymus and liver cells. On incubation of the isolated nuclei with [2-14C]acetate and [1-14C]glycerol, the label was intensively incorporated into phospholipids and with a significantly lower intensity into fatty acids and cholesterol. Only trace amounts of radioactivity were detected in the lipids of chromatin prepared from isolated thymus nuclei after their incubation, and this suggested that lipids were mainly synthesized on the nuclear membrane. On the preincubation of thymus tissue homogenate with [2-14C]acetate and the subsequent isolation of the nuclei and chromatin, the radioactivity of chromatin lipids was comparable to the radioactivity of nuclear lipids. The findings suggested that in the isolated nuclei the newly synthesized lipids were not transported into chromatin from the nuclear membrane. The specific radioactivities of individual phospholipids and fatty acids were different in the isolated nuclei and in nuclei obtained from preincubated homogenate. Mechanisms of lipid synthesis in isolated nuclei and causes of the different radioactivities of lipids in the isolated nuclei and in the nuclei obtained from the preincubated homogenate are discussed.

Animals↗
PubMed1980

Microbial metabolism of amino alcohols. Biosynthetic utilization of ethanolamine for lipid synthesis by bacteria.

1. Ten bacteria utilizing [2-14C]ethanol-2-amine as the sole or major source of nitrogen for growth on glycerol + salts medium incorporated radioactivity into a variety of bacterial substances. A high proportion was commonly found in lipid fractions, particularly in the case of Erwinia carotovora. 2. Detailed studies of [14C]ethanolamine incorporation into lipids by five bacteria, including E. carotovora, showed that all detectable lipids were labelled. Even where phosphatidylethanolamine was the major lipid labelled, radioactivity was predominantly in the fatty acid rather than the base moiety. The labelled fatty acids were identified in each case. 3. The addition of acetate to growth media decreased the incorporation of radioactivity from ethanolamine into both fatty acid and phosphatidyl-base fragments of lipids from all the bacteria except Mycobacterium smegmatis. Experiments with [3H]ethanolamine and [14C]acetate confirmed that unlabelled acetate decreased the incorporation of both radioactive isotopes into lipids, except in the case of M. smegmatis. 4. Enzyme studies suggested one of two metabolic routes between ethanolamine and acetyl-CoA for each of four bacteria. A role for ethanolamine O-phosphate was not obligatory for the incorporation of [14C]ethanolamine into phospholipids, but correlated with CoA-independent aldehyde dehydrogenase activity.

Acetates↗
PubMed1977

Methods for the rapid separation and estimation of the major lipids of arteries and other tissues by thin-layer chromatography on small plates followed by microchemical assays.

Methods are described for the rapid separation of the major individual phospholipids and neutral lipids of tissues by thin-layer chromatography on small glass plates (75 X 75 mm), and for the specific microchemical estimation of separated lipids and for determination of fatty acid composition and radioactivity. The overall method, involving tissues extraction, thin-layer chromatographic separation and assay has been evaluated using pure standards and biological samples and gives good reproducibility and almost complete recovery of lipids.

Arteries↗
PubMed1993

Copper deficiency increases in vivo hepatic synthesis of fatty acids, triacylglycerols, and phospholipids in rats.

This study examined the influence of dietary copper status on the in vivo hepatic fatty acid synthesis and the incorporation of nascent fatty acids into various hepatic lipid classes. Fifty weanling male Sprague-Dawley rats were assigned to two dietary treatments, copper deficient (5.4 nmol/g of diet) and copper adequate (102 nmol/g of diet). After 7 weeks of treatment, rats were injected with 0.111 MBq of [1-14C] acetate (1.85 GBq/mM)/100 g body wt through the femoral vein. Five rats from each treatment were sacrificed at 3, 6, 9, 12, and 24 min after injection. Radioactivities of nascent fatty acid samples were used to determine relative rates of fatty acid synthesis and their assembly into triacylglycerols and phospholipids. Linear increases were observed up to 12 min after injection for total hepatic fatty acid synthesis and their assembly into triacylglycerols and phospholipids for both treatments. In addition, 46% and 30% of total fatty acid synthesized were assembled into triacylglycerols and phospholipids, respectively, for both groups. Furthermore, hepatic fatty acid synthesis and assembly into triacylglycerols and phospholipids were enhanced more than 2-fold by copper deficiency when the data were expressed as per liver per 100 g body weight.

Animals↗
PubMed1990

Erucic acid metabolism in rat heart. A combined biochemical and radioautographical study.

Metabolism of Erucic Acid was studied in rat heart in comparison with that of oleic acid, particularly in relation with diet lipids. Rats were fed for 3 or 60 days a diet containing 30% of the calories of either Rapessed Oil, rich in erucic acid or sunflower seed oil rich in linoleic acid. They were I.V. injected with tritiated erucic or oleic acid. After 1 or 15 min the radioactivity recovered in heart lipids was very low whatever the diet (1 to 2%). One minute after injection of erucic acid the radioactivity was mainly recovered in the free fatty acid fraction and as untransformed erucic acid. After 15 min the major part of radioactivity was recovered in the triacylglycerol fraction which contained a high proportion of labelled oleic acid formed by shortening of erucic acid. When oleic was injected, the radioactivity was principally recovered in triacylglycerols as untransformed oleic acid whatever the experimental conditions. Electron microscopy showed that a much higher proportion of peroxisomes, was present in heart cells, following sunflower seed oil diet as compared to rapeseed oil diet. In all cases mitochondria supported the greater part of radioactivity, especially when erucic acid was injected in rats fed rapeseed oil. After sunflower seed oil, a noticeable radioactivity was observed in peroxisomes, most of them containing silver grains, especially when oleic acid was injected. According to the data reported, peroxisomes do not seem more implicated than mitochondria in the metabolism of erucic acid in myocardium.

Animals↗
PubMed1985

Chain-shortening of erucic acid and microperoxisomal beta-oxidation in rat small intestine.

The ability of rat small intestine to chain-shorten C22:1 fatty acids was investigated. Radioactive chain-shortened products, mainly C18:1, were demonstrated in intestinal-lymph lipids after intraluminal injection of [14-14C]erucic acid. Chain-elongation to C24:1 was also observed. Adaptation to a diet containing C22:1 fatty acids (partially hydrogenated-marine-oil diet) slightly increased the percentage of chain-shortened products. Microperoxisomal beta-oxidation activity, measured as CN(-)-insensitive palmitoyl-CoA-dependent NAD+ reduction, was detected in a microperoxisome-enriched fraction from mucosal scrapings. This activity was increased 1.9-fold by a soya-bean-oil diet, and 2.7-fold by a diet containing partially hydrogenated marine oil.

Animals↗
PubMed1975

The incorporation of orally fed radioactive gamma-linolenic acid and linoleic acid into the liver and brain lipids of suckling rats.

The incorporation of radioactivity from orally administered gamma-linolenic acid-1--14C and linoleic acid--3H into the liver, plasma, and brain lipids of suckling rats was studied. Significantly more radioactivity from the former compound was incorporated into the liver and brain lipids 22 hr after dosing. The distribution of the radioactivity in the fatty acids of the liver and brain lipids was different for each isotope. Most of the -3H was still associated with linoeic acid, whereas most of the -14C was in the 20:3 and 20:4omega6 fractions. These results suggest that the desaturation of linoleic to gamma-linolenic acid in vivo is a rate-limiting step in the conversion of linoleic to arachidonic acid.

Administration, Oral↗
PubMedDate not supplied

Incorporation of 14C stearic acid in lens organ culture.

14C stearic acid was incubated in organ culture of rabbit lens for 6, 12 and 20 hours. Stearic acid did penetrate the lens and was incorporated into triacyl-glycerides and phospholipids. It also served as a carbon source for cholesterol synthesis. Individual fatty acids were isolated and radioactivity measurements were made. Incorporation of label was found in C12:0, C14:0 and C16:0 fatty acids, indicating de novo synthesis in the lens; incorporation was found in C16:1 and C18:1, indicating delta 9 desaturase activity, and incorporation was found in the fatty acids of 20 carbons or longer, indicating the presence of a fatty acid elongation system in the lens.

Animals↗
PubMed1998

Role of acetyl-L-carnitine in rat brain lipogenesis: implications for polyunsaturated fatty acid biosynthesis.

This study was undertaken to explore the metabolic fate of acetyl-L-carnitine in rat brain. To measure the flux of carbon atoms into anabolic processes occurring at regional levels, we have injected [1-(14)C]acetyl-L-carnitine into the lateral brain ventricle of conscious rats. After injection of [1-(14)C]acetyl-L-carnitine, the majority of radioactivity was recovered as 14CO2 expired (60% of that injected). The percentage of radioactivity recovered in brain was 1.95, 1.60, 1.30, and 0.93% at 1, 3, 6, and 22 h, respectively. Radioactivity distribution in various lipid components indicated that the fatty acid moiety of phospholipid contained the majority of radioactivity. The radioactive profile of these fatty acids showed that the acetyl moiety of acetyl-L-carnitine was incorporated into saturated (60%), monounsaturated (15%), and polyunsaturated (25%) fatty acids [mainly present in 20:4 (5.2%) and 22:6 (7.8%)]. Injection in the brain ventricle of radioactive glucose, the major source of acetyl-CoA in the CNS, revealed that glucose was a precursor of saturated (85%) and monounsaturated (15%) but not of polyunsaturated fatty acids. Thus, this study demonstrated distinct fates of glucose and acetyl-L-carnitine following intracerebroventricular injection. In summary, these data implicate acetyl-L-carnitine as an important member of a complex acetate trafficking system in brain lipid metabolism.

Acetylcarnitine↗
PubMed1978

Oxidation of [U-14C] palmitic acid by cock spermatozoa.

When washed cock spermatozoa were incubated with [U-14 C] palmitic acid at 37 C for 2 hr under aerobic conditions, radioactivity was recovered as carbon dioxide indicating the fatty acid was oxidized. Little if any radioactivity was recovered as carbon dioxide when spermatozoa were killed by boiling, indicating the necessity for viable spermatozoa for the successful incorporation and oxidation of palmitic acid. To determine whether the oxidation of palmitic acid could serve as a source of energy for cock spermatozoa, ATP concentrations of spermatozoa were compared immediately following ejaculation and after 1,2, and 3 hr of incubation (37 C) with and without the addition of palmitic acid. At 1,2, and 3 hr of incubation, spermatozoa with palmitic acid as a substrate produced significantly (P less than or equal to .03) more ATP (2.62, 2.24, and 1.26 micrograms ATP/10(9) cells, respectively) than did spermatozoa without palmitic acid (1.62, 1.11, and .79 micrograms ATP/10(9) cells, respectively). These findings indicate that palmitic acid, one of the most abundant long-chain saturated fatty acids in cock spermatozoa, can be utilized as a source of energy.

Adenosine Triphosphate↗
PubMed1996

Nonoxidative metabolism of 2-butoxyethanol via fatty acid conjugation in Fischer 344 rats.

Nonoxidative metabolism of ethylene glycol monobutyl ether (2-butoxyethanol or BE) via fatty acid conjugation was studied in the liver of Fischer 344 male rats following a single oral administration of 500 mg/kg body weight [ethyl-1,2-14C]BE (70 microCi/kg). Animals were killed 2 h after the treatment, hepatic lipids extracted, and the neutral lipids were separated using solid-phase extraction. The neutral lipid fraction was subjected to preparative thin-layer chromatography, and the esters corresponding to the relative flow of authentic fatty acid 2-butoxyethyl esters were recovered and analyzed by reversed-phase high-performance liquid chromatography (HPLC) using methanol-water (37:3, v/v) as solvent. Approximately 85% of the 14C label present in the ester fraction was coeluted at retention times corresponding to the different fatty acid 2-butoxyethyl ester standards. The radioactive fractions were analyzed by electron impact mass spectrometry. Molecular ion peaks and fragmentation patterns similar to that of 16:0, 18:0, 18:1, 18:2, and 20:4 fatty acid 2-butoxyethyl ester standards were detected in the corresponding radioactive HPLC fractions. Fatty acid ethyl ester synthase (FAEES), purified from the rat liver microsomal fraction, was also found to catalyze the formation of 18:1 fatty acid 2-butoxyethyl ester. These studies demonstrate that BE is metabolized nonoxidatively via conjugation with long-chain fatty acids, and the formation of these esters appears to be catalyzed by the enzyme(s) involved in fatty acid conjugation of xenobiotic alcohols. However, the biological significance of BE conjugation with fatty acids remains to be investigated.

Acyltransferases↗
PubMed1978

Cholesterol arachidonate as a prostaglandin precursor in adrenocortical cells.

Rat adrenocortical cells were incubated with labeled arachidonate, and the radioactivity in unesterified fatty acids was reduced by washing with 2% albumin solutions. These cells were then incubated for two hours in the absence and presence of 7.1 x 10(-10)M ACTH. During subsequent incubation of prelabeled cells with ACTH, both the mass and radioactivity of arachidonate in adrenocortical cholesteryl esters was depleted to the same extent (30--32%). The released arachidonate was in part incorporated into phospholipids, and there was also a significant increase in unesterified arachidonic acid. During this period, there was also increased incorporation of arachidonate into labeled prostaglandins. Of this increase, 92% by isotope analysis, and 88% by radioimmunoassay techniques was attributable to prostaglandins of the E pathway. These data demonstrate that prostaglandin E synthesis is specifically increased during ACTH stimulation of rat adrenocortical cells and suggest that a major source of the arachidonate substrate for this synthesis is derived from hormone-dependent hydrolysis of cortical cholesteryl esters.

Adrenal Cortex↗
PubMed1998

Characterization of lipolytic activity associated with a Vibrio species of bacterium isolated from fish intestines.

Cultures of a species of Vibrio isolated from fish intestines and known to synthesize the polyunsaturated fatty acid eicosapentaenoic acid (20:5n-3) were incubated with di-[1-14C]palmitoyl phosphatidylcholine to determine if lipolytic enzymes are produced by the bacteria. After two days of culture, most radioactivity was recovered in the phospholipids of the bacterial cells. When supernatants from cultures of the Vibrio were incubated with either di-[1-14C]palmitoyl phosphatidylcholine, 1-palmitoyl-2-[1-14C]linoleoyl phosphatidylcholine or 1-[1-14C]palmitoyl lysophosphatidylcholine almost all radioactivity was recovered in the free fatty acid fraction after 24 h. Only very small levels of radioactivity from di-[1-14C]palmitoyl phosphatidylcholine were recovered in diacylglycerols and phosphatidic acid. Over the same incubation period 61% and 5% of the radioactivity originally present in glycerol tri-[1-14C]oleate and cholesteroyl [1-14C]oleate, respectively, was released to free fatty acids. Soybean phosphatidylcholine and cod roe phosphatidylcholine, which differed in polyunsaturated fatty acid profile, were both hydrolyzed by culture supernatant. The results suggest that the Vibrio species examined produces a phospholipase B capable of hydrolyzing both intact phospholipids and lysophospholipids.

Journal Article↗