Labeled oxidation products from [1-14C], [U-14C] and [16-14C]-palmitate in hepatocytes and mitochondria

When [1-¹⁴C], [U-¹⁴C], and [16-¹⁴C]palmitate were oxidized by isolated rat hepatocytes, there was a differential distribution of label as a percent of total oxidized products, such that¹⁴CO₂ from [1-¹⁴C]>[U-¹⁴C]>[16-¹⁴C]-palmitate and acid-soluble radioactivity from [16-¹⁴C]>[U-¹⁴C]>[1-¹⁴C]palmitate. The oxidation of [2,3-¹⁴C]succinate to¹⁴CO₂ by isolated hepatocytes was only 9.1% of that from [1,4-¹⁴C]succinate, demonstrating that the differences in distribution of labeled products are in part due to less¹⁴CO₂ production from label in the even carbon positions entering the citric acid cycle. Apparent total ketone body production from [16-¹⁴C]palmitate was markedly higher than [1-¹⁴C], and [U-¹⁴C]palmitate. In addition, the¹⁴C-acetone:¹⁴CO₂ ratio derived from decarboxylation of labeled acetoacetate from [1-¹⁴C]palmitate was less than 1 and positively correlated to the rate of fatty acid oxidation in hepatocytes. These findings indicate that the known preferential incorporation of the omega-C₂ unit of fatty acids into¹⁴C-ketone bodies also contributed to the differential distribution of labeled products and that this contribution was greatest at the lower rates of fatty acid oxidation. In isolated mitochondria, the distribution of label to¹⁴CO₂ and acid-soluble radioactivity from [1-¹⁴C], [U-¹⁴C] and [16-¹⁴C]palmitate was qualitatively similar to that seen with hepatocytes. The distribution of label from [1-¹⁴C]acetylcarnitine to¹⁴CO₂ and¹⁴C-ketone bodies by mitochondria was identical to that observed from [1-¹⁴C]palmitate, indicating that the higher rates of¹⁴CO₂ production from [1-¹⁴C]palmitate cannot be explained by a preferential oxidation in the citric acid cycle of either extramitochondrial acetyl-CoA (generated in peroxisomes) or the carboxyl terminal of the fatty acid. As shown by others in cell-free systems, we observed that the total oxidation of [16-¹⁴C]palmitate by hepatocytes and mitochondria was significantly less than [1-¹⁴C] and [U-¹⁴C]palmitate, suggesting either incomplete mitochondrial β-oxidation or incomplete degradation of peroxisomal oxidation products. The data indicate that this incomplete oxidation does not, however, contribute to the differential distribution of label to oxidized products.     

Regulation of the metabolism of linoleic acid to arachidonic acid in rat hepatocytes

When 5×10⁶ hepatocytes were incubated for 40 min with from 0.15 to 0.60 mM [1-¹⁴C]linoleic acid, [1-¹⁴C]6,9,12-octadecatrienoic acid, or [1-¹⁴C]8,11,14-eicosatrienoic acid, there was a concentration-dependent acylation of radioactive metabolites into both triglycerides and phospholipids. When the concentration of either [1-¹⁴C]linoleic acid or [1-¹⁴C]8,11,14-eicosatrienoic acid exceeded 0.3 mM, there was no further increase in the metabolism of either fatty acid to other (n−6) metabolites. When the concentration of [1-¹⁴C]6,9,12-octadecatrienoic acid exceeded 0.15 mM, there was an apparent substrate-induced inhibition in its metabolism to 8,11,14-eicosatrienoic acid. With all three substrates (0.3 mM), there was time-dependent metabolism to other (n−6) acids. Cells then were incubated simultaneously with 0.3 mM [1-¹⁴C]linoleic acid along with 0.15 to 0.45 mM 6,9,12-octadecatrienoic acid or 8,11,14-eicosatrienoic acid. These exogenous nonradioactive (n−6) acids suppressed but did not abolish the conversion of [1-¹⁴C]linoleate to radioactive arachidonate. These findings suggest that some linoleate is converted to arachidonate without intracellular mixing of 6,8,12-octadecatrienoic or 8,11,14-eicosatrienoic acids. This hypothesis is supported by the finding that exogenous linoleate did not markedly affect the metabolism of [1-¹⁴C]6,9,12-octadecatrienoic or [1-¹⁴C]8,11,14-eicosatrienoic acid by microsomal chain elongating or desaturating enzymes.     

Modulation of fatty acid incorporation and desaturation by trifluoperazine in fungi

The effects of trifluoperazine (TFP) on [1-¹⁴C]fatty acid incorporation into the lipids ofMortierella ramanniana var.angulispora were studied. TFP decreased [1-¹⁴C]-fatty acid incorporation into phosphatidylcholine, phosphatidylethanolamine and triacylglycerol, but greatly increased¹⁴C-labeling in phosphatidic acid. These changes in [1-¹⁴C]fatty acid incorporation induced by TFP were accompanied by a decrease in desaturation of some [1-¹⁴C]fatty acids taken up by the fungal cells. When [1-¹⁴C]lioleic acid (LA) was incubated with the fungal cells, total γ-linolenic acid (GLA) formation from incorporated [1-¹⁴C]LA decreased, but the¹⁴C-labeled GLA conent in individual lipid classes was essentially unchanged. This suggests that the site of the TFP effect on GLA formation from [1-¹⁴C]LA taken up from the medium is not the desaturase acting on LA linked to complex lipids. On the other hand, GLA formation from [1-¹⁴C]oleic acid was much less susceptile to TFP, which suggests that in this fungus Δ6 desaturation to GLA has at least two different pathways with different degrees of susceptibility to TFP.     

Lipid products formed during desaturation of [1-14C] stearyl CoA by hen liver microsomes

A number of lipid products are formed during the desaturation of stearyl CoA by hen liver microsomes. This article presents an analysis of the products formed when [1-¹⁴C] stearyl CoA is incubated with hen liver microsomes for various time periods. [1-¹⁴C] Oleyl CoA was the first radioactive unsaturated product formed. Synthesis of phospholipids containing [1-¹⁴C] oleic acid occurs only after the desaturase activity has begun to decline. The specific radioactivity of [1-¹⁴C] oleyl CoA was similar to the specific radioactivity of [1-¹⁴C] stearyl CoA at all time periods tested. The specific radioactivities of [1-¹⁴C] oleic acid and phospholipids containing [1-¹⁴C] oleic acid were much lower than that of the [1-¹⁴C] stearyl CoA.     

The biosynthesis of cyclopropane and cyclopropene fatty acids in higher plants (Malvaceae)

The biosynthesis of cyclopropane and cyclopropene fatty acids has been investigated in immature seeds, leaves and callus tissue cultures of several species of Malvaceae. Chemical characterization of labeled cyclopropane and cyclopropene fatty acids obtained from incubations withl-[¹⁴CH₃] methionine confirmed that the ring methylene group was derived from the methyl group of methionine. The variation with time in the distribution of radioactivity in the products of incubations with [¹⁴CH₃] methionine and [2-¹⁴C] acetate suggested that the pathway involved initial formation of dihydrosterculic acid from oleic acid with subsequent desaturation to sterculic acid and α-oxidation to malvalic and dihydromalvalic acids. Direct evidence in favor of this pathway was provided by the conversion of [1-¹⁴C] oleic acid to dihydrosterculic and sterculic acids and by the desaturation of [1-¹⁴C] dihydrosterculic acid to sterculic acid, the first time that these processes have been demonstrated in higher plants. No conversion of [1-¹⁴C] stearolic acid to sterculic acid could be obtained under the same conditions. The presence of an active fatty acid α-oxidation system was demonstrated in the callus cultures.     

Intravenously injected [1-14C]arachidonic acid targets phospholipids, and [1-14C]palmitic acid targets neutral lipids in hearts of awake rats

The differential uptake and targeting of intravenously infused [1-¹⁴C]palmitic ([1-¹⁴C] 16∶0) and [1-¹⁴C]arachidonic ([1-¹⁴C]20∶4n−6) acids into heart lipid pools were determined in awake adult male rats. The fatty acid tracers were infused (170 μCi/kg) through the femoral vein at a constant rate of 0.4 mL/min over 5 min. At 10 min postinfusion, the rats were killed using pentobarbital. The hearts were rapidly removed, washed free of exogenous blood, and frozen in dry ice. Arterial blood was withdrawn over the course of the experiment to determine plasma radiotracer levels. Lipids were extracted from heart tissue using a two-phase system, and total radioactivity was measured in the nonvolatile aqueous and organic fractions. Both fatty acid tracers had similar plasma curves, but were differentially distributed into heart lipid compartments. The extent of [1-¹⁴C]20∶4n−6 esterification into heart phospholipids, primarily choline glycerophospholipids, was elevated 3.5-fold compared to [1-¹⁴C]16∶0. The unilateral incorporation coefficient, k ^*, which represents tissue radioactivity divided by the integrated plasma radioactivity for heart phospholipid, was sevenfold greater for [1-¹⁴C]20∶4n−6 than for [1-¹⁴C]16∶0. In contrast, [1-¹⁴C]16∶0 was esterified mainly into heart neutral lipids, primarily triacylglycerols (TG), and was also found in the nonvolatile aqueous compartment. Thus, in rat heart, [1-¹⁴C]20∶4n−6 was primarily targeted for esterification into phospholipids, while [1-¹⁴C]16∶0 was targeted for esterification into TG or metabolized into nonvolatile aqueous components.     

Metabolism of long-chain fatty acids,alcohols and alkylglycerols in the fish parasiteParatenuisentis ambiguus (Acanthocephala)

Specific differences between the acyl composition of lipids on the helminthParatenuisentis ambiguus and its host eel, as shown previously, prompted us to study the lipid metabolism in this intestinal fish parasite. Adults and larvae ofP. ambiguus were fed various lipid precursors, e.g., fatty acids, long-chain alcohols and 1-O-alkylglycerols, which may occur as common nutrients of intestinal parasites. Incorporation of [1-¹⁴C]palmitic acid into neutral and polar lipids was found to be similar under aerobic and near-anaerobic conditions. In adult parasites maintained in culture medium supplemented with glucose, [1-¹⁴C]palmitic acid was incorporated mainly into triacylglycerols and phosphatidylcholines, whereas [1-¹⁴C]oleic acid was incorporated preferentially into triacylglycerols. In fasted adults, as well as in larvae, [1-¹⁴C]oleic acid was mainly transferred to phosphatidylcholines. Lipolytic activity was detected in adult parasites that had been incubated with radioactive trioleoylglycerol. [1-¹⁴C]Hexadecan-1-ol was oxidized inP. ambiguus at a high rate to labeled palmitic acid, which was incorporated into various lipid classes ofP. ambiguus. Small but significant proportions of radioactivity from hexadecan-1-ol were incorporated into ether glycerolipids of the parasite. A more direct precursor in ether glycerolipid metabolism, i.e.,rac-1-O-[1′-¹⁴C] hexadecylglycerol, was incorporated into alkyl and 1′-alkenyl moieties of choline and etha-nolamine etherglycerophospholipids ofP. ambiguus in high yield. High proportions of labeled diacylglycerols, triacylglycerols and steryl esters were detected in surface lipids as well as lipid extracts of the culture media after incubation ofP. ambiguus with [1-¹⁴C]palmitic or [1-¹⁴C]oleic acids. The results suggest that palmitic acid and oleic acid are incorporated into neutral and polar lipids ofP. ambiguus maintained in glucose medium quite differently with oleic acid showing a strong preference for triacylglycerols. However, the incorporation of palmitic acid in glucose-fed parasites was similar to that of oleic acid in fasted parasites, as well as in larvae. This may be explained by partial fatty acid depletion in fasted worms and rapid cell division in larvae, respectively.     

Asymmetric distribution of decanoate in triacylglycerol synthesized in vitro by mammary glands of lactating mice

Slices, prepared from the mammary glands of lactating mice, were incubated with either [1-¹⁴C]acetate, [U-¹⁴C]glucose, or [1-¹⁴C]decanoate. From all 3 substrates, radioactivity in the synthesized lipids was found mainly in triacylglycerols (TG). When acetate or glucose served as substrate, decanoate (C₁₀) accounted for 24% of the fatty acids in TG. Hydrolysis of the TG by pancreatic lipase yielded [¹⁴C] fatty acids which had relatively more C₁₀ (38%) than did either of the other hydrolysis products mono- or diacylglycerol (14–17%). However, when TG produced by slices from C₁₀ were hydrolyzed, the acid was found to be esterified equally at the C-1, C-2 and C-3 of glycerol. Thus, when fatty acids are synthesized de novo and are converted to TG by gland slices, C₁₀ is predominantly located in the C-3 position, a finding in accord with the situation in milk TG, although such preferential incorporation does not occur when the free acid is presented to the tissue slices.     

Tissue culture of cocoa beans (Theobroma cacao L.): Incorporation of acetate and laurate into lipids of cultured cells

Suspension cultures of cocoa bean tissue readily incorporated exogenous acetate into lipids. The distribution of radioactivity from acetate in individual lipid classes after 48 hr was 20, 5, 1, 15, 25, and 35% in triglycerides, diglycerides, free fatty acids, sterol esters, sterols and polar lipids, respectively. The labeled acetate was rapidly incorporated into various fatty acids within 2 hr. The [1-¹⁴C] saturated fatty acids declined slightly after 4 hr, whereas [1-¹⁴C] oleate declined significantly after 2 hr. There was a concomitant increase in [1-¹⁴C] linoleate. The radioactivity associated with linolenate was relatively high up to 4 hr, declined by 24 hr, and then increased again. The kinetics of fatty acid labeling suggested that biosynthesis of linolenic acid in cocoa bean suspension culture may occur via the desaturation of linoleic acid and the chain elongation of dodecatrienoic acid. The patterns of fatty acid radiolabeling following incubation of cells with [1-¹⁴C] laurate was consistent with this mechanism.     

Differential biohydrogenation and isomerization of [U-13C]Oleic and [1-13C]Oleic acids by mixed ruminal microbes

The additional mass associated with ¹³C in metabolic tracers may interfere with their metabolism. The comparative isomerization and biohydrogenation of oleic, [1-¹³C]oleic, and [U-¹³C]oleic acids by mixed ruminal microbes was used to evaluate this effect. The percent of stearic, cis-14 and- 15, and trans-9 to-16 18∶1 originating from oleic acid was decreased for [U-¹³C]oleic acid compared with [1-¹³C]oleic acid. Conversely, microbial utilization of [U-¹³C]oleic acid resulted in more of the ¹³C label in cis-9 18∶1 compared with [1-¹³C]oleic acid (53.7 vs. 40.1%). The isomerization and biohydrogenation of oleic acid by ruminal microbes is affected by the mass of the labeled tracer.     

Synthesis and characterization of [1-13C]-and d8-arachidonic acid

Methyld ₈- and [1-¹³C] 5,8,11,14-eicosatetraenoate (arachidonate) were prepared from a common synthetic precursor, 4,7,10,13-nonadecatetrayn-1-ol. The purified products were characterized by gas chromatography-mass spectrometry. Mass spectra oft-butyldimethylsilyl esters ofd ₈-and [1-¹³C]-arachidonic acid showed a most intense [M-57]⁺ peak at high mass. The isotopic purity of methyl [1-¹³C] arachidonate was 99% and that of methyld ₈-arachidonate was 56%. Whend ₈-arachidonic acid was prepared by direct deuteration of 5,8,11,14-eicosatetraynoic acid, the isotopic purity of the sample was 86%.     

Methodology for the In Vivo Measurement of the Δ<Superscript>9</Superscript>-Desaturation of Myristic,Palmitic, and Stearic Acids in Lactating Dairy Cattle

There is limited methodology available to quantitatively assess the activity of the Δ⁹-desaturase enzyme in vivo without chemically inhibiting the enzyme or using radioactively labeled substrates. The objective of these experiments was to develop methodology to determine the incorporation and desaturation of ¹³C-labeled fatty acids into milk lipids. In a preliminary experiment, 3.7 g [1-¹³C]myristic acid ([1-¹³C]14:0), 19.5 g [1-¹³C]palmitic acid ([1-¹³C]16:0), 20.0 g [1-¹³C]stearic acid ([1-¹³C]18:0) were combined and infused into the duodenum of a cow over 24 h. In a following experiment, 5.0 g [1-¹³C]14:0, 40.0 g [1-¹³C]16:0, and 50.0 g [1-¹³C]18:0 were infused into the abomasums of separate cows as a bolus over 20 min or continuously over 24 h. Milk fat was extracted using chloroform:methanol. Fatty acids were methylated, and fatty acid methyl esters (FAME) were converted to dimethyl disulfide derivatives (DMDS). The FAME and DMDS were analyzed by gas chromatography mass spectrometry. In the preliminary experiment, ¹³C enrichment in 14:0 but not 16:0 or 18:0 was observed. When dosage amounts were increased in the following experiment, peak enrichments from the bolus infusion were observed at 8 h. Enrichments for continuous infusion peaked at 16 h for 14:0 and 18:0, and at 24 h for 16:0. The Δ⁹-desaturase products of these fatty acids were estimated to be 90% of cis-9 14:1, 50% of cis-9 16:1, and 59% of cis-9 18:1. This study demonstrates that ¹³C-labeled fatty acids may be utilized in vivo to measure the activity of the Δ⁹-desaturase enzyme.     

Lipid composition and erucic acid in rat liver cells in culture

Erucic acid (Δ¹³-docosenoic acid) was added to fetal calf serum, then fed to rat liver epithelial cells in culture, and uptake measured at intervals over 24 hr. During the first 6 hr. of incubation, uptake of the docosenoic acid was 21 nmoles/hr/mg protein in 7-day cells, and 15 mmoles/hr/mg protein in 14-day cells. Of¹⁴C-labeled erucic acid taken up by the cells in 24 hr, radioactivity measurements showed 60% of the total lipid¹⁴C activity derived from [1-¹⁴C] 22∶1 in neutral lipid (NL) and 40% in phospholipid (PL); whereas 55% of lipid¹⁴C activity was in NL and 45% in PL when the substrate was [14-¹⁴C] 22∶1. Within the NL fraction, 75% of¹⁴C activity derived from [1-¹⁴C] 22∶1 was in triglyceride (TG) and 11% in cholesterol (CHL), while 79% was in TG and 6.5% in CHL when the substrate was [14-¹⁴C] 22∶1. Triglycerides and cholesteryl esters accumulated in the cells during incubation with erucic acid. Among phospholipids separated by thin layer chromatography, 75% of¹⁴C activity was in lecithin (PC), 10% in phosphatidylethanolamine (PE), 5% in sphingomyelin (SPH), and 1% or less in cardiolipin (DPG). The highest specific activity (SA) was in PC, followed by SPH and PE. Incubation with erucic acid altered fatty acid composition of PC, PE, and SPH, although amounts of phospholipids were unaffected. Gas liquid chromatography analyses detected 18% erucic acid in PC, 2% in PE, and 4–5% in SPH.     

Metabolism of erucic acid in adipocytes isolated from rat epididymal fat

The metabolism of [14-¹⁴C]erucic acid and [U-¹⁴C]palmitic acid has been investigated in adipocytes isolated from rat epididymal fat. The rate of acylation of [¹⁴C]erucic acid in cellular lipids and oxidation to CO₂ and acid-soluble activity was ca. 1/3 of the rate with [¹⁴C]palmitic acid as substrate. A maximal incorporation of fatty acids in triacylglycerol was found at a fatty acid concentration of 0.8 mM in the medium, both with [¹⁴C]erucic acid and [¹⁴C]palmitic acid as substrate. Glucose added to the medium increased the esterification and decreased the oxidation of both fatty acids. No significant chain-shortening of [¹⁴C]erucic acid to shorter monoenes was identified in the fat cells. Increasing concentrations of unlabeled palmitic acid in the incubation medium markedly inhibited the esterification of [¹⁴C]erucic acid, whereas unlabeled erucic acid had little effect on the rate of esterification of [¹⁴C]palmitic acid.     

Effects of phorbol esters,A23187 and vasopressin on oleate metabolism in isolated rat hepatocytes

Studies were conducted to compare the metabolic effects of vasopressin, 4β-phorbol-12-myristate-13-acetate (PMA) and A23187 on ketogenesis and oleate metabolism in isolated hepatocytes from fed rats. Vasopressin inhibited the formation of acid-soluble products from [1-¹⁴C]oleate (0.25 mM, 0.5 mM and 1 mM), the inhibition being most marked at low (0.25 mM) concentration of oleate. Conversion of [1-¹⁴C]oleate into¹⁴CO₂ and esterified products was stimulated by vasopressin. The stimulatory effect of this hormone on¹⁴CO₂ production was most marked at high (1 mM) concentration of oleate, whereas that on [1-¹⁴C]-oleate esterification was most marked at low (0.25 mM) concentration of oleate. These vasopressin actions were abolished when hepatocytes were incubated in the absence of calcium in the medium. Our results strongly suggest that both increase in esterification and increase in oxidation to CO₂ contribute to the anti-ketogenic action of vasopressin when oleate is added as substrate, although the relative extent of their contribution varies according to the oleate concentration. The anti-ketogenic action of vasopressin was mimicked by PMA but not by A23187. PMA also caused a stimulation of [1-¹⁴C]oleate esterification although the effect was diminished at 1 mM [1-¹⁴C]oleate. A23187 failed to affect [1-¹⁴C]oleate esterification. The metabolic effects of PMA were elicited in the absence of extracellular calcium, too. Conversion of [1-¹⁴C]oleate into¹⁴CO₂ was only slightly increased by both PMA and A23187 when 1 mM [1-¹⁴C]oleate was added as substrate. The marked stimulatory effect of vasopressin on¹⁴CO₂ production from [1-¹⁴C]oleate was not reproduced even by the combination of PMA and A23187. The possible involvement of protein kinase C and calcium mobilization in the regulation of oleate metabolism is discussed.     

Oleate hydratase: Studies of substrate specificity

This work concerns studies of the substrate specificity of an enzyme preparation from a pseudomonad which catalyzes the stereospecific hydration of the Δ⁹-double bond of oleic acid. [5DL-³H]-5DL-hydroxystearic acid, [6DL-³H]-6DL-hydroxystearic acid, [7DL-³H]-7DL-hydroxystearic acid, [9DL-³H]-9DL-hydroxystearic acid, [10DL-³H]-10DL-hydroxystearic acid, [11DL-³H]-11DL-hydroxystearic acid, [14DL-³H]-14DL-hydroxystearic acid, [15DL-³H]-15DL-hydroxystearic acid, and [1-¹⁴C]-stearolic acid were prepared and incubated with the enzyme. Only the 10-hydroxysteric acid served as a substrate for the enzyme. The findings reported in this study, in conjunction with those previously reported, indicate that only the D-isomer of 10-hydroxystearic acid serves as a substrate for the enzyme.     

Metabolism of [1-14C]docosahexaenoate (22∶6n−3), [1-14C]eicosapentaenoate (20∶5n−3) and [1-14C]linolenate (18∶3n−3) in brain cells from juvenile turbotScophthalmus maximus

The incorporation of [1-¹⁴C]18∶3n−3, (LNA) and [1-¹⁴C]-22∶6n−3 (DHA), and the metabolismvia the desaturase/elongase pathways of [1-¹⁴C]LNA, and [1-¹⁴C]20∶5n−3 (EPA) were studied in brain cells from newly-weaned (1-month-old) and 4-month-old turbot. The rank order of the extent of net incorporation of both LNA and DHA into glycerophospholipids was total diradyl glycerophosphocholines (CPL)> total diradyl glycerophosphoethanolamines (EPL)> phosphatidylserine (PS) and phosphatidylinositol (PI) and was independent of the polyunsaturated fatty acid added, the age of the fish and the time of incubation. However, the rate of incorporation of LNA into total lipid, CPL, EPL and PS was significantly greater than the rate of incorporation of DHA, and there was a significantly greater amount of DHA incorporated into EPL than LNA. There was no significant difference between the amounts of LNA and DHA incorporated into total lipid, CPL, PS and PI. Therefore, little preferential uptake and incorporation of DHA into brain cells was apparent. In 24-h incubations, on average 1.1% and 8.5% of radioactivity from [1-¹⁴C]LNA and [1-¹⁴C]EPA, respectively, were recovered in the DHA fraction. Therefore, LNA cannot contribute significantly to brain DHA levels in the turbot but EPA can. There were no significant differences between the amounts of radioactivity from either [1-¹⁴C]LNA or [1-¹⁴C]EPA recovered in the individual products/intermediates of the desaturase pathways in brain cells from 30-day-old and 120-day-old turbot.     

Saturated fatty acids >C20 are not activated by acid:CoA ligase in rat brain or liver?

The formation of long-chain saturated acyl-[³H]CoA and [1-¹⁴C] acyl-[³H]CoA by rat brain microsomes and rat liver was examined. Acyl-CoA formation was markedly decreased as fatty-acid chain length increased from C₁₆ to C₂₀. No biosynthesis of behenyl-[³H] CoA or [1-¹⁴C] lignoceryl-[³H] CoA was observed. The results suggest that long-chain saturated fatty acids >20 carbons in length are not activated by acid:CoA ligase to form acyl-CoA.     

Fatty acid metabolism of the calanoid copepodParacalanus parvus: 2. Palmitate,stearate, oleate and acetate

The de novo biosynthesis of fatty acids in the wild, calanoid copepodParacalanus parvus was studied. The incubation of labeled acetate proved the de novo biosynthesis of saturated and monounsaturated even fatty acids from 14 to 20 carbons and the 22∶1 acid. Saturated and monounsaturated uneven fatty acids from 15 to 21 carbons were also synthesized. The copepod could not synthesize linoleic and α-linolenic acids. By administration of [1-¹⁴C]palmitate, [1-¹⁴C] stearate and [1-¹⁴C]oleate, it was possible to elucidate the general pattern of the de novo biosynthesis of fatty acids in the wildP. parvus.     

Effect of the Δ<Superscript>6</Superscript>-desaturase inhibitor SC-26196 on PUFA metabolism in human cellsinhibitor SC-26196 on PUFA metabolism in human cells

The objective of this study was to determine the effect of 2,2-diphenyl-5-(4-{[(1E)-pyridin-3-yl-methylidene]-amino}piperazin-1-yl)pentanenitrile (SC-26196), a Δ⁶-desaturase inhibitor, on PUFA metabolism in human cells. SC-26196 inhibited the desaturation of 2 μM [1-¹⁴C] 18∶2n−6 by 87–95% in cultured human skin fibroblasts, coronary artery smooth muscle cells, and astrocytes. By contrast, SC-26196 did not affect the conversion of [1-¹⁴C]20∶3n−6 to 20∶4 in the fibroblasts, demonstrating that it is selective for Δ⁶-desaturase. The IC₅₀ values for inhibition of the desaturation of 2 μM [1-¹⁴C] 18∶3n−3 and [3-¹⁴C]24∶5n−3 in the fibroblasts, 0.2–0.4 μM, were similar to those for the inhibition of [1-¹⁴C] 18∶2n−6 desaturation, and the rates of recovery of [1-¹⁴C] 18∶2n−6 and [3-¹⁴C] 24∶5n−3 desaturation after removal of SC-26196 from the culture medium also were similar. SC-26196 reduced the conversion of [3-¹⁴C] 22∶5n−3 and [3-¹⁴C] 24∶5n−3 to DHA by 75 and 84%, respectively, but it had no effect on the retroconversion of [3-¹⁴C] 24∶6n−3 to DHA. These results demonstrate that SC-26196 effectively inhibits the desaturation of 18- and 24-carbon PUFA and, therefore, decreases the synthesis of arachidonic acid, EPA, and DHA in human cells. Furthermore, they provide additional evidence that the conversion of 22∶5n−3 to DHA involves Δ⁶-desaturation.