Incorporation of linoleic acid and its conversion to λ-linolenic acid in fungi

The incorporation of [1-¹⁴C]linoleic acid (LA) into lipids ofMortierella ramanniana var.angulispora was studied to determine which lipid classes participated in the δ6-desaturation of [1-¹⁴C]LA. [1-¹⁴C]LA was rapidly taken up into fungal cells and esterified into various lipids. Comparison of the profile of [1-¹⁴C]LA incorporation between fungal cells at the exponential growth phase and the stationary growth phase showed that [1-¹⁴C]LA incorporation into most lipids—except for triacylglycerol (TG) and phosphatidylcholine (PC)—were greatly reduced at the stationary growth phase. Desaturation of [1-¹⁴C]LA into λ-linolenic acid (GLA) readily occurred at the exponential growth phase, but was greatly decreased at the stationary growth phase. Moreover, pulse-chase experiments revealed that the radiolabel incorporated into phosphatidylserine (PS) and PC rapidly turned over, while that in TG and diacylglycerol (DG) accumulated after the 4 hr chase. In addition to the change of the radiolabel in individual lipids, the content of radiolabeled GLA converted from [1-¹⁴C]LA varied with individual lipids. In phospholipids such as PC, phosphatidylethanolamine (PE) and PS, radiolabeled GLA rapidly increased after 1 hr and then decreased after 4 hr. On the other hand, a gradual increase in radiolabeled GLA until 4 hr was observed in TG. These results suggest that LA, which has been esterified into phospholipids such as PC, PE and PS, is readily desaturated to GLA, which is then transferred to TG. These differences in the fate of GLA derived from LA between phospholipids and neutral lipids may be reflected in the GLA content in the individual lipids.     

Microbial conversion of an oil containing α-linolenic acid to an oil containing eicosapentaenoic acid

Mycelia of arachidonic acid-producing fungi belonging to the genusMortierella were found to convert an oil containing α-linolenic acid to an oil containing 5,8,11,14,17-cis-eicosapentaenoic acid (EPA). This conversion was observed when they were grown in a medium containing the oil, glucose and yeast extract at 28 C. On the screening of various oils, linseed oil, in which α-linolenic acid amounts to about 60% of the total fatty acids, was found to be the most suitable for EPA production. Under the optimal culture conditions, a selected strain,Mortierella alpina 20-17, converted 5.1% of the α-linolenic acid in the added oil into EPA, the EPA production reaching 1.35 g/l of culture broth (41.5 mg/g dry mycelia). This value corresponded to 7.1% (by weight) of the total fatty acids in the extracted lipids. The lipid was also found to be rich in arachidonic acid (12.3%). Other major fatty acids in the lipid were palmitic acid (4.4%), stearic acid (3.2%), oleic acid (13.5%), linoleic acid (13.7%), α-linolenic acid (38.5%) and γ-linolenic acid (0.9%).     

Eskimo plasma constituents,dihomo-γ-linolenic acid,eicosapentaenoic acid and docosahexaenoic acid inhibit the release of atherogenic mitogens

Studies in man and laboratory animals suggest that ω3 polyunsaturated fatty acid consituents of fish oils have antiatherosclerotic properties. We have studied the effects of several such polyunsaturated fatty acids for ability to modify the in vitro release of mitogens from human platelets. Such mitogens may produce the fibroproliferative component of atherosclerotic plaques. Both 5,8,11,14,17-eicosapentaenoic acid (20∶5ω3) and 4,7,10,13,-16,19-docosahexaenoic acid (22∶6ω3), major constituents of fish oils, inhibited adenosine diphosphate-induced aggregation of platelets and the accompanying release of mitogens. These effects are dose dependent. Linolenic acid (18∶3ω3), the biosynthetic precursor of eicosapentaenoic acid, also inhibited platelet aggregation and mitogen release. Eicosapentaenoic acid also inhibited mitogen release from human monocyte-derived macrophages, which, in vivo, are an additional source of mitogens during atherogenesis. Potent inhibition of human platelet aggregation and mitogen release was also seen with dihomo-γ-linolenic acid (8,11,14-eicosatrienoic acid 20∶3ω6), whose levels are reportedly elevated in Eskimos subsisting on marine diets. We conclude that diets that elevate plasma and/or tissue levels of eicosapentaenoic acid, docosahexaenoic acid and dihomo-γ-linolenic acid precursor γ-linolenic acid (18∶3ω6) may exert antiatherosclerotic effects by inhibiting the release of mitogens from platelets and other cells.     

Metabolic transformation of intracraneally injected [1-14C]linoleic and [1-14C]α-linolenic acids in malnourished developing rats

The effect of a low protein diet during pregnancy and lactation on the fatty acid composition of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) from brains of ten-day-old rats was studied. The results indicated that partial deprivation of protein during early development was associated with an increase in the fatty acids of the n−9 family in PC. The fatty acids of the linoleic acid series decreased in PE but were not modified in PC. These minor changes did not affect the double bond index values either in PC or in PE. The effect of protein depletion on thein vivo metabolic transformation of intracraneally injected [1-¹⁴C]linoleic and [1-¹⁴C]α-linolenic acids was also studied. The percentage distribution of the labeled precursors and their derivatives among PC and PE differed from that of mass distribution. These results indicate that the direct uptake of polyunsaturated fatty acids from the blood and/or the low turnover rate of these acids incorporated into PC and PE might be involved in maintaining the fatty acid pattern of these brain lipids.     

Incorporation of linolenic-1-C14 acid into eicosapentaenoic and docosahexaenoic acids in fish

Following intraperitoneal injection of methyl linolenate-1-C¹⁴ into kelp bass,Paralablax clathratus, the highly polyunsaturated fatty acids of their body fats were concentrated by low temperature crystallization from acetone, and eicosapentaenoic and docosahexaenoic acids were isolated from the concentrate by reversed-phase chromatography and hydrogenated. The resulting arachidic and behenic acids were degraded stepwise to margaric acid, and the distribution of activity was determined. The results indicate that the injected linolenic acid was converted to eicosapentaenoic acid and the latter incorporated into docosahexaenoic acid. A probable conversion pathway is linolenic acid→6,9,12,15-octadecatetraenoic acid→8,11,14,17-eicosatetraenoic acid→5,8,11,14,17-eicosapentaenoic acid→7,10,13,16,-19-docosapentaenoic acid→4,7,10,13,16,19-docosahexaenoic acid. Supported by a training grant from the National Heart Institute, Bethesda, Md. This paper based partially on work performed under Contract AT(04-1)GEN-12 between the Atomic Energy Commission and the University of California.     

Linoleic acid uptake by isolated enterocytes: Influence of α-linolenic acid on absorption

In a previous study we showed that intestinal uptake of α-linolenic acid (18∶3n−3) was carrier-mediated and we suggested that a plasma membrane fatty acid protein was involved in the transport of long-chain fatty acids. To further test this hypothesis, the mechanism of linoleic acid (18∶2n−6) uptake by isolated intestinal cells was examined using a rapid filtration method and 20 mM sodium taurocholate as solubilizing agent. Under these experimental conditions transport of [1-¹⁴C]linoleic acid monomers in the concentration range of 2 to 2220 nM was saturable with a V_m of 5.1±0.6 nmol/mg protein/min and a K_m of 183±7 nM. Experiments carried out in the presence of metabolic inhibitors, such as 2,4-dinitrophenol and antimycin A, suggested that an active, carriermediated mechanism was involved in the intestinal uptake of this essential fatty acid. The addition of excess unlabeled linoleic acid to the incubation medium led to a 89% decrease in the uptake of [1-¹⁴C]linoleic acid, whiled-glucose did not compete for transport into the cell. Other long-chain polyunsaturated fatty acids added to the incubation mixture inhibited linoleic acid uptake by more than 80%. The presence of α-linolenic acid (18∶3n−3) in the incubation medium caused the competitive inhibition (K_i=353 nM) of linoleic acid uptake. The data are compatible with the hypothesis that intestinal uptake of both linoleic, and α-linolenic acid is mediated by a membrane carrier common to long-chain fatty acids.     

Involvement of cytochrome b5 in the oxidative desaturation of linoleic acid to γ-linolenic acid in rat liver microsomes

The effects of antibodies against microsomal electron-transport components on the in vitro activity of Δ⁶-desaturation of linoleic acid to γ-linolenic acid have been studied in intact microsomal membranes of rat liver. Reduced nicotinamide adenine dinucleotide (NADH) or reduced nicotinamide adenine dinucleotide phosphate (NADPH) (0.87 mM) served as electron donors, and effectively prompted the Δ⁶-desaturase activities with yields of about 1.1 to 1.3 nmol per mg of protein in 10 min. Of the two antibodies studied under the same in vitro conditions, i.e., rabbit antisera preparations against rat liver microsomal hydrophilic parts of cytochrome b₅ and NADPH-cytochrome c reductase, only the antibody against cytochrome b₅ demonstrated a marked ability to inhibit the Δ⁶-desaturase activity. This evidence supports a participation of cytochrome b₅ in the Δ⁶-desaturation of linoleic acid and suggests a pathway analogous to the Δ⁹-desaturation of stearyl-CoA.     

The effect of dietary α-linolenic acid in the rat on fatty acid profiles of immunocompetent cell populations

Analysis of diet-induced fatty acid changes in the major phospholipids of various immune cell populations has not been previously documented, particularly modifications induced by dietary α-linolenic acid. Rats were fed purified diets containing either 10% corn oil (CO), 10% linseed oil (LO) or 10% soybean oil-linseed mixture (SL) for 8 weeks. The α-linolenic to linoleic acid ratios of the diets were 1∶32, 1∶1 and 3∶1, respectively. Fatty acid analysis of cell populations isolated from the spleen, thymus, thoracic cavity and peripheral blood phospholipids showed increases in ω3 fatty acids accompanied by decreases in the ω6 fatty acids when diets high in α-linolenic to linoleic acid ratios were fed. The extent of change observed was dependent on the magnitude of the α-linolenic to linoleic acid ratio. Both magnitude of change and the specific fatty acids altered varied with the cell population examined.     

Enzymatic fractionation of fatty acids: Enrichment of γ-linolenic acid and docosahexaenoic acid by selective esterification catalyzed by lipases

Immobilized lipase preparations from seedlings of rape (Brassica napus L.) andMucor miehei (lipozyme) used as biocatalysts in esterification and hydrolysis reactions discriminate strongly against γ-linolenic and docosahexaenoic acids/acyl moieties. Utilizing this property, γ-linolenic acid contained in fatty acids of evening primrose oil has been enriched seven to nine-fold, from 9.5 to almost 85% by selective esterification of the other fatty acids with butanol. Similarly, docosahexaenoic acid of cod liver oil has been enriched four to five-fold, from 9.4 to 45% by selective esterification of fatty acids (other than docosahexaenoic acid) with butanol. As long as the reaction is stopped before reaching equilibrium, very little of either γ-linolenic acid or docosahexaenoic acid are converted to butyl esters, which results in high yields of these acids in the unesterified fatty acid fraction.     

The incorporation of orally administered radiolabeled dihomo γ-linolenic acid (20∶3ω6) into rat tissue lipids and its conversion to arachidonic acid

Radioactivity from orally administered radiolabeled dihomo-γ-linolenic acid (20∶3ω6) was recovered from the liver, plasma and brain lipid fractions. After administration the fatty acid was metabolized to arachidonic acid, the 22 carbon chain length fatty acid, and was also β-oxidized. However, 22 hr after administration of [1-¹⁴C]20∶3 between one-third and one-half of the recovered radioactivity was still associated with dihomo-γ-linolenic acid in the liver and plasma lipid fractions. Orally administered dihomo-γ-linolenic acid is incorporated into lipid fractions and is, therefore, available in the metabolic pool for PGE₁ synthesis.     

Influence of dietary fat on metabolism of (14-14C)erucic acid in the perfused rat liver. Distribution of metabolites in lipid classes

Two groups of rats were fed diets containing 20% by weight of either partially hydrogenated marine oil supplemented with sunflower seed oil (PHMO) or palm oil (PO) for 8 wk. Using a liver perfusion system, the effect of dietary long chain monoenoic fatty acids on the uptake and metabolism of [14-¹⁴C]erucic acid was studied. The perfusion times were 15 and 60 min, respectively. The two groups showed equal ability for erucic acid uptake in the liver but differed in the channeling of the fatty acids into various metabolic pathways. A higher metabolic turnover of 22∶1 in the PHMO livers relative to the PO livers was demonstrated by an increased recovery of total [¹⁴C]labeling in the triglyceride (TG) and phospholipid (PL) fractions, already evident after 15 min of perfusion. The chainshortening capacity was highest in the PHMO group, reflected by a higher [¹⁴C]18∶1 incorporation in both TG and PL, and increasing from 15 to 60 min of perfusion. The amount of [¹⁴C]18∶1 found in PL and TG after 60 min of perfusion of livers from rats fed PO corresponded to that shown for the PHMO group after 15 min. The PL demonstrated a discrimination against 22∶1 compared to TG, and, when available, 18∶1 was highly preferred for PL-synthesis. The total fatty acid distribution in the TG, as determined by gas liquid chromatography (GLC), reflected the composition of the dietary fats. In the total liver PL, 22∶1 and 20∶1 were present in negligible amounts, although the PHMO diet contained 12–13% of both 22∶1 and 20∶1. In the free fatty acid fraction (FFA), the major part of the radioactivity (≈80%) was [14-¹⁴C]erucic acid, and only small amounts of [¹⁴C]18∶1(<2%) were presents, even after 60 min of perfusion. The shortened-chain 18∶1 was readily removed from the FFA pool and preferentially used for lipid esterification.     

Effect of dietary α-linolenic acid intake on incorporation of docosahexaenoic and arachidonic acids into plasma phospholipids of term infants

The fractional conversion rates of plasma phospholipid α-linolenic acid (18:3n-3) and linoleic acid (18:2n-6) to docosahexaenoic acid (22:6n-3) and arachidonic acid (20:4n-6), respectively, and the fractional rates of incorporation of 22:6n-3 and 20:4n-6 into plasma phospholipids were determined in 27 healthy 3-wk-old term infants who had received formulas with ≈16% of fat as 18:2n-6 and 0.4% (n=6), 1.0% (n=11), or 3.2% (n=10) as 18:3n-3 from birth. The infants were given a single dose of both [U-¹³C] 18:2n-6 and [U-¹³C]18:3n-3 with a feeding, and blood samples were collected 8, 12, and 24 h afterward for determination of the isotopic enrichments of the [M+18] isotopomers of plasma phospholipid fatty acids by negative chemical ionization gas chromatography/mass spectrometry. A simple precursor/product compartmental model was used to estimate fractional rates of conversion and incorporation. All infants converted 18:3n-3 to 22:6n-3 and 18:2n-6 to 20:4n-6. Although the fractional rate of conversion of 18:3n-3 to 22:6n-3 did not differ among groups, the fractional rate of incorporation of 22:6n-3 into the plasma phospholipid fraction was greater in infants who received 3.2% vs. 0.4% or 1.0% 18:3n-3 (4.1±2.2 vs 1.6±1.5 or 2.0±1.0% of the plasma phospholipid 22:6n-3 pool daily). The fractional rate of conversion of 18:2n-6 to 20:4n-6 was less in infants who received the 3.2% 18:3n-3 intake (0.4±0.3% of the plasma phospholipid 18:2n-6 pool daily vs. 1.1±0.7% and 0.8±0.5% in those who received 0.4 and 1.0% 18:3n-3, respectively). The fractional rate of incorporation of 20:4n-6 into plasma phospholipid also was less in the 3.2% vs. the 0.4 and 1.0% 18:3n-3 groups (2.7±1.4% vs. 5.9±2.6 and 4.4±1.7%, respectively, of the plasma phospholipid 20:4n-6 pool daily).     

[3-13C] γ-linolenic acid: A new probe for 13C nuclear magnetic resonance studies of arachidonic acid synthesis in the suckling rat

Our objective was to develop a suitable probe to study metabolism of polyunsaturated fatty acids by ¹³C nuclear magnetic resonance (NMR) in the suckling rat pup. [3-¹³C] γ-Linolenic acid was chemically synthesized, and a 20 mg (Experiment 1) or 5 mg (Experiment 2) dose was injected into the stomachs of 6–10-day-old suckling rat pups that were then killed over a 192 h (8 d) time course. ¹³C NMR showed that ¹³C in γ-linolenate peaked in liver total lipids by 12-h post-dosing and that [5-¹³C]-arachidonic acid peaked in both brain and liver total lipids 48–96 h post-dosing. ¹³C enrichment in brain γ-linolenic acid was not detected by NMR, but gas chromatography-combustion-isotope ratio mass spectrometry showed that its mass enrichment in brain phospholipids at 48–96 h post-dosing was 1–2% of that in brain arachidonic acid. ¹³C was present in liver and brain cholesterol and in perchloric acid-extractable water-soluble metabolites in the brain, liver and carcass. We conclude that low but measurable amounts of exogenous γ-linolenic acid do access the suckling rat brain in vivo. The slow time course of [5-¹³C] arachidonic acid appearance in the brain suggests most of it was probably transported there after synthesis elsewhere, probably in the liver. Some carbon from γ-linolenic acid is also incorporated into lipid products other than n−6 long-chain polyunsaturated fatty acids.     

Natural and accelerated docosahexaenoic acid accumulation in the prenatal rat brain

The quantity and distribution of docosahexaenoic acid (DHA) in major brain phospholipids (PL) was examined in the fetal rat brain before birth, using thin-layer and capillary column gas chromatography. A rapid increment of DHA content of about 187 μg/g brain/day was observed between 17 to 20 days gestation, as opposed to 39.3±2.9 μg/g brain/day prior to that. Single intraamniotic injections of 5 μL ethyl-docosahexaenoate (Et-DHA) 12 μM, 4.25 mg) administered to 17-day-old fetuses were used to examine the uptake of DHA into brain PL. Three days following injection, the amount of n-3 polyunsaturated fatty acids increased by 28% compared to ethyl-oleate (Et-Ole) injected fetuses. Compared to the n-6 fatty acid family, the relative amount of DHA increased in the phosphoatidylserine (PS), phosphatidylethanolamine and phosphatidylcholine (PC) lipids by 15 (P=0.02), 13 and 14%, respectively. A major increase in the pool size of phosphatidylinositol and PS (110 and 50.3%, respectively), and a decrease in PC (8.2%) were observed 3 d after Et-DHA as compared to Et-Ole administration. The data suggest that a single intraamniotic administration of Et-DHA can modulate membrane PL content and alter PUFA composition.     

Differential effects of dietary linoleic and α-linolenic acid on lipid metabolism in rat tissues

Comparative effects of feeding dietary linoleic (safflower oil) and α-linolenic (linseed oil) acids on the cholesterol content and fatty acid composition of plasma, liver, heart and epididymal fat pads of rats were examined. Animals fed hydrogenated beef tallow were used as isocaloric controls. Plasma cholesterol concentration was lower and the cholesterol level in liver increased in animals fed the safflower oil diet. Feeding the linseed oil diet was more effective in lowering plasma cholesterol content and did not result in cholesterol accumulation in the liver. The cholesterol concentration in heart and the epididymal fat pad was not affected by the type of dietary fatty acid fed. Arachidonic acid content of plasma lipids was significantly elevated in animals fed the safflower oil diet and remained unchanged by feeding the linseed oil diet, when compared with the isocaloric control animals fed hydrogenated beef tallow. Arachidonic acid content of liver and heart lipids was lower in animals fed diets containing safflower oil or linseed oil. Replacement of 50% of the safflower oil in the diet with linseed oil increased α-linolenic, docosapentaenoic and docosahexaenoic acids in plasma, liver, heart and epididymal fat pad lipids. These results suggest that dietary 18∶2ω6 shifts cholesterol from plasma to liver pools followed by redistribution of 20∶4ω6 from tissue to plasma pools. This redistribution pattern was not apparent when 18∶3ω3 was included in the diet.     

Fatty acid synthesis in rat testes injected intratesticularly or incubated with 1-14C acetate

Fatty acid synthesis was studied in testes of young and adult rats either injected intratesticularly or incubated with 1-¹⁴C acetate. The pattern of¹⁴C incorporation into lipids and individual fatty acids in the two age groups was similar but results obtained with intratesticular injection differed considerably from those obtained in the in vitro studies. In the former more than 70% of the¹⁴C incorporated in total lipids was in phosphatides, with about 15% in triglycerides and only minor amounts in cholesteryl esters and free fatty acids. Most of the¹⁴C incorporated into total fatty acids was in saturated acids (predominantly 16∶0). A small amount of¹⁴C was in the higher polyenes and there was a progressive increase with time after acetate injection in the¹⁴C content of 22∶5 W6. In testes incubated with 1-¹⁴C acetate, the phosphatide, triglyceride, and free fatty acid fractions had similar amounts of¹⁴C. In the total fatty acid fraction about 40% of the incorporated¹⁴C was in saturated acids (predominantly 14∶0 and 16∶0) and about 50% in the higher polyenes. Twenty carbon polyenes and 22∶5 W6 had significant¹⁴C incorporation, but most of the¹⁴C was in 22∶4 W6. About 80% of the¹⁴C in the latter compound was in the carboxyl carbon, indicating its origin from endogenous 20∶4 W6 elongated by the added 1-¹⁴C acetate used as substrate. The¹⁴C 22∶4 was present predominantly in the triglyceride and phosphatide fractions with minor amounts in other lipids.¹⁴C-labeled compounds of retention time greater than 22∶5 were also present in all lipid fractions. Presented at the ISF-AOCS World Congress Chicago, September 1970.     

Increased hepatic β-oxidation of docosahexaenoic acid, elongation of eicosapentaenoic acid, and acylation of lysophosphatidate in rats fed a docosahexaenoic acid-enriched diet

Rats were fed a diet supplemented with corn oil (n-3 deficient), soy oil, or a mixture containing 8% 22∶6n-3 ethyl ester for 6 wk. The hepatic capacities for the β-oxidation and synthesis of 22∶6n-3, in addition to the acylation of lysophosphatidate, were tested in vitro. In rats that were fed a 22∶6n-3-enriched diet, both the β-oxidation of 22∶6n-3 and elongation of 20∶5n-3 were enhanced compared to those in rats fed the other diets. Acylation of lysophosphatidate was also enhanced in rats fed a 22∶6n-3-enriched diet, while the rate of dephosphorylation of phosphatidate was not changed. The amount of 22∶6n-3 in the liver was much less than that consumed in a docosahexaenoic acid-enriched diet. These results suggest that a significant amount of dietary 22∶6n-3 was degraded via β-oxidation, and that a portion of the retroconverted 20∶5n-3 was recycled for the synthesis of 22∶6n-3. The recycling of 20∶5n-3 might contribute to the low level of 22∶6n-3 in rats fed an n-3-deficient diet.     

Production of dihomo-γ-linolenic acid byMortierella alpina 1S-4

The mycelial dihomo-γ-linolenic acid content of an arachidonic acid-producing fungus,Mortierella alpina 1S-4, was found to increase, with an accompanying marked decrease in its arachidonic acid content, on cultivation with sesame oil. The resultant mycelia were found to be a rich source of dihomo-γ-linolenic acid. This unique phenomenon was suggested to be due to specific repression of the conversion of dihomo-γ-linolenic acid to arachidonic acid by the oil. After fractionation of the oil with acetone into oil and non-oil fractions, it was found that the effective factor(s) was present in the non-oil fraction. In a study on optimization of the culture conditions for the production of dihomo-γ-linolenic acid byM. alpina 1S-4, a medium containing glucose, yeast extract and the non-oil fraction was found to be suitable for the production. Under the optimal conditions in a 50-1 fermentor, the fungus produced 107 mg of dihomo-γ-linolenic acid/g dry mycelia (2.17 g/l of culture broth). This value accounted for 23.1% of the total fatty acids in the lipids extracted from the mycelia. The mycelia were also rich in arachidonic acid (53.5 mg/g dry mycelia, 11.2%). Other major fatty acids in the lipids were palmitic acid (24.1%), stearic acid (7.0), oleic acid (20.1), linoleic acid (6.6) and γ-linolenic acid (4.1). On leave from Suntory Ltd.     

The in vivo incorporation of labeled linoleic acid, α-linolenic acid and arachidonic acid into rat liver lipids

The incorporation of 1-¹⁴C-linoleic acid, 1-¹⁴C-α-linolenic acid and 1-¹⁴C-arachidonic acid into rat liver lipids was measured and the per cent distribution of radioactivity into the different lipid fractions determined. Normal rats were injected into the portal vein with the labeled solutions during a one minute period. Livers were quickly frozen, pulverized, and the lipids extracted and fractioned by thin layer chromatography. No significant differences were observed in the amounts of labeled fatty acids incorporated per gram of rat liver. While 1-¹⁴C-linoleic acid and 1-¹⁴C-α-linolenic acid were found in appreciable amounts in the 1,2 diacylglycerol fraction, about one fifth as much 1-¹⁴C-arachidonic acid was esterified in this fraction. 1-¹⁴C-arachidonic acid was the leading acid esterified in the phospholipid fractions.     

The metabolism of 1-14C-palmitic acid in the testis of the rat

The metabolism of 1-¹⁴C-palmitic acid was studied in rat testis at various intervals after intratesticular injection and after a 2 hr incubation period. Significant catabolism occurred as evidenced by production of¹⁴CO₂ in both in vivo and in vitro experiments. Of the¹⁴C activity retained in the injected testis, less than 15% remained as free fatty acid after 2 hr and less than 5% by the end of 2 weeks. Activity of¹⁴C appeared in phosphatides faster than in the triglyceride fraction, and with time (1–2 weeks) the activity in phosphatides decreased relative to that in triglycerides. In phosphatides of these experiments ca. 80% of the¹⁴C was in the palmitic acid fraction and the balance predominantly (98%) in palmitate with the balance in stearate and oleate. By the first and second weeks almost half of the¹⁴C-palmitate present in the testes had been newly synthesized from¹⁴C-acetyl CoA resulting from oxidation of the administered 1-¹⁴C-palmitate. In the incubated samples the only fatty acid with¹⁴C activity in any lipid fraction was palmitate. In these experiments ca 90% of the¹⁴C was in the free fatty acid fraction, 7% in the phosphatides and 2% in the triglycerides.