Triacylglycerol biosynthesis in everted sacs of rat intestinal mucosa.

The molecular specificity of the biosynthesis of triacylglycerols by rat intestinal mucosa was examined by means of radioactive and mass tracers, and thin-layer chromatography with silver nitrate and gas-liquid chromatography with radioactivity monitoring. Bile salt micelles of alternately labeled monoacylglycerols and free fatty acids were incubated with everted sacs of intestinal mucosa for various periods of time and the triacylglycerols isolated by solvent extraction and thin-layer chromatography. Analyses of the molecular species of the triacylglycerols labeled from monoacylglycerols showed that the 2-monoacylglycerol pathway was responsible for the biosynthesis of a maximum of 90% and the X-1-monoacylglycerol pathway for about 10% of the total radioactive triacylglycerols. Detailed analyses of the molecular species of triacylglycerols labeled fro free fatty acids showed that the phosphatidic acid pathway contributed a minimum of 20-30% of the total labeled triacylglycerol formed. There was a preferential utilization in triacylglycerol biosynthesis of the more unsaturated diacylglycerols arising from the monoacylglycerol pathway and of the more saturated diacylglycerols originating from the phosphatidic acid pathway. The above experiments do not allow a demonstration of the utilization of the sn-2,3-diacylglycerols in triacylglycerol biosynthesis but are not inconsistent with it.     

SELECTIVE SYNTHESIS OF MOLECULAR CLASSES OF PHOSPHATIDIC ACID, DIACYLGLYCEROL AND PHOSPHATIDYLINOSITOL IN RAT BRAIN

—Phosphatidic acids of rat brain were shown to be predominantly of the monoenoic class while diacylglycerols and phosphatidylinositols were constituted mainly by tetraenes. Metabolic inter-relationships were examined after intraventricular injection of [¹⁴C]glycerol, [³H]arachidonate and [9,10-³H₂]stearate. In each case, diacylglycerols were most highly labelled, although a small pool of arachidonate was located in brain triacylglycerols, mainly esterified to a primary carbinol, with extremely high turnover rate. Fractionation of the lipids showed a preferential synthesis of disaturated, monoenoic and polyenoic classes (>4 double bonds) of phosphatidic acid, diacylglycerol and phosphatidylinositol. The high flux of [³H]stearate through disaturated species of phosphatidic acid and diacylglycerol could be partially suppressed by simultaneous injections of unsaturated fatty acids, both probably consequences of perturbing the very small brain pool of free fatty acids. Kinetics of labelling of phosphatidylinositols were consistent with formation of arachidonoyl-containing species by acyl transfer mechanisms with disaturated and oligoenoic classes serving as precursors. Although the profile of molecular classes of diacylglycerol and phosphatidylinositol strongly suggests a metabolic relation, there was no obvious evidence for this in the kinetic studies of the whole brain lipids. Such relation, however, may have been masked by the rapid flow of radioactivity from phosphatidic acids to diacylglycerols.     

THE METABOLISM OF [1-14C]ARACHIDONIC ACID IN THE NEUTRAL GLYCERIDES AND PHOSPHOGLYCERIDES OF MOUSE BRAIN1

Abstract— [1-¹⁴C]Arachidonic acid was incorporated into brain lipids with a half-life of approx. 5 min. Within 40 min after intra-cerebral injection, radioactivity was distributed mainly among the diacyl-sn-glycero-3-phosphorylcholine (45 per cent), diacyl-sn-glycero-3-phosphorylinositol (22 per cent), diacyl-sn-glycero-3-phosphorylethanolamine (14 per cent) and triacylglycerols (9 per cent). At comparable times, the proportions of radioactivity distributed in diacyl-sn-glycero-3-phosphorylserines and alkenylacyl-sn-glycero-3-phosphorylethanolamines were relatively small. Radioactivity was initially incorporated into the phosphatidio acids and diacylglycerols before labelling of the triacylglycerols and other phosphogly-cerides. The relative specific activity of diacylglycerols was maximum between 3–6 min after injection. Due to the small level of diacyl-sn-3-phosphorylinositol present in brain, its relative specific radioactivity was higher than other types of brain phosphoglycerides. Results of the experiment thus indicate that labelled arachidonic acid is an excellent precursor for metabolic studies with regard to acyl groups present in the 2-position of the phosphoglyceride molecules. Furthermore, this labelled precursor is specially useful in studies related to metabolism of diacyl-sn-glycero-3-phosphorylinositol in brain.     

The distribution of 14C-arachidonic acid in hamster lungs is sensitive to quinacrine

Isolated hamster lungs were labelled with ¹⁴C-arachidonic acid. When the lungs were ventillated with a respirator only a small amount of radioactivity was released to the perfusion effluent. This release was not changed significantly by pulmonary infusion of quicacrine (0.5 mM), a known inhibitor of phospholipase A₂. After the perfusion about 75% of the radioactivity in the lungs was in phospholipids, mainly in phosphatidylcholine, phosphatidylethanolamine and phosphatidylinostil and to a lesser degree in phosphatidylserine and phosphatidic acid. About one fourth of the radioactivity was in neutral lipids (tri- and diacylglycerols) and as free unmetabolized ¹⁴C-arachiodonic acid. Pulmonary infusion of quinacrine increased the amount of radioactivity in diacylglycerols and phosphatidylinositol but had no effect on that in phosphatidylcholine, phosphatidylserine, phosphatidic acid and triacylglycerols. The amount of radioactivity in phosphatidylethanolamine was decreased by quinacrine and increased in the vicinity of an unidentified phospholipid-quinacrine complex. The present study indicates that the distribution of ¹⁴C-arachidonic acid in hamster lung lipids is sensitive to quinacrine. The detected changes can, however, not be explained by an overall inhibition of phospholipase A₂ activities.     

The distribution of C-arachidonic acid in hamster lungs is sensitive to quinacrine

Isolated hamster lungs were labelled with ¹⁴C-arachidonic acid. When the lungs were ventillated with a respirator only a small amount of radioactivity was released to the perfusion effluent. This release was not changed significantly by pulmonary infusion of quicacrine (0.5 mM), a known inhibitor of phospholipase A₂. After the perfusion about 75% of the radioactivity in the lungs was in phospholipids, mainly in phosphatidylcholine, phosphatidylethanolamine and phosphatidylinostil and to a lesser degree in phosphatidylserine and phosphatidic acid. About one fourth of the radioactivity was in neutral lipids (tri- and diacylglycerols) and as free unmetabolized ¹⁴C-arachiodonic acid. Pulmonary infusion of quinacrine increased the amount of radioactivity in diacylglycerols and phosphatidylinositol but had no effect on that in phosphatidylcholine, phosphatidylserine, phosphatidic acid and triacylglycerols. The amount of radioactivity in phosphatidylethanolamine was decreased by quinacrine and increased in the vicinity of an unidentified phospholipid-quinacrine complex. The present study indicates that the distribution of ¹⁴C-arachidonic acid in hamster lung lipids is sensitive to quinacrine. The detected changes can, however, not be explained by an overall inhibition of phospholipase A₂ activities.     

Glycerolipid synthesis by homogenate and oil bodies from developing mustard (Sinapis alba L.) seed

[1-¹⁴C]Oleic acid and [14-¹⁴C]erucic acid were converted to their acyl-CoA derivatives and incorporated into acyl lipids by a homogenate from developing mustard (Sinapis alba L.) seed and oil bodies, as well as supernatant isolated by centrifugation at 20000 g. In both homogenate and oil bodies, the oleoyl moieties from exogenous [1-¹⁴C]oleoyl-CoA were most extensively incorporated into phosphatidic acids, but very little into phosphatidylcholines. The pattern of labelling of acyl lipids by oleoyl versus erucoyl moieties from either of the corresponding fatty acids, added individually or as a mixed substrate, indicates that oleoyl-CoA directly acylates sn-glycerol-3-phosphate to yield lysophosphatidic acids and phosphatidic acids that are subsequently converted to mono- and diacylglycerols. In contrast, erucoyl-CoA predominantly acylates preformed mono-and diacylglycerols containing oleoyl moieties to yield triacylglycerols containing erucoyl moieties.     

Fatty Acid Incorporation in Normal and Degenerating Rat Sciatic Nerve In Vivo

Abstract: Labeled palmitic acid ([16-¹⁴C]palmitate) (0).5 μCi) was injected into rat sciatic nerves in vivo to characterize thc incorporation of this fatty acid into complex peripheral nerve lipids after time lapses of 1 min to 2 weeks. For the first 30 min after intraneural injection, the label was concentrated in nerve diglycerides. Thereafter, the relative diglyccride label declined rapidly, and phospholipid radioactivity rose steadily. After 120 min, phospholipids contained over 70% of the total lipid radioactivity. Among the phospholipids, phosphatidylcholine had the largest percentage of total phospholipid label, and acylation of lysophosphatidylcholine accounted for approximately 75% of this label. With time, there was conversion of [16-¹⁴C]palmitate to other long-chain fatty acids by elongation and desaturation. Phosphatidic acid was labeled also, suggesting the operation of the de novo biosynthetic mechanism. However, the specific radioactivity of 1,2-diacylglycerol was much higher than that of phosphatidic acid, suggesting phosphorylation of diglycerides by diglyceride kinase. After nerve section and survival of 2 h to 50 days, the injection of [16-¹⁴C]palmitate into the degenerating distal segment revealed an overall decline of phospholipid labeling and a commensurate increase of triglyceride radioactivity. Phosphatidylcholine in degenerating nerve contained a larger percentage of the fatty acid label than that in normal nerve. Almost all of the labeling was due to acylation of lysophosphatidylcholine, implying a much smaller contribution of the de novo pathway. Phosphatidylethanolamine and phosphatidylserine showed a relative loss of radioactivity. The changes were apparent at 1 day, but not at 2 h, suggesting loss of homeostatic control, presumably by interruption of axonal flow. An incidental observation was the stimulation of phosphatidylcholine biosynthesis by acylation of lysophosphatidylcholine in the contralateral unoperated sciatic nerve.     

Phospholipid biosynthesis in the lungs in a chronic inflammatory bronchopulmonary process

Glycerokinase, glycerophosphate dehydrogenase, glycerophosphate acyltransferase activity, glycerophosphate, dioxiacetonphosphate level and in vivo incorporation of (U-14C)-glucose into the lung phospholipid structure were studied in normal rats and in conditions of chronic bronchopulmonary inflammation. The inhibition of glycerokinase and glycolytic ways of glycerophosphate formation was demonstrated. The data obtained have shown that glucose incorporation into phosphatidyl cholines, phosphatidyl glycerols and phosphatidyl ethanolamines was decreased, while the incorporation of radioactivity into the sphingomyelins and lysophosphatidyl cholines was significantly activated.     

Incorporation of [1-14C]Palmitic Acid into Neutral Lipids and Phospholipids of Rat Cerebral Cortex In Vitro

Abstract: Incorporation of [1-¹⁴C]palmitic acid into neutral lipids and phospholipids of rat cerebral cortex was examined in vitro in normal Krebs-Ringer bicarbonate buffer containing 3% (wthol) albumin and 0.75 mM palmitic acid. Under standard assay conditions, radioactivity in the triacylglycerol fraction increased rapidly during the first 30 min, and then decreased after 60 min, with corresponding increase in radioactivity in phosphatidyl choline, phosphatidyl ethanolamine, and a fraction of phosphatidyl inositol plus phosphatidyl serine. Diacylglycerol was shown to be an intermediate metabolite. Radioactivity increased in triacylglycerol, and decreased in phosphatidyl choline and phosphatidyl ethanolamine throughout incubation under NZ gas. In the fraction of phosphatidyl inositol plus phosphatidyl serine, radioactivity decreased after 30 min during incubation under N, gas. A possible acylation-deacylation cycle, in which triacylglycerol could be a source of free fatty acids for phospholipids, is discussed.     

Lipid utilization in adult Pieris brassicae with special reference to the rôle of linolenic acid

Fatty acids of adult Pieris brassicae and the incorporation of dietary linolenic acid-1-¹⁴C into adult (and egg) lipids were analysed 1 and 9 days after ecdysis. Females grown on a leaf diet retained palmitic, palmitoleic, and oleic acids but lost linoleic and linolenic acids during adult life, while males utilized their fatty acids more evenly. On an artificial diet both sexes retained palmitic acid but utilized palmitoleic and oleic acids. In both cases females laid eggs with a high palmitic and oleic acid content.Analysis of thorax flight muscles (artificial diet) revealed that 67·9% of the lipids in 1-day females and 83·6% in 9-day females was phospholipid (PL). During adult life linolenic acid increased in thorax neutral lipids (NL) from 14·6 to 20·0% in females and from 18·5 to 30·0% in males. Males incorporated more linolenic acid-1-¹⁴C especially in fat body and flight muscle PL than females. The majority of this was recovered from phosphatidyl cholines (PTC) in 1-day adults whereas in 9-day adults phosphatidyl ethanolamines (PTE) and another compound, most likely cardiolipin, contained more label (29·0% in PTC, 33·1% in PTE, 34·9% in cardiolipin, and 2·0% in sphingomyelin in the thorax of females). The females also incorporated the label into egg lipids (42·2% in PL, 57·8% in NL). There was recovered from PTC 54·5% of the label in egg PL.Most of the label in thorax NL was found to be in free fatty acids (FFA). The label disappeared from triglycerides during adult life and tended to accumulate in FFA (82·7% in 9-day females) while in diglycerides the label did not vary during adult life (17·2% in 9-day post-emergence females). PTC apparently is a fairly labile PL type which is utilized in muscle whereas PTE and cardiolipin may be more structural in function and accumulated more label from linolenic acid with increasing adult age. Linolenic acid, then, essentially is a structural fatty acid and its rôle appears to be mainly in the structures of flight muscle membranes and organelles.     

Lipid biosynthesis in developing mustard seed: formation of triacylglycerols from endogenous and exogenous Fatty acids

Cotyledons of developing mustard (Sinapis alba L.) seed have been found to synthesize lipids containing the common plant fatty acids and very long-chain monounsaturated (icosenoic, erucic, and tetracosenic) and saturated (icosanoic, docosanoic, and tetracosanoic) fatty acids from various radioactive precursors. The in vivo pattern of labeling of acyl lipids, either from fatty acids synthesized `endogenously' from radioactive acetate or malonate, or from radioactive fatty acids added `exogenously', indicates the involvement of the following pathways in the biosynthesis of triacylglycerols. Palmitic, stearic, and oleic acid, synthesized in the acyl carrier protein-track, are channeled to the Coenzyme A (CoA)-track and converted to triacylglycerols via the glycerol-3-phosphate pathway. Pools of stearoyl-CoA and oleoyl-CoA are elongated to very long-chain saturated and monounsaturated acyl-CoA, respectively. Most of the very long-chain saturated acyl-CoAs acylate preformed diacylglycerols. Very long-chain monounsaturated acyl-CoAs are converted to triacylglycerols, partly via phosphatidic acids and diacylglycerols, and partly by acylation of preformed diacylglycerols.     

Solvent-free glycerolysis of palm and palm kernel oils catalyzed by commercial 1,3-specific lipase from Humicola lanuginosa and composition of glycerolysis products

Glycerolysis of palm and palm kernel oils were conducted using a commercial 1,3-specific lipase from Humicola lanuginosa (trade name: SP 398) as catalyst (500 units lipase g^–1 oil) at 40°C and oil:glycerol (1:2 mol mol^–1) in a solvent-free system. After 24 h, the glycerolysis products of palm and palm kernel oils consisted of 23% triacylglycerols, 18% monoacylglycerols, 38% diacylglycerols and 18% triacylglycerols, 31% monoacylglycerols, 42% diacylglycerols, respectively. The monoacylglycerol fraction of the glycerolysis product of palm oil was enriched in oleic acid. Palmitic acid content of the monoacylglycerol fraction of the same product was less than that of the original oil. Under the same conditions, monacylglycerol fraction of the palm kernel oil glycerolysis product was enriched in palmitic, stearic and oleic acids.     

Biosynthesis of triacylglycerols containing very long chain monounsaturated acyl moieties in developing seeds

Particulate (15,000g) fractions from developing seeds of honesty (Lunaria annua L.) and mustard (Sinapis alba L.) synthesize radioactive very long chain monounsaturated fatty acids (gadoleic, erucic, and nervonic) from [1-¹⁴C]oleoyl-CoA and malonyl-CoA or from oleoyl-CoA and [2-¹⁴C]malonyl-CoA. The very long chain monounsaturated fatty acids are rapidly channeled to triacylglycerois and other acyl lipids without intermediate accumulation of their CoA thioesters. When [1-¹⁴C]oleoyl-CoA is used as the radioactive substrate, phosphatidylcholines and other phospholipids are most extensively radiolabeled by oleoyl moieties rather than by very long chain monounsaturated acyl moieties. When [2-¹⁴C]malonyl-CoA is used as the radioactive substrate, no radioactive oleic acid is formed and the newly synthesized very long chain monounsaturated fatty acids are extensively incorporated into phosphatidylcholines and other phospholipids as well as triacylglycerols. The pattern of labeling of the key intermediates of the Kennedy pathway, e.g. lysophosphatidic acids, phosphatidic acids, and diacylglycerols by the newly synthesized very long chain monounsaturated fatty acids is consistent with the operation of this pathway in the biosynthesis of triacylglycerols.     

Diaglycerol biosynthesis in everted sacs of rat intestinal mucosa.

The molecular specificity in the biosynthesis of diacylglycerols by rat intestinal mucosa was examined by means of radioactive markers, thin-layer chromatography with silver nitrate and gas-liquid chromatography with radioactivity monitoring. Bile salt micelles of alternately labeled monoacyglycerols and free fatty acids were incubated with everted sacs of intestinal mucosa for various periods of time and the diacylglycerols were isolated by solvent extraction and thin-layer chromatography. Sterospecific analyses of the X-1,2-diacylglycerols labeled from 2-monoacylglycerols showed that the sn-1,2-isomers (45-55%) were slightly in excess of the sn-2,3-isomers (34-45%) with the X-1,3-diacylglycerols accounting for the rest of the radioactivity (5-10%). This suggests that racemic diacylglycerols may be intermediates in the resynthesis of dietary fat in rat intestinal mucosa. Detailed analyses of the molecular species of the sn-1,2-diacylglycerols labeled from free fatty acids revealed that 10-45% of the total did not contain the acid present in the 2-monoacylglycerol supplied, and therefore had originated from the phosphatidic acid pathway. These findings are at variance with those obtained in isolated microsomes, which have suggested an inhibition of the phosphatidic acid pathway by monoacylglycerols as well as have given evidence of an exclusive synthesis of sn-1,2-diacylglycerols from 2-monoacylglycerols.     

Effect of sex and bezafibrate on incorporation of blood borne palmitate into lipids of rat liver nuclei

The aim of the present study was to investigate whether lipid metabolism in the nuclei is affected by changes in the metabolism of free fatty acids in the liver. The experiments were carried out on 3 groups of rats: 1 - control-male, 2 - female, and 3 - male, treated with bezafibrate (a peroxisome proliferator). The rats received ¹⁴C-palmitic acid intravenously. Thirty min later liver samples and blood from the abdominal aorta were taken. The liver nuclei were isolated in sucrose gradient. Lipids were extracted from the nuclei and the liver homogenate and subsequently separated into the following fractions: phospholipids, mono-, di- and triacylglycerols, free fatty acids, cholesterol and cholesterol esters. The radioactivity of each fraction was counted. Furthermore, the content of free fatty acids and the fatty acid binding proteins was measured. It was found that radioactivity was present in each lipid fraction obtained from the liver homogenate and from the nuclei. In the female group, the total radioactivity of lipids in the liver homogenate was lower, whereas in the nuclei it was higher in comparison to the male group. The reduction in the radioactivity in the liver was mostly accounted for by decreased radioactivity in the fraction of triacylglycerols and phospholipids. In the nuclei, the radioactivity of the fraction of phospholipids, free fatty acids and diacylglycerols was elevated. Bezafibrate did not affect the total radioactivity of lipids in the liver and reduced it in the nuclei. In the liver, the drug increased radioactivity mostly in the fraction of phospholipids and reduced it mainly in the fraction of triacylglycerols. In the nuclei, the radioactivity of each lipid fraction examined was reduced. The content of the fraction of free fatty acids in the liver and in the nuclei in the female and in the bezafibrate-treated groups did not differ from the respective value in the control group. The content of fatty acid binding proteins in the nuclei of the female and bezafibrate-treated groups increased in parallel to the elevation in their content in the cytosol. It is concluded that the female sex hormones and bezafibrate influence the transport of selected lipids into the nuclei. The effects seem to be a consequence of the action of these factors directly on the nucleus.     

DIFFERENTIAL DISTRIBUTION OF [U-14C]GLUCOSE AND [U-14C]GLYCEROL AMONG MOLECULAR SPECIES OF PHOSPHATIDYL CHOLINE,PHOSPHATIDYL ETHANOLAMINE AND 1,2-DIACYLGLYCEROL IN RABBIT BRAIN12

Specific radioactivities of molecular species of phosphatidyl choline(PC), phosphatidyl ethanolamine(PE) and 1,2-diacylglycerol were determined in rabbit brain 15 and 30 min after intraventricular injection of 10OpCi of either [U-¹⁴C]glucose or [U-¹⁴C]glycerol. The rate of de nouo synthesis of glycerophospholipids and their molecular species could be determined after glycerol labelling, since 94.0–99.7% of ¹⁴C activity was recovered in glyceryl moieties of brain lipids. After injection of glucose radioactivity was measured in both glyccrol and acyl residues of lipids. High incorporation rates were measured in species of PC, PE and 1,2-diacylglycerol with oleic acid in position 2 and with palmitic, stearic or oleic acids in position 1. The conclusion may therefore be drawn that these molecular species were preferably synthesized de novo by selective acylation of glycerol 3-phosphate. The lowest specific activities were observed for 1,2-dipalmitoyl- and l-stearoyl-2- arachidonoyl-glycerol, -PC and -PE. These turnover rates point to incorporation of arachidonate, and probably also of palmitate in dipalmitoyl-PC, amounting to 20% of total PC, via deacylation-acylation- cycle.     

The distribution of 14C-arachidonic acid in hamster lungs is sensitive to quinacrine

Isolated hamster lungs were labelled with 14C-arachidonic acid. When the lungs were ventilated with a respirator only a small amount of radioactivity was released to the perfusion effluent. This release was not changed significantly by pulmonary infusion of quinacrine (0.5 mM), a known inhibitor of phospholipase A2. After the perfusion about 75% of the radioactivity in the lungs was in phospholipids, mainly in phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol and to a lesser degree in phosphatidylserine and phosphatidic acid. About one fourth of the radioactivity was in neutral lipids (tri- and diacylglycerols) and as free unmetabolized 14C-arachidonic acid. Pulmonary infusion of quinacrine increased the amount of radioactivity in diacylglycerols and phosphatidylinositol but had no effect on that in phosphatidylcholine, phosphatidylserine, phosphatidic acid and triacylglycerols. The amount of radioactivity in phosphatidylethanolamine was decreased by quinacrine and increased in the vicinity of an unidentified phospholipid-quinacrine complex. The present study indicates that the distribution of 14C-arachidonic acid in hamster lung lipids is sensitive to quinacrine. The detected changes can, however, not be explained by an overall inhibition of phospholipase A2 activities.     

Biochemistry of development in insects. Incorporation of fatty acids into different lipid classes.

1. To study the different metabolic behaviour of various stages of development of the insect Ceratitis capitata, the incorporation of labelled decanoic, lauric, myristic, palmitic, stearic, oleic, and linoleic acids into triacylglycerols by insect homogenates was investigated. The time-course of incorporation of labelled fatty acids was firstly studied by using oleic acid; it showed that after 10 min of incubation the levels of radioactivity incorporated into triacylglycerols and those remaining in the free fatty acids were practically unchanged. 2. All labelled fatty acids were efficiently incorporated by larval homogenates; however, most of the radioactivity remained as free fatty acids in the presence of pharate adult homogenates, palmitic, and stearic acids being the most scarcely incorporated by this stage of development of the insect. 3. Plots of triacylglycerol and free fatty acid radioactivites versus the stage of development defined a crossing-zone in coincidence with the larval-pupal apolysis. This metabolic difference between larval and pharate adult homogenates could not be explained through differences in the acyl-CoA synthetase activity of the insect; this enzyme activity was notably higher in pharate adult homogenates than in the larval homogenates whatever would be the nature of the fatty acid. 4. [14C]Triolein was scarcely hydrolyzed by both larval and pharate adult homogenates. 5. Double-label experiments were carried out by incorporating either [3H]oleic acid or [3H]-palmitic acid and [14C]glycerol 3-phosphate by larval and pharate adult homogenates at different incubation intervals. Diacylglycerols, triacylglycerols, and phosphoglycerides were isolated and the 14C/3H molar ratio calculated. Results suggest the existence of a different acyltransferase activity in the different stages of development of the insect.     

THE METABOLIC TURNOVER OF MOLECULAR SPECIES OF PHOSPHATIDYLINOSITOL and ITS PRECURSOR PHOSPHATIDIC ACID IN GUINEA-PIG CEREBRAL HEMISPHERES

Abstract– The molecular species composition of phosphatidylinositol from guinea-pig cerebral hemispheres was studied and found similar to that of phosphatidylinositol from ox cerebral hemispheres. In both cases the tetraenoic species was predominant. Phosphatidic acid from guinea-pig cerebral hemispheres contained two major molecular species; the monoenoic and hexaenoic (33.4 and 24 mol/100 mol respectively). In order to study the metabolism of molecular species of phosphatidic acid and phosphatidylinositol in the cerebral hemispheres, guinea-pigs were injected intracisternally with ³²P_i and [U-¹⁴C]glucose. After 5 min of isotopic exchange, the specific radioactivity of ³²P in phosphatidylinositol was nearly equal to that in phosphatidic acid, whereas specific radioactivity of ¹⁴C in the glycerol was 1.4 times and in the fatty acids nearly 0.5 times that in the phosphatidic acid respectively, indicating metabolic heterogeneity of both phospholipids. The glycerol specific radioactivity was different in all the molecular species of phosphatidic acid being greatest in the monoenoic and least in the tetranenoic species. When the molecular species were arranged in this way, the order was representative of their relative rates of synthesis by acylation of glycerol-3-phosphate. An almost opposite order was obtained when the molecular species were arranged according to their phosphate/glycerol radioactivity ratios, indicating the relative contribution of the diacylglycerol kinase pathway to their formation. When the specific radioactivity values and ratios of phosphatidylinositol were similarly considered, the orders of the molecular species were, on the whole, similar to that of phosphatidic acid. This indicated that synthesis de novo (Paulus & Kennedy , 1960) was operative in the formation of most of its molecular species, but due to other considerations it was concluded that part of the tetraenoic, and probably the whole of saturated phosphatidylinositol may be formed by transacylation reactions. The results are discussed in terms of the experimental limitations of previous and present techniques for the analysis of phospholipid molecular species.     

Biosynthesis of triacylglycerols by rat intestinal mucosa in vivo.

The structure of mucosal triacylglycerols was studied in rat intestinal mucosa in vivo during the absorption of a low molecular weight fraction of butter oil and of the corresponding free fatty acids of medium and long chain length. The mucosal lipids were isolated by solvent extraction and the acylglycerol structures were determined by combined AgNO3- thin-layer chromatography and gas-liquid chromatography techniques and stereospecific analysis. Evidence was obtained for a rapid biosynthesis of triacylglycerols from diacylglycerols arising from the operation of both the monoacylglycerol and the phosphatidic acid biosynthetic pathways. Both sn-1,2- and sn-2,3-diacylglycerols appeared to be converted to triacylglycerols at significant rates, but a preferential utilization of sn-1,2-diacylglycerols could not be excluded. Endogenous dilution varied from a miniumum of 5% during triacylglycerol biosynthesis from monoacylglycerols to 15% during their synthesis from free fatty acids, and was characterized by a preferential placement of the endogenous acids in the sn-3 and 2 positions of the triacylglycerol molecules. Exogenous myristic acid was preferentially associated with the sn-3 position, and stearic acid became preferentially bound to the sn-1 position. The complexity of the triacylglycerol end products prevented an exact estimate of the contribution of the phosphatidic acid pathway, but the acylglycerol structures were compatible with a minimum of 20% of total triacylglycerol yield at all times.