The phospholipid composition of pig lung surfactant

Surface active material (surfactant), which contained 80% lipid material, was isolated from pig lungs by the saline perfusion procedure. The lipids were analyzed by column, thin layer and paper chromatography, followed by gas liquid chromatography to determine the fatty acid composition of the major phospholipid fractions. It was found that this pig lung surfactant contained the phospholipid constituents known to be present in mammalian tissues and in addition, phosphatidylglycerol (1.5%) and lyso-bisphosphatidic acid (2.0%) were also found. These two components occurred in higher proportions in the surfactant lipids than in the lipids of the whole lung tissues.     

The phospholipid composition of plasma in various mammalian species

Plasma phospholipids in several common mammalian species, including cat, cow, dog, goat, guinea pig, horse, pig, rabbit, rat, and sheep, were analyzed by using chromatographic and spectrophotometric methods. Lipids were extracted from plasma with chloroform-methanol 2∶1 (v/v) and freed of nonlipid material by passage through a Sephadex column. The phospholipids were separated by two-dimensional thin-layer chromatography (TLC). Spots were identified by spray reagents, also by infrared spectrophotometry. The relative distribution of the phospholipids was determined by phosphorus analysis on the spot scraped off the TLC plate. Lecithin, lysolecithin, and sphingomyelin were found in the plasma of all species and accounted for more than 95% of the phospholipids except in the rodents. Lecithin was without exception the major phospholipid in plasma (56 to 83%). Lysolecithin and sphingomyelin content varied between 8 and 23% and 6 and 15% respectively. Phosphatidyl ethanolamine and phosphatidyl inositol were the only noncholine-containing phospholipids detected (detection limits 0.2%) in the plasma of these species. Together these compounds usually made up less than 5% of the total phospholipid. Rodents were an exception, especially the guinea pig, which had 21.7% phosphatidyl ethanolamine.     

Progressive gross changes in renal medullary composition in pregnant rats

The biochemical composition of the renal medulla and the blood pressures of pregnant rats and nonpregnant controls were compared on days 15, 18, 20, and 22 of pregnancy. Deoxyribonucleic acid and protein content of the renal medulla changed together with the tissue weight (wet weight) during gestation except on day 20 when a slight increase in protein was observed. However, the glycerophospholipids (phosphatidyl ethanolamine, phosphatidyl choline, and phosphatidyl serine) and sphingomyelin showed significant increases throughout the latter stages of pregnancy with the maxima in most cases occurring on day 20 of the gestation period. Medullary hypertrophy and increased lipid content per cell coexist with the decrease in blood pressure and may indicate peak activity in production of antihypertensive renal lipids. Authorized for publication on 3/17/76 as paper No. 5040 in journal series of Pa. Ag. Exp. Station. Submitted to the Graduate School of The Pennsylvania State University in partial fulfillment of the requirements for the Ph.D.     

Phosphatidylethanolamine metabolism in rats fed a low methionine,choline-deficient diet

The metabolism of phosphatidylethanolamine (PE) was studied in male rats fed a low methionine diet for 7 days with or without supplemental choline. Groups of animals were injected with 2-¹⁴C-ethanolamine and killed at intervals thereafter up to 72 hr. Liver phospholipids were isolated, and PE and phosphatidylcholine (PC) were separated by argentation chromatography into diene (18∶2), tetraene (20∶4) and hexaene (22∶6) fractions. Fatty acid composition and the distribution of radioactivity and specific activity in the total phospholipids and in the fractions were determined. Choline deficiency did not affect total liver phospholipid, but it did increase the amount of PE and decreased that of PC. The major effect of the deficiency on phospholipid fatty acids was to decrease the proportion of PE arachidonate and to increase that of docosahexaenoate. Ethanolamine incorporation into liver PE of deficient rats was only slightly less than in the controls, but loss of the radioactivity from the PE was slower. Ethanolamine radioactivity appearing in the PC of deficient rats was about half that of the controls, even though specific activities of the PE precursors were similar to the control rats. Choline deficiency increased the biological half-lives of the total PE and its fractions. Although the proportion of PE tetraenoic fraction was reduced, the total amount of this liver PE fraction in deficient rats was not affected. However the amount of hexaenoic fraction was doubled, and it accounted for most of the increased quantity of liver PE seen in deficient animals. The results suggested that in choline deficiency PE synthesis was delayed but not appreciably suppressed, and that limited availability of methionine for methylating the PE fractions in their proper proportions affected the concentrations of the PE fractions and impaired their normal conversion to PC. Presented in part at the AOCS Meeting, Houston, May 1971.     

Compartmental study of rat renal phospholipid metabolism

Phospholipid content and metabolism were studied in rat renal papillary, medullary and cortical slices. The highest concentration of phospholipids was found in cortex and the lowest in papilla samples (ratio cortex/medulla, 1.3; cortex/papilla, 3.7). The profile of the various phospholipids was different depending on the zone. The most important difference was the relative concentrations of sphingomyelin (CerPCho) and phosphatidylinositol (PtdIns) with ratios for PtdIns/CerPCho of 5.0, 3.3 and 2.5 in papilla, medulla, and cortex, respectively. In the three zones, PtdIns showed the highest specific activity for [2-¹⁴C]glycerol and [1-¹⁴C]arachidonic acid incorporation. By contrast, a higher amount of [1-¹⁴C]palmitic acid was incorporated into phosphatidylcholine than into any other phospholipid. The various radioactive precursors were only poorly incorporated into phosphatidylethanolamine. No radioactivity was associated with phosphatidylserine. The papilla possesses the most active phospholipid metabolism of all the pathways studied.     

Lipid profiles of plasma lipoproteins of fasted and fed normal and choline-deficient rats

Three major density classes of lipoproteins and a residual protein (d>1.21) were isolated by ultracentrifugation from plasma of fasted, fed normal, and choline-deficient rats. Lipid extracts were obtained from total plasma and the various density classes of lipoproteins, and each extract was examined in detail by thin layer and gas chromatographies. The results indicated essentially identical compositions of molecular species of phosphatidyl choline, which suggested their rapid equilibration among the different plasma lipoprotein classes. In contrast, the molecular species of the triacylglycerols and cholesteryl esters showed significant differences among the chylomicrons, very low and low, and high density lipoproteins, which excluded the possibility of their ready equilibration in vivo. Omission of choline from diet resulted in a sharp and statistically significant decrease in all lipid components of the very low and low density lipoproteins within 2 days. After 10 days of choline deficiency, the lipid levels of chylomicrons and very low and low density lipoproteins were ca. one-half the levels found in the choline supplemented animals, and there were discernible distortions in their lipid composition. Reintroduction of choline led to a prompt return to normal levels and lipid composition of both chylomicron and very low and low density lipoprotein fractions. The lack of equilibration of the triacylglycerols among the lipoprotein classes under normal conditions and in choline deficiency demonstrates an as yet unrecognized source of compartmentation of plasma lipids.     

The influence of n-6 and n-3 fatty acids on kidney phospholipid composition and on eicosanoid production in aging rats

Changes in eicosanoid production may contribute to some of the complications of the aging process such as atherosclerosis and glomerular sclerosis. Polyunsaturated fatty acids of the n-6 and n-3 series are precursors of eicosanoids. We fed diets containing safflower oil as a source of n-6 fatty acids, fish oil as a source of n-3 fatty acids or beef tallow as a source of saturated fats to three groups of normal rats from 2–18 months of age. We demonstrated incorporation of the n-3 fatty acids, 20:5n-3 and 22:6n-3 into kidney phospholipids. Feeding of the diet containing n-3 fatty acids was associated with a markedly decreased giomerular production of PGE, 6-keto-PGF_1α and TXB₂. It also decreased the aortic production of 6-keto-PGF_1α and platelet production of TXB₂. No significant effect of n-6 fatty acids on dienoic eicosanoid production was observed. There were no adverse effects on kidney function as measured by urinary protein excretion and serum creatinine levels or on renal morphology by any diet. A diet enriched in n-3 fatty acids for 18 months remains effective in decreasing dienoic eicosanoids in the aging rat.     

Liver phospholipids of rats fed a choline-deficient diet supplemented with choline or methionine

The low amount of arachidonic acid in the total phospholipids in the liver of rats fed a standard type of choline-deficient diet was corrected by either choline or methionine, which also increased food intake. Choline increased the content of this fatty acid in the phosphatidyl ethanolamine but not in the phosphatidyl choline. Methionine increased both the amount of phosphatidyl choline and its content of arachidonic acid.     

Dietary fiber supplements: Effects on serum and liver lipids and on liver phospholipid composition in rats

Rats (6 per group) were fed semipurified diets containing either particulate fibers (alfalfa, 10%; cellulose, 10%; bran, 10%), a soluble ionic fiber (pectin 5%), soluble, nonionic fibers (guar gum, 5%; Metamucil, 10%), a mixed fiber preparation (Fibyrax, 10%, or an insoluble, ionic bile acid-binding resin (cholestyramine, 2%). The control group was fed the unsupplemented diet. The feeding period, during which diet and water were provided ad libitum, was 28 days. Compared with the control group, serum total cholesterol levels were increased by more than 10% in rats fed alfalfa and decreased by more than 10% in rats fed cellulose, guar gum, Fibyrax and cholestyramine. There were no significant differences in percentage of plasma HDL cholesterol. Serum triglycerides were elevated in the groups fed alfalfa, pectin, guar gum or Fibyrax and reduced in the group fed Metamucil. Plasma phospholipids were elevated in rats fed alfalfa or bran, unaffected in rats fed pectin or Metamucil and reduced in the other groups. Liver total cholesterol was elevated in all groups but those fed wheat bran and cholestyramine. The percentage of liver cholesterol present as ester was elevated in every group except that fed cholestyramine. Liver triglycerides were reduced in rats fed guar gum or Metamucil and elevated in those fed alfalfa. Liver phospho-lipids were lowered in the group fed cellulose. Liver phospholipids were fractionated by thin layer chromatography to give phosphatidylcholine (PC), phosphatidylethanolamine (PE), sphingomyelin (Sph), lysophosphatidylcholine (LPC) and phosphatidylinositol plus phosphatidylserine (PI+PS). PC was elevated in all test groups (7–25%); PE levels ranged from 14% below to 0.3% above controls; Sph levels were sharply lower (20–53%) in all groups. LPC and PI+PS levels were close to the control value in all test groups. The results demonstrate that different dietary fibers can affect liver phospholipid composition. In view of the critical roles of phospholipids in many biological reactions, it will be interesting to survey the influence of dietary fiber on phospholipid spectra of other tissues. Deceased.     

Activity of fatty acyl CoA-lysophospholipid acyltransferases in liver microsomes of rats fed a choline-deficient diet

Feeding a choline-deficient diet to rats has no effect on the activity of fatty acyl CoA-lysophospholipid acyltransferases, even though liver microsomes are severely depleted of lecithins. Thus lecithins appear to have no functional role in the activity of these transferases. It can be excluded that the latter enzymes are involved in the production of the changes in the fatty acid composition of liver phospholipids occurring in choline-deficient rats. These changes result most likely from alterations in the biosynthesis of liver lecithins and cephalins.     

Dietary phospholipid alters biliary lipid composition in formula-fed piglets

Plasma cholesterol, arachidonic acid (AA, 20∶4n−6), and docosahexaenoic acid (DHA, 22∶6n−3) are higher in breast-fed infants than in infants fed formula without cholesterol, AA, or DHA. This study investigated differences in plasma, hepatic, and bile lipids and phospholipid fatty acids, and expression of hepatic proteins involved in sterol metabolism that result from feeding formula with cholesterol with egg phospholipid to provide AA and DHA. For this study, three groups of piglets were evaluated: piglets fed formula with 0.65 mmol/L cholesterol, the same formula with 0.8% AA and 0.2% DHA from egg phospholipid, and piglets fed sow milk. Piglets fed the formula with phospholipid AA and DHA had higher plasma high density lipoprotein, but not apoprotein (apo) B cholesterol or triglyceride; higher bile acid and phospholipid concentrations in bile; and higher liver and bile phospholipid AA and DHA than piglets fed formula without AA and DHA (P<0.05). Hydroxy methylglutaryl (HMG)-CoA reductase and 7-α-hydroxylase, the rate-limiting enzymes of cholesterol and bile acid synthesis, respectively, and low density lipoprotein receptor mRNA levels were not different between piglets fed formula without and with phospholipid AA and DHA, but HMG-CoA reductase and 7α-hydroxylase mRNA were higher, and plasma apo B containing lipoprotein cholesterol was lower in all piglets fed formula than in piglets fed milk. These studies show that supplementing formula with AA and DHA from egg phospholipid alters bile metabolism by increasing the bile AA and DHA, and bile acid and phospholipid.     

Tissue phospholipid fatty acid composition in the diabetic rat

Tissue phospholipid fatty acid compositions in streptozotocin-induced diabetic rats were studied. The major changes in liver, plasma, erythrocyte and heart were increased proportions of linoleic and dihomo-γ-linolenic acids and a decreased proportion of aracchidonic acid. The latter was not significantly changed in phospholipids of kidney, adrenal gland and testis. Skin fatty acids in diabetic rats showed an increase in the proportion of arachidonic acid and a reduction in the proportion of linoleic acid. The fatty acid desaturating activity in diabetes may be regulated differently in different tissues.     

Altered microsomal phospholipid composition in the streptozotocin diabetic rat

Streptozotocin diabetes in the rat alters liver microsomal membrane fatty acid composition. The present study was undertaken to determine if such changes in fatty acid composition were due to changes in the amount of individual phosphoglycerides or to disproportionate changes in fatty acid composition in any of the individual phosphoglycerides. The diabetic animals showed a small increase in total microsomal phospholipid, which is due to a selective increase in the phosphatidylethanolamine fraction. The changes in fatty acid composition in the total lipid extract (decreased palmitoleic, oleic and arachidonic acids and increased linoleic and docosahexaenoic acids) from the diabetic animals were present in both the major phosphoglycerides, phosphatidylcholine and phosphatidylethanolamine, with very little change in fatty acid composition in the phosphatidylserine and inositol fraction. Further studies are necessary to delineate the cause of the abnormal membrane phospholipid composition in the diabetic animal. Abbreviations: The abbreviated fatty acid nomenclature refers to the number of carbon atoms in the chain, the number of unsaturated bonds, and the position of the first unsaturated bond counting from the terminal methyl group; thus arachidonic acid, 5,8,11,14-eicosatetraenoic acid, is 20∶4ω6.     

The effect of diet on the phospholipid composition of the red blood cells of man

Short term (16 day) controlled fat (formula type diet) feeding to 10 healthy adult males led to no detectable change in the total amt or the relative proportions of the individual phospholipids of the red blood cells, although limited changes did occur in the fatty acids of certain of the phospholipids. The total phospholipid content of the red blood cells was 315±10 mg/100 ml (average of 20 samples). Lecithin accounted for 34% of the total, with sphingomyelin, phosphatidyl ethanolamine and phosphatidyl serine representing 25, 25 and 16%, respectively. Approx 36% of the phosphatidyl ethanolamine, 4% of the phosphatidyl serine and 6% of the lecithin was present in the plasmalogen form. Each phospholipid class was found to have a distinctive fatty acid spectrum. The M ratio of saturated to unsaturated fatty acids in all three phosphoglycerides was nearly 1:1. Behenic, lignoceric and nervonic acids made up almost half of the sphingomyelin fatty acids, and the M ratio of saturated to unsaturated fatty acids in this lipid was 3:1. When compared with red cells from subjects consuming a diet with a high butter fat content, red cells from subjects on a diet rich in corn oil were found to contain higher levels of linoleic acid in the lecithin and phosphatidyl serine fractions, and lower levels of oleic acid in the lecithin fraction. No changes were observed in the fatty acids of the phosphatidyl ethanolamine and sphingomyelin fractions. It is probable that these alterations represent the result of highly specific exchanges with plasma fatty acids, and it is suggested that three levels of specificity are involved: class of phospholipid, type of fatty acid, and specific fatty acid.     

The effect of temperature on the fatty acid and phospholipid composition ofCephalosporium falciforme andCephalosporium kiliense

The effect of temperature on the lipid composition ofCephalosporium falciforme andCephalosporium kiliense, causative agents of maduromycosis, was investigated. The fungi were grown at 28.5 C and 37 C in a chemically defined medium. The lipids were solvent extracted, purified on Sephadex, and separated into their component classes by silicic acid column chromatography. Five lipid classes were found: (a) sterol esters, (b) triacylglycerides, (c) free fatty acids, (d) sterols, and (e) phospholipids. Fatty acids were analyzed by gas liquid chromatography and phospholipids by thin layer chromatography. Temperature induced changes of varying degrees occurred in both the fatty acid phospholipid fractions of each organism.     

Ethanolamine kinase activity and compositions of diacylglycerols,phosphatidylcholines and phosphatidylethanolamines in livers of choline-deficient rats

These experiments were performed to find the reasons for the increased concentrations of docosahexaenoyl phosphatidylethanolamines (PE) in livers of choline-deficient rats. We measured the activity of ethanolamine kinase, which catalyzes the first step in PE formation. We also measured the compositions of PE and phosphatidylcholines (PC) and concentrations and fatty acid compositions of diacylglycerols (DG), which are precursors of PE. Young male rats were fed for one week a low-methionine, choline-deficient diet, or the same diet supplemented with choline. Ethanolamine kinase activity was measured in liver cytosol (100,000 g supernatant). Fatty acids were measured in total liver diacylglycerols and in microsomal PE and PC. Ethanolamine kinase activities were equal in choline-deficient and choline-supplemented rats. Concentrations of DG were elevated 6-fold by choline deficiency. The percentage of docosahexaenoic acid (22∶6n−3) in microsomal PE was nearly doubled by choline deficiency. Although the increased concentrations of PE in choline-deficient livers cannot be attributed to increased activity of ethanolamine kinase, the rate of PE formation probably was increased by increases in concentrations of its precursors, including DG. The disproportionate increase in 22∶6n−3 PE probably was caused by a selective formation of PE from DG that contain 22∶6n−3.     

The phospholipid composition of human placenta,endometrium and amniotic fluid: A comparative study

The contents of lecithin in human lung, liver, spleen, kidney, heart muscle, skeletal muscle, placenta, endometrium, amniotic flud, and brain gray and white matter are 52.1–54.2%, 44.2–45.8%, 42.6–44.5%, 35.1–37.2%, 42.5–43.0%, 48.0–50.0%, 45.5–46.4%, 42.0–43.0%, 65.0–66.0%, 23.9–24.3% and 24.7–24.9% respectively in total phospholipids. The level of diphosphatidyl glycerol is much lower in placenta, lung and spleen, and almost absent in amniotic fluid and brain; the heart muscle has 8.6%. The content of phosphatidyl serine is high in endometrium and brain. The content of sphingomyelin is high in lung, spleen, kidney and brain as compared to liver, heart muscle, skeletal muscle and endometrium. It is suggested that the phospholipid composition of various human tissues is characteristic for that class.     

Effect of tumor necrosis factor-α on triglyceride and phospholipid content and fatty acid composition of liver and carcass in rats

We have previously shown that a continuous infusion of tumor necrosis factor-α (TNF-α) in rats results in an increase in plasma triglyceride (TG), liver protein and DNA, and at the same time a reduction in muscle protein. However, there is no information on the associated effects of TNF-α on liver and muscle lipids. The present study, therefore, determined the effect of TNF-α on the TG and phospholipid (PL) content and their fatty acid composition, in the liver and carcass of rats and correlated with the plasma levels of insulin, corticosterone, and catecholamaines. Total parenteral nutrition that met the daily nutrient requirements was continuously infused for 10 d through a central vein catheter in two groups of rats, one receiving infused TNF-α (100 μg/kg/d) and a control group. Hepatic TG and PL, expressed either as mg/g dry weight or total organ content, were significantly increased in the TNF-α-infused group compared with controls. Livers of TNF-α infused rats contained significantly less saturated and monounsaturated fatty acids and significantly more polyunsaturated fatty acids (PUFA) of the ω3 and ω6 series than controls. The carcass, on the other hand, contained significantly more monounsaturated and significantly less polyunsaturated fatty acids of the ω6 series than controls. There were no changes in corticosterone level. Although plasma glucose levels were comparable between control and TNF-α infused rats, the latter had a marked increase in insulin levels, demonstrating insulin resistance. In addition TNF-α infused rats had raised norepinephrine levels. The fall in carcass and rise in liver TG and PL are consistent with the presence of insulin resistance and raised norepinephrine levels. However, the increase in PUFA content of the liver, especially the increase in 20∶4ω6 levels in liver PL and TG, is likely to be the effect of TNF-α and suggests increased activity of Δ5-desaturase.