Distribution and fatty acid composition of phosphoglycerides in normal human brain

A thin-layer chromatographic procedure for the isolation of tissue phospholipids and their subsequent analysis is described. The method has been applied to the determination of the fatty acids of phosphoglycerides in human brain from the early fetal stage to old age. The study shows changes in the distribution and fatty acid composition of each phosphoglyceride in normal brain, although they are quite small after early childhood. A lipid-specific fatty acid pattern for each of the four major phosphoglycerides was found. Besides this, the pronounced differences between fatty acids of the lipids from the cerebral cortex and from the adjacent white matter justify speaking of a tissue-specific fatty acid pattern for brain phosphoglycerides. The phospholipids of cerebral white matter contained more monoenoic acid but much less polyunsaturated fatty acid than those of cerebral cortex. The brain phosphoglycerides also showed an age-dependent fatty acid pattern. With increasing age the concentration of the fatty acids of the linoleate family diminished while that of the linolenate family increased. Brain inositol phosphoglycerides, the fatty acid composition of which has not been studied systematically before, were characterized by a large concentration of arachidonate which was nearly as high for white as for gray matter and showed only small changes with age.     

Fatty acid composition of cholesteryl esters of human meibomian gland secretions

Very long chain cholesteryl esters (CE) are a major group of lipids found in meibomian gland secretions (MGS, also called meibum). MGS are produced by the meibomian glands of human and animal eyelids. They are a critical part of the tear film which covers the exposed ocular surface and serves various physiological roles. The composition of CE of MGS is complex, and still remains poorly understood. Here, a liquid chromatography-ion trap mass spectrometry (LC-MS) procedure developed to analyze CE is described, and a detailed composition of human meibomian CE is reported.MGS were collected from donors, analyzed without any modifications by LC-MS in positive and negative ion modes (PIM and NIM), and quantified using lipid standards where available.CE comprised about 30% of human meibum by mass. More than 40 individual CE species were found and characterized. In PIM, CE were observed as spontaneously in-source generated product ions m/z 369. The signals of the proton adducts of intact CE (M+H)⁺ were of very low intensity. In NIM, all tested CE spontaneously fragmented in-source producing signals of their respective FA. By combining the LC and MS information, the most abundant CE were found to be based on FA ranging from C₁₆ to at least C₃₂ in the following order C_26:0 > C_25:0 > C_24:0 > C_27:0 > C_24:1 = C_18:1 = C_20:0 > other CE.We conclude that the FA composition of CE can be successfully established in LC-MS experiments conducted in NIM. Meibomian CE have a large presence of both saturated and unsaturated FA with an average molar ratio of 4 to 1, respectively.     

Concentration and fatty acid composition of cerebrosides and sulfatides in mature and immature human brain

The fatty acid composition of cerebrosides and sulfatides from frontal lobe gray and white matter was determined for five fresh and four formalinized adult brains and for eight infants. Fatty acid patterns were unaffected by formalinization, but varied considerably from one another in the proportion of saturated to unsaturated fatty acids. The percentages of 24:0 and 24:1 increased with age. Cerebrosides obtained from areas such as the brainstem and cerebellum, where myelination was more advanced, tended to have a larger proportion of long-chain fatty acids than samples extracted from frontal or parietal lobe white matter. Hydroxy fatty acids showed an adult pattern in all instances in which amounts sufficient for accurate quantification could be isolated. Lipid hexose, cerebroside + sulfatide hexose, and methanoleluted hexose were measured in the brains of 12 infants ranging in age from a 4 month fetus to 2 yr. In the most immature, the majority of lipid hexose was in the form of glycolipids more polar than cerebrosides and sulfatides. These have tentatively been identified as hematosides and globosides. With maturation, cerebrosides and sulfatides increased progressively, but the amounts of the more polar glycolipids remained constant in relation to the total lipid content of tissue.     

Cholesterol esters in developing rat brain: concentration and fatty acid composition

Abstract— The contents and the fatty acid composition of cholesterol esters were analysed in developing rat brain. The total content did not exceed 20 μg/brain throughout development. Elimination of serum by adequate perfusion was essential for accurate results. Two separate events appeared to affect the levels of cholesterol esters in developing rat brain, one probably reflecting general developmental changes and the other apparently related to myelination. On either a unit weight or a whole brain basis, the curves appeared to be a superimposition of the two events. There was an underlying developmental change, which was characterized on a unit weight basis by the highest level of cholesterol esters immediately after birth and a steady decline to the adult level by 30 days of age or which on the basis of whole brain was characterized by a steady increase throughout the development. A period of transient increase was superimposed on this underlying developmental change between the ages of 7 and 27 days and corresponded to the period of active myelination. The major fatty acids of rat brain cholesterol esters were palmitic, palmitoleic, oleic and arachidonic acids. Palmitic and palmitoleic acids decreased in proportion while oleic acid increased, as the animal matured. The fatty acid composition of serum cholesterol esters was distinctly different from that of brain cholesterol esters; those from serum contained much higher proportions of linoleic and arachidonic acids and much less palmitoleic and oleic acids.     

Metabolism of ethanol by human brain to fatty acid ethyl esters

Although the most prominent acute and chronic effect of alcohol ingestion in man is alteration of brain function, metabolism of ethanol by human brain has not been documented. This study was designed to detect and localize a new family of nonoxidative ethanol metabolites, fatty acid ethyl esters, in human brain and characterize their synthetic pathways. Fatty acid ethyl ester synthase activity was present in 10 different locations in human brain, with gray matter containing more activity than white matter (0.53 nmol of ethyl oleate/mg of protein/h and 0.25 nmol of ethyl oleate/mg of protein/h, respectively). Two forms of this synthase, present in cytosol or loosely bound to membrane fractions, were isolated from human gray and white matter and then partially purified by ion-exchange chromatography. Both were active at low ethanol concentrations easily attained in vivo in man. Importantly, fatty acid ethyl esters were also detected in brains of individuals dying while intoxicated; only small amounts were present in control subjects at autopsy. Thus, alcohol metabolism in human brain has been documented for the first time by identifying both fatty acid ethyl esters and their synthases in this important target-organ of alcohol abuse.     

Changes in fatty acid composition of human brain myelin lipids during maturation

Abstract— The variation with age of the fatty acid composition of the major lipids in human brain myelin was compared with that of cerebral white matter from the same region. The myelin was isolated from the semiovale centre of the cerebrum of 27 subjects neonatal to old aged. The phospholipid, cholesterol and galactolipid concentrations were determined in all the samples, as were the proportions of the major phospholipid classes. The proportions of cholesterol and especially of the galactolipids increased in myelin during the first 6 months, and in cerebral white matter up to 2 years. During this period the individual phospholipids also varied substantially. Serine phosphoglycerides and especially sphingomyelins increased, and choline phosphoglycerides decreased. The fatty acid patterns of ethanolamine phosphoglycerides (EPG) and sphingomyelins underwent the largest changes. The proportions of saturated fatty acids in EPG diminished rapidly, and there was an increase of monoenoic acids. Fatty acids of the linoleic acid series showed a peak between 4 and 12 months, after which time their proportion slowly diminished to old age. The major fatty acid of this series was docosatetraenoic acid, 22:4 (n-6), which constituted more than 25% of total fatty acids at the maximum level. The fatty acid changes were larger in cerebral white matter, but from 2 years of age the EPG fatty acid pattern in myelin was similar to that in white matter. The fatty acid changes in serine and choline phosphoglycerides of myelin with maturation were much less striking than in EPG but of a similar type. In myelin sphingomyelin the proportion of saturated long-chain fatty acids, C₁₆-C₂₂, diminished, while that of monoenoic acids increased and continued to do so up to old age. From 2 years of age the fatty acid patterns in myelin and cerebral white matter were quite similar. Also the fatty acid patterns of cerebrosides and sulphatides in cerebral white matter and myelin were the same except for the first 2 months of life. The same fatty acid changes occurred in cerebrosides and sulphatides as in the sphingomyelins, i.e. increased proportions of unsaturated (monoenoic) acids. The proportions of 24:1 and 24h:1 and of the odd-numbered fatty acids 25:1 and 23h:1 continued to increase to old age. The variations of the individual lipid fatty acid patterns were small except in the youngest age classes, in which the variations were presumably ascribable to the difficulty in determining the gestational age.     

Fatty acid composition of phospholipids and cholesteryl esters in maternal serum in the early puerperium

The fatty acid composition of serum phospholipids (PL) and cholesteryl esters (CE) in 26 healthy pregnant women at the end of term and 1 and 3 days after delivery was analysed in order to determine whether the maternal serum fatty acid composition changes in the early puerperium. The composition of the saturated fatty acids significantly changes in the PL fraction: 16:0 decreased and 18:0 increased. Both 20:4n-6 and 20:5 n-3 significantly increased after parturition in serum PL while 22:6n-3 remained constant at the three sampling time points. The sum of HUFA was slightly higher 3 days postpartum compared to the prepartum data. The essential fatty acid index significantly increased after delivery. In the CE fraction too differences occurred during puerperium: 18:2n-6 and 20:4n-6 increased and 18:1n-9 decreased after parturition. The sum of the n-3 fatty acids in CE remained unaltered. The EFA index significantly improved both in PL as in CE after delivery.In conclusion, the previously reported changes in the fatty acid composition of PL and CE during normal pregnancy diminish shortly after delivery. In fact, very soon after delivery the maternal fatty acid composition returns to more normal values.     

Steroidal fatty acid esters

Several years ago we discovered an unexpected family of steroidal metabolites, steroidal fatty acid esters. We found that fatty acid esters of 5-ene-3β-hydroxysteroids, pregnenolone and dehydroisoandrosterone are present in the adrenal. Subsequently, others have shown the existence of these non-polar 5-ene-3β-hydroxysteroidal esters in blood, brain and ovaries. Currently, almost every family of steroid hormone is known to occur in esterified form. We have studied the esters of the estrogens and glucocorticoids in some detail, and have found that these two steroidal families are esterified by separate enzymes. In a biosynthetic experiment performed simultaneously with estrodiol and corticosterone, we established that the fatty acid composition of the steroidal esters is quite different. The corticoid is composed predominantly of one fatty acid, oleate, while the estradiol esters are extremely heterogeneous. Our studies have demonstrated that the estrogens are extremely long-lived hormones, that they are protected by the fatty acid from metabolism. They are extremely potent estrogens, with prolonged activity. Esterification appears to be the only form of metabolism that does not deactivate the biological effects of estradiol. We have demonstrated the biosynthesis of fatty acid esters of estriol, monoesters at both C-16 and C-17β. They too are very potent estrogens. These fatty acid esters of the estrogens are the endogenous analogs of estrogen esters, like benzoate, cypionate, etc., which have been used for decades, pharmacologically because of their prolonged therapeutic potency. We have found that the estradiol esters are located predominantly in hydrophobic tissues, such as fat. Sequestered in these tissues, they are an obvious reservoir of estrogenic reserve, requiring only an esterase for activation. To the contrary the biological activity of the fatty acid esters of the glucocorticoid, corticosterone, is not different from that of its free parent steroid. We have shown that the rapid kinetics of its induction of gluconeogenic responses is caused by its labile C-21 ester which is rapidly hydrolyzed by esterase enzymes. While it appears that the physiological role of the estrogen esters may be related to their long-lived hormonal activity, the role of the other families of steroidal esters is not yet apparent. They, and perhaps the estrogen esters as well, must serve other purposes. Indeed they may serve important biological functions beyond those which we ordinarily associate with steroid hormones.     

Incorporation of deuterium-labeled fatty acids into human milk, plasma, and lipoprotein phospholipids and cholesteryl esters

Fatty acid metabolism and the contribution of dietary fatty acids to milk cholesteryl ester (CE) and phospholipid (PL) were investigated in normal lactating mothers. The approach used was to feed mixtures of triglycerides containing deuterium-labeled palmitic acid (16:0-2H2), oleic acid (18:1-2H6), and linoleic acid (18:2-2H4). Milk and plasma samples were collected for 72 hr. Triglyceride (TG), CE, and PL fractions from milk, plasma, and lipoprotein were isolated and analyzed by gas-liquid chromatography and mass spectrometry. Data for the milk CE and PL fractions showed a definite selectivity for incorporation of 16:0-2H2 and 18:1-2H6 relative to 18:2-2H4. Based on the ratios of the deuterated fatty acids incorporated into the milk CE and PL samples, their incorporation times and isotopic enrichment data, it appears that these fatty acids are supplied mainly by the TG derived from chylomicrons and very low density lipoproteins. Plasma and lipoprotein CE data showed a progressive increase in 18:2-2H4 content, with 16:0-2H2 and 18:1-2H4 remaining relatively constant over the collection period. Plasma and lipoprotein PL data showed a higher rate for incorporation of 18:2-2H4 than 16:0-2H2 and 18:1-2H6 over the course of the sampling period. Comparison to previous data from adult males indicates lactation does not have a major effect on the general metabolism of these fatty acids. An increase with time in the isotopic enrichment of 18:2-2H4 in the plasma and lipoprotein CE and PL samples was observed which is consistent with in vitro selectivities reported for lecithin:cholesterol acyltransferase and phosphatidylcholine acyltransferase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Preparation of autoxidized cholesterol mixtures from [4-14C] cholesterol and cholesteryl fatty acid esters

Autoxidation of non-esterified cholesterol, in the solid state, at 100 degrees C, is known to be a relatively slow reaction. The presence of carefully chosen cholesteryl esters considerably increases the ratio of autoxidation. Using this method, mixtures of autoxidized free cholesterol (oxycholesterol) labelled on carbon 4 can be obtained almost quantitatively, in the presence of benzoyl peroxide. The ratio of [4-14C] ester produced by transesterification at the end of the reaction is about 10%.     

Correlation of the fatty acid composition and fluid property of the cholesteryl esters in the serum of Nigerian children with sickle cell disease and healthy controls

In a previous study conducted in Nigeria, we found that children with sickle cell disease (SCD) had exceedingly low total serum cholesterol levels (mean=100-102mg/dl). The fact that significant reductions in the levels of certain polyunsaturated fatty acids (PUFA) have been documented in the serum phospholipids of these same SCD subjects led us to inquire as to the fatty acid composition of the cholesteryl esters (CE) in their serum. Lecithin:cholesterol acyl transferase (LCAT), the enzyme in blood that catalyzes the reaction in which tissue cholesterol is acylated prior to its removal from cell membranes, is relatively specific for certain PUFA. CE in blood serum from 43 male and 42 female children with SCD, ages 4-18 years, and equal numbers of age- and gender-matched controls were analyzed for their fatty acid composition. Relative to the non-SCD controls, the CE of the SCD subjects contained 9% less linoleic acid, 16% less arachidonic acid, 40% less alpha-linolenic acid, 50% less eicosapentaenoic acid, and 36% less docosahexaenoic acid, but 15% more palmitic acid and 10% more oleic acid. Overall, the acyl chains of the CE of the SCD subjects were less fluid than those of the controls, as determined by comparison of their mean melting points (MMP) and double bond indices (DBI). MMP and DBI were both estimated from the individual constituent fatty acids comprising the CE acyl chains. The strongest correlations between MMP and fatty acid mole percent were seen with palmitic acid and linoleic acid. These results show that the fatty acid composition of the serum CE of children with SCD is abnormal relative to controls who do not have this hematologic disorder. We speculate that suboptimal fatty acid nutrition in Nigerian children with SCD compromises their ability to remove cholesterol from their tissues due to preference of the LCAT enzyme for PUFA, thereby accounting, in part at least, for the low total serum cholesterol levels one finds in children with SCD.     

A comparison of human and canine platelet fatty acid composition

1. Human and canine platelets were analysed for lipid composition by gas chromatography. 2. In comparing the two species' platelets, there were several lipids for which there were significant differences in lipid concentration. 3. The results indicate that human and canine platelets are quite similar in platelet lipid composition; however, the lipids for which there are significant differences may contribute to variations between species in platelet responses to stimulation.