Adult rat brain synaptic vesicles II. Lipid composition

The lipid composition of synaptic vesicles isolated from adult rat brain was determined. Vesicles contained cholesterol and phospholipid but very little ganglioside, galactolipid, free fatty acid and triglyceride was detected. Ethanolamine and choline phosphoglycerides were the dominant phospholipids. Lysophosphatidyl choline was present in very low amounts. The fatty acid composition of the phosphoglycerides was characterized by high levels of docosahexaenoic acid in the ethanolamine and serine phosphoglycerides, and the absence of long chain fatty acids from the sphingomyelins. All the characteristic features of the lipid composition of the synaptosomal plasma membrane (with the exception of the ganglioside content) were seen in the synaptic vesicle lipids. The results are discussed in terms of the exocytosis mechanism of transmitter release.     

Effect of temperature acclimatization on the fatty acid composition of goldfish intestinal lipids

1. The fatty acid composition of whole goldfish, whole-intestinal mucosa, intestinal mucosal membranes and individual phospholipids extracted from mucosal membranes were measured, fish adapted to different temperatures being used. 2. Alterations of the adaptation temperature did not noticeably affect the fatty acid composition of the whole-fish lipids, but there were marked changes in the fatty acids of lipids extracted from homogenates of goldfish intestinal mucosa. These changes were more pronounced in a membrane fraction prepared from these homogenates. Raising the adaptation temperature by 20 degrees C halved the percentage of C(20:1), C(20:4) and C(22:6) fatty acids and nearly doubled the percentage of C(18:0) and C(20:3) fatty acids recovered. 3. Choline phosphoglycerides constituted about one-half and ethanolamine phosphoglycerides about one-quarter of the total membrane phospholipids. 4. The fatty acids of choline and ethanolamine phosphoglycerides were more susceptible to temperature-dependent changes than were the phosphoglycerides of inositol or serine. 5. The increase in C(18:0) fatty acid that occurred in membranes of warm-adapted fish was greatest for ethanolamine phosphoglycerides, but increases also occurred in other phospholipid fractions and in membrane neutral lipids.     

Content of fatty acids bound to proteins and of cholesterol in neuroblastoma C1300 N18 cells during differentiation

The content and concentration of fatty acids lightly and tightly bound with proteins and the concentration of cholesterol were studied in differentiated and undifferentiated neuroblastoma C1300 N18 cells. Lightly bound lipids were extracted by the method of Blight and Dyer with subsequent additional rinsing by chloroform-methanol (1:1) and methanol extractions. The remaining protein-bound lipid was cleaved by mild alkaline hydrolysis in the methanol medium. Methyl esters of fatty acid were the fraction tightly bound with proteins. The main components in the fractions were fatty acids 16:0, 18:0, 18:1 omega 9, 20:4 omega 6. Cell differentiation caused changes essential in the content and concentration of fatty acids in the both fractions: the total quantity of saturated fatty acids was found to increase, the relative level of saturated fatty acids was higher in the tightly bound lipid fraction. During cell differentiation the level of cholesterol increased per 1 mg of protein in the lightly bound lipid fraction. In the tightly bound lipid fraction the cholesterol level per 1 mg of protein was unchanged.     

Lipid and fatty acid composition of canine lipoproteins

Lipid classes and their fatty acids were studied in the major lipoprotein fractions from canine, in comparison with human, plasma. In dogs, high-density-lipoprotein (HDL), the main carrier of plasma phospholipid (PL), cholesterol ester (CE) and free cholesterol, was the most abundant lipoprotein, followed by low and very-low density lipoproteins (LDL and VLDL). Notably, LDL and VLDL contributed similarly to the total dog plasma triacylglycerol (TG). The PL composition was similar in all three lipoproteins, dominated by phosphatidylcholine (PC). Even though the content and composition of lipids within and among lipoproteins differed markedly between dog and man, the total amount of circulating lipid was similar. All canine lipoproteins were relatively richer than those from humans in long-chain (C20-C22) n-6 and n-3 polyunsaturated fatty acids (PUFA) but had comparable proportions of total saturated and monoenoic fatty acids, with 18:2n-6 being the main PUFA in both mammals. The fatty acid profile of canine and human lipoproteins differed because they had distinct proportions of their major lipids. There were more n-3 and n-6 long-chain PUFA in canine than in human plasma, because dogs had more HDL, their HDL had more PC and CE, and both these lipids were richer in such PUFA.     

Age-related changes of lipid fractions and total fatty acids in the serum of female and male rats aged from 37 to 1200 days

1. Total lipids and the lipid fractions cholesterol ester, triacylglycerols, free cholesterol, free fatty acids and phospholipids as well as the fatty acid patterns of total lipids were measured in the serum of female and male rats (Wistar SPF, strain Hannover) aged 37-1213 days. 2. All parameters were monitored every 49th day. Five female and five male animals were used in each experiment. 3. All lipid fractions showed a significant positive linear regression vs age in both sexes. There were multiple significant differences in lipid fractions between the sexes. 4. Among the fatty acids only docosahexaenoic acid shows a significant correlation with age in both sexes.     

13C NMR, GC and HPLC characterization of lipid components of the salted and dried mullet (Mugil cephalus) roe "bottarga"

13C NMR spectroscopy, in conjunction with HPLC and GC techniques, has been used to study the molecular composition of lipids extracted from commercial products of bottarga. To this goal, both the saponifiable and unsaponifiable fractions of lipid extracts were also examined by 13C NMR. Among the major lipid classes wax esters (WE) showed a concentration of more than 50mol%, triacylglycerols (TAG) and phospholipids (PL) represented a minor fraction. Concentrations up to 29mol% of free fatty acids (FFA) were found. The most represented fatty alcohol was 16:0 that accounted for more than 50%, among fatty acids the most represented were 16:1 n-7, 22:6 n-3, 18:1 n-9, 16:0, and 20:5 n-3, in particular the n-3 polyunsaturated fatty acids (PUFA) averaged 40mg/g of the edible portion. 13C NMR spectroscopy put in evidence that cholesterol was present in its free and esterified forms and its total content was measured as ca. 10mg/g of the edible portion.     

The fatty acids of Trypanosoma vivax

A study was carried out to determine the lipid composition of the blood-stream form of the African trypanosome. Trypanosoma vivax. Data from thin layer chromatography showed that the major polar lipids were lysophosphatidylcholine, sphingomyelin, phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine and diphosphatidylglycerol. The major neutral lipids were sterol, monoacylglycerol, diacylglycerol, free fatty acid and triacyglycerol. 16:0, 18:0, 18:1 and 18:2 constituted the major fatty acids of both the polar and neutral lipid fractions. The work constituted the first detailed study on the fatty acid composition of this African trypanosome.     

Incorporation of 2-14C-acetate into isolated nuclei and cytosolic lipids of rat thymus cells

It is generally recognized nowadays that active lipid metabolism takes place in the nucleus of a mammalian cell. Experimental data testify to the biosynthesis of polyphosphoinositides and phosphatidylcholine and reveal corresponding enzymes within nuclei of mammalian cells. These findings suggest that lipidmediated signaling pathways in nuclei operate independently of lipid-mediated regulatory mechanisms functioning in membranes and cytosol. To explore the pathways of intranuclear lipid biosynthesis, we studied incorporation of 2-¹⁴C-acetate into lipids of cytosol and isolated nuclei of rat thymus cells after separate and combined incubation with the labeled precursor. The most efficient incorporation of 2-¹⁴C-acetate into lipids (cholesterol, free fatty acids, and phospholipids) was observed in a reaction mixture containing cytosol. When the reaction mixture contained only nuclei, incorporation of the radioactive precursor into lipids also took place, but specific radioactivity of the lipids was essentially lower than in the cytosol. In both cases, 2-¹⁴C-acetate incorporated into phosphatidylethanolamine, sphingomyelin, phosphatidylserine, phosphatidylinositol, and cardiolipin. Phosphatidylcholine, the most abundant membrane phospholipid, demonstrated the lowest radioactivity, which was significantly lower than that of phosphatidylethanolamine. Incorporation of newly synthesized free fatty acids in nuclear phospholipids was inhibited, if nuclei were incubated with cytosol. As a result, radioactive free fatty acids were accumulated in nuclei, while in cytosol they were efficiently incorporated into phospholipids. The levels of phospholipids and cholesterol remained constant regardless of incubation protocol, while the overall yield of free fatty acids decreased after combined incubation of nuclear and cytosolic fractions or after incubation of cytosol without nuclei. Putative mechanisms underlying the appearance of radioactive lipids in isolated nuclei of thymus cells are discussed.     

Characterization of membrane components of the flask-shaped mycoplasma Mycoplasma mobile

The cell membrane of Mycoplasma mobile was isolated by either ultrasonic or French press treatment of intact cells. The membrane fraction contained all of the cellular lipids, but only one-third of cellular proteins and had a density of 1.14 g ml-1. The soluble fraction contained the NADH dehydrogenase activity of the cells, as well as a protein with an apparent molecular mass of 55 kDa that was phosphorylated in the presence of ATP. Lipid analyses of M. mobile membranes revealed that membrane lipid could be labelled by radioactive glycerol, oleate and to a much higher extent by palmitate but not by acetic acid. The membrane lipid fraction was composed of 54% neutral and 46% polar lipid. The major constituents of the neutral lipid fraction were free fatty acid, free cholesterol and cholesterol esters (45, 25 and 20%, respectively, of total neutral lipid fraction). The free cholesterol count was 13% (w/w) of total membrane lipids with a cholesterol:phospholipid molar ratio of about 0.9. Among the polar lipids, both phospho- and glycolipids were detected. The phospholipid fraction consisted of a major de novo-synthesized phosphatidylglycerol (approximately 63% of total phospholipids), plus exogenous phosphatidylcholine and sphingomyelin incorporated in an unchanged form from the growth medium. The glycolipid fraction was dominated by a single glycolipid (approximately 90% of total glycolipids) that was preferentially labelled by palmitic acid and showed a very high saturated:unsaturated fatty acids ratio.     

Dietary polyunsaturated n-6 lipids effects on the growth and fatty acid composition of rat mammary tumors

The aim of this study was to analyze the effects of a polyunsaturated n-6 high-fat diet on rat DMBA-induced breast cancer at different stages of the carcinogenesis and to investigate if changes in the tumor fatty acid composition are one of the mechanisms by which dietary lipids could exert their effects. 14 fatty acids were evaluated in 6 lipid fractions. The results firstly showed that this high-fat diet stimulated the malignant mammary tumor growth, mainly all in the promotion group. The tumor lipid analysis indicated: 1) that each lipid fraction presented distinct major fatty acids (>5%) which were not the most abundant in the diet, except in the case of the triacylglicerides, suggesting the different resistance to dietary fatty acid modification of the tumor lipid fractions; 2) a higher arachidonic acid content in the fractions with less linoleic acid, above all in phospholipids, particularly in the phosphatidylethanolamine, indicating a different efficiency of conversion; 3) the three most abundant fatty acids in the dietary lipid (18:2n-6, 18:1n-9 and 16:0) were those which essentially displayed the differences between groups; thus, the high-fat diet changed the tumor lipid profile, increasing the 18:2n-6 relative content and decreasing that of the 18:1n-9; differences were significant in phosphatidylcholine, free fatty acids and triacylglycerides. Any change was obtained in the phosphatidylinositol. The greatest number of differences was found in the promotion group. Taken as a whole, our results suggest the different roles of lipid fractions in breast cancer cells and an association between cancer malignancy and the content of linoleic and oleic acids.     

Membrane lipids of human peripheral nerve and spinal cord.

Major membrane lipids were determined in specimens of human peripheral nerve (cauda equina) and spinal cord of 10 subjects aged 20-70 years. The same lipids were also assayed in myelin from the same tissues isolated with two different procedures and in myelin of cauda equina from 3 subjects aged 17-91 years isolated with a third method. The concentrations (mean and standard deviation) of phospholipids were 90 +/- 11 and 96 +/- 9 nmol/g fresh weight; of cholesterol 70 +/- 15 and 101 +/- 16; of cerebroside 19 +/- 3 and 41 +/- 7; of sulfatide 10 +/- 1 and 11 +/- l; and of gangliosides 0.80 +/- 0.08 and 0.40 +/- 0.05 N in cauda equina and spinal cord, respectively. The proportion of ethanolamine phosphoglyceride was lower and that of sphingomyelin higher in cauda equina than in spinal cord. The myelin of peripheral nerve and spinal cord contained almost the same proportions of lipids as the whole tissue. The protein-bound sialic acid content was 3-fold higher than the lipid-bound sialic acid content in cauda myelin. The fatty acid patterns of choline, ethanolamine, inositol and serine phosphoglycerides of spinal cord and its myelin, were very similar to those of cerebral white matter, while the phosphoglycerides of cauda equina had higher proportions of monoenoic acids and lower proportions of polyunsaturated fatty acids. The fatty acid patterns of sphingomyelin, cerebroside and sulfatide of spinal cord were similar to those of cerebral white matter, while those of cauda equina contained significantly more saturated fatty acids. This suggests that the lipid and fatty acid compositions of peripheral nerve are particularly suitable for the formation of a tightly packed myelin membrane which can be a powerful shield against infections and other injuries.     

Lipid synthesis in the adult dog heartworm, Dirofilaria immitis.

1. Incorporation studies with three labelled substrates--[14C]2-glycerol, [14C]1-acetate and [14C]1-oleic acid--demonstrated that adult dog heartworms can synthesize all classes of complex lipids present, including free cholesterol. 2. Diacylglycerols and phosphoglycerides were most rapidly labelled regardless of the precursor employed. 3. 14C from glycerol was found in the aqueous phase of saponified lipids, whereas that from oleic acid was in the fatty acid portion. 4. Tag from acetate was predominantly in the fatty acid portion of saponified lipids and also occurred in the unesterified fatty acids. 5. Acetate and unesterified fatty acids, as represented by oleic acid, were more readily used for lipid synthesis than was glycerol.     

Isolation and properties of two classes of low-density vesicles from Saccharomyces cerevisiae.

A mixture of small (0.43-mum diameter) and large (0.62-mum diameter) low-density vesicles from spheroplasts of Saccharomyces cerevisiae was fractionated by rate centrifugation in a gradient of 0 to 8% (wt/vol) Ficoll to yield fractions rich (90 to 95%) in small or large vesicles. The large, but not small, vesicles swelled when diluted into mannitol solutions containing less than 0.4 M mannitol. The pH-electrophoretic mobility curve of the large vesicles showed that they are probably enclosed in a phospholipid-protein membrane. The dyes neutral red and toluidine blue, accumulated into large vesicles by intact cells and spheroplasts, were largely lost from large vesicles when these were separated from stained spheroplasts. Sudan black III stained small and large vesicles, both classes of vesicle retaining the stain on separation. Fractions rich in large vesicles contained proportionately more phospholipid and less free sterols, diacylglycerols, and free fatty acids compared with those enriched in small vesicles. The two classes of vesicles contained about the same proportions of esterified sterols and triacylglycerols. The free fatty acids in both small and large vesicles were free from unsaturated fatty-acyl residues; diacylglycerols and triacylglycerols contained appreciable proportions of unsaturated fatty-acyl residues. Small vesicles were richer in lipase activity, whereas the larger vesicles contained greater beta-glucanase and alpha-mannosidase activities. Phospholipase activity could not be detected in any of the fractions.     

Human prostasome membranes exhibit very high cholesterol/phospholipid ratios yielding high molecular ordering

Lipid analysis and ESR studies were carried out on prostasomes isolated from human semen. Cholesterol plus phospholipids amounted to approximately 0.80 mumol per mg protein with a striking quantitative domination of cholesterol over the phospholipids, the molar ratios of cholesterol/sphingomyelin/glycerophospholipids being 4:1:1. Saturated and monounsaturated fatty acids were dominating both in the glycerophospholipids and in sphingomyelin. The order parameters, S, deduced from ESR spectra of spin-labelled fatty acids incorporated into prostasome membranes order parameters, S, deduced from ESR spectra of spin-labelled fatty acids incorporated into prostasome membranes were very high, viz. 0.75 for 5-doxylstearic acid and 0.30 for 16-doxylstearic acid at 25 degrees C. Slightly lower values were obtained for the spin-labelled fatty acids when they were incorporated into dispersions of extracted prostasome lipids or into synthetic lipid mixtures of similar composition. The highly ordered lipids in the prostasome membrane thus seemed to be minimally perturbed by proteins in the membrane and ESR spectra showed no signs of immobilized lipids.     

Lipid metabolism in brain tissue explants

Abstract— Tissue explants from frontal lobes of rat brain were used for the study of cerebral fatty acid metabolism. After tissues had been maintained in serum-supplemented medium, a lipid-free medium was substituted and metabolic studies were carried out. Under these conditions explants continued to take up lipid precursors for at least 48 h, as judged by incorporation of dl -[2-¹⁴C]mevalonic acid into cellular lipids. [l-¹⁴C]Stearic acid and [l-¹⁴C]palmitic acid were bound to cells as the free fatty acids, or incorporated into neutral lipids (particularly triglycerides), glycolipids and phospholipids. In the galactolipid fraction, cerebrosides were the principal radioactive lipids. Choline phosphoglycerides, ethanolamine phosphoglycerides, inositol phosphoglycerides and serine phosphoglycerides were the principal radioactive phospholipids. Fatty acids were incorporated into cellular lipids either unchanged or after desaturation, chain elongation, or both. Maximum incorporation of stearate occurred in tissues derived from 3-day-old animals. With increasing age the uptake of fatty acid dropped sharply. When the labelling of lipids as a function of time was followed in 3-day-old animals, triglycerides and choline phosphoglycerides were the first fractions to take up labelled stearate. Labelling of cerebrosides occurred slowly, only becoming evident after 24 h. These studies exemplify the usefulness of tissue explants for prolonged metabolic studies in normal and pathological specimens of brain.     

Age-related changes of lipid fractions and total fatty acids in liver lipids and heart lipids of female and male rats aged 37-1200 days (liver) and 331-1200 days (heart)

1. Total lipids and the lipid fractions cholesterol ester, triacylglycerol, free cholesterol, free fatty acids and phospholipids, as well as the fatty acid patterns of total lipids, were measured in liver homogenates of female and male rats (Wistar SPF, strain Hannover) aged 37-1213 days. 2. The same parameters were measured in the apex of the heart in female and male rats aged 331-1213 days. 3. All parameters were monitored every 49th day. Five female and five male animals were used in each experiment. 4. The lipid fractions in liver showed a positive linear regression vs age, whereas all lipids in rat heart showed a negative regression vs age in both sexes. 5. The significance of regression vs age of fatty acids was much less than that in the lipid fractions of liver and heart of these animals.     

Variation in plasma lipids during the reproductive cycle of male and female desert tortoises,Gopherus agassizii

Plasma triacylglycerol, phospholipid, cholesterol, cholesterol esters, fatty acids, and total lipids were measured in 30 female and 20 male desert tortoises (Gopherus agassizii) during the annual reproductive cycle in the eastern Mojave desert, Nevada. Blood samples were collected at monthly intervals from April to October. All lipid fractions, with the exception of free fatty acids, were significantly higher in female plasma than in male plasma in all months of the year. In contrast, free fatty acids were higher in male plasma than in female plasma in all months. The seasonal pattern in estradiol secretion mirrored that of triacylglycerol, phospholipid, cholesterol, and total lipid, all of which showed a significant correlation with the hormone. Estradiol and the vitellogenesis-associated lipids were all significantly higher in August, September, October, and April than in June. The seasonal variation in cholesterol ester levels in females did not correlate with any of the reproductive events and did not appear to be involved in yolk precursor formation. Total lipid in males showed a negative correlation with testosterone and spermatogenesis. Individual fatty acids in the June and August samples (at the highest and lowest estradiol levels) were compared in male and female plasma. The percent of C18:3n3, C18:2n6, C18:1n9, C20:5n3, and C22:5 were significantly higher in the June female plasma sample than in the August sample. Docosahexanoic (C22:6n3) acid was barely detectable in female plasma in either month.     

The relationship between plasma membrane lipid composition and physical-chemical properties. III. Detailed physical and biochemical analysis of fatty acid-substituted EL4 plasma membranes

Murine leukemia EL4 cells were modified by supplementation of culture media with fatty acids for 24 h. A plasma membrane-enriched fraction was prepared from substituted and normal cells. Analyses were performed to determine fatty acyl composition, phospholipid headgroup composition and cholesterol content. The two major membrane phospholipids, phosphatidylethanolamine (PE) and phosphatidylcholine (PC) were isolated by thin-layer chromatography and ESR measurements were done on liposomes prepared from these lipids as well as on the intact plasma membrane preparations. Slight perturbations in overall plasma membrane lipid composition were observed when EL4 cells were supplemented with a single exogenous fatty acid. This may be consistent with the idea that the incorporation of exogenous fatty acid induces compensatory changes in membrane lipid composition. On the other hand, we observed no significant difference in two ESR motional parameters between the unsubstituted control and various fatty acid-substituted plasma membranes. ESR measurements carried out on PE and PC liposomes derived from 17:0- and 18:2c-substituted membranes also failed to detect major differences between these liposomes and those made from normal EL4 phospholipids. In the case of liposomes prepared from 18:2t,-substituted membranes, the order parameter was significantly changed from the normal. However, the change was in opposite directions in PE and PC, perhaps accounting for the fact that no change parameter is seen in intact 18:2t-substituted plasma membrane. Measurements of order parameter (S) in mixed lipid vesicles showed that at up to 50 mol% mixture of a synthetic PC with plasma membrane PC, the value of S was only marginally different from that of the plasma membrane PC vesicles. We interpret these data as an indication that the two ESR parameters used are not sufficiently sensitive to detect changes due to modifications of the acyl chain composition of a complex biological membrane.     

Phospholipid and fatty acid composition of tetrodotoxin receptor-rich membrane fragments from Electrophorus electricus

To study the interaction of voltage-sensitive Na+-channels with membrane lipids, the phospholipid and fatty acid composition of highly purified membrane fragments from the remarkably differentiated plasma membrane of Electrophorus electricus has been analyzed. After density gradient fractionation and carrier free electrophoresis, fractions with up to 30 pmol tetrodotoxin binding/mg protein can be obtained, which may correspond to a 50% pure preparation of the extrasynaptic part of the excitable face. Phospholipid classes and cholesterol are separated by one-dimensional thin-layer chromatography in acidic and alkaline solvent systems. The following mean molar contents are found: 40% phosphatidylcholine, 23% phosphatidylserine, 30% phosphatidylethanolamine and 7% sphingomyelin. In a series of 11 animals, significant deviations from these mean values have been observed. The fatty acid composition of the phospholipids has been determined by gas chromatography. Phosphatidylcholine contains more than 50% 16:0, and about 20% unsaturated fatty acids in the C-18 group. Compared to other plasma membrane fractions, this phospholipid is the least differentiated. By contrast, phosphatidylethanolamine and phosphatidylserine show many characteristics in different membrane fractions, especially in their unsaturated components representing more than 50%. 22:6, as the major constituent in these fractions, accounts for a quarter to a third of all fatty acids in these fractions. 18:0 is the main saturated component in these two phospholipids with abundances of typically a quarter or less of all fatty acids. Knowledge of the lipid composition of these excitable membranes may help to conserve binding and structural properties when analyzing lipid-sensitive Na+-channels in vitro. It is also useful as a guideline for systematic reconstitution studies.     

Composition of the protoplast membrane from Saccharomyces cerevisiae

1. Protoplasts of Saccharomyces cerevisiae N.C.Y.C. 366 were prepared by incubating washed exponential-phase cells in buffered mannitol (0.8m) containing 10mm-magnesium chloride and snail gut juice (about 8mg. of protein/ml. of reaction mixture). Protoplast membranes were obtained by bursting protoplasts in ice-cold phosphate buffer (pH7.0) containing 10mm-magnesium chloride. 2. Protoplast membranes accounted for 13-20% of the dry weight of the yeast cell. They contained on a weight basis about 39% of lipid, 49% of protein, 6% of sterol (assayed spectrophotometrically) and traces of RNA and carbohydrate (glucan+mannan). 3. The principal fatty acids in membrane lipids were C(16:0), C(16:1) and C(18:1) acids. Whole cells contained a slightly greater proportion of C(16:0) and a somewhat smaller proportion of C(18:1) acids. Membrane and whole-cell lipids included monoglycerides, diglycerides, triglycerides, sterols, sterol esters, phosphatidylcholine, lysophosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol+phosphatidylserine. Phosphorus analyses on phospholipid fractions from membranes and whole cells showed that membranes contained proportionately more phosphatidylethanolamine and phosphatidylinositol+phosphatidylserine than whole cells, which in turn were richer in phosphatidylcholine. Phospholipid fractions from membranes and whole cells had similar fatty acid compositions. 4. Membranes and whole cells contained two major and three minor sterol components. Gas-liquid chromatography, mass spectrometry and u.v. and i.r. spectra indicated that the major components were probably Delta(5,7,22,24(28))-ergostatetraen-3beta-ol and zymosterol. The minor sterol components in whole cells were probably episterol (or fecosterol), ergosterol and a C(29) di-unsaturated sterol. 5. Defatted whole cells contained slightly more glutamate and ornithine and slightly less leucine and isoleucine than membranes. Otherwise, no major differences were detected in the amino acid compositions of defatted whole cells and membranes.