Lipid analysis of the plasma membrane and mitochondria of brewer's yeast.

The plasma membrane and mitochondria of bottom fermenting brewer's yeast obtained as a by-product of industrial beer production were isolated and the lipid fraction was analyzed. The phospholipid content accounted for 78 mg/g protein in the plasma membrane and 59 mg/g protein in the mitochondria. Major phospholipids in both preparations were phosphatidylinositol, phosphatidylcholine and phosphatidylethanolamine but their proportions differed significantly. In the plasma membrane phosphatidylinositol, and in the mitochondria phosphatidylcholine were present in the highest concentration (37 and 30%, respectively). The main classes of neutral lipids (triacylglycerols, ergosterol, squalene and steryl esters) were twice more abundant in the plasma membrane than in the mitochondria (61 and 33 mg/g protein, respectively). A characteristic of the neutral lipid composition of both organelles was the low content of ergosterol (12 and 7 mg/g protein, respectively) and a high content of squalene (25 and 22 mg/g protein). The main feature of the fatty acid composition of both organelles was the preponderance of saturated fatty acids (78 and 79%, respectively), among which palmitic acid was the principal one. The most expressed characteristics of lipid fractions of the analyzed plasma membranes and mitochondria, high concentration of squalene and preponderance of saturated fatty acids are the consequences of anaerobic growth conditions. The lack of oxygen had possibly the strongest effect on the lipid composition of the plasma membranes and mitochondria of bottom fermenting brewer's yeast.

Cerebrosides of Candida lipolytica yeast.

Candida lipolytica yeast was grown batchwise on glucose medium. Cerebrosides were isolated from the sphingolipid fraction of total lipids using column chromatography and separated into two compounds by high-performance thin-layer chromatography. Glucose was detected as the sole sugar constituent in cerebrosides. The fatty acid composition of cerebrosides was characterised by a predominance of saturated fatty acids and by a high proportion of fatty acids with 16 carbon atoms. The dominant fatty acid was h16:0. The principal long-chain base components of both cerebroside species were trihydroxy bases, 18- and 20-phytosphinosine. The unique characteristic of cerebrosides was the presence of a high proportion of sphingosine (one-fourth of the total long-chain bases), which is a common characteristic of mammalian sphingolipids and rarely occurs in yeast cerebrosides. The ceramide moiety profile of cerebrosides is similar to that of epidermal ceramides, which implies a possibility for their application in care cosmetics.

Effect of growth phase on the content and composition of ceramides of the hydrocarbon-assimilating yeast Candida lipolytica.

Candida lipolytica yeast was grown batchwise on n-hexadecane as the carbon and energy source. Ceramides were quantitatively isolated from total lipids of exponential and stationary phase cells by a combination of column chromatography and preparative high-performance thin-layer chromatography. After acid methanolysis their composition was analyzed by gas-liquid chromatography. The ceramide content of the exponential phase cells was two times higher than the one of the stationary phase cells. The composition of long-chain base moiety of ceramides did not change significantly during the growth. In both growth phases 19-phytosphingosine was the major long-chain base. However, the fatty acid composition of ceramides changed greatly during the growth. In the exponential growth phase, ceramides contained predominantly fatty acids greater than 20 carbon atoms, while fatty acids shorter than 20 atoms predominated in ceramides of the stationary phase, 16:0 being the main one. In the exponential growth phase fatty acid moiety of ceramides was characterized by unusually high degree of unsaturation and relatively high proportion of odd-numbered fatty acids. However, the proportion of both, unsaturated and odd-numbered fatty acid decreased significantly in ceramides of the stationary phase. The unexpected finding was the absence of fatty acid hydroxylation of ceramides in the exponential phase cells and unusually low degree of hydroxylation in the stationary phase.

The influence of carbon source on the level and composition of ceramides of the Candida lipolytica yeast.

Candida lipolytica yeast was grown batchwise on two different carbon sources, glucose and n-hexadecane. Free ceramides were quantitatively isolated from sphingolipid fractions of total lipids by a combination of column chromatography and preparative thin-layer chromatography. Their composition, after acid methanolysis, was analysed by gas-liquid chromatography. The ceramide content accounted for 2.6% of the total cell lipids in hexadecane-grown cells, which was 1.5 times higher than in glucose-grown cells. The fatty acid composition of ceramides was characterized by the predominance of fatty acids shorter than 20 carbon atoms and by high concentrations of fatty acids with 16 carbon atoms after growth on both carbon sources. The dominant fatty acid was hydroxylated 16:0 in the glucose-grown cells and 16:0 in the hexadecane-grown cells. The striking finding was the low degree of fatty acid hydroxylation and relatively high proportion of odd-numbered fatty acids in ceramide of the n-hexadecane-grown cells. The ceramides contained an unusual long-chain base composition. In hexadecane-grown cells more than 60% of the long-chain bases were C19 phytosphingosine. In glucose-grown cells more than one-half of the total long-chain bases were tetrahydroxy bases, 4,5-dihydroxysphinganine and 4,5-dihydroxyeicosasphinganine.

Isolation and chemical composition of the ceramide of the Candida lipolytica yeast.

Candida lipolytica yeast was grown batchwise on glucose medium. Ceramide was quantitatively isolated from sphingolipid fractions of total lipids by a combination of column chromatography and preparative thin-layer chromatography. Fatty acids and long-chain bases, obtained after acid methanolysis, were analyzed by gas-liquid chromatography. Fatty acid composition was characterized by predominance of fatty acids shorter than 20 carbon atoms (72.2% of total fatty acids) and by high concentration of fatty acids with 16 carbon atoms (43.3% of total fatty acids). The dominant fatty acids were h16:0 and 18:1. The major fatty acid with more than 20 carbon atoms was h24:0. Hydroxylated fatty acids quantitatively represented almost half of the total fatty acids. The ceramide contained an unusual long-chain base composition. Besides the well-known trihydroxy bases, 18- and 20-phytosphingosine, which is a common characteristic for yeast sphingolipids, a high proportion of more polar bases than the trihydroxy ones, was found. The structures of 4,5-dihydroxyicosasphinanine (1,3,4,5-tetrahydroxy-2-aminooctadecane) and 4,5-dihydroxyicosasphinganine (1,3,4,5-tetrahydroxy-2-aminoeicosane) were tentatively assigned to these bases. Their chromatographic properties were identical to those of synthetic bases.

Tissue zinc dynamics during the immune reaction in mice.

Owing to the importance of zinc for the functioning of the immune system, the role of endogenous Zn, located both in lymphoid and nonlymphoid organs, was investigated during the standard humoral and cellular types of immune response. For this purpose, the dynamics of hepatic, thymic, splenic, and renal Zn content was determined in mice sensitized with (a) sheep red blood cells and (b) semiallogeneic lymphocytes during the local host vs graft reaction (HVGR). The data obtained by ion-coupled plasma spectrometry revealed that the humoral type of immunity is characterized by a significant increase of Zn concentration in the liver and in the thymus. Simultaneously, linear regression analysis showed that the generation of plaque-forming cells in the individual mouse was highly positively correlated with Zn concentration in the liver (r=0.897), and spleen (r=0.833), and negatively with Zn concentration in the thymus (r=-0.624). Similar relationships between the intensity of local immune reaction and tissue Zn levels were found in local HVGR at the fifth day in the liver and spleen (r=0.861 and r=0.695, respectively), at the seventh day in the thymus (r=-0.797), and at the tenth day in the liver (r=-0.859). The data emphasize the necessity of Zn for the development of normal immune response and point to the existence of a Zn-dependent hepato-thymic axis during the humoral and cellular types of immune reactivity.

Cell lipids of the Candida lipolytica yeast grown on methanol.

Candida lipolytica yeast, grown on 1% methanol as the only carbon and energy source, synthesized 4.9% of dry cell mass as lipids, 52.3% of which were polar lipids. Polar lipids consisted mainly of phospholipids and sphingolipids as their minor components. The total long-chain bases content has been found to account only for 0.7% of the polar lipids. The long-chain bases composition determined by thin-layer and gas chromatography shows a preponderance of trihydroxy bases and a small amount of dihydroxy bases. The striking finding was the high content of 19-phytosphingosine (90.8% of total long-chain bases). Fatty acid (FA) composition of polar lipids was characterized by the relatively high concentration of unsaturated fatty acids (66.4% of total FA) and by the predominance of fatty acids with 16 carbon atoms (85.0% of total FA).