Unusual fatty acids in turkeys fed a diet containing "Toprina". I. Composition of the higher fatty acids in "Toprina", the diet, feces and blood]

Totals lipids (TL) phospholipids (PL) and fatty acid composition of TL and PL (the latter in blood only) were estimated in Candida lipolytica cultivated on n-alkanes by industrial method ("Toprina"), in 0%, 10% and 15% "Toprina" diets and in faeces and blood of turkeys, males and females, fed these diets. With increasing concentrations of dietary "Toprina" phospholipids and 15:0 and 17:0 fatty acids increased, 17:1 and 17:2, typical of the product, appeared and increased in diets and blood. Other lipid and fatty acids values were not affected by dietary yeast.     

Different fatty acids in the egg, blood and feces of laying hens fed a diet containing "Toprina"

Total lipids (TL, phospholipids (PL) and fatty acid composition of TL and PL (the latter in blood and eggs only) were estimated in Candida lipolytica cultivated on n-alkanes by industrial method ("Toprina") in 0%, 10% and 15% "Toprina" diets and in eggs, blood and faeces of laying hens fed these diets. With increasing concentrations of dietary "Toprina" 15:0 and 17:0 fatty acids increased, 17:1 and 17:2, typical of the product, appeared and increased in diets, eggs, blood and faeces. Other fatty acid values were not affected by dietary yeast.     

Unusual fatty acids in turkeys fed a diet containing "Toprina". III. Composition of higher fatty acids in the heart and skeletal muscle

Total lipids (TL), phospholipids (PL) and fatty acid composition of TL and PL were estimated in breast, leg and cardiac muscles of turkeys, males and females, fede diets containing 0%, 10% and 15% of Candida lipolytica cultivated on n-alkanes by industrial method ("Toprina"). LT and PL were also valued in "Toprina" and diets. With increasing concentration of dietary yeasts it was observed that: a) Pl increased in diets, breast and leg muscles; b) fatty acids 15:0 and 17:0 increased, 17:1 and 17:2, typical of the product, appeared and increased in the tissues.     

Lipids of Candida lipolytica cultivated in n-alkanes by the industrial method ("Toprina")]

The following lipidic classes are examined in the present study: Total Lipids, Phospholipids, Neutral Lipids, Sterols, and Carotenoid Pigments from dried biomasses of Candida lipolytica grown on n-alkanes by industrial process following the BP technique ("Toprina"). The composition of the lipid classes examined in "Toprina" agree generally with bibliographic data about n-alkanes grown Candida lipolytica in batch cultures.     

Control of fatty-acid synthetase levels by exogeneous long-chain fatty acids in the yeasts Candida lipolytica and Saccharomyces cerevisiae.

Endogeneous fatty acid biosynthesis in the two yeast species, Saccharomyces cerevisiae and Candida lipolytica is completely repressed by the addition of long-chain fatty acids to the growth medium. In Candida lipolytica, this repression is accompanied by a corresponding loss of fatty acid synthetase activity in the cell homogenate, when the cells were grown on fatty acids as the sole carbon source. The activity of the Saccharomyces cerevisiae fatty acid synthetase, however, remains unaffected by the addition of fatty acids to a glucose-containing growth medium. From fatty-acid-grown Candida lipolytica cells no fatty acid synthetase complex can be isolated, nor is there any immunologically cross-reacting fatty acid synthetase protein detectable in the crude cell extract. From this it is concluded that Candida lipolytica, but not Saccharomyces cerevisiae, is able to adapt to the growth on fatty acids either by repression of fatty acid synthetase biosynthesis or by a fatty-acid-induced proteolytic degradation of the multienzyme complex. Similarly, the fatty acid synthetase complex disappears rapidly from stationary phase Candida lipolytica cells even after growth in fatty-acid-free medium. Finally, it was found that the fatty acid synthetase complexes from Saccharomyces cerevisiae and Candida lipolytica, though very similar in size and subunit composition, were immunologically different and had no common antigenic determinants.     

Assimilation of chlorinated alkanes by hydrocarbon-utilizing fungi.

The fatty acid compositions of two filamentous fungi (Cunninghamella elegans and Penicillium zonatum) and a yeast (Candida lipolytica) were determined after the organisms were grown on 1-chlorohexadecane or 1-chlorooctadecane. These organisms utilized the chlorinated alkanes as sole sources of carbon and energy. Analyses of the fatty acids present after growth on the chlorinated alkanes indicated that 60 to 70% of the total fatty acids in C. elegans were chlorinated. Approximately 50% of the fatty acids in C. lipolytica were also chlorinated. P. zonatum contained 20% 1-chlorohexadecanoic acid after growth on either substrate but did not incorporate C18 chlorinated fatty acids.     

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.     

Production of extracellular higher fatty acids by yeasts grown on hexadecanoic acid

Production of exocellular higher fatty acids by Candida yeasts was studied during their growth in a mineral medium with hexadecane. The qualitative and quantitative composition of exocellular higher fatty acids was investigated during cultivation of Candida lipolytica VCM Y 2378(695) under the conditions of different aeration (5 and 80% saturation of the medium with oxygen). Palmitic (C16:0), palmitooleic (C16:1), oleic (C18:1) and linoleic (C18:2) acids predominated among other higher fatty acids. The overall amount of fatty acids increased and the content of unsaturated fatty acids decreased when the yeast growth was limited with oxygen.     

Lipids of Pseudomonas aeruginosa Cells Grown on Hydrocarbons and on Trypticase Soy Broth

Lipids were extracted from cells of Pseudomonas aeruginosa grown on a pure hydrocarbon (tridecane), mixed hydrocarbons (JP-4 jet fuel), and on Trypticase Soy Broth. Total lipids produced from each substrate represented from 7.1 to 8.2% of cellular dry weight, of which 5.0 to 6.4% were obtained before cellular hydrolysis (free lipids) and 1.7 to 2.0% were extracted after cellular hydrolysis (bound lipids). Free lipids from cells grown on each medium were separated into four fractions by thin-layer chromatography. All fractions were present in cells from each type of medium, and the "neutral fraction" constituted the largest fraction. The fatty acid composition of free lipids was determined by gas-liquid chromatography. Cells grown on each medium contained saturated and unsaturated C(14) to C(20) fatty acids. Trace amounts of C(13) fatty acids were found in tridecane-grown cells. Saturated C(16) and C(18) were the major acids present in all cells. Quantitative differences were found in fatty acids produced on the three media, but specific correlations between substrate carbon sources and fatty acid content of cells were not evident. Tridecane-grown cells contained only traces of C(13) acid and small amounts of C(15) and C(17) acids, suggesting that the organism's fatty acids were derived from de novo synthesis rather than by direct incorporation of the hydrocarbon.     

Plasma membrane from Candida tropicalis grown on glucose or hexadecane. II. Biochemical properties and substrate-induced alterations.

Isolated plasma membranes from the yeast Candida tropicalis grown on two different carbon sources (glucose or hexadecane), had similar contents of protein (60% of total dry weight), lipid (21-24%) and carbohydrates (16-21%). Sodium dodecyl sulphate gel electrophoresis of the membrane proteins revealed 17 and 19 protein bands, respectively, for glucose and hexadecane grown cells. There were marked differences in RF values and relative peak heights between the two gels. Sterols and free fatty acids were the major components of the plasma membrane lipids. Phospholipid content was less than 2% of total plasma membrane lipids. Membrane microviscosity, as determined by fluorescence polarization, was very high (16.6 P). Fatty acid determination of membrane lipids by gas chromatography showed a significant increase of C16 fatty acids in plasma membranes of cells grown on hexadecane. Reduced-oxidized difference spectra demonstrated the presence of a b-type cytochrome in both Saccharomyces cerevisiae and C. tropicalis plasma membranes. Its concentration in C. tropicalis plasma membranes was three-fold greater in cells grown on hexadecane than in glucose grown cells.     

Toxic effects of fatty acids on yeast cells: dependence of inhibitory effects on fatty acid concentration.

The yeasts Saccharomyces cerevisiae, Candida utilis, and Candida lipolytica were used to investigate the action of different concentrations of fatty acids (from acetic to myristic acid) on cell growth, division, uptake of inorganic phosphate, and substrate oxidation. The former two yeasts were found to undergo an inhibition of growth, cell division, and phosphate uptake at lower acid concentrations and to experience the inhibition of substrate oxidation at higher acid concentrations. The concentration dependence of the action of fatty acids can be classified into four categories: 1) subthreshold concentrations which do not inhibit growth and have either no effect on, or stimulate, oxygen consumption; 2) threshold concentrations which lower the rate of growth, cell division, and phosphate uptake but do not inhibit the oxidation of carbon substrate; 3) above-threshold concentrations which inhibit partially even substrate oxidation, and 4) microbicide concentrations. Candida lipolytica displays the same sensitivity toward the action of fatty acids as the above yeast species; however, the threshold concentrations are higher and can be quickly lowered owing to oxidation by the yeast. The concentrations of fatty acids found in the medium after cultivations of yeast with n-alkanes are of the same order as limiting concentrations; the formation of acids with twelve and less carbons in the molecule can thus be assumed to be one of the basic reasons for lowering of biomass yields during cultivations on these hydrocarbons.     

Degradation of Hydrocarbons by Members of the Genus Candida III. Oxidative Intermediates from 1-Hexadecene and 1-Heptadecene by Candida lipolytica

Candida lipolytica (Phaff) was grown in a mineral-salts medium amended with either 1-hexadecene or 1-heptadecene as substrate. Intermediates of the same chain length as the substrate were isolated and identified by various analytical procedures. The following intermediates of 16 and 17 carbon atoms were identified: omega-unsaturated acids, omega-unsaturated primary and secondary alcohols, 1,2-epoxides, 1,2-diols, and 2-hydroxy acids. Based on the chemical structure of these compounds, three oxidative mechanisms are proposed for the degradation of long-chain 1-alkenes by this yeast: (i) methyl-group oxidation, (ii) double-bond oxidation, and (iii) subterminal oxidation.     

Single cell oil production by Yarrowia lipolytica growing on an industrial derivative of animal fat in batch cultures

The growth of an oleaginous strain of Yarrowia lipolytica on an industrial fat composed of saturated free fatty acids (stearin) was studied. Lipid accumulation during primary anabolic growth was critically influenced by the medium pH and the incubation temperature. This process was independent of the nitrogen concentration in the culture medium, but was favored at a high carbon substrate level and at a low aeration rate. At pH 6 and a temperature of 28-33 degrees C, 9-12 g/l of dry biomass was produced, whereas significant quantities of lipids were accumulated inside the yeast cells (0.44-0.54 g of lipid per gram of biomass). The strain showed the tendency to degrade its storage lipids, although significant amounts of substrate fat, rich in stearic acid, remained unconsumed in the culture medium. Y. lipolytica presented a strong fatty acid specificity. The fatty acids C12:0, C14:0, and C16:0 were rapidly incorporated and mainly used for growth needs, while C18:0 was incorporated with reduced rates and was mainly accumulated as storage material. Reserve lipids, principally composed of triacylglycerols (55% w/w of total lipids) and free fatty acids (35% w/w), were rich in stearic acid (80% w/w), while negligible amounts of unsaturated fatty acids were detected. When industrial glycerol was used as co-substrate, together with stearin, unsaturated fatty acid concentration in the reserve lipid increased.     

Lipid hydrogenation induces elevated 18:1-CoA desaturase activity in Candida lipolytica microsomes.

Microsomal membranes prepared from the mesophilic yeast Candida lipolytica grown at 10 degrees C were hydrogenated by the homogeneous Pd-catalyst, palladium di (sodium alizarine sulfonate) (Pd(QS)2). After hydrogenation to various levels, the microsomes were washed free of the Pd-complex and transferred to a reaction mixture (containing NADH, MgCl2, ATP, CoA and [14C]18:1-CoA) for assay of 18:1-CoA desaturase activity. Microviscosity alterations were also followed by measuring changes in DPH fluorescence polarization. Rapid catalytic hydrogenation of unsaturated fatty acids of the lipids occurred within 20-120 s, resulting in large increases in 16:0, 18:0 and 18:1 acids and decreases in 18:2 acid. In the range 7-20% 18:0 content, a pronounced increase in desaturase activity was observed, with a maximum of greater than 2-fold at a 18:0 content of 12%, followed by a decrease to the initial activity at 33% 18:0 content. These changes were well-correlated with changes in microviscosity, maximal desaturase activity occurring in the DPH fluorescence anisotropy range of 0.23-0.24; above and below this range, desaturase activities were close to the initial control values. It is suggested that the hydrogenation-induced increase in the formation of 18:2 from 18:1-CoA (proceeding partly through direct desaturation of PC) may be due to changes in conformation of the membrane-bound desaturase enzyme complex as a result of controlled rigidification of the surrounding lipids. The operation of such a self-regulating control mechanism would be consistent with a previously proposed model for microsomal desaturase action.     

Biodegradation of solar by Candida guilliermondii strain 1

Summary Candida guilliermondii strain 1 was grown on solar as a sole carbon source for 14 days, and the lipid classes were investigated. The yeast showed high affinity towards hydrocarbons of short chain length, and within 6 days the cellular lipid classes represented 39.69%, 27.50%, 15.35%, 2.23%, 16.20% of hydrocarbons, neutral lipids, free fatty acids, sterols and polar lipids respectively. Undecanoic and hexadecanoic acids were the major fatty acids of the cellular neutral lipids and oleic acid was the major component of the polar lipids.     

Mixed cultures of different yeasts species and yeasts with filamentous fungi in the SCP production. I. Production of single cell protein by mixed cultures Candida lipolytica and Candida tropicalis.

The aim of this study was to determine the application of mixed cultures Candida lipolytica and Candida tropicalis in the SCP production. N-paraffin fraction of crude oil and individual n-alkanes C:7--C:17 and glucose were used as carbon sources. The cultures were grown on laboratory scale in shaking flasks and in a 7 1 fermentor. It was found that the mixed cultures gave about 18% higher yield of biomass than the individual cultures.     

解脂耶罗维亚酵母工程菌合成超长链脂肪酸及温度的影响

[背景]解脂耶罗维亚酵母属于产油微生物,大量研究表明该酵母能够高产长链脂肪酸和油脂,但是应用该酵母合成超长链脂肪酸仍待研究。[目的]工程化解脂耶罗维亚酵母合成高值超长链脂肪酸,并研究温度对脂肪酸合成的影响。[方法]合成密码子优化的拟南芥(Arabidopsis thaliana)延长酶基因AtFAE1、非洲芥菜(Brassica tournefortii)延长酶基因BtFAE1和碎米芥属植物Cardamine graeca的延长酶基因CgKCS,分别构建质粒pYLEX1-AtFAE1、pYLEX1-BtFAE1、pYLEX1-CgKCS和pYLEX1-AtFAE1-BtFAE1-CgKCS。以解脂耶罗维亚酵母菌株Po1g为宿主,通过化学法分别转化上述4个质粒,获得工程菌Po1g-AtFAE1、Po1g-BtFAE1、Po1g-CgKCS和Po1g-AtFAE1-BtFAE1-CgKCS,比较评价超长链脂肪酸的合成。在此基础上,过表达内源二酯酰甘油酰基转移酶基因DGAT1(diacylglycerol acyltransferase)提高产油量,并研究温度对生物量、产油、脂肪酸组成的影响...     

Comparative lipid content ofCandida lipolytica grown on glucose and onn-hexadecane

Candida lipolytica, grown onn-hexadecane as the sole source of carbon and energy, contained 17.1% lipids in the logarithmic phase of growth, and 7.3% lipids in the stationary phase of growth. When the yeast was grown on glucose, it contained 6.2% lipids in the logarithmic phase of growth, and 3.6% lipids in the stationary phase of growth. Fatty acids, that could be extracted by petroleum ether after saponification, constituted the major part of the fatty acids ofC. lipolytica in its logarithmic phase of growth on glucose. They constituted only a minor amount of the fatty acids in the stationary phase of growth on glucose. The reverse was true when the yeast was grown onn-hexadecane. The broth contained more free, petroleum ether-soluble fatty acids when the cellular lipid content was high than when it was low. Overnight starvation ofC. lipolytica grown onn-hexadecane in a carbon-free nutrient medium, removed the residual cell-bound hydrocarbon, increased the cell population by one half and decreased the cellular lipid content (as % of dry yeast) by one third. Various methods for the determination of lipids, described as appropriate for yeasts were compared. The highest yields were obtained by extraction of the freeze-dried paste, at room temperature, with a 1:1 chloroform-methanol mixture.     

Analysis of fatty acid composition of Candida species by gas-liquid chromatography using a polar column

The fatty acid composition of representative Candida species was examined by gas-liquid chromatography (GLC) using a polar column. The major fatty acids were C14:0, C16:0, C18:0 saturated, C16:1 and C18:1 monoenoic series, with or without C18 polyunsaturated acids (C18:2 and C18:3). In Torulopsis glabrata and Saccharomyces cerevisiae the C18:2 and C18:3 acids were not found, but the C10:0 and C12:0 acids were detected in S. cerevisiae. These results indicated that the Candida genus could be distinguished from Torulopsis and Saccharomyces genera by GLC analysis of fatty acids. Quantitative differences in the fatty acid composition between cells grown at high temperature (37 degrees C) and low temperature (25 degrees C) were found generally in Candida species, and the amounts of C18 polyunsaturated acids (C18:2 and C18:3) increased in the cells grown at 25 degrees C. Each Candida species showed a characteristic profile in fatty acid composition. Determination of the cellular fatty acid composition in Candida species is likely to be useful for the grouping or chemotaxonomy of newer isolates of Candida species.     

A survey of yeasts in traditional sausages of southern Italy

The evolution of the yeast population during manufacturing and ripening of 'salsiccia sotto sugna', a typical salami of the Lucania region (southern Italy), was investigated. Four different batches, produced in four farms in Lucania, were studied. Each batch showed a specific yeast population, and the most frequently isolated yeasts belonged to Debaryomyces hansenii and its anamorph Candida famata, and Rhodotorula mucilaginosa. Yarrowia lipolytica was isolated from three sausage batches. The Y. lipolytica isolates were further characterised, in particular for their lipolytic activity on pork fat. Lipolytic activity was maximal at pH 5.5, with oleic and palmitic acids as major free fatty acids produced. The use of randomly amplified polymorphic DNA-polymerase chain reaction allowed the detection of a high genetic heterogeneity among the isolates phenotypically assigned to the species Y. lipolytica.