Oxygen pressure,fatty acid composition and ergosterol level in Rhodotorula gracilis

Cells of Rhodotorula gracilis grown for 6 hrs at 2 mm Hg O2 pressure when compared with cells grown for 6 hrs at 140 mm Hg, show: a) a large decrease in the level of ergosterol, b) a significant increase in the level of oleic acid and a decrease in the levels of linoleic and linolenic acids, both in fatty acid fraction of the phospholipids and in the free fatty acids and neutral fat fractions. The results suggest that the dehydrogenation of oleic acid to linoleic acid is preferentially inhibited at low O2 pressure. The possibility is considered that these changes of lipid metabolism might be causally related with decrease of the growth rate observed at low O2 pressure.     

Control of acyl lipid desaturation in the yeast Rhodotorula gracilis via the use of the cyclopropenoid fatty acid,sterculate

Summary The effect of the cyclopropene fatty acid, sterculic acid, on fatty acid metabolism in the oleaginous yeast Rhodotorula gracilis (CBS 3043) has been studied. Sterculate caused approximately 90% inhibition of [1-¹⁴C]sterate desaturation but only about 25% inhibition when [1-¹⁴C]acetate was used as precursor. Measurement of acyl-coenzyme A (CoA) pool labelling and the pattern of distribution of radioactivity within lipid classes suggested that the high rate of inhibition of stearate desaturation was due to sterculate reducing the formation of stearoyl-CoA. In agreement with previous suggestions for plant and animal systems, experiments with a post-mitochondrial (20 000 g × 30 min) supernatant suggested that sterculoyl-CoA was the active inhibitor. The action of sterculate on endogenous lipid composition was most marked in stationary-phase cells. In such cells an increase in stearate was seen especially in the triacylglycerol pools, while -linolenate was increased predominantly in the phospholipid fractions. The increased flux of carbon to -linolenate at the same time that stearate desaturation appeared to be inhibited by sterculate, indicated that that two separate pathways for desaturation, employing different substrates, may operate in R. gracilis. We suggest that sterculate inhibits the stearoyl-(acyl-)CoA-dependent pathway but has little effect when phospholipids are used as substrates for acyl chain desaturation. Offprint requests to: J. L. Harwood     

Kinetics of growth and fatty acid production of Rhodotorula glutinis

Summary The effect of culture conditions, especially nitrogen/carbon (N/C) ratio and temperature, on acyl lipid profiles in the oleaginous yeast Rhodotorula glutinis NRRL Y 1091 have been investigated. Cultivation of the microorganism under N-limited conditions (N/C < 0.14 g/g) resulted in enhanced fatty acid (FA) cell content but a reduced relative amount of polyunsaturated fatty acids (PUFA). The maximal FA productivity was obtained for a 0.025 N/C ratio resulting from the arrangement between the specific rate of FA synthesis and the concentration of lipid-free biomass. Under nitrogen-non-limiting conditions, cells grown at lower temperatures had a higher PUFA content and the maximal productivity of -linolenic acid was obtained by shifting the temperature of the culture from 30° C to 25° C. Offprint requests to: A. Pareilleux     

Effects of altering dietary fatty acid composition on prostaglandin synthesis and fertility

Several studies over the past 20 years have demonstrated that subjects on diets composed of substances with high levels of n-3 polyunsaturated fatty acids (PUFAs) (e.g. fish) have a decreased incidence of heart disease. On this basis, a recent report from the Department of Health has advised UK consumers to decrease the proportion of saturated as opposed to unsaturated fats in their diet and to increase the ratio of n-3 to n-6 PUFAs. This could be achieved by altering the amounts of these constituents in milk and meat. n-3 Fatty acids can most easily be added to animal feed as either fish oil or linseed oil and can be increased in the blood and milk of ruminants following protection to avoid hydrogenation in the rumen. In western countries the ratio of consumption of n-6 to n-3 PUFAs is greater than 10 and current evidence tends to suggest that a ratio nearer 5 would be more desirable and compatible with cardiovascular well being. As fertility in the UK dairy herd is already poor, it is important to establish whether alterations in dietary n-3 and n-6 PUFAs affects herd fertility before widespread changes in animal diets are recommended. Therefore, this review considers the role played by PUFAs and eicosanoids in fertility, with particular reference to the implications for farm livestock production. The evidence reviewed shows that alteration of the concentration and ratio of n-6 and n-3 PUFAs in feeds can influence prostaglandin synthesis/metabolism in a number of mammalian systems. The changed patterns of prostaglandin synthesis can as a consequence, affect the diverse functions (e.g. hormone secretion) that are normally mediated via prostaglandins. Similarly, changes in prostaglandin synthesis effected through manipulation of PUFAs has a major bearing on fertility (as PGs affect many reproductive parameters, e.g. ovulation). Several studies in cattle and other mammals, show that feeding or infusing different types of fat with varying PUFA content to females can alter: the number and size of ovarian follicles, the ovulation rate, progesterone production by the corpus luteum, the timing of luteolysis and gestational length. In the male most recent work has focussed on sperm production and experiments in fowl have demonstrated clear effects of dietary PUFAs on both the sperm membrane phospholipid composition and on fertilizing ability.     

Effect of growth rate on lipid and lipoteichoic acid composition in Streptococcus faecium.

The lipid composition of Streptococcus faecium (S. faecalis ATCC 9790) was analyzed at various growth rates. Diphosphatidylglycerol and the non-ionic lipid fraction containing diacylglycerols and neutral glycolipids appeared to accumulate relative to cellular mass as the culture mass doubling time increased from 30 to 80 min. Within the same range of doubling times the non-ionic lipid fraction appeared to become substantially enriched with diacylglycerols. All lipid species and cellular lipoteichoic acid accumulated relative to the cellular mass at doubling times exceeding 80 min, although diacylglycerol accumulation exceeded that of all other compounds studied.     

镉胁迫对平邑甜茶脂肪酸构成及脂质过氧化的影响

以平邑甜茶幼苗为试材,研究了镉胁迫下幼苗叶片和根系膜脂肪酸构成、活性氧、脂氧合酶和丙二醛含量的变化.结果表明：氯化镉处理后7～12 h,脂肪酸种类及其相对含量变化最为明显.处理后7 h,叶片和根系脂肪酸不饱和水平升至最高,含量分别达8282%和7243%;叶片可检测到的脂肪酸在处理后12 h由11种增至14种,根系则在处理17 h后由4种增至6种.O₂^.-产生速率在处理3 h、H₂O₂含量在处理7 h时升至最高,丙二醛含量和脂氧合酶活性则随着处理时间的延长逐渐增加.镉胁迫通过诱导活性氧和脂氧合酶来改变平邑甜茶脂肪酸构成,并引起脂质过氧化;镉处理12 h前,脂质过氧化是活性氧和脂氧合酶的共同结果;但处理12 h后,脂质过氧化加剧主要在于脂氧合酶活性的持续增加.     

Linear growth and lipid synthesis in the oleaginous yeastRhodotorula gracilis

A kinetic analysis was made of batch cultures of an industrially important yeastRhodotorula gracilis, using a nitrogen-limited medium. The exponential phase of growth lasted 15 h, followed by a linear phase up to 36 h. The generation time was 2.8 h (15–36 h of fermentation) which corresponded to a μ of 0.248/h. The lipid synthesis was partially growth-associated with the linear growth phase (15–36 h) and the early stationary phase (36–60 h). The rate of linear growth was estimated as 0.267 g cell per L per h and the rate for lipid synthesis over the period of 15–60 h was 0.17 g lipid per L per h. More than 50% of the total supplied nitrogen was assimilated by the organism and the rest remained in the medium. Sugar assimilation was nearly 100% by the end of 60-h fermentation. At 0 h the intracellular protein was 3.3% and it significantly increased to 11.7% by the end of 12 h. Morphological and physiological characteristics were also found to change during different stages of growth.     

碳源种类及碳氮比对眼点拟微绿球藻生长密度、油脂含量和脂肪酸组成的影响

研究了三种碳源Na2CO3、NaHCO3、葡萄糖对眼点拟微绿球藻生长密度和油旨含量的影响,实验结果表明相对于葡萄糖,无机碳源NaHCO3更利于眼点拟微绿球藻的生长.以NaHCO3为碳源,研究了在不同的接种密度、NaNO3浓度下,C/N对眼点拟微绿球藻生长密度和油脂含量的影响.实验结果表明,C/N对眼点拟微绿球藻生长密度的影响与接种密度和NaNO3浓度有关,在高的NaNO3浓度时,C/N对眼点拟微绿球藻生长密度的影响很小;在低的NaNO3浓度时,随着C/N比的增加,微绿球藻的生长密度先增加后下降,存在最佳的C/N比.最佳的C/N比随接种密度而变化,在接种密度为OD440=0.10时,最佳C/N比为3,当接种密度提高到OD440=0.70时,最佳C/N比增加到5.NaNO3浓度和C/N对微藻油脂含量均有较大影响,在不同的接种密度和NaNO3浓度下都表现为C/N=1时最利于微藻油脂的积累,这与卡尔文循环过程中核酮糖-1,5-二磷酸羧化酶/加氧酶的活性有关.本实验的最佳产油培养条件为以NaHCO3为碳源,初始接种密度为OD440=0.70,C/N=1∶1,CNaNO3=0.225g/L,此时油脂产率为56.7 mg/(L·d),EPA产率为6.5 mg/(L·d).     

The effect of glycerol as a sole and secondary substrate on the growth and fatty acid composition of Rhodotorula glutinis

Rhodotorula glutinis is a yeast that produces copious quantities of lipids in the form of triacylglycerols (TAG) and can be used to make biodiesel via a transesterification process. The ester bonds in the TAG are broken leaving behind two products: fatty acid methyl esters and glycerol that could provide an inexpensive carbon source to grow oleaginous yeast R. glutinis. Described here are the effects of different growth substrates on TAG accumulation and fatty acids produced by R. glutinis. Yeast cultured 24h on medium containing dextrose, xylose, glycerol, dextrose and xylose, xylose and glycerol, or dextrose and glycerol accumulated 16, 12, 25, 10, 21, and 34% TAG on a dry weight basis, respectively. Lipids were extracted from R. glutinis culture and transesterified to form fatty acid methyl esters. The results show a difference in the degree of saturation for the carbon sources tested. Cells cultivated on glycerol alone had the highest degree of unsaturated fatty acids at 53% while xylose had the lowest at 25%. R. glutinis can be cultivated on all sugars tested as single carbon substrates or in mixtures. Glycerol may be used as secondary or primary carbon substrate.     

不同饵料对卤虫生长、总脂含量及脂肪酸组成的影响

为了提高养殖卤虫的饵料营养价值,了解其不同生长阶段营养成分变化情况,采用单因子试验研究了8种饵料（三角褐指藻、小球藻、微绿球藻、酵母液、三角褐指藻＋小球藻＋微绿球藻、三角褐指藻＋酵母液、小球藻＋酵母液和微绿球藻＋酵母液）对卤虫生长、总脂含量及脂肪酸组成的影响,结果表明：不同饵料种类对卤虫生长、总脂含量及脂肪酸组成的影响显著（P＜0.05）,增长率,以三角褐指藻＋酵母液最优;总脂含量、以三角褐指藻最优（19.67%）,除酵母液外,与其它饵料相差不显著（P＞0.05）;脂肪酸组成效果,以微绿球藻组最优（EPA：18.01%,DNA：0.55%,（n-3）HUFA：19.08%）,与三角褐指藻组相差不大（P＞0.05）,显著高于其它各组（P＞0.05）.同时以三角褐指藻为饵料,研究了卤虫不同生长阶段（体长2、4、6、8、10 mm）总脂含量、脂肪酸组成变化,结果表明：卤虫体长2～10 mm总脂含量为14.27%～20.93%,随体长的增长降低;EPA、DHA及（n-3）HUFA的含量,均随体长的增长降低,EPA含量为：10.47%～20.77%,DNA含量为：0～0.70%,（n-3）HUFA含量为：10.85%～22.01%.结论认为,卤虫以三角褐指藻或三角褐指藻＋酵母液为饵料培养营养价值最佳,其体长小于6 mm营养价值较佳.     

Effects of nitrogen source and concentration on growth rate and fatty acid composition of Ellipsoidion sp. (Eustigmatophyta)

In order to study the effects of different nitrogen source and concentrationon the growth rate and fatty acid composition, a marine microalga Ellipsoidion sp. with a high content of eicosapentaenoic acid (EPA) wascultured in media with different nitrogen sources and concentrations.During the pre-logarithmic phase, the alga grew faster with ammoniumas N source than with nitrate, but the reverse applied during thepost-logarithmic phase. The alga grew poorly in N-free mediumor medium with urea as the sole N source. In the same growth phase,ammonium medium resulted in higher yield of total lipid, but the EPA yielddid not differ significantly different from that using nitrate medium. Themaximum growth rate occurred in medium containing 1.28 mmolL^-1 sodium nitrate, while maximum EPA and total lipid contents werereached at 1.92 mmol L^-1, when EPA accounted for 27.9% totalfatty acids. The growth rate kept stable when NH₄Cl ranged from0.64 to 2.56 mmol L^-1, and the maximum content of total lipidand EPA occurred in the medium with 2.56 mmol L^-1NH₄Cl. The EPA content was higher in the pre- thanpost-logarithmic phase, though the total lipid content was lower. Thehighest EPA content expressed as percent total fatty acid was 27.9% innitrate medium and and 39.0% in ammonium medium.     

Effects of hyperoxia on composition and rate of synthesis of fatty acids in Escherichia coli

Growth of and fatty acid synthesis in Escherichia coli were inhibited by oxygen at partial pressures above 1 atm and were prevented by exposure to oxygen at 4.2 atm on membranes incubated on a minimal medium. Growth and fatty acid synthesis returned to control rates when cells were removed from hyperoxia to air. The spectrum of fatty acids produced was unchanged by oxygen at pressures which reduced the rate of synthesis. In situ fatty acids were stable to oxygen at pressures which prevented growth and synthesis. Reinitiation of synthesis after complete inhibition in hyperoxia occurred without production of aberrant fatty acids. Fatty acid synthetase specific activity was virtually unchanged, compared with air controls, in cells exposed either to 3.2 or to 15.2 atm of oxygen. The spectrum of fatty acids synthesized by cell-free extracts during incubation in 4.2 atm of oxygen was not different from air-incubated controls. Synthetase assays included added NADPH, acyl carrier protein, mercaptoethanol, and malonyl coenzyme A; hence, damage, other than reversible sulfhydryl oxidation, to the apoenzymes of synthetase was ruled out.     

Effects of unsaturated fatty acid deprivation on neutral lipid synthesis in Saccharomyces cerevisiae

The effects of unsaturated fatty acid deprivation on lipid synthesis in Saccharomyces cerevisiae strain GL7 were determined by following the incorporation of [14C]acetate. Compared to yeast cells grown with oleic acid, unsaturated fatty acid-deprived cells contained 200 times as much 14C label in squalene, with correspondingly less label in 2,3-oxidosqualene and 2,3;22,23-dioxidosqualene. Cells deprived of either methionine or cholesterol did not accumulate squalene, demonstrating that the effect of unsaturated fatty acid starvation on squalene oxidation was not due to an inhibition of cell growth. Cells deprived of olefinic supplements displayed additional changes in lipid metabolism: (i) an increase in 14C-labeled diacylglycerides, (ii) a decrease in 14C-labeled triacylglycerides, and (iii) increased levels of 14C-labeled decanoic and dodecanoic fatty acids. The changes in squalene oxidation and acylglyceride metabolism in unsaturated fatty acid-deprived cells were readily reversed by adding oleic acid. Pulse-chase studies demonstrated that the [14C]squalene and 14C-labeled diacylglycerides which accumulated during starvation were further metabolized when cells were resupplemented with oleic acid. These results demonstrate that unsaturated fatty acids are essential for normal lipid metabolism in yeasts.     

Effect of hydrogen peroxide on d-amino acid oxidase from Rhodotorula gracilis

D-amino acid oxidase from Rhodotorula gracilis is a FAD-containing enzyme that belongs to the oxidase class that is characterized by the ability of the reduced flavin to react quickly with oxygen, yielding hydrogen peroxide and the oxidized cofactor. Hydrogen peroxide, necessary for the production of glutaryl-7-ACA from cephalosporin C had a deleterious effect on the enzyme. H(2)O(2) induced the oxidation of tryptophan and cysteine residues of the protein that could be involved in the dimerization process, required for the attainment of a fully competent enzyme. H(2)O(2) had also a kinetic effect on the reaction catalyzed by D-amino acid oxidase. It was a pure noncompetitive inhibitor; the corresponding inhibition constants were K(is) = 0.52 mM and K(ii) = 0.70 mM.     

Lipid growth requirement and influence of lipid supplement on fatty acid and aldehyde composition of Syntrophococcus sucromutans

Results concerning the ruminal fluid growth requirement of the ruminal acetogen, Syntrophococcus sucromutans, indicate that octadecenoic acid isomers satisfy this essential requirement. Complex lipids, such as triglycerides and phospholipids, can also support growth. The cellular fatty acid and aldehyde composition closely reflects that of the lipid supplement provided to the cells. Up to 98% of the fatty acids and 80% of the fatty aldehydes are identical in chain length and degree of unsaturation to the octadecenoic acid supplement provided in the medium. S. sucromutans shows a tendency to have a greater proportion of the aldehyde form among its 18 carbon chains than it does with the shorter-chain simple lipids, which may be interpreted as a strategy to maintain membrane fluidity. 14C labeling showed that most of the oleic acid taken up from the medium was incorporated into the membrane fraction of the cells.     

Brugia malayi: microfilarial polyunsaturated fatty acid composition and synthesis

The polyunsaturated fatty acid composition of Brugia malayi microfilariae was analyzed by gas chromatography and compared to that of sera from B. malayi-infected jirds. The essential fatty acid, linoleic acid (18:2 omega 6), was the most abundant fatty acid present in both microfilarial total lipids and phospholipids as well as in jird sera. In contrast, arachidonic acid (20:4 omega 6), as well as the 18:3 omega 6, 20:2 omega 6, and 20:3 omega 6 intermediates that are formed in the enzymatic conversion of linoleic acid to arachidonic acid, were proportionally more abundant in microfilariae than in jird sera. To assess the capacity of microfilariae to transform linoleic acid into arachidonic acid, B. malayi microfilariae were incubated with [14C]linoleic acid. Microfilarial lipids were extracted and resolved by high-pressure liquid chromatography and thin-layer chromatography. A portion of [14C]linoleic acid incorporated by microfilariae was converted to [14C]arachidonic acid. Thus, microfilariae can not only incorporate exogenous arachidonic acid, as previously demonstrated, but can also synthesize arachidonic acid from exogenous linoleic acid. The capacity of microfilariae to utilize specific host polyunsaturated fatty acids raises the possibility that intravascular filarial parasites may synthesize eicosanoid metabolites of arachidonic acid that could mediate filarial-host cell interactions.     

Ethanol-induced changes in lipid composition of Escherichia coli: Inhibition of saturated fatty acid synthesis in vitro

Growth of Escherichia coli in the presence of ethanol results in the synthesis of lipids containing elevated proportions of unsaturated fatty acids. Previous in vivo experiments indicated that the ethanol-induced changes in fatty acid composition result from a preferential inhibition of saturated fatty acid synthesis. In this study, the inhibition of saturated fatty acid synthesis by ethanol was confirmed in vitro. This inhibition was not membrane mediated and resulted from a direct action of ethanol on the soluble enzymes of fatty acid synthesis. The addition of ethanol resulted in a decrease in chain length of both saturated and unsaturated acyl products in vitro. Experiments with enzymes prepared from several fatty acid synthesis mutants of E. coli indicate that β-hydroxydecanoyl-acyl carrier protein dehydrase is not the site of the ethanol inhibition of saturated fatty acid synthesis. The two condensing enzymes are the probable sites for inhibition by ethanol.