Inhibitory effect of medium-chain-length fatty acids on synthesis of polyhydroxyalkanoates from volatile fatty acids by Ralstonia eutropha

Medium-chain-length fatty acids, such as nonanoic (9:0) and octanoic (8:0) acids, are more toxic to Ralstonia eutropha than volatile fatty acids such as acetic, propionic and butyric acids. Nonanoic acid was degraded to acetic and propionic acids via -oxidation by Ralstonia eutropha for cell growth and synthesis of polyhydroxyalkanoates (PHAs). In a mixture of the fatty acids, utilization of nonanoic acid was depressed by acetic and propionic acids, and vice versa. The PHA accumulation from the volatile fatty acids was decreased from 53% (w/w) of dry cell mass to 23% due to the nonanoic acid. Similar phenomena were also observed with octanoic acid and its metabolic intermediates, acetic and butyric acids.     

Cellular lipid accumulation by Pseudomonas aeruginosa 44T1

Summary Growth of Pseudomonas aeruginosa strain 44T1 on glucose, an n-alkane mixture or olive oil was characterized by the formation of intracellular lipid inclusions and extracellular accumulation of rhamnolipids. Maximum values of cellular lipid accumulation were obtained in olive-oil-grown cells and reached up to 38% w/w of its dry biomass. The principal fatty acids of cellular lipids drived from P. aeruginosa cultures varied with the carbon source employed. The major fatty acids detected were palmitic and trans-oleic acids. Arachidonic acid was only found in medium containing glucose or the n-alkane mixture. Offprint requests to: A. Manresa     

Changes of chemical and nutrient composition of porcine blood during fermentation by <Emphasis Type="Italic">Aspergillus oryzae</Emphasis>

A solid-state fermentation (SSF) of a mixture of porcine blood and wheat bran with a ratio of 8:1 by Aspergillus oryzae was investigated. Water content, pH, crude protein, heme and total iron, free amino acids (FAA) and total fatty acids (TFA) of the fermented mixture were determined at 0, 40, 80 and 120 h, respectively, and protein hydrolysis were analyzed with SDS-PAGE accordingly. The results showed that, during the fermentation, water contents and pH decreased significantly (P < 0.05) from 68.33% to 59.94%, 7.18% to 4.48%, respectively. Heme iron content changed slightly (P > 0.05). With the degradation of large protein molecules, free amino acids in the mixture increased from 872.83 mg l⁻¹ to 11560.94 mg l⁻¹ (P < 0.05). Content of free isoleucine, methionine and cystine, deficient in fresh porcine blood increased (P < 0.05) after fermentation. Percentages of saturated fatty acids such as C14:0, C17:0 and C18:0 in the total fatty acids decreased significantly (P < 0.05), and those of polyunsaturated fatty acids C18:1, C18:2, C18:3, C20:1 and C20:2 increased significantly from 27.06% to 47.90% (P < 0.05). The results indicated that Aspergillus oryzae could ferment porcine blood and bran mixture and change its chemical and nutrient composition.     

Trapping of 2,7-Dichlorodibenzo-<Emphasis Type="Italic">p</Emphasis>-Dioxin in Aqueous Solution by Enzymatic Reaction of Fungal Manganese Peroxidase in the Presence of Polyunsaturated Fatty Acids

Abstract:In the presence of polyunsaturated fatty acids, including cis-4,7,10,13,16,19-docosahexaenoic acid (DHA), 2,7-dichlorodibenzo-p-dioxin (DCDD) was treated with manganese peroxidase (MnP) from white rot basidiomycete Phanerochaete sordida YK-624. After incubation with MnP, DCDD could not be extracted from the reaction mixture with n-hexane and was trapped in the water layer. DCDD was released by alkalification of the water layer. DCDD was also trapped after treatment with lipoxidase, which produces hydroperoxides from unsaturated lipids. The amounts of thiobarbituric acid-reactive substances produced in the MnP reactions with three highly unsaturated fatty acids were higher than the amounts produced with three fatty acids with a lower degree of unsaturation. These results suggest that a DCDD-trapping compound may be produced by peroxidation of the polyunsaturated fatty acids.Received: 22 October 2002 / Accepted: 6 December 2002     

Fatty Acid Specificity in Lipase-Catalyzed Synthesis of Glucoside Esters

The fatty acid specificity of the B-lipase derived from Candida antarctica was investigated in the synthesis of esters of ethyl D-glucopyranoside. The specificity was almost identical with respect to straight-chain fatty acids with 10 to 18 carbon atoms. However, lower fatty acids such as hexanoic and octanoic acid and the unsaturated 9-cis-octadecenoic acid were found to be poor substrates of the enzyme. As a consequence of this selectivity, these fatty acids were accumulated in the unconverted fraction when ethyl D-glucopyranoside was esterified with an excess of a mixture of fatty acids. This accumulation can reduce the overall effectiveness of the process as the activity of the lipase was found to be reduced when exposed to high concentrations of short-chain fatty acids. Finally, using a simplified experimental set-up, the specificity of the C. antarctica B-lipase was compared to the specificity of lipases derived from C. rugosa, Mucor miehei, Humicola, and Pseudomonas. Apart from the C. rugosa lipase, which exhibited a very poor performance, all the enzymes showed a very similar specificity with respect to fatty acids longer than octanoic acid while only the C. antarctica B-lipase showed activity towards sort-chain fatty acids.     

Aptitude of cheese bacteria for volatile S-methyl thioester synthesis I. Effect of substrates and pH on their formation by Brevibacterium linens GC171

The aptitude of resting cells of Brevibacterium linens G171 to synthesize S-methyl thioesters was studied in presence of methanethiol and nine short-chain fatty acids individually or as a mixture. Esterification of acetic, propionic and methyl branched-chain acids occurred with methanethiol alone and was enhanced by fatty acid addition. Addition of n-chain, 3-hydroxybutyric and 2-hydroxyvaleric acids allowed synthesis of n-chain thioesters up to thiocaproate. The kinetics of production and the effect of concentrations of both substrates and of cells were tested. The optimum pH for synthesis varied according to the kind of thioesters produced. Results suggested that thioesters were derived mainly from acyl-CoA from different metabolic breakdowns, such as the degradation of fatty acids or some amino acids, and that several acyltransferases could be involved. Received: 1 August 1996 / Received revision: 10 October 1996 / Accepted: 18 October 1996     

Isolation and Characterization of a Neutral Glycosphingolipid from the Epimastigote Form of Trypanosoma mega1

ABSTRACT. A glycosphingolipid fraction from Trypanosoma mega was isolated after acetylation and was further purified on a silicic acid column. Final purification was by preparative thin-layer chromatography. The carbohydrate components of the glycolipid were fucose and galactose in approximately equimolar amounts. The neutral glycolipid of T. mega has a sphingosine base composition that consists of sphingosine and traces of dihydrosphingosine. Fatty acids forming amide groups with the sphingosine bases were analyzed by gas-liquid chromatography-mass spectrometry and are a mixture of normal and α-hydroxy fatty acids. Normal C_16:0, C_18:0, and 2-hydroxy C_18:0 are the predominant fatty acids.     

Lipid Auxotrophy and the Effect on Lipid Composition of Entamoeba invadens*†

SYNOPSIS. A medium for the axenic cultivation of Entamoeba invadens has been developed. Serum, an essential constituent of conventional media, has been replaced by a mixture of albumin, unsaturated fatty acids, Tween, and cholesterol to control the lipid composition of the medium. Entamoeba invadens requires both cholesterol and unsaturated fatty acids for growth. The fatty acid composition of the phospholipids of the ameba reflects that of the medium to a great extent, especially with regard to the unsaturated fatty acids. The amount of membrane bounded cholesterol depends on the cholesterol concentration in the medium.     

Initiation of anaerobic growth of Saccharomyces cerevisiae by amino acids or nucleic acid bases: ergosterol and unsaturated fatty acids cannot replace oxygen in minimal media

Nine out of ten industrially important strains of Saccharomyces cerevisiae did not grow in minimal media under anaerobic conditions even when ergosterol and unsaturated fatty acids were provided. Anaerobiosis was maintained either by flushing the culture flasks with prepurified nitrogen or by incubating the flasks in an anaerobic chamber. Traces of oxygen present in ‘prepurified nitrogen gas’ were sufficient to initiate yeast growth and on removal of the oxygen by catalytic means the yeasts failed to grow. The yeast grew very well anaerobically if the medium was supplemented with a mixture of amino acids or with a mixture of purines and pyrimidines. The growth initiated by including a mixture of amino acids was further enhanced when the medium was supplemented with ergosterol and an unsaturated fatty acid. Since no oxygen requirement for the synthesis of amino acids or purines and pyrimidines has been demonstrated, growth promotion by these compounds under anaerobic conditions is most likely not by eliminating the need for oxygen for their synthesis. We suggest that the amino acids and the nucleic acid bases yielded, through some hitherto unknown reactions, small amounts of a molecular or usable form of oxygen which allowed key reactions essential for ‘anaerobic’ growth to proceed. Received 16 July 1998/ Accepted in revised form 8 October 1998     

Detection of DNA Damage in Cultured Human Fibroblasts Induced by Methyl Linoleate Hydroperoxide

To establish a strategy to generate N-acylated proteins modified with fatty acids having a specific chain length, tGelsolin-streptag, an epitope-tagged model protein having an N-myristoylation motif, was synthesized using an insect cell-free protein synthesis system in the presence of acyl-CoA with various fatty acid chain lengths. It was found that the fatty acid species attached to the N-termini fully depended on the acyl-CoA species added to the reaction mixture. N-Acylated proteins with fatty acid chain lengths of 8, 10, 12, and 14 were generated successfully.     

Cerulenin inhibits de novo sophorolipid synthesis of Candida bombicola

Synthesis of medium-chain sophorolipids by Candida bombicola is a challenging objective. One of the difficulties is that the obtained sophorolipids always represent a mixture of medium-chain and native de novo formed or long-chain sophorolipids. The fatty acid moiety of de novo sophorolipids is derived from the de novo synthesis of fatty acids. Fatty acid synthesis can be blocked by the antifungal agent cerulenin, an inhibitor if the fatty acid synthase (FAS) complex acting on the β-ketoacyl thioester synthetase reaction. The toxic effect of cerulenin on C. bombicola was evaluated and 20 mg/ml was added in the stationary growth phase. No de novo formed sophorolipids were observed when the cells were cultured on merely glucose. Also when the hydrophilic substrate, 1,12-dodedanediol, was added, no de novo formed sophorolipids were detected, leading to a reduced complexity of the sophorolipid mixture.     

Simultaneous conversion of free fatty acids and triglycerides to biodiesel by immobilized Aspergillus oryzae expressing Fusarium heterosporum lipase

The presence of high levels of free fatty acids (FFA) in oil is a barrier to one‐step biodiesel production. Undesirable soaps are formed during conventional chemical methods, and enzyme deactivation occurs when enzymatic methods are used. This work investigates an efficient technique to simultaneously convert a mixture of free fatty acids and triglycerides (TAG). A partial soybean hydrolysate containing 73.04% free fatty acids and 24.81% triglycerides was used as a substrate for the enzymatic production of fatty acid methyl ester (FAME). Whole‐cell Candida antarctica lipase B‐expressing Aspergillus oryzae, and Novozym 435 produced only 75.2 and 73.5% FAME, respectively. Fusarium heterosporum lipase‐expressing A. oryzae produced more than 93% FAME in 72 h using three molar equivalents of methanol. FFA and TAG were converted simultaneously in the presence of increasing water content that resulted from esterification. Therefore, F. heterosporum lipase with a noted high level of tolerance of water could be useful in the industrial production of biodiesel from feedstock that has high proportion of free fatty acids.     

Fatty Acid Compositions of Triglyceride and Phospholipid from Candida tropicalis Grown on n-Alkanes

The content of total cellular lipid of Candida tropicalis grown on a mixture of n-alkanes (C₁₀–C₁₈) was about 20% of the dry cell weight at the exponential growth phase and 14% at the early stationary phase. Phospholipid corresponded to approximately 70 % of the total lipid independent of the growth phases. The composition of cellular lipid classes did not change significantly during the growth. On the other hand, a drastic time-course change in fatty acid composition was observed. The proportion of odd-chain fatty acids, one of the most specific cellular components of the yeast grown on the n-alkane mixture, increased in both phospholipid and triglyceride along with the yeast growth. In the meantime, the proportion of polyunsaturated fatty acids varied markedly during the course of cultivation, showing a peak at the early growth phase. The high content of polyunsaturated fatty acids at the early stages of growth correlated to the contents of these acids in phospholipid rather than in triglyceride.     

ALLELOCHEMICALS OF BOTRYOCOCCUS BRAUNII (CHLOROPHYCEAE)1

The alga Botryococcus braunii Kützing (Chlorophyceae) present in Liyu Lake (Huanlien County, Taiwan) has toxic effects on a variety of aquatic organisms. Blooms of this alga, which typically occur in autumn, are associated with fish deaths in this lake. Experiments using 15 phytoplankton and 5 zooplankton isolated from Liyu Lake indicate that these plankton exhibit various susceptibilities to B. braunii. A close correlation between the degree of susceptibility tested in the laboratory and the absence of certain phytoplankton during B. braunii blooms in the lake was observed, suggesting allelopathic effects. Isolation, identification, and verification with authentic compounds indicated that allelochemicals were a mixture of free fatty acids, including α‐linolenic, oleic, linolic, and palmitic acids. Compared with other phytoplankton isolates, B. braunii produced significantly higher amounts of free fatty acids, particularly of oleic and α‐linolenic acids. The role of these fatty acids in favoring dominance of B. braunii in the natural environment was elucidated.     

Oviposition deterrents from the egg masses of the adzuki bean borer, Ostrinia scapulalis and Asian corn borer, O. furnacalis

Bioassays using gravid females of the adzuki bean borer, Ostrinia scapulalis (Walker), and the Asian corn borer, O. furnacalis (Guenée) (Lepidoptera: Crambidae), showed that the presence of an egg mass of a conspecific deters oviposition. Volatile chemicals emanating from the egg mass were responsible for the deterrence, and these deterrents could be extracted from the egg mass with hexane. When fractionated using a Sep‐Pak® Plus NH₂ cartridge, the deterrents were eluted with a 98 : 2 mixture of diethyl ether and acetic acid (polar lipid fraction). The polar lipid fraction contained free fatty acids with 14–20 carbons, and palmitic acid, palmitoleic acid, and oleic acid were predominant. A blend of all identified fatty acids, a blend of six major fatty acids (palmitic, palmitoleic, stearic, oleic, linoleic, and linolenic acids), a blend of the two Z‐9‐alkenoic acids (palmitoleic and oleic acids), palmitoleic acid alone, and oleic acid alone showed deterrence against O. scapulalis which was comparable to that provoked by the full egg extract. The dose‐dependency of the deterrent effects of palmitoleic acid and oleic acid was verified in O. scapulalis. The binary blend of palmitoleic acid and oleic acid was also confirmed to deter oviposition in O. furnacalis.     

Secondary metabolites and lipids in Chara globularis Thuill

Sterols, volatiles and lipid fatty acids were analysed in the fresh-water alga Chara globularis. Five sterols, two of them new for the genus, were identified, together with 12 fatty acids from lipids. The volatiles appeared to form a complex mixture, containing mainly acids (part of them chlorinated), terpenoids, ketones, hydrocarbons, etc. No flavonoids have been found. Antibacterial activity of extracts has been tested, unpolar extracts showed moderate activity.     

The influence of carbohydrate dissimilation on the fatty acid metabolism of Monascus purpureus

The formation of methyl ketones from fatty acids is known as a detoxifying mechanism and takes place during the lag period of growth. Fermentations of fatty acid and saccharides mixture led to two-phase growth kinetics. It was found that first the fatty acid is oxidized before saccharide dissimilation and growth occurs. This principle can also be observed in fermentation processes with precursor mixtures. Precursor-induced breakdown happens due to the toxicity of fatty acids and methyl ketones. Thereby the fatty acid is metabolized prior to the methyl ketone. After this, saccharide dissimilation and growth take place. Correspondence to: N. Peters     

Biosurfactants from Bacillus licheniformis: structural analysis and characterization

Summary The surface-active compounds of the strain Bacillus licheniformis were isolated and their structure was elucidated. The high surface-active capacity of the crude extract was basically due to traces of long-chain saturated fatty acids, especially of palmitic and stearic acids, to a mixture of small amounts of hydrocarbons with chain lengths of 20 and 22 carbons, and mainly to appreciable amounts of four slightly different lipopeptides. The lipopeptides were found to be a mixture of four closely related compounds. The lipophilic part consisting of i-, n-C₁₄ or i-, ai-C₁₅ -OH fatty acids was linked to the hydrophilic peptide moiety, which contained seven amino acids (Glu, Asp, Val, three Leu and Ile) by a lactone linkage. Fifteen milligrams per litre of the purified lipopeptide product decreased the surface tension of water from 72 mN m^–1 to 27 mN m^–1, characterizing the product as a powerful surface-active agent that compares favourably to other (bio)surfactants. Antibiotic activity was demonstrated against bacteria and yeasts. Offsprint requests to: O. Käppeli     

Acidolysis between Triolein and Short-Chain Fatty Acid by Lipase in Organic Solvents

Ten kinds of lipases were examined as biocatalysts for the incorporation of short-chain fatty acids (acetic, propionic, and butyric acids) into triolein in order to produce one kind of reduced-calorie structured lipids. Trans-esterification (acidolysis) was successfully done in n-hexane by several microbial lipases. Among them, lipase from Aspergillus oryzae was used to investigate the effects of incubation time, substrate molar ratio, and water content on acidolysis. Finally, more than 80% of triolein was incorporated by butyric acid (molar ratio of triolein to butyric acid, 1:10) in the dried n-hexane at 52 °C for 72 h. More than 90% of the products was monosubstituent, which was esterified with this short chain fatty acid at the 1-position of the glycerol moiety of triolein. These results suggest that A. oryzae lipase would be a powerful biocatalyst for the synthesis of low caloric oil, such as triacylglycerol containing a mixture of long- and short-chain aliphatic acids.     

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.