Добірка наукової літератури з теми "Acides gras cyclopropanes"

Examination of seven strains of gram-negative bacteria isolated from the rhizosphere of the rape plant showed profiles of fatty acids, ester-linked to their phospholipids that were distinctly different from sterile roots. The bacteria were enriched in short and branched saturated, cyclopropane and monoenoic fatty acids in which the unsaturation was formed by the anerobic desaturase pathway when growth was on a medium simulating root exudates. This suite of fatty acids was significantly increased in sand and in roots of rape plants grown from seeds in sands inoculated with these organisms at the start of the experiments. Some of the bacteria formed the endogenous storage polymer poly β-hydroxybutyrate in monocultures. Poly β-hydroxybutyrate was formed by the bacteria in the inoculated sand in large amounts but not by the bacteria that were recovered from the roots. This suggests that bacteria associated with the roots were in balanced growth, whereas the bacteria in the sand were lacking essential nutrients and showed an unbalanced growth response. These chemical analyses provided estimates of the bacterial biomass (from the sum of bacterial ester-linked phospholipid fatty acids) and the metabolic status (poly β-hydroxybutyrate to phospholipid fatty acid ratio) in this experiment. With further definition of lipid biomarkers of soil microorganisms these experimental techniques may be extended to the rhizosphere.

The effect of NaCl concentration on the growth rate and phospholipid composition of a moderately halophilic Gram-negative rod-shaped bacterium has been studied. This bacterium, designated HX, grows between 0.5 and 4.25 M NaCl at 20, 30, and 37 °C. It has a broad optimum salt concentration for growth of 1.0–2.0 M NaCl at 20 and 30 °C and 1.5–2.8 M NaCl at 37 °C. Its major lipids are phosphatidylethanolamine and phosphatidylglycerol, which constitutes >90% of the total lipid, together with smaller amounts of diphosphatidylglycerol; there are traces of an unidentified glycolipid, lysophosphatidylethanolamine, and phosphatidylserine. Lipid composition depends on NaCl concentration of the culture medium: at higher salinities the proportion of anionic phospholipids, mainly phosphatidylglycerol, is greater relative to the proportion of zwitterionic phospholipids, mainly phosphatidylethanolamine. The major fatty acids are 16:0, 16:1, 18:0, 18:1, 17:0cyc, and 19:0cyc; the proportions of the cyclopropane fatty acids are greater in cultures grown at higher salinities and this increase is due to NaCl concentration, not growth rate. Following a sudden increase in NaCl concentration of the culture medium (shift up), there are concomitant changes in growth rate and phospholipid composition. A lag period in growth is observed only when the magnitude of the salinity shift up is at least threefold. After a salinity shift up, there is a rapid decrease in the ratio of phosphatidylethanolamine to phosphatidylglycerol. It appears that the alterations in growth rate and lipid composition are a physiological response for adaptation to a change in salinity.Key words: halophilic bacteria, haloadaptation, membranes, lipids, salinity.

A novel Gram-positive, catalase negative, rod-shaped strain, FI11369T, was isolated from gari, a traditional West African fermented food derived from cassava. Based on 16S rRNA gene sequence similarity, the closest type strains were Lactobacillus xiangfangensis LMG 26013T (99.4 % similarity), Lactobacillus plajomi NBRC 107333T (99.1 %), Lactobacillus paraplantarum DSM 10667T (99.1 %), Lactobacillus pentosus DSM 20314T (99.0 %), Lactobacillus plantarum subsp. plantarum ATCC 14917T (99.0 %), Lactobacillus modestisalitolerans NBRC 107235T (98.9 %), Lactobacillus plantarum subsp. argentoratensis DSM 16365T (98.9 %) and Lactobacillus daowaiensis NCIMB 15183T (98.8 %). The genome of strain FI11369T was sequenced and the average nucleotide identity (ANI) was compared with its closest relatives. ANI analysis showed that the closest relative, L. xiangfangensis DSM 27103T, had only a 82.4 % similarity. The main fatty acids of FI11369T were saturated C16 : 0 (18.2 %), unsaturated C18 : 1 ω9c (43.8 %) and cyclopropane C19 : 0 cyclo (ω10c and/or ω6; 22.5 %). Based on the genotypic and phenotypic data obtained in this study, a novel Lactobacillus species, Lactobacillus garii sp. nov., with the type strain FI11369T (=NCIMB 15148=DSM 108249), is proposed.

The Montreal Biodome operates a methanol-fed denitrification system that treats the water in its three million litre marine mesocosm. An unknown bacterium, named strain NL21T, was isolated from this system on TSA and R2A agar. The organism is a Gram-negative, rod-shaped (1×3 μm) facultative aerobe. Optimal growth conditions on R2A agar are 30–35 °C, pH 7–7·5 and 1 % (w/w) NaCl. Phylogenetic analysis of the 16S rDNA sequence reveals that strain NL21T forms a novel lineage in the family ‘Phyllobacteriaceae’ within the α2 subgroup of the Proteobacteria. The closest related genera are Aminobacter, Pseudaminobacter, Mesorhizobium and Defluvibacter. Major cellular fatty acids are C18 : 1 ω7c (75 %), C19 : 0 ω8c cyclopropane (9·4 %) and C18 : 0 (4·2 %). The DNA G+C content of strain NL21T (57 mol%) differs from those of all other described members of the ‘Phyllobacteriaceae’ (60–64 mol%). Strain NL21T reduces nitrate to nitrite, but does not reduce nitrite to nitrogen gas. Only a few sugars and amino acids can serve as carbon sources. Strain NL21T is able to grow without salt and tolerates up to 5 % NaCl. Phylogenetic analysis, as well as physiological and biochemical tests, showed that strain NL21T was different from all other members of the ‘Phyllobacteriaceae’ with validly published names. Strain NL21T therefore represents a novel genus, for which the name Nitratireductor aquibiodomus gen. nov., sp. nov. is proposed, with the type strain NL21T (=DSM 15645T=ATCC BAA-762T).

With the aim to introduce fragment of cyclopropane and fragments of p-, m-, o-fluorophenyls into the structures of N-ethoxyethylpiperidines, acylation of oxime and phenylacetylenic alcohol of 1-(2-ethoxyethyl)-4-ketopiperidine by cyclopropanecarbonylchloride was carried out; on the basis of 1-(2-ethoxyethyl)-4-ethynyl-4-hydroxypiperidine (cascaine alcohol), acylation by 4-fluoro-, 3-fluoro-, 2-fluorobenzoylchlorides was carried out with formation of the corresponding piperidine containing hydrochlorides of cyclopropanecarboxylic acid esters and para-, meta-, ortho-fluorobenzoic esters. Acylation reaction on the hydroxyl group of compounds is carried out in absolute dioxane, the acylating agents are cyclopropanecarbonylchloride, p-, m-, o-fluorobenzoyl chlorides taken in excess. The obtained esters of cyclopropanecarboxylic and para-, meta-, ortho-fluorobenzoic acids are crystalline substances with a clear melting point, well soluble in water, ethanol, acetone. P-fluorobenzoates are obtained with better yields, m-fluorobenzoates occupy an intermediate position, and o-fluorobenzoates are formed with the lowest yields. The best yields of fluorobenzoates are obtained using dioxane as a solvent. Para-, meta-, ortho-fluorobenzoic esters of 1-(2-ethoxyethyl)-4-ethynyl-4-hydroxypiperidine coded A-4 – A-6 were studied for the presence of antimicrobial activity, the actions of these preparations were evaluated in vitro in relation to strains of gram-positive bacteria Staphylococcus aureus, Bacillus subtilis, gram-negative strains of Escheriсhia coli, Pseudomonas aeruginosa and to yeast fungus Сandida albicans by the diffusion method into agar (holes). Introduction of fluorine atom into the structure of cascaine lead to manifestation of antimicrobial activity.

Two Gram-positive, rod-shaped, moderately halophilic bacteria were isolated from a deep-sea carbonate rock at a methane cold seep in Kuroshima Knoll, Japan. These bacteria, strains IS-Hb4T and IS-Hb7T, were spore-forming and non-motile. They were able to grow at temperatures as low as 9 °C and hydrostatic pressures up to 30 MPa. Based on high sequence similarity of their 16S rRNA genes to those of type strains of the genus Halobacillus, from 96.4 % (strain IS-Hb7T to Halobacillus halophilus NCIMB 9251T) to 99.4 % (strain IS-Hb4T to Halobacillus dabanensis D-8T), the strains were shown to belong to this genus. DNA–DNA relatedness values of 49.5 % and 1.0–33.0 %, respectively, were determined between strains IS-Hb4T and IS-Hb7T and between these strains and other Halobacillus type strains. Both strains showed the major menaquinone MK7 and l-orn–d-Asp cell-wall peptidoglycan type. Straight-chain C16 : 0, unsaturated C16 : 1 ω7c alcohol and C18 : 1 ω7c and cyclopropane C19 : 0 cyc fatty acids were predominant in both strains. The DNA G+C contents of IS-Hb4T and IS-Hb7T were respectively 43.3 and 42.1 mol%. Physiological and biochemical analyses combined with DNA–DNA hybridization results allowed us to place strains IS-Hb4T (=JCM 14154T=DSM 18394T) and IS-Hb7T (=JCM 14155T=DSM 18393T) in the genus Halobacillus as the respective type strains of the novel species Halobacillus profundi sp. nov. and Halobacillus kuroshimensis sp. nov.

A novel lactic acid-producing, Gram-stain-positive, catalase-negative and rod-shaped strain, designated as strain C06_No.73T, was isolated from a traditional Japanese fermented beverage called kôso. According to the results of phylogenetic analysis based on 16S rRNA gene sequences, strain C06_No.73T belongs to the genus Lentilactobacillus . The closest type strain was Lentilactobacillus curieae CCTCC M 2011381T, with a sequence identity of 98.1 %. The identity values with other strains were all below 97 %. The isolate propagated under the conditions of 18–39 °C (optimum, 27 °C for 48 h incubation) and pH 4.0–7.0 (optimum, pH 6.5). The G+C content of its genomic DNA was determined to be 37.9 mol%. The main fatty acids were C16 : 0, C18 : 1 ω7c, C18 : 1 ω9c and C19 : 0 cyclopropane 11,12. The major polar lipid was identified as phosphatidylglycerol. No isoprenoid quinone was detected. The predominant cell-wall amino acids were lysine, alanine, glutamic acid and aspartic acid. Neither meso-diaminopimelic acid nor ornithine were detected. On the basis of this polyphasic taxonomic study, the isolate is concluded to represent a novel species, for which the name Lentilactobacillus kosonis sp. nov. is proposed. The type strain is C06_No.73T (=NBRC 111893T=BCRC 81282T).

Enterococcus faecalis , a multi-antibiotic-resistant Gram-positive bacterium, has emerged as a serious nosocomial pathogen. Here, we used a genetic approach to characterize the strategies used by E. faecalis to fulfill its requirements for endogenous fatty acid (FA) synthesis in vitro and in vivo . The FA synthesis (FASII) pathway is encoded by two operons and two monocistronic genes. Expression of all these genes is repressed by exogenous FAs, which are incorporated in the E. faecalis membrane and modify its composition. Deletion of nine genes of the 12-gene operon abolished growth in a FA-free medium. Addition of serum, which is lipid-rich, restored growth. Interestingly, the E. faecalis membrane contains cyclic fatty acids that modify membrane properties, but are unavailable in host serum. The cfa gene that encodes the cyclopropanation process, is located in a locus independent of the FASII genes. Its deletion did not alter growth under the conditions tested, but yielded bacteria devoid of cyclic FAs. No differences were observed between mice infected with wild-type, or FASII or cyclopropanation mutant strains, in terms of bacterial loads in blood, liver, spleen or kidneys. We conclude that in E. faecalis , neither FASII nor cyclopropanation enzymes are suitable antibiotic targets. Importance Membrane lipid homeostasis is crucial for bacterial physiology, adaptation, and virulence. Fatty acids are constituents of the phospholipids that are essential membrane components. Most bacteria incorporate exogenous fatty acids into their membranes. Enterococcus faecalis has emerged as a serious nosocomial pathogen, which is responsible for urinary tract infections, bacteremia and endocarditis, and is intrinsically resistant to numerous antibiotics. E. faecalis synthesizes saturated and unsaturated fatty acids, but also cyclic fatty acids that are not found in the human host. We characterized mutant strains deficient in fatty acid synthesis and modification using genetic, biochemical, and in vivo approaches. We conclude that neither the fatty acid synthesis pathway nor the cyclopropanation enzyme are suitable targets for E. faecalis antibiotic development.

Les acides linoléiques conjugués (CLA) et les acides gras cyclopropanes (CFA) sont deux types d’acides gras à haute valeur ajoutée. Ils présentent un grand intérêt dans divers domaines agroalimentaire ou industriel. Les CLA sont connus pour leurs effets bénéfiques pour la santé, alors que les CFA constituent une bonne source pour la fabrication de lubrifiants, plastiques…après leur hydrogénation. Les CLA sont synthétisés par voie chimique qui génère différents types d’isomères. Les CFA sont produits par les plantes dépendantes du climat et les bactéries présentant naturellement de faibles productivités. Il semble donc intéressant de chercher des alternatives pour leurs productions. Y. lipolytica constitue une bonne alternative. Ceci passe par des modifications génétiques pour permettre ces synthèses et l’étude de l’influence des conditions de culture. Pour cela, les voies de synthèse de ces deux acides gras ainsi que les éléments qui peuvent influencer leurs productions sont à acquérir. Dans ce contexte, nous avons testé plusieurs souches possédant différentes modifications génétiques en erlenmeyer et en fermenteur sur le milieu de néo-synthèse et de bioconversion. Une souche a été sélectionnée permettant la diminution de la dégradation des CLA par le blocage de la voie de la β-oxydation. La meilleure production de CLA en fermenteur a été obtenue en présence d’huile de soja et grâce à la surexpression du gène FAD2 (302 mg. L-1). L’influence de la composition et des paramètres de culture a été étudiée. Les résultats ont montré une meilleure assimilation de la peptone de Soja Dynamis riche en acides aminés libres par la levure qui conduit à une productivité améliorée. Ceci par son effet positif sur la croissance et les productions. De plus, la stratégie Fed-batch en fermenteur a permis la production optimale de CLA (0,4 g.L-1), la limitation en azote et phosphore semble affecter leur accumulation. D’autres souches dites de premières générations productrices de CFA grâce à l’expression du gène de la CFAs d’E.coli ont été testées sur différents milieux de culture. Deux souches (GY1005 et GY1070) ont été retenues pour leur production maximale d’huile et/ou de CFA sur le milieu de néo-synthèse en fermenteur. Ceci grâce à la surexpression des gènes DGA2 et GPD1 permettant une forte accumulation de lipides et l’absence de remobilisation et de dégradation des acides gras. Les paramètres et la composition du milieu de culture tels qu’une forte agitation (1 000 rpm) et une stratégie Fed-batch en utilisant une solution de même composition que le milieu de départ ont permis de maximiser la production de CFA (1,2 g.L-1 pour GY1070 et 0,9 g.L-1 pour GY1005). Une souche dite de deuxième génération (JMY5578) a été optimisée génétiquement (expression du gène de la CFAs sous le contrôle d’un promoteur plus fort et l’expression du gène LRO1). Cette souche a été testée en erlenmeyer et en fermenteur dans le but d’évaluer l’influence des composés du milieu de culture sur la production de CFA. Les résultats ont montré un gain de 30% de CFA et 6% de biomasse en présence des chlorures de magnésium et des vitamines dans le milieu de culture. De plus, un ajout en continu de dextrose et un milieu de départ riche en azote et phosphore a permis une production de 3 g.L-1 de CFA. Nous avons étudié l’impact des promoteurs sur l’expression du gène de la CFAs dans des souches de 2éme génération. L’expression du gène sous le contrôle du promoteur php8d a donné le meilleur résultat en termes d’accumulation des CFA dans la souche JMY6068 (46% dans les lipides totaux) ; il semble être le promoteur le plus fort en comparaison avec TEF
Conjugated linoleic acids (CLA) and cyclopropane fatty acids (CFA) are two high value added products. They are of a great interest in various agri-food or industrial fields. CLAs are known for their health benefits, while CFAs, after their hydrogenation are a good source for the manufacture of lubricants, plastics.... CLAs are synthesized chemically which generates different types of isomers. CFAs are produced by climate-dependent plants and by bacteria with low productivity. It seems therefore interesting to look for alternatives for their production. Y. lipolytica is a good alternative. This involves genetic modifications to enable these syntheses and the study of the influence of culture conditions. For this purpose, synthetic pathways of these two fatty acids as well as the elements that can influence their production are to be acquired. In this context, we tested several strains with different genetic background in flask and fermenter on neo-synthesis and bioconversion media. One strain was selected allowing reduction of CLA degradation by blocking the β-oxidation pathway and the better production of CLAs in fermenter with soybean oil by overexpression of FAD2 (302 mg. L-1). The influence of media composition was studied. The results showed better assimilation of Dynamis soy peptone rich in free amino acids by this yeast. This has a positive effect on growth and productions. In addition, the Fed-batch fermenter strategy allowed the optimal production of CLA (0.4 g. L-1), and nitrogen and phosphorus limitations seems to affect their accumulation. First generation strains producing CFA by expression of the CFA gene of E. coli were studied. They were tested on different culture media. Two strains (GY1005 and GY1070) were retained for their maximum oil production and/or CFA accumulation on neosynthesis medium in fermenter. This is due to DGA2 and GPD1 genes overexpression allowing a strong lipid accumulation and in the absence of remobilization and degradation of fatty acids. Media composition and culture parameters, such as strong stirring (1 000 rpm) and a Fed-batch strategy using a solution with the same composition as the starting medium, made it possible to maximize CFA production (1.2 g. L-1 for GY1070 and 0.9 g. L-1 for GY1005). A so-called second generation strain (JMY5578) was genetically optimized (CFA gene was expressed under the control of a stronger promoter and LRO1 over-expression gene). This strain was tested in flask and fermenter in order to evaluate the influence of culture medium compounds on CFA production. The results showed a gain of 30% CFA and 6% biomass in the presence of magnesium chloride and vitamins in the culture medium. In addition, a continuous addition of dextrose in fermenter and a nitrogen and phosphorus-rich starting medium allowed production of 3 g.L-1 of CFA. We also looked at the effect of promoters for the CFA gene expression in secondgeneration strains. Expression of the gene under the control of the php8d promoter gave the best result in terms of CFA accumulation in JMY6068 strain (46% in total lipids) and appears to be the strongest promoter in comparison with TEF

Ce travail de recherche portait sur l'extraction d'huile microbienne, notamment de certains actifs inhabituels comme les acides gras cyclopropanes, à partir de la levure oléagineuse Yarrowia lipolytica. L'objectif était de développer un itinéraire d'extraction en voie humide afin de réduire le coût global de production de cette huile microbienne. Pour cela, des pré-traitements de la matière ont été mis en œuvre (pressage, champs électriques pulsés, décharges électriques de haute tension, ultrasons et homogénéisation haute pression) afin de lyser les cellules. L'impact de ces technologies sur le rendement en huile extraite et le profil en acides gras a été étudié. Le criblage, en voie sèche, de ces différents procédés a permis de sélectionner les deux techniques les plus efficaces à approfondir : les ultrasons et l'homogénéisation haute pression (HHP). Afin d'améliorer leur efficacité, les ultrasons ont été appliqués directement en voie humide en utilisant une suspension de levures dans un solvant. Les paramètres clés du traitement ont été optimisés. Ainsi, le même rendement d'extraction d'huile a pu être obtenu en voie humide comparativement à la voie sèche. L'HHP a tout d'abord été étudiée en voie sèche afin de déterminer les valeurs optimales des paramètres opératoires. Par la suite, la faisabilité d'une extraction en voie humide a été démontrée. Une suspension a subi un pré-traitement à 150 MPa à température ambiante pendant 5 passages puis a été extraite dans un solvant à l'aide d'un disperseur à haute vitesse. Le rendement maximal obtenu par cette méthode était très proche de celui de l'extraction voie sèche qui permettait de récupérer la totalité de l'huile
This research work was focused on microbial oil extraction, especially unusual molecules such as cyclopropane fatty acids, from the oleaginous yeast Yarrowia lipolytica. The aim was to develop a wet extraction route in order to reduce the overall cost of microbial oil production. Therefore, biomass pretreatments were studied (mechanical pressing, pulsed electric fields, high voltage electrical discharges, ultrasounds and high pressure homogenization) in order to disrupt the cell wall. The effect of these technologies on oil extraction and fatty acid profile was investigated. Screening of these technologies in dry route allowed the selection of two effective techniques; ultrasounds and high pressure homogenization, which were further studied. ln order to enhance their efficiency, ultrasounds were directly applied in a wet route (yeast suspension in an extracting solvent). The key parameters were optimized and, thus, the same yield was obtained for both wet and dry routes. High pressure homogenization was studied in dry route in order to determine the optimum values for pressure, the number of passes and the temperature. The feasibility of a wet route extraction has been then demonstrated. A yeast suspension was treated by high pressure homogenization at 150 MPa, room temperature, and for 5 passes. Oil was then extracted in a solvent using a high speed disperser. Following this methodology, the maximum oil recovery yield was very close to the yield reached in the dry route