Tricine, 0.1 m sorbitol, 0.5 m M N a2H P 0 4, pH 7.8.
Results and Discussion
The fattyacid pattern of the main cellular and cya- nelle lipids of Cyanophora p aradoxa is very unusual, as can be seen from Table I. W ithin the typical cya- nelle lipids phosphatidylglycerol, monogalactosyl- diacylglycerol and digalactosyldiacylglycerol 16:0 and 20:4 acids predom inate, w hereas phosphatidyl- ethanolam ine which occurs exclusively in cellular m em branes contains 16:0, 18:0, 18:1, 20:3 and 22:0. Phosphatidylcholine which is found in both, cellular and cyanelle m em branes, shows an interm ediate p at tern. From these results a direct relationship to the fattyacid pattern of any cyanobacterial or higher plant cell so far analyzed can not be established, whereas some marine and freshw ater algae do con tain minor am ounts of 20:3 and 20:4 acids (for review see ). A certain parallelism to plant cells, how ever, may be stated in that the most unsaturated acids, 20:4 in C yanophora and 18:3 in plant cells, are most abundant within the cyanelle and chloroplast lipids, respectively. The unexpected findings pres
Dominic Helm 8 , Bernhard Küster 8 , Jan Krumsiek 1,9,11 & Josef Ecker 6
Interactions between the gut microbial ecosystem and host lipid homeostasis are highly relevant to host physiology and metabolic diseases. We present a comprehensive multi- omics view of the effect of intestinal microbial colonization on hepatic lipid metabolism, integrating transcriptomic, proteomic, phosphoproteomic, and lipidomic analyses of liver and plasma samples from germfree and speci ﬁc pathogen-free mice. Microbes induce mono- unsaturated fattyacid generation by stearoyl-CoA desaturase 1 and polyunsaturated fattyacid elongation by fattyacid elongase 5, leading to signi ﬁcant alterations in glyceropho- spholipid acyl-chain pro ﬁles. A composite classiﬁcation score calculated from the observed alterations in fattyacid pro ﬁles in germfree mice clearly differentiates antibiotic-treated mice from untreated controls with high sensitivity. Mechanistic investigations reveal that acetate originating from gut microbial degradation of dietary ﬁber serves as precursor for hepatic synthesis of C16 and C18 fatty acids and their related glycerophospholipid species that are also released into the circulation.
Four decimicrolitres of the fatty acids’ methyl esters were injected onto a G C -R IA Shimadzu with R P F R -G 1 processor, FD I detector and cap illary column A T -W A X (Superox II) 30 m x 0.25 mm ID. Column tem perature was set from 180 to 240° C, with nitrogen as gas carrier. The analysis was run three times for each sample. Fattyacid identification was done by comparison, with a standard mixture of fatty acids methyl esters. Theoretical iodine absorption value was calculated based on the percentages of enoic fatty acids (C ar reras et al., 1989).
In non-green tissues a deficient conversion o f triose- into hexose m o n o p h o sp h ates by lack o f fru cto se 1,6-bisphosphatase has been observed, b u t c a n n o t be generalized . In plastids, hexose m o n o p h o sp h a te s are required b o th fo r starch syn thesis a n d as substrates for the pentose p h o sp h ate p ath w ay . O u r results concerning the stim ulation o f A T P -driven fattyacid synthesis by sugar p h o s p h ates (Table II) suggest th a t G lc 6 P and F r u 6 P are cap ab le o f m aintaining the plastidial pyridine- nucleotide com plem ent in their reduced form s. T he low er effect o f D H A P and F ru B P suggests an effective u p tak e o f G lc 6 P an d F r u 6 P w hich m ight be m ediated by a p h o sp h ate tra n slo c a to r capable o f tra n sp o rtin g hexose m o n o p h o sp h ates .
Polymer nanoparticles have been prepared by an optimized interfacial deposition method . As in the commonly used procedure, surfactants were needed to stabilize the nanoparticles of PGA-fattyacid esters with high esterification degrees , the preparation method has been modified in this study. It has been discovered that the implementation of a heating and cooling step (as described in 2.2.1) lead to stable, self- stabilizing nanoparticles with all tested Lauroyl-, Stearoyl- and Oleoyl-PGA polyesters. For Behenoyl-PGAs, which are insoluble in acetone, further modification was necessary and another applicable organic solvent had to be found. The interfacial deposition method requires a semi-polar, water miscible solvent with adequate solubility of the polymer and rapid diffusion into water. Decreased miscibility with water increases the interfacial tension and thus increases the resulting size of the particles . The tested solvents dichloromethane/acetone (volume ratio 1/6) , THF and THF/ethanol (3/2)  all showed sufficient solubility of Behenoyl-PGA. But due to the reduced miscibility with water they lead to bigger particles (in descending order). Therefore, the amount of ethanol in the mixture with THF was increased. The ratio THF/ethanol 1:1 was found to be the best compromise between solubility of polymer (owing to THF) and rapid diffusion of the solvent into water (enhanced by ethanol). With this solvent mixture and the optimized preparation method, Behenoyl-PGA nanoparticles could be prepared for the first time.
Wenn man das Gesamtkollektiv (n = 101) betrachtet, finden sich keine signifikant höheren Glukose-Werte (gemessen vom Zentrallabor) während des OGTT bei den für das Thr54-Allel homozygoten Probanden (n = 10, siehe Abb. 13). Im Gegenteil, sie liegen am niedrigsten. Zwar liegen die Mittelwerte der für die Mutation heterozygoten über denen des Wildtyps, jedoch ohne dabei statistisch signifikant zu sein. Auch wenn man die Untergruppen (FFA-Kollektiv, FFA-Kollektiv mit IGT/NGT) betrachtet (siehe Abb. 14 - 16), entdeckt man keine signifikanten Unterschiede zwischen den untersuchten Gruppen. Wenn man die hetero- und homozygoten Träger des Codon 54-Polymorphismus (Ala/Thr und Thr/Thr) unter der Annahme eines dominanten Rollenmodels zusammenfasst, bevor man sie mit der Wildtypgruppe (Ala/Ala) vergleicht, findet man außer für den Nüchternglukose- Wert ebenfalls keine Signifikanzen (siehe Abb. 15 und 16). Ein dominantes Rollenmodell wird unter der Vorstellung angenommen, dass auch das polymorphe Codon Ala54Thr zur Synthese eines lipophileren intestinalen fattyacid binding protein (IFABP) führt [Baier et al. 1995].
Fats contain 80 % of the heat content of diesel fuels but they show severe disadvantages when used in engines like oil deterioration and incomplete combustion. These problems and possible solutions are discussed in detail in 2 . The European Union aims to substitute mineral oil based fuels. In 2010 the proportion of natural fuels should exceed 5.75 %, in 2020 20 % 3 . To enhance the fuel properties and to reduce the viscosity fats are converted to fattyacid esters of short chain alcohols. Fattyacid methyl esters (FAME) are favored because of the low price of methanol and the easier processing. In this area research is done because new processes are expected to show less energy consumption compared to traditional processes 1 at temperatures of 240 °C and pressures of 9 MPa. One approach is the optimization of reaction conditions at low temperatures and atmospheric pressure with alkaline catalysts. Examples are found in literature for rapeseed oil 4 , for soybean oil 5, 6 and for Pongamia Oil 7 . As acidic catalysis is much slower, this way only makes sense for special cases like used oil 3 which contains high fractions of free fatty acids that interfere with the use of alkaline catalysts.
A. stellata chloroplasts is essential. Researchers
have shown that fattyacid composition of lipids comprising biological membranes, such as the gly- colipids and phospholipids of chloroplast thyla- koids, can be effected by environmental conditions (Thompson, Jr., 1996; Harwood, 1996; Pohl and Zurheide, 1979). Decreased temperatures have long been demonstrated to result in increased levels of fattyacid unsaturation (reviewed in Har wood et al., 1994; Marr, 1962). Growth limiting conditions such as decreased light intensity, increased salinity, increased cell concentration, su- boptimal pH, and suboptimal temperature have also been indicated to alter algal fattyacid profiles (Cohen et al., 1988). Preference towards PUFAs with fewer points of unsaturation has been ob served under suboptimal growth conditions, as de monstrated in the red alga Porphyridium cruentum by a decrease in eicosapentaenoic acid and concur rent increase in arachidonate (Cohen et al., 1988). Because environmental conditions can have such a significant impact on the fattyacid content of algal lipids, the contrarieties observed amongst fattyacid profiles obtained from species specific algal collections have often been attributed to these external factors, rather than the cause of ge netic differences. For example, the controversial reports on the fattyacid content of the red algae
natural phylogenetic system, fattyacid composition will possibly prove to be a further useful tool in ex ploring the relationships between C hlorophyta. In comparison to the o th er classes of green algae more data for lipid research in Chlorophyta are available especially from the orders of the Volvocales and the Chlorococcales. A p art from some disagreem ents re garding the occurrence of 16:4 and 18:4 acids, the special position of the thus far investigated Chlorococcales, containing the simplest fattyacid composition within the green algae and the only identical characteristics to higher plant leaves, is con firmed by all authors ,
Table I. Influence o f cycloxydim on the incorporation of [ l4C ]acetate into the fattyacid fraction o f chloroplasts and etiop lasts isolated from primary leaves of sensitive oat plants and tolerant pea seedlings. The incorporation rate is expressed in the case o f chloroplasts as kBq per mg chlorophyll a + b and also per mg carotenoids (x + c) and in the case o f etioplasts per mg carotenoids (x + c). Incorpora tion tim e 20 min. M ean o f 4 determ inations with standard deviation (S D ). The chloroplast pigm ent ratio (chloro phylls to carotenoids) was found to be 4.9 ( ± 0.3) for chloroplasts o f both plants.
The NRPSs of the lipopeptide antibiotics from the daptomycin-class do not contain integrated PKS elements. Instead, different biosynthetic pathways for fattyacid activation and modification can be deduced from their biosynthetic gene clusters [9, 104, 105]. In the daptomycin biosynthetic system, the stand-alone proteins DptE and DptF are a putative fatty acyl-AMP ligase and an ACP, respectively, responsible for activation of the fattyacid building blocks [9, 95]. In analogy to the above discussed AL domain of MycA, DptE is likely to recruit fatty acids provided by primary metabolism and to activate them as acyl-adenylates, which are subsequently transferred to the carrier protein DptF. A fused equivalent of DptE and DptF is present in the A54145 NRPS system, where LptEF was proposed to carry out the same biosynthetic function during lipoinitiation. After fattyacid activation and attachment to the respective ACP, the activated intermediates are possibly transferred to the amino acid that is bound to the first module of the daptomycin or A54145 peptide synthetase (Figure 2.18).
Clr transformations were applied to fattyacid data to account for compositionality. Clr was calculated as the natural log of the quotient of the individual fattyacid concentration over the geometric mean of all fattyacid concentrations within a sample [ 22 ]. The geometric mean was used as the denominator as it is a meaningful measure of central tendency for variables with skewed distributions. Principal component analysis (PCA) was used to evaluate correlations between fattyacid prior to generating compositional biplots. Fattyacid groups/clusters were selected based on collinear links of a sub-composition showing a one-dimensional pattern in the compositional biplots for 12 months human milk samples. These fattyacid groups/clusters were calculated by adding the individual crude fattyacid concentrations, after which this sum was clr transformed for inclusion into the analysis models. A general linear model was used to assess the impact of maternal pre-pregnancy BMI, change in weight status between pregnancy and at 6 weeks, age, education, occupation, parity, delivery mode, and gestational age on individual human milk fatty acids. Maternal age was included because of its close association with maternal education, less exposure to cigarette smoking, and (prolonged) breastfeeding [ 17 ], thus potentially being an indicator of aspects of life style that might influence human milk fattyacid composition. Additionally, education and occupation were used as proxies for socio-economic status. A general linear model based repeated measure analysis was used to evaluate changes in human milk fattyacid constituents during lactation, adjusting for lifestyle-related factors. Bonferroni adjustment was applied to account for multiple testing. All statistical analyses were performed with SAS version 9.4 (The SAS Institute, Cary, NC, USA).
Besides the positive effects on the general health constitu- tion, traditional oil pulling is also suggested to prevent tooth decay, bleeding gums, and halitosis [ 47 , 50 , 70 , 135 – 137 ]. Patients repeatedly described a positive impact on their oral tissues and although there is no clear scientific explanation to this phenomenon yet, several popular scientific sources promote the procedure as a daily supplement of oral hy- giene. It was to be expected that there are numerous research findings on the effects of edible oils in preventive dentistry but there are only a few scientific studies. Some mouth care preparations, predominantly in the field of natural cosmet- ics, contain vegetable oils as ingredients, varying from “mouth-oil” to toothpaste or saliva substitutes [ 138 ]. It is important to note that their distribution is rather based on an empirical application, as previous findings and literature regarding the efficacy of vegetable oils are controversial and require further investigation. Nevertheless, a few studies attempt to scientifically prove a potential of vegetable oils to inhibit oral biofilm accumulation and to have a protective impact on the oral hard and soft tissue [ 48 , 49 , 70 , 135 ]. The following will point out beneficial results that were obtained in vitro as well as in vivo and are derived by lipid compo- nents; however, further scientific data is needed to compre- hend molecular interactions. For instance, a positive correlation between dietary fats and the fattyacid composi- tion of the few enamel lipids was noticed in vitro [ 139 , 140 ]. Certain fatty acids were shown to disturb bacterial adhesion as an in vitro study revealed for S. aureus under the influ- ence of oleic acid [ 141 ].
de novo fattyacid biosynthesis is very close to the in vivo situation. The lowest ho -values we always
found in the chloroplast test system. In a direct as say of the ACC target enzyme, the ho -values are about 5 to 10 times higher or even more, depend ing upon the plant and the isolation procedures applied. The results obtained in isolated intact chloroplasts (or etioplasts) thus fully reflect the binding and inhibition characteristics of the target enzyme acetyl-CoA carboxylase. This is of great im portance for the screening of possible differ ences in sensitivity and tolerance towards both herbicide classes, which in our experience is best performed at the level o f isolated chloroplasts.
Nowadays, consumers are increasingly aware of health. Commercial trend has changed to improve the health aspect of dairy products. Milk fattyacid (FA) compositions have many effects on human health (Parodi, 1997; German et al., 2009) and each FA has different effects (Palmquis et al., 1993; Mensink et al., 2003; German and Dillard, 2006). Triglyceride affects the flavor of cheese and the melting point of butter (Jensen, 2002). The ratio of saturated to unsaturated FA reflects butterfat hardness. Conjugated linoleic acid posses several health benefits. Zock et al. (1994) found that myristic acid (C14:0) and palmitic acid (C16:0) caused high LDL cholesterol. Double bond FA in trans form raise LDL and lower HDL cholesterol (Brouwer et al., 2010). Hence improvement of milk FA is interesting.
Fettsäuren haben im menschlichen Körper unterschiedlichste Funktionen. Sie sind wichtige Energieträger, werden zum Aufbau komplexer Proteine (z.B. Phospholipide, Cholesterol) benötigt und sind in die Signalwirkung von Hormonen und Signalkaskaden eingebunden. Sie werden täglich über die Nahrung aufgenommen und zusätzlich aus Glukose in den Leberzellen hergestellt. Zum Transport der hydrophoben Moleküle werden Transportmoleküle wie Albumin, Lipocaline oder Fatty-acid-binding Proteine benötigt. Die FABPs liegen sowohl membran-assoziiert als auch frei im Zytoplasma vor. Die langkettigen Fettsäuren sind insbesondere im Herzen ein wichtiger Energieträger. Unter normaler Arbeitsbelastung wird der Hauptteil der benötigten Energie aus der Oxidation langkettiger Fettsäuren gewonnen. Da nur eine limitierte Menge der benötigten Substrate gespeichert werden kann, ist eine kontinuierliche Zufuhr notwendig.
De novo synthesis of fatty acids involves a set of conserved reactions for the cyclic elongation of precursors and requires a multienzyme com- plex. First, there is the acyl transfer from acetyl and malonyl coenzyme A to the prosthetic phos- phopantetheine group of the acyl carrier protein (ACP) by an acetyl transferase (AT) and malonyl transferase (MT). The enzyme-bound substrates are then condensed by the ketoacyl synthase (KS) by decarboxylation of the malonyl to acetoacetyl- ACP, which is further reduced by a ketoacyl re- ductase (KR), dehydrated by a dehydratase (DH) and reduced by an enoyl reductase (ER) to form butyryl-ACP. These reactions are repeated six times. The mammalian fattyacid synthases (FASs) release the newly formed fattyacid by a thioeste- rase (TE) whereas in fungi it is transferred back from the ACP to coenzyme A by an MT (Jenni et al., 2006). The fatty acids are further elongated by elongases (Rawlings, 1998).
The combination of the structural information from NR and morphological features from AFM results in the following uniﬁed picture: thin polysaccharide/lipid ﬁlms, with an ordered multi- layer structure, can be prepared in a single and rapid step, by spin-coating a solution of spontaneously co-assembled multilayer vesicles onto a solid surface. The hydrophilic regions of the ﬁlm, namely, the chitosan chains and the fattyacid PEGylated head groups, selectively and reversibly absorb water as a function of the RH, resulting in a remarkable color change of the ﬁlm, thereby allowing for optical sensing of humidity and the design of humidity-triggered optical ﬁlters. Moreover, in dry conditions, the ﬁlm switches to a polycrystalline morphology, with partially ordered platelets formed by crystalline chitosan/lipid domains. A schematic representation of the process is given in Fig. 7 . A slow phase separation in the ﬁlm is observed over a time scale of several weeks, which is strongly dependent on the humidity conditions at which the ﬁlms are stored.
Fattyacid biotransformation activity of E. coli BL21(DE3) pCOLA-RcsB-DsrA
The effect of the GDAR system engineering on the whole-cell fattyacid biotransformation activity was investigated by conducting the bioconversion of ricinoleic acid (i.e., 12-hydroxyoc- tadec-9-enoic acid (1)) into n-heptanoic acid (5) and 11-hydroxyundec-9-enoic acid (4) ( S1 Scheme ), which was described in our previous study [ 15 ]. When ricinoleic acid was added into the culture broth of the recombinant E. coli BL21(DE3) pACYC-ADH, pET-BVMO, pCOLA or E. coli BL21(DE3) pACYC-ADH, pET-BVMO, pCOLA-RcsB-DsrA expressing the alcohol dehydrogenase (ADH) of Micrococcus luteus and the Baeyer-Villiger monooxygenase (BVMO) of Pseudomonas putida, the final product formation rates of the both cells were similar to at t < 4 h ( Fig 6 ). However, the final product formation rate of E. coli BL21(DE3) pACYC-ADH, pET-BVMO, pCOLA ceased resulting in an accumulation of the reaction intermediate (2) in the culture medium at t > 4 h, when the product concentration and bioconversion yield reached over 6 mM and 45%, respectively ( Fig 6A ). This might be ascribed to the toxicity of n- heptanoic acid ( Fig 5B ). In contrast, the final product formation rate of E. coli BL21(DE3) pACYC-ADH, pET-BVMO, pCOLA-RcsB-DsrA was further maintained resulting in a final product concentration of over 10 mM (conversion yield, 68%), which was 1.6-fold higher as compared to the E. coli BL21(DE3) pACYC-ADH, pET-BVMO, pCOLA. This value was also Fig 4. The relationship between GadA/B activity and cultivation pH (A) or n-heptanoic acid concentration (B). The GadA/B
2.3. Free fattyacid analysis
Federation of Cocoa Commerce (FCC) recommended method was used for the FFA analysis and double extraction was carried out. Round bottomed flasks of 250 mL were dried in the oven at 105 o C, cooled in the desiccator, weighed and 180 mL of hexane was measured into the round bottomed flasks. Each test sample of 10 g was measured into a thimble and was set up for extraction for two hours using the Soxhlet apparatus. The set up was allowed to cool and the solvent drained into the round bottomed flask. Each sample was ground with sand and set up for two 2 h again. The solvent was concentrated into fat by evaporating the hexane using the rotary evaporator. The fat content was dried in the oven for 2 h and cooled in the desiccator. Weight of the extract and the flask were taken and recorded.