Presence Of Long-chain Fatty Acids Research Articles

Bioassay-guided detection, identification and assessment of antibacterial and anti-inflammatory compounds from olive tree flower extracts by high-performance thin-layer chromatography linked to spectroscopy

Olive trees are one of the most widely cultivated fruit trees in the world. The chemical compositions and biological activities of olive tree fruit and leaves have been extensively researched for their nutritional and health-promoting properties. In contrast, limited data have been reported on olive flowers. The present study aimed to analyse bioactive compounds in olive flower extracts and the effect of fermentation-assisted extraction on phenolic content and antioxidant activity. High-performance thin-layer chromatography (HPTLC) hyphenated with the bioassay-guided detection and spectroscopic identification of bioactive compounds was used for the analysis. Enzymatic and bacterial in situ bioassays were used to detect COX-1 enzyme inhibition and antibacterial activity. Multiple zones of antibacterial activity and one zone of COX-1 inhibition were detected in both, non-fermented and fermented, extracts. A newly developed HPTLC-based experimental protocol was used to measure the high-maximal inhibitory concentrations (IC50) for the assessment of the relative potency of the extracts in inhibiting COX-1 enzyme and antibacterial activity. Strong antibacterial activities detected in zones 4 and 7 were significantly higher in comparison to ampicillin, as confirmed by low IC50 values (IC50 = 57–58 µg in zone 4 and IC50 = 157–167 µg in zone 7) compared to the ampicillin IC50 value (IC50 = 495 µg). The COX-1 inhibition by the extract (IC50 = 76–98 µg) was also strong compared to that of salicylic acid (IC50 = 557 µg). By comparing the locations of the bands to coeluted standards, compounds from detected bioactive bands were tentatively identified. The eluates from bioactive HPTLC zones were further analysed by FTIR NMR, and LC-MS spectroscopy. Multiple zones of antibacterial activity were associated with the presence of triterpenoid acids, while COX-1 inhibition was related to the presence of long-chain fatty acids.

Screening of Selected Cultivars of Sweet Sorghum for Phytochemicals and In vitro Evaluation of Their Antihyperglycemic and Cytotoxic Activity

Background: Sorghum, a wonder millet, is well known for its beneficial phytochemical profiles. In comparative terms, juice from sweet sorghum has better commercial potential as syrup in several food-based applications. Various sweet sorghum differs in their profile of various phytochemicals, which can impact the commercial potential of sweet sorghum juice. Methods: Our previous works on cultivars developed at ICAR-IIMR were screened for phytochemical, sugar and mineral profile. To give a holistic view of the phytochemical profile of sweet sorghum varieties, the present study is attempted to evaluate the total phenolic and flavonoid content, antihyperglycaemic and cytotoxic profile of the components present in the varieties CSV19SS, SSV84, SSV74. The phytochemical footprint of sweet sorghum juice was studied through HRLCMS. Results: The results showed ethyl acetate extract of SSV84 having potential antihyperglycemic effects with an IC50 of 22.156 ± 0.9 µg/ml (α-glucosidase) and 0.070 ± 0.02 mg/ml (α-amylase) with a comparatively higher phenolic (232.6 ± 1 mg GAE/g) and flavonoid (138.18 ± 0.9 mg QE/g) content. The chloroform extract of SSV 84 showed a higher cytotoxic effect at an IC50 of 165.502 ± 7 µg/ml (HeLa cells) and 237.895 ± 15 µg/ml (Hep G2 cells). HRLCMS profile of SSV 84 showed the presence of long-chain fatty acids in hexane extract. Anthraquinones, carotenoids, xanthophylls, cinnamic acid and derivatives, and isoflavones were present in chloroform extract, while Ethyl acetate extract was rich in phenolic acids and also consisted of coumarins, quinones, alkaloids, and terpenoids. Conclusion: The high cytotoxic and antihyperglycemic activities of extracts can be attributed to the presence of these phytochemicals.

Insights into the organic matter composition of soda lakes in the Pantanal, Brazil, through fatty acids analysis in sediments.

We conducted a geochemical analysis on three sediment cores collected from soda and freshwater lakes in the Pantanal region, in Brazil. Our objective was to identify the primary sources of organic matter associated with the Pleistocene-Holocene transition, as previously documented in that area. The Nhecolandia region, located in the Pantanal, encompasses over 10,000 lakes, with approximately 10% of them exhibiting alkaline characteristics. Soda lakes became highly saline and alkaline after ~ 910calyr BP, which influences biogeochemistry and aquatic ecology. They have high electrical conductivity and pH can reach 10.5. In contrast to freshwater, soda lakes are absent of surrounding vegetation. Literature suggests a strong influence by the Last Glacial Maximum on the region. We hypothesized that periods of aridity and increased precipitation influenced the composition of organic matter present in sediments and preserved within these cores. Our analysis focused on examining the presence and distribution of fatty acids, organic carbon content, and total nitrogen. In general, the cores exhibited two distinct parts in terms of organic matter sources: the upper sections of the cores were primarily composed by terrestrial sources, identified by the presence of long-chain fatty acids, while the deeper sections were dominated by aquatic sources, therefore short-chain fatty acids. We did not find significant difference among fatty acid profile that could distinguish freshwater from soda lakes, the only remarkable difference was the occurrence of saturated fatty acids, which is lower in freshwater lake. These findings suggest the occurrence of humid and arid periods in the region. The C/N ratio displayed a similar trend to the fatty acid's profiles, exhibiting an abrupt change that was likely induced by climate variations. Although diagenesis can alter the composition of organic matter and, subsequently, the C/N values, it is noteworthy that the abrupt change observed in the Salina da Ponta (soda lake) core corresponds to 3,200years BP. This coincides with documented climate changes that occurred during the Holocene. Our study revealed the influence of past climatic conditions on the sources and variations of organic matter in sediment cores from the Pantanal's soda and freshwater lakes. Consideration of diagenesis and climate variations is crucial for interpreting sedimentary records.

FA Sliding as the Mechanism for the ANT1-Mediated Fatty Acid Anion Transport in Lipid Bilayers.

Mitochondrial adenine nucleotide translocase (ANT) exchanges ADP for ATP to maintain energy production in the cell. Its protonophoric function in the presence of long-chain fatty acids (FA) is also recognized. Our previous results imply that proton/FA transport can be best described with the FA cycling model, in which protonated FA transports the proton to the mitochondrial matrix. The mechanism by which ANT1 transports FA anions back to the intermembrane space remains unclear. Using a combined approach involving measurements of the current through the planar lipid bilayers reconstituted with ANT1, site-directed mutagenesis and molecular dynamics simulations, we show that the FA anion is first attracted by positively charged arginines or lysines on the matrix side of ANT1 before moving along the positively charged protein-lipid interface and binding to R79, where it is protonated. We show that R79 is also critical for the competitive binding of ANT1 substrates (ADP and ATP) and inhibitors (carboxyatractyloside and bongkrekic acid). The binding sites are well conserved in mitochondrial SLC25 members, suggesting a general mechanism for transporting FA anions across the inner mitochondrial membrane.

Cyclic AMP is a critical mediator of intrinsic drug resistance and fatty acid metabolism in M. tuberculosis.

Cyclic AMP (cAMP) is a ubiquitous second messenger that transduces signals from cellular receptors to downstream effectors. Mycobacterium tuberculosis (Mtb), the etiological agent of tuberculosis, devotes a considerable amount of coding capacity to produce, sense, and degrade cAMP. Despite this fact, our understanding of how cAMP regulates Mtb physiology remains limited. Here, we took a genetic approach to investigate the function of the sole essential adenylate cyclase in Mtb H37Rv, Rv3645. We found that a lack of rv3645 resulted in increased sensitivity to numerous antibiotics by a mechanism independent of substantial increases in envelope permeability. We made the unexpected observation that rv3645 is conditionally essential for Mtb growth only in the presence of long-chain fatty acids, a host-relevant carbon source. A suppressor screen further identified mutations in the atypical cAMP phosphodiesterase rv1339 that suppress both fatty acid and drug sensitivity phenotypes in strains lacking rv3645. Using mass spectrometry, we found that Rv3645 is the dominant source of cAMP under standard laboratory growth conditions, that cAMP production is the essential function of Rv3645 in the presence of long-chain fatty acids, and that reduced cAMP levels result in increased long-chain fatty acid uptake and metabolism and increased antibiotic susceptibility. Our work defines rv3645 and cAMP as central mediators of intrinsic multidrug resistance and fatty acid metabolism in Mtb and highlights the potential utility of small molecule modulators of cAMP signaling.

Biotransformation of grease waste into fatty acid by Penicillium chrysogenum SNP5 through media engineering and artificial neural network.

Degradation of grease waste remains a challenging task. Current work deals with the biotransformation of grease waste into fatty acids under submerged fermentation using Penicillium chrysogenum SNP5 through media formulation and artificial neural network (ANN). Fermentation media was formulated to ameliorate the uptake of hydrocarbon by enhancing alkane hydroxylase (AlkB) activity, extracellular release of fatty acids and inhibiting beta-oxidation of fatty acid by regulating transketolase. Further, the process parameters of fermentation were optimized through Artificial Neural Network (ANN) using three critical variables viz; inoculum size (spores/ml), pH, and incubation time (days) while media engineering was done with the optimal supplementation of various medium components such as glucose, YPD, MnSO4, tetrahydrobiopterin (THB) and phloretin. The maximum conversion of 66.5% of grease waste into fatty acid was achieved at optimum conditions: inoculums size 3.36 × 107 spores/ml, incubation time 11.5days, pH 7.2 along with formulated media composed of 1% grease in czapek-dox medium supplemented with 55.5mM glucose, 0.5% YPD, 16.6mM hexadecane, 1mM MnSO4, 1mM THB, and 1mM phloretin. The presence of long-chain fatty acids in purified extracts such as oleic acid and octadecanoic acid as end products has valued the evolved process as another source of alternative fuel.

Studies on antibacterial and chemotaxis properties of Pseudomonas aeruginosa TEN01 biomass-derived sustainable biosurfactant

Biosurfactant producing bacterial strains were isolated from oil-contaminated sites at Chennai Petroleum Corporation Limited, Chennai, the potential strain was selected and identified as Pseudomonas aeruginosa TEN01 by 16 S rRNA sequencing technique. Biosurfactant was produced from cassava solid waste from the sago industry. Further, it was extracted by solvent extraction and partially purified by column chromatography. The partially purified biosurfactant was qualitatively analyzed by Thin Layer Chromatography (TLC), quantitatively analyzed by anthrone assay and characterized by Fourier Transform Infra-Red Spectroscopy (FT-IR) and Gas Chromatography-Mass Spectrometry (GC-MS). Rf value and chemical groups confirm the presence of glycolipid in the partially purified biosurfactant. GC-MS results confirmed the presence of long-chain fatty acids and carbohydrate which is found to be mainly present in glycolipids. Biosurfactants are surface-active molecules which have been found to be the best alternative to chemical-based surfactants. The present study focuses on modifying the cell surface using a biosurfactant from P. aeruginosa TEN01 to enhance membrane permeabilization. Antibacterial and chemotaxis properties of biosurfactant from P. aeruginosa TEN01 were found to be better towards Xenorhabdus poinarii, a bio-pesticide producing microbial strain, X. poinarii exhibited 81.7% adhesion to hydrocarbons upon biosurfactant treatment as analyzed by Bacterial Adhesion to Hydrocarbon (BATH) assay. The alteration in the membrane permeability was tested in X. poinarii using biosurfactant and chemical surfactants viz. Sodium dodecyl sulfate (SDS) and toluene by estimating the amount of intracellular protein released. High protein recovery (51.55%) was achieved with a biosurfactant. Cell viability in the biosurfactant-treated cells was also high (93.98%) in comparison to cells treated with chemical surfactants. Increased recovery of intracellular protein along with high cell viability makes the biosurfactant a potential candidate for application in numerous environmental fields.

Applications of Transcriptomics and Proteomics for Understanding Dormancy and Resuscitation in Mycobacterium tuberculosis.

Mycobacterium tuberculosis can survive within its host for extended periods of time without any clinical symptoms of disease and reactivate when the immune system is weakened. A detailed understanding of how M. tuberculosis enters into and exits out of dormancy, is necessary in order to develop new strategies for tackling tuberculosis. Omics methodologies are unsupervised and unbiased to any hypothesis, making them useful tools for the discovery of new drug targets. This review summarizes the findings of transcriptomic and proteomic approaches toward understanding dormancy and reactivation of M. tuberculosis. Within the granuloma of latently infected individuals, the bacteria are dormant, with a marked slowdown of growth, division and metabolism. In vitro models have attempted to simulate these features by subjecting the bacterium to hypoxia, nutrient starvation, potassium depletion, growth in the presence of vitamin C, or growth in the presence of long-chain fatty acids. The striking feature of all the models is the upregulation of the DosR regulon, which includes the transcriptional regulator Rv0081, one of the central hubs of dormancy. Also upregulated are chaperone proteins, fatty acid and cholesterol degrading enzymes, the sigma factors SigE and SigB, enzymes of the glyoxylate and the methylcitrate cycle, the Clp proteases and the transcriptional regulator ClgR. Further, there is increased expression of genes involved in mycobactin synthesis, fatty acid degradation, the glyoxylate shunt and gluconeogenesis, in granulomas formed in vitro from peripheral blood mononuclear cells from latently infected individuals compared to naïve individuals. Genes linked to aerobic respiration, replication, transcription, translation and cell division, are downregulated during dormancy in vitro, but upregulated during reactivation. Resuscitation in vitro is associated with upregulation of genes linked to the synthesis of mycolic acids, phthiocerol mycocerosate (PDIM) and sulfolipids; ribosome biosynthesis, replication, transcription and translation, cell division, and genes encoding the five resuscitation promoting factors (Rpfs). The expression of proteases, transposases and insertion sequences, suggests genome reorganization during reactivation.

ANT1 Activation and Inhibition Patterns Support the Fatty Acid Cycling Mechanism for Proton Transport.

Adenine nucleotide translocase (ANT) is a well-known mitochondrial exchanger of ATP against ADP. In contrast, few studies have shown that ANT also mediates proton transport across the inner mitochondrial membrane. The results of these studies are controversial and lead to different hypotheses about molecular transport mechanisms. We hypothesized that the H+-transport mediated by ANT and uncoupling proteins (UCP) has a similar regulation pattern and can be explained by the fatty acid cycling concept. The reconstitution of purified recombinant ANT1 in the planar lipid bilayers allowed us to measure the membrane current after the direct application of transmembrane potential ΔΨ, which would correspond to the mitochondrial states III and IV. Experimental results reveal that ANT1 does not contribute to a basal proton leak. Instead, it mediates H+ transport only in the presence of long-chain fatty acids (FA), as already known for UCPs. It depends on FA chain length and saturation, implying that FA’s transport is confined to the lipid-protein interface. Purine nucleotides with the preference for ATP and ADP inhibited H+ transport. Specific inhibitors of ATP/ADP transport, carboxyatractyloside or bongkrekic acid, also decreased proton transport. The H+ turnover number was calculated based on ANT1 concentration determined by fluorescence correlation spectroscopy and is equal to 14.6 ± 2.5 s−1. Molecular dynamic simulations revealed a large positively charged area at the protein/lipid interface that might facilitate FA anion’s transport across the membrane. ANT’s dual function—ADP/ATP and H+ transport in the presence of FA—may be important for the regulation of mitochondrial membrane potential and thus for potential-dependent processes in mitochondria. Moreover, the expansion of proton-transport modulating drug targets to ANT1 may improve the therapy of obesity, cancer, steatosis, cardiovascular and neurodegenerative diseases.

Structural models of mitochondrial uncoupling proteins obtained in DPC micelles are not functionally relevant.

Uncoupling protein 1 (UCP1) is found in the inner mitochondrial membrane of brown adipocytes. In the presence of long-chain fatty acids (LCFAs), UCP1 increases the proton conductance, which, in turn, increases fatty acid oxidation and energy release as heat. Atomic models of UCP1 and UCP2 have been generated based on the NMR backbone structure of UCP2 in dodecylphosphocholine (DPC), a detergent known to inactivate UCP1. Based on NMR titration experiments on UCP1 with LCFA, it has been proposed that K56 and K269 are crucial for LCFA binding and UCP1 activation. Given the numerous controversies on the use of DPC for structure-function analyses of membrane proteins, we revisited those UCP1 mutants in a more physiological context by expressing them in the mitochondria of Saccharomycescerevisiae. Mitochondrial respiration, assayed on permeabilized spheroplasts, enables the determination of UCP1 activation and inhibition. The K56S, K269S, and K56S/K269S mutants did not display any default in activation, which shows that the NMR titration experiments in DPC detergent are not relevant to UCP1 function.

Bidens pilosa (Black Jack) Standardized Extract Ameliorates Acute TNBS-induced Intestinal Inflammation in Rats.

Bidens pilosa is an herb popularly used to treat inflammation, hemorrhoids, fever, and gastric ulcers with reported pharmacological activities and chemical composition that sustain its selection as a potential intestinal anti-inflammatory product. Based on this, we examined the effects of a B. pilosa fatty acid-standardized supercritical preparation on the intestinal inflammatory process induced by trinitrobenzenesulphonic acid in rats, using either preventative or curative treatments. We also investigated the safety of plant extract by acute and sub-chronic toxicological analysis. The intestinal anti-inflammatory activity was related to modulation of the immune response, increasing IL-10 production and reducing IL-1β, IL-6, and TNF-α level, the oxidative stress, and the MUC production in the inflamed colon. Optic, scanning, and transmission electron microscopy (TEM) analysis supported the beneficial effects promoted by B. pilosa, which were closely related to downregulation of heparanase, Hsp70, Mapk 3, and NF-κB signaling and with the presence of long-chain fatty acids in extract. Our data suggest that B. pilosa supercritical preparation is a chemically standardized preparation potentially useful as complementary intestinal anti-inflammatory agent to treat inflammatory bowel disease.

In situ acid catalysed transesterification of biodiesel production from Sterculia foetida oil and seed

ABSTRACTThe quantity of fossil fuel is declining year by year, and the pollutant also released to the environment which insist on the exploration of new alternative fuel. Biodiesel from non-edible seeds is of primary concern for increasing prices of crude oil and thereof preventing greenhouse gas emissions. In the present study, the production of biodiesel by In-situ acid catalyzed transesterification from Sterculia foetida oil and seeds were reported. GC-FID characterization of oil sample revealed the presence of long-chain fatty acids with a higher proportion of sterculic acid (32%). The operating parameters in acid transesterification process viz., the weight of the oil and biomass, solvent volume, acid volume, temperature and time were optimized using Placket-Burman design. The maximum biodiesel yield obtained for sterculia oil and seeds were 91.58% and 95.91% exhibiting volume of oil being the critical factor based on Plackett-Burman design. One-way ANOVA analysis showed that chosen parameters were statistically significant (P < .05) except for temperature. FT-IR analysis confirms the presence of biodiesel (1741 cm–1) obtained from the transesterification process.

The Mechanism FA-Dependent H+ Transport by UCP1.

Uncoupling protein 1 (UCP1) is an integral protein of the inner mitochondrial membrane (IMM) that is expressed specifically in brown and beige fat depots. UCP1 is responsible for the production of heat to control core body temperature, the regulation of fat metabolism, and the energy balance. As an uncoupling protein, UCP1 transports H+ across the IMM in presence of long-chain fatty acids (FA), which makes brown fat mitochondria produce heat at the expense of ATP. However, the exact mechanism of UCP1 action has remained difficult to elucidate, because direct methods for studying currents generated by UCP1 were unavailable. Recently, the patch-clamp technique was successfully applied to brown and beige fat mitochondria to directly study H+ currents across the IMM and characterize UCP1 function. A new model of the UCP1 mechanism was proposed based on the patch-clamp analysis. In this model, both FA anions (FA-) and H+ are transport substrates of UCP1, and UCP1 operates as a non-canonical FA-/H+ symporter. Here, we summarize recent findings obtained with the patch-clamp technique that describe how UCP1 can transport not only H+ but also FA-.

Small-Molecule Uncoupling Protein Mimics: Synthetic Anion Receptors as Fatty Acid-Activated Proton Transporters.

Uncoupling proteins (UCPs) regulate energy expenditure in living cells by inducing proton leakage across the mitochondrial inner membrane, thereby uncoupling adenosine diphosphate phosphorylation from nutrient oxidation. The proton transport activity of UCP1 and UCP2 requires activation by fatty acids. We report here the first examples of synthetic neutral anion receptors performing this biologically important fatty acid-activated function in phospholipid bilayers. We have shown that a tripodal thiourea possesses poor H+/OH- transport activity without fatty acids, but in the presence of long-chain fatty acids is "switched on" as a proton transporter with an activity close to that of a commonly used protonophore. The fatty acid-enhanced proton transport was also observed for other hydrogen and halogen bond-based synthetic anion transporters. We propose that these compounds induce proton permeability by catalyzing transbilayer movement ("flip-flop") of anionic forms of fatty acids, so allowing the fatty acids to complete a proton transport cycle. Several lines of evidence have been provided to support such a fatty acid cycling mechanism. Our findings open up new applications of anion receptor chemistry and provide important clues for understanding biological activities of synthetic anion transporters and potentially the uncoupling mechanism of naturally occurring membrane proteins.

The rational design of biomimetic skin barrier lipid formulations using biophysical methods.

The focus of this communication was to study phospholipid-structured emulsions whose phase behaviour is modified with monoalkyl fatty amphiphiles. Ideally, these systems would mimic key physical and structural attributes observed in human stratum corneum (SC) so that they better alleviate xerotic skin conditions. Phosphatidylcholine-structured emulsions were prepared, and their phase behaviour modified with monoalkyl fatty amphiphiles. The effect of molecular volume, acyl chain length and head-group interactions was studied using a combination of physical methods. Water vapour transmission rate (WVTR) was used as a primary test to assess occlusive character. Changes in the vibrational modes observed in Fourier transform infrared (FTIR) spectroscopy and bilayer spacing measured by X-ray diffraction (XRD) were then applied to elucidate the lateral and lamellar microstructural characteristics in the systems. Water vapour transmission rate demonstrated that as the phosphatidylcholine acyl chain length increased from C14, to C18, to C22, there was a corresponding increase in occlusive character. The addition of monoalkyl fatty amphiphiles such as behenic acid, behenyl alcohol or cetostearyl alcohol to a base formulation incorporating dipalmitoyl and distearoylphosphatidylcholine (C18) was seen to further increase barrier characteristics of the emulsions. FTIR methods used to probe lipid-chain conformational ordering demonstrated that as phosphatidylcholine acyl chain lengths increased, there was a corresponding improvement in acyl chain ordering, with an increase in thermal transition temperatures. The addition of a monoalkyl fatty amphiphile resulted in conformational order and thermal transition temperature improvements trending towards those observed in stratum corneum. FTIR also demonstrated that systems containing behenic acid or behenyl alcohol exhibited features associated with orthorhombic character. X-ray diffraction data showed that addition of monoalkyl fatty amphiphile also resulted in thicker lamellar structures than when those agents are not present. The generalized approach described herein is shown to mechanistically describe the occlusive character of phospholipid-structured formulations in the presence of long-chain fatty acids or alcohols and that they exhibit characteristics mimicking those found in human SC lipids.

Inhibitory Effect of Long-Chain Fatty Acids on Biogas Production and the Protective Effect of Membrane Bioreactor.

Anaerobic digestion of lipid-containing wastes for biogas production is often hampered by the inhibitory effect of long-chain fatty acids (LCFAs). In this study, the inhibitory effects of LCFAs (palmitic, stearic, and oleic acid) on biogas production as well as the protective effect of a membrane bioreactor (MBR) against LCFAs were examined in thermophilic batch digesters. The results showed that palmitic and oleic acid with concentrations of 3.0 and 4.5 g/L resulted in >50% inhibition on the biogas production, while stearic acid had an even stronger inhibitory effect. The encased cells in the MBR system were able to perform better in the presence of LCFAs. This system exhibited a significantly lower percentage of inhibition than the free cell system, not reaching over 50% at any LCFA concentration tested.

Cooperative effects of fatty acids and n-butanol on lipid membrane phase behavior

Biodiesel-derived crude glycerol can be fermented to produce n-butanol, which is a platform chemical for biorefining and a biofuel. One limitation to crude glycerol fermentation is the presence of long-chain fatty acids (FAs) that can partition into cellular membranes, leading to membrane fluidization and interdigitation, which can inhibit cellular function. In this work, we have examined the phase behavior of dipalmitoylphosphatidylcholine (DPPC, C16:0) membranes and the membrane partitioning of n-butanol as a function of FA degree of unsaturation (steric, oleic, and linoleic acids) using differential scanning calorimetry (DSC) and monolayer surface pressure studies. All three FAs at 15mol% (85mol% DPPC) prevented interdigitation by n-butanol based on the DSC results. n-Butanol partitioning and membrane expansion was greatest for DPPC/oleic acid membranes, where monounsaturated oleic acid (OA, C18:1) was miscible in gel and fluid phase DPPC. Saturated steric acid (SA, C18:0), which ordered the membranes and yielded a SA-rich phase during melting, led to a modest increase in n-butanol partitioning compared to DPPC alone. Di-unsaturated linoleic acid (LA, C18:2), which disordered the membranes and phase separated, had little affect on n-butanol partitioning into the DPPC-rich phases. The effects of OA and LA are attributed to the additional interfacial area provided by these FAs due to acyl tail ‘kinks’ at the carbon double bonds. These results show that exogenous FAs can partition into membranes, impacting n-butanol partitioning and acting cooperatively with n-butanol to alter membrane structure.

Regulation of alkane degradation pathway by a TetR family repressor via an autoregulation positive feedback mechanism in a Gram-positive Dietzia bacterium.

n-Alkanes are ubiquitous in nature and serve as important carbon sources for both Gram-positive and Gram-negative bacteria. Hydroxylation of n-alkanes by alkane monooxygenases is the first and most critical step in n-alkane metabolism. However, regulation of alkane degradation genes in Gram-positive bacteria remains poorly characterized. We therefore explored the transcriptional regulation of an alkB-type alkane hydroxylase-rubredoxin fusion gene, alkW1, from Dietzia sp. DQ12-45-1b. The alkW1 promoter was characterized and so was the putative TetR family regulator, AlkX, located downstream of alkW1 gene. We further identified an unusually long 48 bp inverted repeat upstream of alkW1 and demonstrated the binding of AlkX to this operator. Analytical ultracentrifugation and microcalorimetric results indicated that AlkX formed stable dimers in solution and two dimers bound to one operator in a positive cooperative fashion characterized by a Hill coefficient of 1.64 (± 0.03) [k(D) = 1.06 (± 0.16) μM, k(D) ' = 0.05 (± 0.01) μM]. However, the DNA-binding affinity was disrupted in the presence of long-chain fatty acids (C10-C24), suggesting that AlkX can sense the concentrations of n-alkane degradation metabolites. A model was therefore proposed where AlkX controls alkW1 expression in a metabolite-dependent manner. Bioinformatic analysis revealed that the alkane hydroxylase gene regulation mechanism may be common among Actinobacteria.

Thermophilic anaerobic co-digestion of sewage sludge with grease waste: Effect of long chain fatty acids in the methane yield and its dewatering properties

Thermophilic co-digestion of sewage sludge with three different doses of trapped grease waste (GW) from the pre-treatment of a WWTP has been assessed in a CSTR bench-scale reactor. After adding 12% and 27% of grease waste (on COD basis), the organic loading rate increased from 2.2 to 2.3 and 2.8kgCODm−3d−1 respectively, and the methane yield increased 1.2 and 2.2 times. Further GW increase (37% on COD basis) resulted in an unstable methane yield and in long chain fatty acids (LCFA) accumulation. Although this inestability, the presence of volatile fatty acids in the effluent was negligible, showing good adaptation to fats of the thermophilic biomass. Nevertheless, the presence of LCFA in the effluent worsens its dewatering properties. Specific methanogenic activity tests showed that the addition of grease waste ameliorates the acetoclastic activity in detriment of the hydrogenotrophic activity, and suggests that the tolerance to LCFA can be further enhanced by slowly increasing the addition of lipid-rich materials.

Effect of porcine placenta steroid extract on myogenic satellite cell proliferation, transdifferentiation, and lipid accumulation

Intramuscular long-chain fatty acids (LCFAs) play an important role in energy production and initiation of mitochondrial oxidation of lipids. Herein, we report a natural porcine placenta steroid extract (PPSE) that stimulates transdifferentiation and lipid accumulation in bovine myogenic satellite cells (MSCs). The steroids hormones in PPSE were analyzed using enzyme-linked immunosorbant assay and presence of LCFA was established using gas chromatography. At 70% confluent growth, cells were treated with PPSE, LCFAs, transdifferentiation cocktail and commercially available steroid hormones. The working concentrations of all chemicals were manipulated similar to PPSE. The cells were observed for morphological changes and subjected to quantitative analysis of lipid deposition on Days 2, 4, and 6 of treatment. PPSE-treated MSCs exclusively transformed into lipid-accumulated adipose-like cells (ALCs). However, myotubes or adipocytes were formed in cells treated with other chemicals. Expression of different genes was studied to ascertain the molecular mechanism involved in ALC formation. CD36, fatty acid binding protein 4, and peroxisome proliferator-activated receptor-gamma were up-regulated. The expression of CD36 was established through immunocyto-chemical analysis. A viability assay was used to confirm the effect of PPSE on proliferation of MSCs. Hence, a natural steroid extract from porcine was found as a nontoxic mixture, which induces lipid accumulation and transdifferentiation of MSCs to ALCs. From the gene expression studies, it was established that the extract works almost in homogenous manner with other lipid inducers.