Fatty acids,part 54. Some reactions of long-chain oxygenated acids with special reference to those furnishing furanoid acids

C₁₈ furanoid acids are prepared from natural oxygenated acids by palladium (II)-catalysed cyclodehydrogenation, by rearrangement of epoxides with iodopropane-sodium iodide-dimethylsulphoxide, and by dehydration of endoperoxides. Some reactions give mixed products but routes to the individual 10,13-, 9,12- and 8,11-furans are reported. The endoperoxide route leads to speculation about the biosynthesis of natural furanoid acids.     

Polymerase chain reaction optimization for amplification of Guanine-Cytosine rich templates using buccal cell DNA

CONTEXT:

Amplification of Guanine-Cytosine (GC) -rich sequences becomes important in screening and diagnosis of certain genetic diseases such as diseases arising due to expansion of GC-rich trinucleotide repeat regions. However, GC-rich sequences in the genome are refractory to standard polymerase chain reaction (PCR) amplification and require a special reaction conditions and/or modified PCR cycle parameters.

AIM:

Optimize a cost effective PCR assay to amplify the GC-rich DNA templates.

SETTINGS AND DESIGN:

Fragile X mental retardation gene (FMR 1) is an ideal candidate for PCR optimization as its GC content is more than 80%. Primers designed to amplify the GC rich 5’ untranslated region of the FMR 1 gene, was selected for the optimization of amplification using DNA extracted from buccal mucosal cells.

MATERIALS AND METHODS:

A simple and rapid protocol was used to extract DNA from buccal cells. PCR optimization was carried out using three methods, (a) substituting a substrate analog 7-deaza-dGTP to dGTP (b) in the presence of a single PCR additive and (c) using a combination of PCR additives. All PCR amplifications were carried out using a low-cost thermostable polymerase.

RESULTS:

Optimum PCR conditions were achieved when a combination of 1M betaine and 5% dimethyl sulfoxide (DMSO) was used.

CONCLUSIONS:

It was possible to amplify the GC rich region of FMR 1 gene with reproducibility in the presence of betaine and DMSO as additives without the use of commercially available kits for DNA extraction and the expensive thermostable polymerases.     

Nhs: network-based hierarchical segmentation for cryo-electron microscopy density maps

Cryo-electron microscopy (cryo-EM) experiments yield low-resolution (3-30 ?) 3D-density maps of macromolecules. These density maps are segmented to identify structurally distinct proteins, protein domains, and subunits. Such partitioning aids the inference of protein motions and guides fitting of high-resolution atomistic structures. Cryo-EM density map segmentation has traditionally required tedious and subjective manual partitioning or semisupervised computational methods, whereas validation of resulting segmentations has remained an open problem in this field. We introduce a network-based hierarchical segmentation (Nhs) method, that provides a multi-scale partitioning, reflecting local and global clustering, while requiring no user input. This approach models each map as a graph, where map voxels constitute nodes and weighted edges connect neighboring voxels. Nhs initiates Markov diffusion (or random walk) on the weighted graph. As Markov probabilities homogenize through diffusion, an intrinsic segmentation emerges. We validate the segmentations with ground-truth maps based on atomistic models. When implemented on density maps in the 2010 Cryo-EM Modeling Challenge, Nhs efficiently and objectively partitions macromolecules into structurally and functionally relevant subregions at multiple scales.     

The Evolution of Strategic Timing in Collective-Risk Dilemmas

In collective-risk dilemmas, a group needs to collaborate over time to avoid a catastrophic event. This gives rise to a coordination game with many equilibria, including equilibria where no one contributes, and thus no measures against the catastrophe are taken. In this game, the timing of contributions becomes a strategic variable that allows individuals to interact and influence one another. Herein, we use evolutionary game theory to study the impact of strategic timing on equilibrium selection. Depending on the risk of catastrophe, we identify three characteristic regimes. For low risks, defection is the only equilibrium, whereas high risks promote equilibria with sufficient contributions. Intermediate risks pose the biggest challenge for cooperation. In this risk regime, the option to interact over time is critical; if individuals can contribute over several rounds, then the group has a higher chance to succeed, and the expected welfare increases. This positive effect of timing is of particular importance in larger groups, where successful coordination becomes increasingly difficult.     

Conservation of Dynamics Associated with Biological Function in an Enzyme Superfamily

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Analgesic Effect of Piracetam on Peripheral Neuropathic Pain Induced by Chronic Constriction Injury of Sciatic Nerve in Rats

Despite immense advances in the treatment strategies, management of neuropathic pain remains unsatisfactory. Piracetam is a prototype of nootropic drugs, used to improve cognitive impairment. The present study was designed to investigate the effect of piracetam on peripheral neuropathic pain in rats. Neuropathic pain was induced by the chronic constriction injury of the sciatic nerve. Following this, piracetam was intraperitoneally administered for 2 weeks in doses of 50, 100 and 200 mg/kg, and pain was assessed by employing the behavioural tests for thermal hyperalgesia (hot plate and tail flick tests) and cold allodynia (acetone test). After the induction of neuropathic pain, significant development of thermal hyperalgesia and cold allodynia was observed. The administration of piracetam (50 mg/kg) did not have any significant effect on all the behavioural tests. Further, piracetam (100 mg/kg) also had no effect on the hot plate and tail flick tests; however it significantly decreased the paw withdrawal duration in the acetone test. Piracetam in a dose of 200 mg/kg significantly modulated neuropathic pain as observed from the increased hot plate and tail flick latencies, and decreased paw withdrawal duration (in acetone test). Therefore, the present study suggests the potential use of piracetam in the treatment of neuropathic pain, which merits further clinical investigation.     

Cascade Screening for Fragile X Syndrome/CGG Repeat Expansions in Children Attending Special Education in Sri Lanka

Fragile X syndrome (FXS) is the commonest cause of inherited mental retardation and clinically presents with learning, emotional and behaviour problems. FXS is caused by expansion of cytosine-guanine-guanine (CGG) repeats present in the 5’ untranslated region of the FMR1 gene. The aim of this study was to screen children attending special education institutions in Sri Lanka to estimate the prevalence of CGG repeat expansions. The study population comprised a representative national sample of 850 children (540 males, 310 females) with 5 to 18 years of age from moderate to severe mental retardation of wide ranging aetiology. Screening for CGG repeat expansion was carried out on DNA extracted from buccal cells using 3’ direct triplet primed PCR followed by melting curve analysis. To identify the expanded status of screened positive samples, capillary electrophoresis, methylation specific PCR and Southern hybridization were carried out using venous blood samples. Prevalence of CGG repeat expansions was 2.2%. Further classification of the positive samples into FXS full mutation, pre-mutation and grey zone gave prevalence of 1.3%, 0.8% and 0.1% respectively. All positive cases were male. No females with FXS were detected in our study may have been due to the small sample size.     

Practical synthesis of 1,3-oleoyl 2-docosahexaenoylglycerol by lipase-catalyzed reactions: An evaluation of different reaction routes

Three new synthetic routes were critically evaluated for the lipase-catalyzed production of 1,3-oleoyl-2 docosahexaenoylglycerol (ODO) in relatively large-scale (approximately 200 g). First, the production of 1,3-diolein by the reaction of glycerol and oleic acid followed by incorporation of docosahexaenoic (DHA) ethyl ester at the sn-2 position was studied. 1,3-Diolein was produced in 68.3% and 84.6% yield when stoichiometric amounts of the substrates were reacted at 25 °C for 8 h in the presence of 10% Novozym 435 and Lipozyme RM IM, respectively. Further increase in reaction temperature and time led to decrease in the 1,3-diolein yield. However, only a 9.4% yield of triacylglycerol was obtained in the subsequent reaction step when the 1,3-diolein was reacted with DHA ethyl ester in the presence of Novozym 435. Secondly, the feasibility of direct acidolysis was studied. Acidolysis of single cell oil (SCO) in excess oleic acid using Novozym 435 as the catalyst occurred twice as fast in solvent (tert-butanol) compared to a solvent-free system, and 63% oleic acid was incorporated into SCO. However, the regio-isomeric purity of the product was poor. Finally, the ethanolysis of SCO to produce DHA-enriched 2-monoacylglycerol followed by esterification with oleic acid or ethyl oleate was investigated. ODO was obtained in 50.9% regio-purity by Lipozyme RM IM-catalyzed esterification. The latter method was the most feasible for preparing ODO in large-scale. This synthetic route could be adapted for related triacylglycerols containing highly polyunsaturated when their productions in large-scale and high regio-purity are required.     

The action of lipoxygenase-1 on furan derivatives.

Several 2,5-disubstituted furans, which are known to react with peroxyacids, singlet oxygen and other active forms of oxygen were tested as potential inhibitors, co-oxidants, or substrates for soybean lipoxygenase. The furan, 10,13-epoxy-octadeca-10,12-dienoic acid, methyl ester (IV) was converted by lipoxygenase or singlet oxygen or peroxyacid to the acyclic product, methyl 10,13-dioxo-octadec-11-enoate. Apparently furan IV is able to interact with an active site of lipoxygenase (Km = 220 microM). 2,5-Dimethylfuran (I), 2,5-diphenylfuran (II) and 3-(5'-methyl-2'-furyl)propenoic acid (III) were neither substrates nor inhibitors of lipoxygenase activity. Lipoxygenase-catalyzed oxidation of furan (IV), which is inhibited by hydroquinone, is explained by a mechanism involving lipoxygenase-superoxide complex and furan-radical intermediates. Also described is the selective cleavage of furan rings by m-chloroperoxybenzoic acid to yield the 1,4-diketoethylene functional system.