An efficient method for the application of PHA‐poor solvents to extract polyhydroxybutyrate from Cupriavidus necator

There are many published studies presenting ethanol and acetone as PHAs‐poor solvents, where these two solvents are shown to dissolve <2% (w/v) of PHAs at low temperatures. In this study, the suitability of ethanol and acetone for the recovery of PHB at different temperatures (from room temperature to near boiling point) in Cupriavidus necator was investigated. Experiments were performed using response surface methodology to examine the effects of different temperatures and heating incubation times on recovery percentage using the two solvents. The highest recovery percentage (92.3%) and product purity (up to 99%) were obtained with ethanol‐assisted extraction at 76°C for 32 min of incubation time. Under these conditions the extracted PHB exhibited a molecular mass of 1.2 × 10⁶. The present strategy showed that at temperatures near its boiling point, ethanol, as a nonhalogenated solvent, represents a good alternative to halogenated solvents, like chloroform, when PHB recovery is concerned. DSC analysis showed good thermal properties for ethanol‐ and acetone‐extracted biopolymers. GC and ¹H NMR analysis confirmed the extracted biopolymer to be polyhydroxybutyrate of good purity. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1480–1486, 2016     

Combination effects of mesenchymal stem cells transplantation and anodal transcranial direct current stimulation on a cuprizone-induced mouse model of multiple sclerosis

Journal of Molecular Histology - Multiple sclerosis (MS) has no absolute treatment, and researchers are still exploring to introduce promising therapy for MS. Transcranial direct current...     

Injectable Supramolecular Hydrogel from Insulin-Loaded Triblock PCL-PEG-PCL Copolymer and γ-Cyclodextrin with Sustained-Release Property

Supramolecular hydrogels formed by cyclodextrins and polymers have been widely investigated as a biocompatible, biodegradable and controllable drug delivery system. In this study, a supramolecular hydrogel based on biodegradable poly(caprolactone)–poly(ethylene glycol)–poly(caprolactone) (PCL-PEG-PCL) triblock copolymers and γ-cyclodextrin (γ-CD) was prepared through inclusion complexation as an injectable, sustained-release vehicle for insulin. The triblock copolymer PCL-PEG-PCL was synthesised by the ring-opening polymerisation method, using microwave irradiation. The polymerisation reaction and the copolymer structures were evaluated by nuclear magnetic resonance (NMR) and gel permeation chromatography (GPC). The supramolecular hydrogel was prepared in aqueous solution by blending an aqueous γ-CD solution with an aqueous solution of PCL-PEG-PCL triblock copolymer at room temperature. In vitro insulin release through the hydrogel system was studied. The relative surface hydrophobicity of standard and released insulin from the SMGel was estimated using 8-anilino-1-naphthalene sulfonic acid (ANS). Results of ¹HNMR and gel permeation chromatography revealed that microwave irradiation is a simple and reliable method for synthesis of PCL-PEG-PCL copolymer. Gelation occurred within a minute. The supramolecular hydrogel obtained by mixing 10.54% (w/v) γ-CD and 2.5% (w/v) copolymer had an excellent syringeability. Insulin was released up to 80% over a period of 20 days. Insulin kept its initial folding after formulating and releasing from SMGel. A supramolecular hydrogel based on complexation of triblock PCL-PEG-PCL copolymer with γ-cyclodextrin is a suitable system for providing sustained release of therapeutic proteins, with desirable flow behaviour.Key words: insulin, PCL-PEG-PCL, supramolecular hydrogel, triblock copolymer, γ-CD     

Prevalence of oral mucosal lesions in institutionalized elderly people in Mashhad, Northeast Iran

doi: 10.1111/j.1741‐2358.2011.00588.x
Prevalence of oral mucosal lesions in institutionalized elderly people in Mashhad, Northeast Iran Objective: The aim of this study was to determine the prevalence of oral mucosal lesions in institutionalized elderly people in Mashhad, northeast of Iran. Background: This study was conducted to assess the prevalence of oral lesions in the institutionalized elderly. Methods: In this survey, we studied all of the nursing homes in Mashhad; a total of five encompassing 237 residents. The subjects were examined and lesions recorded in a designed checklist. T‐test, chi‐square and Fisher’s exact tests were used for the analysis. Results: The mean age of the patients was 79.59 ± 8.88. Ninety‐eight per cent of cases had at least one oral mucosal lesion. The most common lesions were fissured tongue (66.5%), atrophic glossitis (48.8%), sublingual varicosity (42%) and xerostomia (38%). There was no significant difference in the prevalence of denture‐related lesions (DRLs) between men and women (p > 0.05). Xerostomia was more prevalent in 70–79‐year‐old than in 60–69‐year‐old subjects. No case of oral malignant lesion was detected. Discussion: These findings revealed a higher prevalence of oral mucosal lesions in Mashhad city compared with other studies conducted in other major cities in Iran and abroad. The results emphasise the necessity of national programmes towards oral health promotion.     

High-level production of amorpha-4,11-diene in a two-phase partitioning bioreactor of metabolically engineered Escherichia coli

Reconstructing synthetic metabolic pathways in microbes holds great promise for the production of pharmaceuticals in large-scale fermentations. By recreating biosynthetic pathways in bacteria, complex molecules traditionally harvested from scarce natural resources can be produced in microbial cultures. Here we report on a strain of Escherichia coli containing a heterologous, nine-gene biosynthetic pathway for the production of the terpene amorpha-4,11-diene, a precursor to the anti-malarial drug artemisinin. Previous reports have underestimated the productivity of this strain due to the volatility of amorphadiene. Here we show that amorphadiene evaporates from a fermentor with a half-life of about 50 min. Using a condenser, we take advantage of this volatility by trapping the amorphadiene in the off-gas. Amorphadiene was positively identified using nuclear magnetic resonance spectroscopy and determined to be 89% pure as collected. We captured amorphadiene as it was produced in situ by employing a two-phase partitioning bioreactor with a dodecane organic phase. Using a previously characterized caryophyllene standard to calibrate amorphadiene production and capture, the concentration of amorphadiene produced was determined to be 0.5 g/L of culture medium. A standard of amorphadiene collected from the off-gas showed that the caryophyllene standard overestimated amorphadiene production by approximately 30%.     

Terminal Heat Tolerance in Bread Wheat Determined by Agronomical Traits and SSR Markers

Since global warming affects wheat cropping systems, more has yet to be indicated on the parameters, which control terminal heat tolerance, and severely influence wheat (Triticum aestivum L.) productivity. Identification of tolerant wheat genotypes by heat tolerance-linked molecular markers is a rapid and cost-effective screening tool in plant breeding. Accordingly, in a four-year field experiment (2015–2019), 44 wheat genotypes were selected out of 100 genotypes, and were examined in timely and late planting (mid-January resulting in heat stress). Stress decreased yield components, including 1000-kernel weight (TKW), grains per spike, and plants per square meter, and the physiological traits, including days to heading and days to maturity, grain filling duration, and greenness, and eventually decreased grain yield up to?~?28%. The early maturity genotypes resulted in higher yields under stress conditions by a stress-avoidance mechanism. Among 14 SSR markers, GWM577 was positively correlated with yield, and WMS3062, GWM261, and WMS1025 had positive correlations with longevity under stress. Accordingly, WMS3062 and GWM261 can be used to determine high yield and early maturity genotypes. Furthermore, GWM114 showed a positive correlation with TKW, indicating their usefulness for grouping wheat genotypes and for identifying heat-related markers. Since the crossing of the genetically distant genotypes can create more diverse populations, the results could be applied to plan breeding projects to establish more diverse populations for different chromosomal locations and traits under heat stress conditions. Moreover, our findings demonstrated that the morphological and molecular analyses could be useful for describing wheat genetic variation of heat tolerance.

     

The NIaIV restriction and modification genes of Neisseria lactamica are flanked by leucine biosynthesis genes

The genes encoding the Neisseria lactamica restriction endonuclease IV (R.NIaIV) and its cognate DNA methyltransferase (M.NlaIV), both of which recognize the sequence GGNNCC, have been cloned in Escherichia coli and overexpressed using the T7 polymerase/promoter system. Analysis of a sequenced 3.58 kb fragment established the gene order, leuD-M.NlaIV-R.NlaIV-leuB. The predicted primary sequence of M.NlaIV (423 amino acids) shows the highest degree of identity to a pair of cytosine-specific methyltransferases, M.BanI (44.9%) and M.HgiCI (44.3%), which recognize the sequence GGYRCC (Y, pyrimidines; R, purines). In contrast, the R.NlaIV protein sequence (243 amino acids) is unique in the existing database, a situation that holds for most endonucleases. Flanking the NlaIV modification and restriction genes are homologues of the leuD and leuB genes of enteric bacteria, which code for enzymes in the leucine biosynthesis pathway. This gene context implies a possible new mode of gene regulation for the RM.NlaIV system, which would involve a mechanism similar to the recently discovered leucine/Lrp regulon in E. coli.     

Regulation of Msh4-Msh5 association with meiotic chromosomes in budding yeast

In the baker’s yeast Saccharomyces cerevisiae, most of the meiotic crossovers are generated through a pathway involving the highly conserved mismatch repair related Msh4-Msh5 complex. To understand the role of Msh4-Msh5 in meiotic crossing over, we determined its genome wide in vivo binding sites in meiotic cells. We show that Msh5 specifically associates with DSB hotspots, chromosome axes, and centromeres on chromosomes. A basal level of Msh5 association with these chromosomal features is observed even in the absence of DSB formation (spo11Δ mutant) at the early stages of meiosis. But efficient binding to DSB hotspots and chromosome axes requires DSB formation and resection and is enhanced by double Holliday junction structures. Msh5 binding is also correlated to DSB frequency and enhanced on small chromosomes with higher DSB and crossover density. The axis protein Red1 is required for Msh5 association with the chromosome axes and DSB hotspots but not centromeres. Although binding sites of Msh5 and other pro-crossover factors like Zip3 show extensive overlap, Msh5 associates with centromeres independent of Zip3. These results on Msh5 localization in wild type and meiotic mutants have implications for how Msh4-Msh5 works with other pro-crossover factors to ensure crossover formation.