Synthesis of Silver Embedded Poly(o-Anisidine) Molybdophosphate Nano Hybrid Cation-Exchanger Applicable for Membrane Electrode

Poly(o-anisidine) molybdophosphate was expediently obtained by sol-gel mixing of Poly(o-anisidine) into the inorganic matrices of molybdophosphate, which was allowed to react with silver nitrate to the formation of poly(o-anisidine) molybdophosphate embedded silver nano composite. The composite was characterized by Fourier Transform Infrared Spectroscopy, X-ray powder diffraction, UV-Vis Spectrophotometry, Fluorescence Spectroscopy, Scanning Electron Microscopy/Energy-dispersive X-ray Spectroscopy and Thermogravimertic Analysis. Ion exchange capacity and distribution studies were carried out to understand the ion-exchange capabilities of the nano composite. On the basis of highest distribution studies, this nano composite cation exchanger was used as preparation of heavy metal ion selective membrane. Membrane was characterized for its performance as porosity and swelling later on was used for the preparation of membrane electrode for Hg(II), having better linear range, wide working pH range (2–4.5) with fast response in the real environment.     

LEAP2 Is an Endogenous Antagonist of the Ghrelin Receptor

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Rapid multiplex PCR based species identification of wild tigers using non-invasive samples

Conservation and management of rare and elusive species requires accurate data on presence or absence. In such cases, molecular genetics based species identification approaches can prove invaluable, especially in conjuncture with non-invasive DNA sampling. However, non-invasive sources yield DNA in low concentration that is degraded, which could result in false negatives for species identification. In this paper, we developed a set of primers for PCR-based species identification of tigers. Our results reveal high rates (upto 90%) of species identification for both fresh (less than 48 h) and old (between 7 days and 3 months) fecal samples from the field. Experiments reveal that multiplex PCR (amplifying more than one genomic region) results in an increase in conclusive species identification (and a consequent decrease in the number of false negatives) from 55% to 89% for old fecal samples. We demonstrate that this increased success is because we experimentally overcome the problems of low DNA template quantity (using the multiplex PCR kit, increases species identification from 55% to 72%) and low template DNA quality (two sets of primers increase the species identification success from 72% to 89%). We recommend that multiplex PCR based methods be used (in conjuncture with species specific primers) for other rare and elusive species since such methods will potentially significantly decrease error in species identification.     

Multienzyme microbiosensor based on electropolymerized o-phenylenediamine for simultaneous in vitro determination of acetylcholine and choline

The electrochemical biosensors based on poly(o-phenylenediamine) (PoPD) and acetylcholinesterase (AChE) and choline oxidase (ChO) enzymes were fabricated on carbon fibre (CF) substrate. The electropolymerized PoPD was used to reduce the interfering substances. The electrode assembly was completed by depositing functionalized carbon nano tubes (FCNTs) and Nafion (Naf). Amperometric detection of acetylcholine (ACh) and choline (Ch) were realized at an applied potential of +750 mV vs Ag/AgCl (saturated KCl). At pH 7.4, the final assembly, Naf-FCNTs/AChE-ChO((10:1))/PoPD/CF(Elip), was observed to have high sensitivity towards Ch (6.3±0.3 μA mM(-1)) and ACh (5.8±0.3 μA mM(-1)), linear range for Ch (K(M)=0.52±0.03 mM) and ACh (K(M)=0.59±0.07 mM), and for Ch the highest ascorbic acid blocking capacity (97.2±2 1mM AA). It had a response time of <5s and with 0.045 μM limit of detection. Studies on different ratio (ACh/Ch) revealed that 10:1, gave best overall response.     

Discovery of pyrazole carboxylic acids as potent inhibitors of rat long chain L-2-hydroxy acid oxidase

Long chain L-2-hydroxy acid oxidase 2 (Hao2) is a peroxisomal enzyme expressed in the kidney and the liver. Hao2 was identified as a candidate gene for blood pressure (BP) quantitative trait locus (QTL) but the identity of its physiological substrate and its role in vivo remains largely unknown. To define a pharmacological role of this gene product, we report the development of selective inhibitors of Hao2. We identified pyrazole carboxylic acid hits 1 and 2 from screening of a compound library. Lead optimization of these hits led to the discovery of 15-XV and 15-XXXII as potent and selective inhibitors of rat Hao2. This report details the structure activity relationship of the pyrazole carboxylic acids as specific inhibitors of Hao2.     

Comparative cellular and molecular analyses of pooled bone marrow multipotent mesenchymal stromal cells during continuous passaging and after successive cryopreservation

The clinical application of human bone marrow derived multipotent mesenchymal stromal cells (MSC) requires expansion, cryopreservation, and transportation from the laboratory to the site of cell implantation. The cryopreservation and thawing process of MSCs may have important effects on the viability, growth characteristics and functionality of these cells both in vitro and in vivo. More importantly, MSCs after two rounds of cryopreservation have not been as well characterized as fresh MSCs from the transplantation perspective. The objective of this study was to determine if the effect of successive cryopreservation of pooled MSCs during the exponential growth phase could impair their morphology, phenotype, gene expression, and differentiation capabilities. MSCs cryopreserved at passage 3 (cell bank) were thawed and expanded up to passage 4 and cryopreserved for the second time. These cells (passive) were then thawed and cultured up to passage 6, and, at each passage MSCs were characterized. As control, pooled passage 3 cells (active) after one round of cryopreservation were taken all the way to passage 6 without cryopreservation. We determined the growth rate of MSCs for both culture conditions in terms of population doubling number (PDN) and population doubling time (PDT). Gene expression profiles for pluripotency markers and tissue specific markers corresponding to neuroectoderm, mesoderm and endoderm lineages were also analyzed for active and passive cultures of MSC. The results show that in both culture conditions, MSCs exhibited similar growth properties, phenotypes and gene expression patterns as well as similar differentiation potential to osteo‐, chondro‐, and adipo‐lineages in vitro. To conclude, it appears that successive or multiple rounds of cryopreservation of MSCs did not alter the fundamental characteristics of these cells and may be used for clinical therapy. J. Cell. Biochem. 113: 3153–3164, 2012. © 2012 Wiley Periodicals, Inc.     

Suppression of cortical functional hyperemia to vibrissal stimulation in the rat by epoxygenase inhibitors

Application of glutamate to glial cell cultures stimulates the formation and release of epoxyeicosatrienoic acids (EETs) from arachidonic acid by cytochome P-450 epoxygenases. Epoxygenase inhibitors reduce the cerebral vasodilator response to glutamate and N-methyl-D-aspartate. We tested the hypothesis that epoxygenase inhibitors reduce the somatosensory cortical blood flow response to whisker activation. In chloralose-anesthetized rats, percent changes in cortical perfusion over whisker barrel cortex were measured by laser-Doppler flowmetry during whisker stimulation. Two pharmacologically distinct inhibitors were superfused subdurally: 1) N-methylsulfonyl-6-(2-propargyloxyphenyl)hexanamide (MS-PPOH), an epoxygenase substrate inhibitor; and 2) miconazole, a reversible cytochrome P-450 inhibitor acting on the heme moiety. Superfusion with 5 micromol/l MS-PPOH decreased the hyperemic response to whisker stimulation by 28% (from 25 +/- 9 to 18 +/- 7%, means +/- SD, n = 8). With 20 micromol/l MS-PPOH superfusion, the response was decreased by 69% (from 28 +/- 9% to 9 +/- 4%, n = 8). Superfusion with 20 micromol/l miconazole decreased the flow response by 67% (from 31 +/- 6% to 10 +/- 3%, n = 8). Subsequent superfusion with vehicle restored the response to 26 +/- 11%. Indomethacin did not prevent MS-PPOH inhibition of the flow response, suggesting that EET-related vasodilation was not dependent solely on cyclooxygenase metabolism of 5,6-EET. Neither MS-PPOH nor miconazole changed baseline flow, reduced the blood flow response to an adenosine A(2) agonist, or decreased somatosensory evoked potentials. The marked reduction of the cortical flow response to whisker stimulation with two different types of epoxygenase inhibitors indicates that EETs play an important role in the physiological coupling of blood flow to neural activation.     

Alpha-crystallin and ATP facilitate the in vitro renaturation of xylanase: enhancement of refolding by metal ions

Alpha-crystallin is a multimeric protein that functions as a molecular chaperone and shares extensive structural homology to small heat shock proteins. For the functional in vitro analysis of alpha-crystallin, the xylanase Xyl II from alkalophilic thermophilic Bacillus was used as a model system. The mechanism of chaperone action of alpha-crystallin is less investigated. Here we studied the refolding of Gdn HCl-denatured Xyl II in the presence and absence of alpha-crystallin to elucidate the molecular mechanism of chaperone-mediated in vitro folding. Our results, based on intrinsic tryptophan fluorescence and hydrophobic fluorophore 8-anilino-1-naphthalene sulfonate binding studies, suggest that alpha-crystallin formed a complex with a putative molten globule-like intermediate in the refolding pathway of Xyl II. The alpha-crystallin.Xyl II complex exhibited no functional activity. Addition of ATP to the complex initiated the renaturation of Xyl II with 30%-35% recovery of activity. The nonhydrolyzable analog 5'-adenylyl imidodiphosphate (AMP-PNP) was capable of reconstitution of active Xyl II to a lesser extent than ATP. Although the presence of Ca(2+) was not required for the in vitro refolding of Xyl II, the renaturation yield was enhanced in its presence. Experimental evidence indicated that the binding of ATP to the alpha-crystallin.Xyl II complex brought about conformational changes in alpha-crystallin facilitating the dissociation of xylanase molecules. This is the first report of the enhancement of alpha-crystallin chaperone functions by metal ions.