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Polietilenglicoli modificati e loro complessi supramolecolari con macromolecole biologicamente attive
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Effective G-CSF Site Directed PEGylation of Hindered Aminoacids.
No full textA new alkyl maleimido activated monomethoxypoly(ethylene glycol) (PEG-C18-Mal) for site-directed protein conjugation was synthesised and characterized. As compared to the commercial mPEG-maleimido, PEG-C18-Mal was more efficient in the bioconjugation to 17Cys of rh-G-CSF in guanidinium free buffer and gave more stable bioconjugate. Also, the new bioconjugate displayed high biological activity
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Self-assembling nanocomposites for protein delivery: Supramolecular interactions between PEG-cholane and rh-G-CSF.
No full textPEG(5kDa)-cholane, PEG(10kDa)-cholane and PEG(20kDa)-cholane self-assembling polymers have been synthesised by the end-functionalisati`on of 5, 10 and 20kDa linear amino-terminating monomethoxy-poly(ethylene glycol) (PEG-NH(2)) with 5β-cholanic acid. Spectroscopic studies and isothermal titration calorimetry showed that the CMC of the PEG-cholane derivatives increased from 23.5±1.8 to 60.2±2.4μM as the PEG molecular weight increased. Similarly, light scattering analysis showed that the micelle size increased from 15.8±4.9 to 23.2±11.1nm with the PEG molecular weight. Gel permeation studies showed that the polymer bioconjugates associate with recombinant human granulocyte colony stimulating factor (rh-G-CSF) to form supramolecular nanocomposites according to multi-modal association profiles. The protein loadings obtained with PEG(5kDa)-cholane, PEG(10kDa)-cholane and PEG(20kDa)-cholane were 7.4±1.1, 2.7±0.3 and 2.1±0.4% (protein/polymer, w/w %), respectively. Scatchard and Klotz analyses showed that the protein/polymer affinity constant increased and that the number of PEG-cholane molecules associated to rh-G-CSF decreased as the PEG molecular weight increased. Isothermal titration calorimetry confirmed the protein/polymer multi-modal association. Circular dichroism analyses showed that the polymer association alters the secondary structure of the protein. Nevertheless, in vitro studies performed with NFS-60 cells showed that the polymer interaction does not impair the biological activity of the cytokine. In vivo studies performed by intravenous and subcutaneous administrations of rh-G-CSF to rats showed that the association with PEG(5kDa)-cholane prolongs the body exposure of the protein. After subcutaneous administration, the protein t(max) values obtained with rh-G-CSF and 1:14 and 1:21 rh-G-CSF/PEG(5kDa)-cholane (w/w ratio) nanocomplexes were 2, 8 and 24h, respectively. The 1:21 (w/w) rh-G-CSF/PEG(5kDa)-cholane formulation resulted in 149% relative bioavailability, and the pharmacokinetic behaviour was similar to that obtained with an equivalent protein dose of rh-G-CSF chemically conjugated with one linear 20-kDa PEG. A single administration of a 1.5mg/kg dose of a 1:21 (w/w) rh-G-CSF/PEG(5kDa)-cholane formulation induced a high production of white blood cells for 96h
A new site-specific monoPEGylated filgrastim derivative prepared by enzymatic conjugation: Production and physicochemical characterization.
No full textWe describe the preparation and characterisation of a new monoPEGylated derivate of a recombinant form of filgrastim (methionyl human granuolcite colony stimulating factor, rh-Met-G-CSF), BK0026, prepared by enzymatic site-specific 20 kDa PEG conjugation to glutamine 135 residue by microbial transglutaminase catalysed reaction. BK0026 was purified to a clinical grade by a single cation exchange chromatography step and characterized using a panel of physicochemical analyses. NH2-terminal sequence and peptide mapping demonstrated no differences between the primary structure of BK0026 and the non-PEGylated filgrastim. The circular dichroism and fluorescence spectroscopy showed the preservation of high order protein structure. The single conjugation site on glutamine 135 was identified by endoproteinase Glu-C peptide mapping combined with mass spectrometry analysis and NH2-terminal sequence of the PEGylated peptides. BK0026 purity as well as product- and process-related contaminants were determined by several analytical methods, which showed that BK0026 is stable for more than 2 years when stored at 4-8 °C. The advantages of enzymatic PEGylation of filgrastim are the absolute specificity of glutamine 135 conjugation combined with high PEGylation yields under very mild reaction conditions. The new site specific monoPEGylated filgrastim is a promising candidate for preclinical and clinical studies aimed at developing a long-lasting treatment of neutropenia in oncological patients under chemotherapy treatment
A New Monopegylated Filgrastim Prepared By Transglutaminase Catalyzed Site Specific Reaction.
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