1,721,023 research outputs found
From batch to flow synthesis of purine ribonucleosides by enzymatic transglycosylation
No full textPurine nucleoside phosphorylases (PNPs, EC 2.4.2.1) catalyze the reversible phosphorolysis of the glycosydic bond of purine nucleosides; upon addition of a second nucleobase, these enzymes may transfer the glycosyl moiety to it, resulting in the chemo-, regio- and stereoselective formation of a new nucleoside (transglycosylation, Scheme 1). This chemoenzymatic process represents an advantageous alternative to conventional chemical strategies which are frequently hampered by several drawbacks such as multistep processes, need for protecting groups, low chemo-, regio- and stereoselectivity.
A PNP from Aeromonas hydrophila (AhPNP) has been recently characterized in terms of substrate specificity [1], also upon immobilization on the inner surface of a silica capillary coupled on-line with a chromatographic column [2].
Because of its wide substrate specificity, AhPNP was then exploited to catalyze the “one-pot, one-enzyme” transglycosylation of 7-methylguanosine iodide with a series of 6-substituted purines, resulting in a moderate to high conversion (18-65%) of the bases into a 23-compound library of 6-substituted purine-9-ribosides. Moreover, AhPNP was covalently immobilized (25 mg immobilized enzyme, 50% yield) in a pre-packed stainless steel column containing aminopropyl silica particles. The resulting AhPNP-IMER (Immobilized Enzyme Reactor) was coupled to a HPLC apparatus containing an analytical or a semi-preparative chromatographic column associated with a UV-visible detector. This system was used to synthesize five 6-substituted purine ribonucleosides at a mg scale by transglycosylation through a “flow-based” approach. Coupling of transglycosylation reaction and product separation resulted in a fast and efficient process (52-89% conversion) with minimized sample handling. To date, AhPNP-IMER completely retained its activity upon 50 reactions in 10 months.
[1] D. Ubiali, C. D. Serra, I. Serra, C. F. Morelli, M. Terreni, A. M. Albertini, P. Manitto, G. Speranza Adv. Synth. Catal. 2012, 354, 96-104
[2] E. Calleri, D. Ubiali, I. Serra, C. Temporini, G. Cattaneo, G. Speranza, C. F. Morelli, G. Massolini J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 2014, 968, 79-8
A fully digital fast convergence algorithm for nonlinearity correction in multistage ADC
No full textThis paper describes a fast digital background calibration algorithm for pipeline or cyclic analog-to-digital converters. The proposed method corrects for the finite gain/bandwidth of the interstage operational amplifier and for capacitor mismatch in multiplying digital-to-analog converters stages. The new algorithm, fast gain error correction, converges more than 100 times faster than other correlation-based correction techniques presented in literature. The high speed of convergence of the error estimation is due to the use of a polynomial interpolation that cancel the interference in the error estimation process caused by the input signal. This new method does not need any added analog circuitry, does not perturb the output samples, and requires only a digital finite-impulse response filter to implement the polynomial interpolation as opposed to existing fast converging correlation techniques
Pullulan as a permanent hydrophilic coating in capillary electrophoresis: applications to penicillin G acylase as chiral selector.
No full textINITIAL CHARACTERIZATION OF STEM BARK EXTRACTS FROM PHYLLANTHUS MUELLERIANUS AS SOURCE OF NEW NATURAL ENTITIES WITH ANTI-CHOLINESTERASE PROPERTIES
No full textThe plant kingdom is an endless source of chemically diverse bioactive compounds, which are used in traditional folk medicine for the treatment of a wide array of diseases. A previous investigation on the bioactive phytocomponents present in the methanol extract of Phyllanthus muellerianus (PMME) demonstrated an interesting activity against C. sporogenes (MIC= 100 μg/ml) and S. pyogenes (MIC= 300 μg/ml) [1], which supported the traditional use of the extract by local populations in Cameroon.
Looking for activity beyond the claimed traditional use [2], PMME was evaluated on human cholinesterase enzymes, namely acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) as selected targets for Alzheimer’s disease (AD) treatment. Indeed AChE inhibitors derived from natural products (galanthamine and rivastigmine) are currently licensed to alleviate cognitive symptoms in dementia. Due to the limited available pharmacological treatments for AD, extensive research has been directed towards the identification of other AChE inhibitors, arising from the plant kingdom. A rational basis for our investigation on the bark extracts of Phyllanthus muellerianus is related to a study carried out in 2007 by Joshi e Parle, in which the antiamnesic potential for Phyllanthus amarus in mice was demonstrated [3].
Anticholinesterase activity was in vitro evaluated by Ellman’s assay [4] using recombinant human AChE and BuChE from human serum.
Due to the interesting activity found for the de-fatted PMME (% inhibition at 100 g mL-1 on hAChE = 52.6 ± 0.6, on hBuChE = 70.1 ± 3.1%, n=4), PMME was fractionated by flash chromatography affording six fractions (PMF1-6); PMF1, PMF2 and PMF4 significantly inhibited human cholinesterases, thus they were further purified by a RP-18 solid phase extraction.
The bio-guided fractionation allowed the identification of three active phytocomponents, with different selectivities against the two ChEs. Two out of the three bioactives have been isolated so far: compound A from PMF1 and compound B from PMF4, both alkaloids. Compound A showed to be a hBuChE selective inhibitor (IC50 = 44.8 ± 3.0 g mL-1 and IC50 = 382 ± 15 g mL-1, for hBuChE and hAChE, respectively) while compound B showed comparable inhibitory potencies against both enzymes (IC50 = 5.45 ± 0.20 g mL-1 and IC50 = 12.6 ± 0.4 g mL-1, for hAChE and BuChE, respectively).
Compared to the commercially available AD drug galanthamine, compound B is one order of magnitude less active on hAChE (galanthamine IC50 = 0.6 g mL-1), representing, however, a potential new natural scaffold to undergo towards optimization.
The chemical structure of the two isolated alkaloids is under investigation by 13C and 1H-NMR. Moreover, the isolation of compound C in suitable amount for the evaluation of the biological profile and its structural elucidation has being carried out.
References
[1] G. Brusotti, I. Cesari, G. Frassa, P. Grisoli, C. Dacarro, G. Caccialanza, J. Ethnopharmacol., 2011, 35, 797–800.
[2] R. Brisson, Etudes pygmées, SELAF n 376, Ed Peeters, 1999, Paris.
[3] H. Joshi and M. Parle, African Journal of Biomedical. Research., 2007, 10, 165–173.
[4] G.L. Ellman, K.D. Courtney, V. Jr. Andres, R.M. Feather-Stone, Biochem. Pharmacol., 1961, 7, 88-9
PURINE NUCLEOSIDE PHOSPHORYLASES AS BIOCATALYSTS AND PHARMACOLOGICAL TARGETS
Full text linkA purine nucleoside phosphorylase from Aeromonas hydrophila (AhPNP) was successfully exploited to catalyze the “one-pot, one-enzyme” regio- and stereoselective transfer of β-D-ribose from a proper sugar donor (7-methylguanosine iodide) to a library of 6-substituted purine acceptors, resulting in the “in batch” synthesis of 24 ribonucleosides. Transglycosylation conversions confirmed the broad tolerance and the potential of AhPNP as biocatalyst, providing the necessary information to undertake the preparative synthesis of 6-modified purine nucleosides. [1]
AhPNP was then immobilized in a stainless steel column resulting in a stable and active bioreactor (AhPNP-IMER, Immobilized Enzyme Reactor) that, upon on-line connection to a semi-preparative HPLC system, was used to run transglycosylations in a flow mode. In such a set-up, biotransformation, on-line monitoring and product purification occurred in a single integrated platform, thus allowing the preparation of five nucleoside analogues at a mg scale (52-89% yield). [2]
As a step forward, a “one-pot, two-enzyme” strategy was applied by coupling AhPNP-IMER with an analogous bioreactor based on a uridine phosphorylase from Clostridium perfringens (CpUP), immobilized in a monolith column. The on-line apparatus obtained by connecting CpUP-IMER and AhPNP-IMER in series was tested in the synthesis of adenosine, 2’-deoxyadenosine and arabinosyladenine from uridine, 2’-deoxyuridine and arabinosyluracyl as sugar donors, respectively. The corresponding nucleobases were transformed into the products in 90-95% conversion over 1 h for the ribosyl and 2’-deoxyribosyl derivatives, and 20% conversion after 5 h for arabinosyladenine. [3]
Furthermore, a new LC-ESI-MS/MS method was set up to evaluate the inhibition activity of 8-substituted purine ribonucleosides toward the PNP from Mycobacterium tuberculosis (MtPNP), as well as the selectivity against the microbial enzyme with respect to the corresponding human one (HsPNP). The corresponding enzymatic assay, based on the phosphorolysis of inosine, proved to be very convenient in terms of time as well as of target amount. A small library of seven 8-substituted purine ribonucleosides were screened, not exerting any significant effect up to 1 mM, with 8-bromoguanosine and 8-methylaminoguanosine being the only exceptions at 500 mM as weak inhibitors. [4]
Finally, the chemical synthesis of a series of 8- and N2-substituted inosinic and guanylic acids as potential ligands of the human GPR17 receptor was carried out, starting from studies aided by molecular modeling on a homology model of the target. The molecules were prepared by 5’-phosphorylation of properly 8- and N2-modified/protected inosine or guanosine. Owing to the scarce nucleophilicity of the exocyclic NH2 group of guanosine, the 2-position of the purine ring was activated as a bromo derivative, whose displacement with the proper amine afforded the desired N2-alkylated products. On the contrary, N2-acylations were carried out through nitrogen functionalization with a proper acyl chloride or anhydride. An additional 2’,3’-O-isopropylidene group was inserted in all the N2-functionalized nucleotides. Binding assays on GPR17 will be carried out.
[1] D. Ubiali, C. F. Morelli, M. Rabuffetti, G. Cattaneo, I. Serra, T. Bavaro, A. M. Albertini, G. Speranza Curr. Org. Chem. 2015, 19, 2220-2225; [2] E. Calleri, G. Cattaneo, M. Rabuffetti, I. Serra, T. Bavaro, G. Massolini, G. Speranza, D. Ubiali Adv. Synth. Catal. 2015, 357, 2520-2528; [3] G. Cattaneo, M. Rabuffetti, G. Speranza, T. Kupfer, B. Peters, G. Massolini, D. Ubiali, E. Calleri Submitted 2017; [4] G. Cattaneo, D. Ubiali, E. Calleri, M. Rabuffetti, G. C. Hofner, K. T. Wanner, M. C. De Moraes, L. K. B Martinelli, D. S. Santos, G. Speranza Anal. Chim. Acta 2016, 943, 89-97
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
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
- …
