Enzyme immobilization on epoxy- and 1,1'-carbonyldiimidazole-activated methacrylate-based monoliths

Monolithic Convective Interaction Media (CIM) have been activated with epoxide and imidazole carbamate functionalities and used as supports for covalent immobilization of protein A, deoxyribonuclease I, and trypsin. The efficiency of immobilization for these proteins was determined from the amount of bound IgG, degradation of DNA, and hydrolysis of Nalpha-benzoyl-L-arginine ethyl ester, respectively. The respective biological activities of trypsin and the binding capacity of protein A immobilized via imidazole carbamate groups were 11.45 and 2.25 times higher than those obtained for epoxide matrix while they were practically equal for deoxyribonuclease I. The kinetics of immobilization was studied in detail for trypsin under dynamic conditions and revealed that the enzyme immobilized via imidazole carbamate groups already reached its highest activity in 5 min. In contrast, a much longer time was required for immobilization via epoxy groups.     

The thermodynamic dissociation constants of ambroxol, antazoline, naphazoline, oxymetazoline and ranitidine by the regression analysis of spectrophotometric data

The mixed dissociation constants of five drug acids—ambroxol, antazoline, naphazoline, oxymetazoline and ranitidine—at various ionic strengths I of range 0.01 and 1.0 and at temperatures of 25 and 37 °C were determined using SQUAD(84) regression analysis of the pH-spectrophotometric titration data. A proposed strategy of efficient experimentation in a protonation constants determination, followed by a computational strategy for the chemical model with a protonation constants determination, is presented on the protonation equilibria of ambroxol. The thermodynamic dissociation constant pK_a^T was estimated by non-linear regression of {pK_a, I} data at 25 and 37 °C: for ambroxol and 8.25 (4), log  and 11.83 (8), for antazoline and 7.83 (6), and 9.55 (2), for naphazoline and 10.63 (1), for oxymethazoline and 10.77 (7), pK_a,2^T=12.03(3) and 11.82 (4) and for ranitidine and 1.77 (1). Goodness-of-fit tests for various regression diagnostics enabled the reliability of the parameter estimates to be found.     

47, 49Ti NMR spectra of half-sandwich titanium(IV) complexes

The 47, 49Ti chemical shifts, resonance line half-widths (Deltanu1/2) and energies of the first electronic charge-transfer transitions (lambdamax1.CT) of Cp'TiX3, where Cp' = eta5-C5H5 (Cp), eta5-C5H4Me (MeCp), eta5-C5HMe4 (Me4Cp), eta5-C5Me5 (Me5Cp), eta5-C5H4SiMe3 (SiCp), eta5-C5H4SnMe3 (SnCp) and eta5-C5H4SiMe2Cl (Si'Cp) and X = Cl, Br, I and OBut, half-sandwich complexes are reported. For the compounds studied, a direct linear relationship between delta(49Ti) and lambdamax1.CT was found.     

Synthesis of 1,3-diazepines and ring contraction to cyanopyrroles

Several tetrazolo[1,5-a]pyridines/2-azidopyridines undergo photochemical nitrogen elimination and ring expansion to 1,3-diazacyclohepta-1,2,4,6-tetraenes, as well as ring cleavage to cyanovinylketenimines, in low temperature Ar matrices. 6,8-Dichlorotetrazolo[1,5-a]pyridine/2-azido-3,5-dichloropridine undergoes ready exchange of the chlorine in position 8 (3) with ROH/RONa. 8-Chloro-6-trifluoromethyltetrazolo[1,5-a]pyridine undergoes solvolysis of the CF(3) group to afford 8-chloro-6-methoxycarbonyltetrazolo[1,5-a]pyridine. Several tetrazolopyridines/2-azidopyridines afford 1H- or 5H-1,3-diazepines in good yields on photolysis in the presence of alcohols or amines. 5-Chlorotetrazolo[1,5-a]pyridines/2-azido-6-chloropyridines and undergo a rearrangement to 1H- and 3H-3-cyanopyrroles and, respectively. The mechanism of this rearrangement was investigated by (15)N-labelling and takes place via transient 1,3-diazepines. The structures of 6,8-dichloro-tetrazolo[1,5-a]pyridine, 6-chloro-8-ethoxytetrazolo[1,5-a]pyridine, dipyrrolylmethane, and 2-isopropoxy-4-dimethylamino-5H-1,3-diazepine were determined by X-ray crystallography. In the latter case, this represents the first reported X-ray crystal structure of a 5H-1,3-diazepine.     

Methacrylate-based short monolithic columns: enabling tools for rapid and efficient analyses of biomolecules and nanoparticles

This review describes the novel chromatography stationary phase--a porous monolithic methacrylate-based polymer--in terms of the design of the columns and some of the features that make these columns attractive for the purification of large biomolecules. We first start with a brief summary of the characteristics of these large molecules (more precisely large proteins like immunoglobulins G and M, plasmid deoxyribonucleic acid (DNA), and viral particles), and a list of some of the problems that were encountered during the development of efficient purification processes. We then briefly describe the structure of the methacrylate-based monolith and emphasize the features which make them more than suitable for dealing with large entities. The highly efficient structure on a small scale can be transferred to a large scale without the need of making column modifications, and the various approaches of how this is accomplished are briefly presented in this paper. This is followed by presenting some of the examples from the bioprocess development schemes, where the implementation of the methacrylate-based monolithic columns has resulted in a very efficient and productive process. Following this, we move back to the analytical scale and demonstrate the efficiency of the monolithic column--where the mass transfer between the stationary and mobile phase is greatly enhanced--for the in-process and final control of the new therapeutics. The combination of an efficient structure and the appropriate hardware results in separations of proteins with residence time less than 0.1 s.     

Additive character of electron donation by methyl substituents within a complete series of polymethylated [1-(η6-Me9n)C6H(6-n))-closo-1,2,3-FeC2B9H11] complexes. Linear correlations of the NMR parameters and Fe(II/III) redox potentials with the number of arene methyls

A systematic method for the incorporation of the {(η(6)-Me(n)C(6)H(6-n))Fe} fragment into the dicarbollide cage was developed based on reactions between [(η(6)-Me(n)C(6)H(6-n))(2)Fe][PF(6)](2) salts (1) and Tl(2)[nido-7,8-C(2)B(9)H(11)]. These reactions proceed with elimination of one arene ligand to generate a complete series of the neutral [1-(η(6)-Me(n)C(6)H(6-n))-closo-1,2,3-FeC(2)B(9)H(11)] (2) complexes with n = 1-6 in yields ranging 15-70% depending on the arene. The structures of mesitylene and pentamethylbenzene complexes were established by X-ray diffraction analyses. All compounds were characterized by (11)B and (1)H NMR measurements, mass spectra, melting points and elemental analyses. Correlations between selected (1)H and (11)B NMR parameters and the Fe(II/III) redox potentials and the number of arene methyls for complexes 2 are linear. These facts establish direct evidence for a strictly additive character of electron donation by the methyl substituents to the arene ring and further to the Fe center and the second (dicarbollide) ligand.Correlations between the number of arene methyls (n) and selected (1)H and (11)B NMR parameters or the Fe(II/III) redox potentials for complexes [1-(η(6)-MenC(6)H(6-n))-closo-1,2,3-FeC(2)B(9)H(11)] are of strictly linear character.     

Structural interactions of conjugacy closed loops

We study conjugacy closed loops by means of their multiplication groups. Let be a conjugacy closed loop, its nucleus, the associator subloop, and and the left and right multiplication groups, respectively. Put . We prove that the cosets of agree with orbits of , that and that one can define an abelian group on . We also explain why the study of finite conjugacy closed loops can be restricted to the case of nilpotent. Group is shown to be a subgroup of a power of (which is abelian), and we prove that can be embedded into . Finally, we describe all conjugacy closed loops of order .

     

Critical speed for the dynamics of truck events on bridges with a smooth road surface

Simple numerical models of point loads are used to represent single and multiple vehicle events on two-lane bridges with a good road profile. While such models are insufficiently complex to calculate dynamic amplification accurately, they are presented here to provide an understanding of the influence of speed and distance between vehicles on the bridge dynamic response. Critical combinations of speed as a function of main bridge natural frequency and meeting point of two vehicles travelling in opposite directions are identified. It is proposed that such simple models can be used to estimate the pattern of critical speeds versus dynamic amplification of the bridge response for trucks on a relatively smooth surface. The crossing of a three-dimensional spring-dashpot truck is simulated over a bridge plate model to test this hypothesis for a range of road roughness. Further validation is carried out using the site-specific mean pattern associated to field measurements of bridge strains when traversed by a truck population. The latter is found to be closely resembled by the theoretical pattern derived from simple point load models.     

Genes within the serotonergic system are differentially expressed in human brain

Background  

Serotonin is an important neurotransmitter with wide-ranging functions throughout the central nervous system. There is strong evidence to suggest that regulation of serotonergic gene expression might be related to genetic variability, and several studies have focused on understanding the functional effects of specific polymorphisms within these genes on expression levels. However, the combination of genotype together with gender and brain region could have an overall effect on gene expression. In this study, we report expression patterns of five serotonergic genes (TPH1, TPH2, 5-HT2A, 5-HT2C, 5-HTT) in seven different human post-mortem brain regions (superior frontal gyrus, superior temporal gyrus, striatum, cerebellum, hippocampus, midbrain and thalamus) using TaqMan™ real-time quantitative PCR. In addition, the effect of genotype and gender on their expression levels was determined.