Towards a reliable molecular mass determination of intact glycoproteins by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Different matrices and sample-matrix preparation procedures have been tested in order to study their influence on the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectra of intact glycoproteins, which present different degrees of glycosylation (human transferrin; bovine fetuin; bovine alpha(1)-acid-glycoprotein; recombinant human erythropoietin; and the novel erythropoiesis stimulating protein). Using sinapinic acid (SA) and the fast evaporation method, the studied glycoproteins became susceptible to fragmentation at any laser intensity, suggesting that this 'hot' matrix is unsuitable for a reliable molecular mass determination of glycosylated compounds. In contrast, 2,5-dihydroxybenzoic acid (DHB) and 6-aza-2-thiothymine (ATT), with an adequate sample-matrix preparation, provided improved results. Samples containing DHB after crystallization by vacuum drying demonstrated the best performance because the labile functional groups from the glycoforms were apparently fragmented to a lower extent. The average molecular masses obtained using this methodology were in all cases a better estimation than those values reported in the literature. The results were reproducible, and sensitivity was similar to that obtained with SA and the fast evaporation method. These excellent results suggest that this MALDI-TOF-MS methodology could be useful for an improved determination of the average molecular mass values of microheterogeneous compounds such as glycoproteins, glycosylated compounds or, in general, molecular mass values of molecules with similar labile functional groups.     

Sterically bulky tris(triazolyl)borate ligands as water-soluble analogues of tris(pyrazolyl)borate

The recently synthesized 3-tert-butyl-5-methyl-1,2,4-triazole reacted with KBH4 to give the new potassium tris(3-tert-butyl-5-methyl-1,2,4-triazolyl)borate K(Ttz(tBu,Me)) ligand. Ttz(tBu,Me) formed a four-coordinate (Ttz(tBu,Me))CoCl complex and five-coordinate (Ttz(tBu,Me))CoNO3 and (Ttz(tBu,Me))ZnOAc complexes. When these complexes were compared to their Tp(tBu,Me) analogues, it was found that Ttz(tBu,Me) resulted in negligible steric differences. K(Ttz(tBu,Me)) is more water-soluble than K(Tp(tBu,Me)), so bulky tris(triazolyl)borate ligands should lead to functional models for enzyme active sites in an aqueous environment and the creation of water-soluble analogues of Tp catalysts.     

Interaction of Tb(III) and hydrolyzed polyacrylamides in solution as a gel and in the solid state

Formation of the HPAM-Tb³ complex in solution was observed in the intensification of Tb(III) fluorescence, viscosity, and light scattering. The interaction is dependent on the degree of hydrolysis of the copolymer and the pH of the solution. As the pH value increases, an intensification of the interaction and formation of a gel is observed. The latter was associated with the appearance of crosslinks in which terbium, or one of its hydrolysis products, works as a binding agent between chains. After drying, the characterization of the residues of the gels was effected by analytical determination (C, H, N, Cl, Tb), fluorescence, and infrared spectrophotometry. The analysis and fluorescence revealed the existence of particles of terbium hydroxide; the infrared spectrophotometry showed the binding of terbium (III) to carboxylate of the copolymer.     

Syntheses and coordination chemistry of methylpyridylphosphine oxide ligands with copper(II)

The coordination properties of three heterofunctional phosphine oxide ligands, 2-methylpyridyldiphenylphosphine oxide (L1), phenylphosphino-bis-2-methylpyridine oxide (L2) and phenylphosphino-bis-2-methylpyridine N,N′,P-trioxide (L3) with Cu(II) is described. The X-ray crystal structures of the compounds display a distorted octahedral geometry, which exhibit Jahn–Teller distortions. In compounds 1 and 2, the L1 and L2 ligands react with Cu(BF₄)₂ in a 2:1 ligand to metal ratio, respectively, with the BF₄ anions interacting with the metal center. L3 reacts with Cu(BF₄)₂ in 1:1 and 2:1 ligand/metal ratios to form compounds 3 and 4, respectively. Addition of either 2,2′-bipyridine or 4,4′-bipyridine to reaction solutions containing Cu(BF₄)₂ and L3 produces a discrete molecule (5) and a polymeric structure (7), respectively. The reaction of both bipyridines in the presence of Cu(BF₄)₂ and L3 gives rise to a discrete molecule (6) characterized by two octahedral coppers interconnected by the 4,4′-bipyridine. The electrochemical and photophysical properties of all compounds were investigated by cyclic voltammetry (CV) and UV–Vis, as they exhibited no emission or excitation in fluorimetric experiments.     

Thermochemistry of new molecular species: SBr and HSBr

Total energies, optimized geometries, and vibrational frequencies of SBr and HSBr have been evaluated at the coupled cluster level of theory with the correlation consistent basis sets. Extrapolated to the complete basis set limit and with corrections for core-valence, scalar relativistic, and spin-orbit effects, atomization energies were computed and then combined with the experimental heats of formation of the atomic species to generate very accurate heats of formation for the species SBr and HSBr. For SBr, we predict 37.45 and 36.07 kcal/mol for DeltaHf(0 K) and DeltaHf (298.15 K), respectively, in very good agreement with the inferred experimental values of 37.98 and 36.15 kcal/mol. For HSBr, the estimate turns out to be 10.38 and 8.29 kcal/mol for DeltaHf (0 K) and DeltaHf (298.15 K), respectively. Using the more recent HBrO experimental heat of formation at 298.15 K of Lock et al., [J. Phys. Chem. 100, 7972 (1996)] the inferred experimental value for HSBr is predicted to be 8.15 kcal/mol, compared with 8.65 kcal/mol derived from the data of Ruscic and Berkowitz [J. Chem. Phys. 101, 7795 (1994)]. Considering the better agreement of the result with that predicted using the experimental value of DeltaHf(298.15 K) of Lock et al., the author also supports the suggestion made by Denis [J. Phys. Chem. A. 110, 5887 (2006)] that the result of Lock et al. should be preferred over the one of Ruscic and Berkowitz. For DeltaHf(0 K), the author found 10.38 and 10.56 kcal/mol, respectively, for the theoretical and inferred experimental estimates.     

Non-destructive method for determination of hydroxyl value of soybean polyol by LS-SVM using HATR/FT-IR

This paper presents the use of least-squares support vector machine (LS-SVM) for quantitative determination of hydroxyl value (OHV) of hydroxylated soybean oils by horizontal attenuated total reflection Fourier transform infrared (HATR/FT-IR) spectroscopy. A least-squares support vector machine (LS-SVM) calibration model for the prediction of hydroxyl value (OHV) was developed using the range 1805.1-649.9 cm(-1). Validation of the method was carried out by comparing the OHV of a series of hydroxylated soybean oil predicted by the LS-SVM model to the values obtained by the AOCS standard method. A correlation coefficient equal to 0.989 and RMSEP = 4.96 mg of KOH/g was obtained. This study demonstrates a better prediction ability of the LS-SVM technique to determine OHV in hydroxylated soybean oil samples by HATR/FT-IR spectra.     

Transformations of diphenylphosphinothioic acid tertiary amides mediated by directed ortho metallation

ortho-Lithiation of N,N-diisopropyl-P,P-diphenylphosphinothioic amide using n-BuLi in the presence of TMEDA in diethyl ether followed by electrophilic trapping is described as an efficient method for the synthesis of ortho-functionalised derivatives in high yields. The structural modification of the phosphinothioic amide includes C-X (X = P, S, Si, Sn, I) and C-C bond forming reactions with a large variety of electrophiles. Additional applications based on functional group transformations are also reported. They include imine formation, desulfurization and Suzuki cross-coupling reactions on selected compounds.     

Novel peptide foldameric motifs: a step forward in our understanding of the fully-extended conformation/3(10)-helix coexistence

The fully-extended, multiple C(5), conformation or 2.0(5) helix is a very appealing peptide secondary structure, in particular for its potential use as a molecular spacer, as it is characterized by the longest elevation (as high as 3.62 ?) between the α-carbon atoms of two consecutive α-amino acids. Despite this intriguing property, however, it is only poorly investigated and understood. Here, using a complete series of C(α,α)-diethylglycine (Deg) homo-oligopeptide esters to the pentamer level, we exploited the properties of a fluorophore and a quencher, synthetically positioned at the N- and C-termini of the main chain, respectively, to check the applicability of the fully-extended conformation as a rigid molecular spacer. The fluorescence study was complemented by FT-IR absorption and NMR conformational investigations. The X-ray diffraction structures of selected compounds are also reported. Unfortunately, we find that, even in a solvent of low polarity, such as chloroform, in this peptide series an equilibrium does take place between the fragile fully-extended conformation and the 3(10)-helical structure, the latter becoming more and more stable as the main chain is elongated. Since the Deg homo-peptide esters lacking any terminal aromatic group, previously investigated, are known to adopt a stable fully-extended conformation in chloroform solution, we tend to attribute the 3D-structure instability observed in this work to the presence of multiple aromatic rings in their blocking groups.     

Epothilone Analogues with Benzimidazole and Quinoline Side Chains: Chemical Synthesis,Antiproliferative Activity,and Interactions with Tubulin

A series of epothilone B and D analogues bearing isomeric quinoline or functionalized benzimidazole side chains has been prepared by chemical synthesis in a highly convergent manner. All analogues have been found to interact with the tubulin/microtubule system and to inhibit human cancer cell proliferation in vitro, albeit with different potencies (IC₅₀ values between 1 and 150 nM ). The affinity of quinoline‐based epothilone B and D analogues for stabilized microtubules clearly depends on the position of the N‐atom in the quinoline system, while the induction of tubulin polymerization in vitro appears to be less sensitive to N‐positioning. The potent inhibition of human cancer cell growth by epothilone analogues bearing functionalized benzimidazole side chains suggests that these systems might be conjugated with tumor‐targeting moieties to form tumor‐targeted prodrugs.