Daniela M. Tomazela

Antimoniais empregados no tratamento da leishmaniose: estado da arte

Antimony preparations are the drugs of choice for the treatment of leishmaniasis over 90 years, a disease that currently affects 12 million people worldwide. Its introduction was based on 19th century concepts of therapeutic effects of metal salts as arsenicals and other metals, most of them abandoned due to toxic effects or better drugs. In the last three decades, there was a great improvement in the knowledge of cell biology and immunology of those infections, but chemotherapy has not been improved in the same strength. The structure and mechanism of action of the two pentavalent antimonial drugs of choice, meglumine antimoniate and sodium stibogluconate, are not well known and the contamination of those pharmaceutical by toxic contaminants have been verified.

Electrospray ionization mass spectrometry fingerprinting of propolis

Crude ethanolic extracts of propolis, a natural resin, have been directly analysed using electrospray ionization mass (ESI-MS) and tandem mass spectrometry (ESI-MS/MS) in the negative ion mode. European, North American and African samples have been analyzed, but emphasis has been given to Brazilian propolis which displays diverse and region-dependent chemical composition. ESI-MS provides characteristic fingerprint mass spectra, with propolis samples being divided into well-defined groups directly related to their geographical origins. Chemometric multivariate analysis statistically demonstrates the reliability of the ESI-MS fingerprinting method for propolis. On-line ESI-MS/MS tandem mass spectrometry of characteristic [M - H](-) ion markers provides an additional dimension of fingerprinting selectivity, while structurally characterizing the ESI-MS marker components of propolis. By comparison with standards, eight such markers have been identified: para-coumaric acid, 3-methoxy-4-hydroxycinnamaldehyde, 2,2-dimethyl-6-carboxyethenyl-2H-1-benzopyran, 3-prenyl-4-hydroxycinnamic acid, chrysin, pinocembrin, 3,5-diprenyl-4-hydroxycinnamic acid and dicaffeoylquinic acid. The negative mode ESI-MS fingerprinting method is capable of discerning distinct composition patterns to typify, to screen the sample origin and to reveal characteristic details of the more polar and acidic chemical components of propolis samples from different regions of the world.

Electrospray ionization mass spectrometry fingerprinting of beer.

After just simple degassing, dilution, pH adjustment and direct flow injection, characteristic fingerprint spectra of beer samples have been obtained by fast (few seconds) electrospray ionization mass spectrometry (ESI-MS) analysis in both the negative and positive ion modes. A total of 29 samples belonging to the two main beer types (lagers and ales) and several beer subtypes from USA, Europe and Brazil could be clearly divided into three groups both by simple visual inspection of their ESI(+)-MS and ESI(-)-MS fingerprints as well as by chemometric treatment of the MS data. Diagnostic ions with contrasting relative abundances in both the positive and negative ion modes allow classification of beers into three major types: P = pale (light) colored (pilsener, pale ale), D = dark colored (bock, stout, porter, mild ale) and M = malt beer. For M beers, samples of a dark and artificially sweetened caramel beer produced in Brazil and known as Malzbiers were used. ESI-MS/MS on these diagnostic beer cations and anions, most of which are characterized as arising from ionization of simple sugars, oligosaccharides, and iso-alpha-acids, yield characteristic tandem mass spectra adding a second and optional MS dimension for improved selectivity for beer characterization by fingerprinting. Direct ESI-MS or ESI-MS/MS analysis can therefore provide fast and reliable fingerprinting characterization of beers, distinguishing between types with different chemical compositions. Other unusual polar components, impurities or additives, as well as fermentation defects or degradation products, could eventually be detected, making the technique promising for beer quality control.

Generic automated method for liquid chromatography-multiple reaction monitoring mass spectrometry based monoclonal antibody quantitation for preclinical pharmacokinetic studies.

Quantitation of therapeutic monoclonal antibodies (mAb) using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for pharmacokinetic (PK) studies is becoming an essential complement to traditional antibody-based ligand binding assays (LBA). Here we show an automated method to perform LC-MS/MS-based quantitation, with IgG1 conserved peptides, a heavy isotope labeled mAb internal standard, and anti-human Fc enrichment. All reagents in the method are commercially available with no requirement to develop novel assay-specific reagents. The method met traditional quantitative LC-MS/MS assay analytical characteristics in terms of precision, accuracy, and specificity. The method was applied to the pharmacokinetic study of a mAb dosed in cynomolgus monkey, and the results were compared with the immunoassay data. This methodology has the potential to benefit and accelerate the early biopharmaceutical development process, particularly by enabling PK analysis across species and candidate molecules with minimal method development.

Keto-Enolic Equilibria of an Isatin-Schiff Base Copper(II) Complex and its Reactivity toward Carbohydrate Oxidation

An interesting isatin-Schiff base copper(II) complex, [Cu(isapn)](ClO4)2 where isapn= N,N′-[bis-(3,3′-indolin-2-one)]-1,3-diiminepropane, was prepared and characterized by different techniques, both in the solid state and in solution, and its reactivity toward carbohydrate oxidation was verified. The positive ion electrospray mass spectrum detects the complex as an isotopologue cluster of singly charged intact isatin-copper(I) ions of m/z 395 (for 65Cu) with an isotopic pattern identical to that calculated for C19H16CuN4O2+. Tandem mass spectrometry reveals an interesting and structurally diagnostic collision-induced dissociation behavior for this ionized complex, which is dominated by the cleavage of the N—(CH2)3—N propylene bridge. In aqueous solution, this complex undergoes a peculiar keto-enolic equilibrium, verified at different pHs by spectroscopic methods (u.v.–vis. and e.p.r.), with a corresponding pKa determined as 9.5. The e.p.r. parameter ratio g‖/A‖ for this complex, in frozen MeOH/H2O (4:1, v/v) solution at 77 K, changes from 188 cm in acidic medium (pH 2.5–3.0) to 118 cm in basic medium (pH 11), indicating a significant structural change from a distorted tetrahedral to a more tetragonal geometry around the copper ion. This compound was shown to catalyze the oxidation of hexoses (glucose, fructose and galactose), in alkaline media, via reactive oxygen species, which were detected by using specific enzymes, and by e.p.r. spin trapping. The reaction was monitored at (25.0 ± 0.1)°C by the consumption of oxygen, and showed first-order dependence on catalyst, followed by a saturation effect. First-order kinetics with respect to [OH−] concentration was also observed, indicating that enolization of the substrate as well as the metal-catalyzed enediol oxidation are the rate-determining steps.

Iridoid glucosides from Randia spinosa (Rubiaceae)

An iridoid glucoside: randinoside, along with five known iridoids: galioside, deacetylasperulosidic acid methyl ester, scandoside methyl ester, geniposide and gardenoside, were isolated from the stems of Randia spinosa. The structures were determined by spectroscopic analysis, including 2D NMR techniques.

Supramolecular conformational effects in the electrocatalytic properties of electrostatic assembled films of meso(3- and 4-pyridyl) isomers of tetraruthenated porphyrins

, have been obtained and characterized by electrochemistry, spectroscopy and mass spectrometry. Layer-by-layer electrostatic assembled films with tetrasulfonated phthalocyaninatecuprate(II) anion, CuTSPc, displayed distinct electrocatalytic activity towards sulfite and nitrite oxidation. In general, the films derived from the M(4-TRPyP) isomers are more effective for the oxidation of nitrite, whereas those containing the M(3-TRPyP) isomers are more efficient for the oxidation of sulfite. The results demonstrated the influence of molecular geometry on the active sites of porphyrinic nanomaterials, enhancing the electron-transfer process in the M(3-TRPyP) derivative. This effect results from changes in the chemical environment around the active sites, induced by the contrasting molecular packing interactions with CuTSPc.

Automated DBS microsampling, microscale automation and microflow LC–MS for therapeutic protein PK

AIM Reduce animal usage for discovery-stage PK studies for biologics programs using microsampling-based approaches and microscale LC-MS. METHODS & RESULTS We report the development of an automated DBS-based serial microsampling approach for studying the PK of therapeutic proteins in mice. Automated sample preparation and microflow LC-MS were used to enable assay miniaturization and improve overall assay throughput. Serial sampling of mice was possible over the full 21-day study period with the first six time points over 24 h being collected using automated DBS sample collection. Overall, this approach demonstrated comparable data to a previous study using single mice per time point liquid samples while reducing animal and compound requirements by 14-fold. CONCLUSION Reduction in animals and drug material is enabled by the use of automated serial DBS microsampling for mice studies in discovery-stage studies of protein therapeutics.