Microscopic Protonation Deprotonation Equilibria of the Antiinflammatory Agent Piroxicam

The microscopic ionization behavior of piroxicam was investigated using two different approaches, i.e., direct UV spectroscopy and an indirect analogue approach (deductive method). The best microscopic pK(a) values (PKa12 = 4.60, pK(a21) = 5.40, pK(a22) = 2.72, and pK(a11) = 1.92) were obtained by the deductive method using as pK(a22) the pK(a) of the enolic O-methylated piroxicam 2. The results show remarkable electrostatic effects in the protonation/deprotonation equilibria, a marked increase in the acidity of the enolic function (2.68 pK(a) units) being caused by the pyridinium group. The electronic structure of piroxicam was studied based on H-1-NMR chemical shifts at various ionization states, indicating an extended electron conjugation through the molecule. The partition measurements in octan-1-ol/H2O of zwitterionic compound 3 (the pyridyl N-methyl derivative of piroxicam (1)) suggest that the two opposite charges in zwitterionic piroxicam are indeed in a close intramolecular proximit

Characterisation of reversed-phase liquid chromatographic columns by chromatographic tests. Rational column classification by a minimal number of column test parameters

The European Pharmacopoeia (Ph. Eur.) and other official compendia give only a general description of the stationary phase in the description of a liquid chromatographic method. Therefore the selection of a column giving suitable selectivity presents difficulties. Earlier, a test procedure was proposed that allows to measure 36 chromatographic parameters which have been described for characterising stationary phases. This procedure was carried out on 69 reversed-phase liquid chromatography (RP-LC) columns. This paper focuses on the classification of RP-LC stationary phases based on chromatographic parameters. A chemometric study was conducted using 24 parameters that could be measured in a repeatable and reproducible way. Principal component analysis was used to classify the columns and to estimate the minimal number of parameters necessary for a rational classification. It is shown that after reducing the number of parameters from 24 to four or three, similar classifications were obtained. The column classifications were compared to the European Pharmacopoeia stationary phase description and to the column properties obtained from the manufacturers</p

Characterisation of reversed-phase liquid chromatographic columns by chromatographic tests. Evaluation of 36 test parameters: repeatability, reproducibility and correlation

The European Pharmacopoeia (Ph. Eur.) or other official compendia give only a general description of the stationary phase in the description of a liquid chromatographic method. Therefore the selection of a column giving suitable selectivity presents difficulties. Earlier, a test procedure was proposed that allows measurement of a number of parameters which are reported to be representative for stationary phase characteristics. This paper describes how the test procedure was applied on 69 RP-LC C18 columns. Chromatographic parameters obtained as test results were evaluated, and their repeatability, reproducibility and correlation were examined</p

Site-specific basicities regulate molecular recognition in receptor binding: in silico docking of thyroid hormones.

Interactions between thyroid hormone alpha and beta receptors and the eight protonation microspecies of each of the main thyroid hormones (thyroxine, liothyronine, and reverse liothyronine) were investigated and quantitated by molecular modeling. Flexible docking of the various protonation forms of thyroid hormones and high-affinity thyromimetics to the two thyroid receptors was carried out. In this method the role of the ionization state of each basic site could be studied in the composite process of molecular recognition. Our results quantitate at the molecular level how the ionization state and the charge distribution influence the protein binding. The anionic form of the carboxyl group (i.e., carboxylate site) is essential for protein binding, whereas the protonated form of amino group worsens the binding. The protonation state of the phenolate plays a less important role in the receptor affinity; its protonation, however, alters the electron density and the concomitant stacking propensity of the aromatic rings, resulting in a different binding score. The combined results of docking and microspeciation studies show that microspecies with the highest concentration at the pH of blood are not the strongest binding ones. The calculated binding free energy values can be well interpreted in terms of the interactions between the actual sites of the microspecies and the receptor amino acids. Our docking results were validated and compared with biological data from the literature. Since the thyroid hormone receptors influence several physiologic functions, such as metabolic rate, cholesterol and triglyceride levels, and heart frequency, our binding results provide a molecular basis for drug design and development in related therapeutic indications