Zoltán Szakács

Protonation microequilibrium treatment of polybasic compounds with any possible symmetry

Microequilibrium treatment that has previously been limited to tridentate ligands is generalized to arbitrary number of functional groups in the molecule and the role of symmetry is also investigated. Cumulative microconstant, a new type of equilibrium parameter, is introduced, allowing an equivalent, but more compact mathematical treatment of large microequilibrium systems. The sufficient number of independent pieces of information for the unambiguous determination of all microconstants is deduced. It has been concluded that even if protonation mole fraction for all the basic sites is available, determinability of all the microconstants is rather the exception than the case, without a priori simplifying assumptions. It has been shown that all microconstants can only be determined from protonation mole fractions for molecules of up to three groups. For molecules of four groups and beyond, only specific symmetry and the concomitant simplification of the microequilibrium system make the strict, complete microspeciation feasible. As a case study, the protonation scheme and the complete microspeciation of a tetradentate ligand is analyzed in detail.

Chiral recognition of imperanene enantiomers by various cyclodextrins: A capillary electrophoresis and NMR spectroscopy study

The enantiomers of imperanene, a novel polyphenolic compound of Imperata cylindrica (L.), were separated via cyclodextrin‐modified capillary electrophoresis. The anionic form of the analyte at pH 9.0 was subject to complexation and enantioseparation CE studies with neutral and charged cyclodextrins. As chiral selectors 27 CDs were applied differing in cavity size, sidechain, degree of substitution (DS) and charge. Three hydroxypropylated and three sulfoalkylated CD preparations provided enantioseparation and the migration order was successfully interpreted in each case in terms of complex mobilities and stability constants. The best enantioresolution (RS = 1.26) was achieved using sulfobutyl‐ether‐γ‐CD (DS ∼4), but it could be enhanced by extensive investigations on dual selector systems. After optimization (CD concentrations and pH) RS = 4.47 was achieved using a 12.5 mM sulfobutyl‐ether‐γ‐CD and 10 mM 6‐monodeoxy‐6‐mono‐(3‐hydroxy)‐propylamino‐β‐cyclodextrin dual system. The average stoichiometry of the complex was determined with Jobs method using NMR‐titration and resulted in a 1:1 complex for both (2‐hydroxy)propyl‐β‐ and sulfobutyl‐ether‐γ‐CD. Further NMR experiments suggest that the coniferyl moiety of imperanene is involved in the host‐guest interaction.

Resolution of carboxylate protonation microequilibria of NTA, EDTA and related complexones

Potentiometric and NMR-pH titrations were carried out on four classical complexones to elucidate their overall and site-specific basicities. NMR-pH profiles and Bjerrums functions were adjusted to compound-specific symmetries and appropriate evaluation methods were developed. Symmetry-modulated relationships between the macro- and microconstants were deduced and self-consistent sets of microconstants were determined. The inherent basicity of carboxylates surrounded by adjacent, intramolecular ammonium and carboxylate sites have been found to be in the range of 1.83-2.02logk units, which are reduced by 0.05-0.12logk units upon a nearby carboxylate protonation.

A novel potentiometric method for the determination of real crosslinking ratio of poly(aspartic acid) gels

In order to obtain nontoxic functional polymer gels for biomedical applications, chemically crosslinked poly(aspartic acid) gels have been prepared using 1,4-diaminobutane as crosslinker. The presence of COOH and amino groups on the network chains renders these gels pH sensitive. Due to the specific hydrophobic-hydrophilic balance, these gels show a significant volume transition at a well-defined pH close to the pK value of uncrosslinked poly(aspartic acid). Since the magnitude of volume change critically depends on the degree of crosslinking, it is an important task to determine the topological characteristics of these networks. A novel method based on potentiometric acid-base titration has been developed to assess the crosslinking ratio, excluding physical crosslinks and entanglements. It turned out that only 25% of all crosslinker molecules forms real crosslinks between the poly(aspartic acid) chains; the rest react with one of its functional groups and forms short pendant side chains. At a nominal crosslinking ratio of 0.1, the number average molecular mass between crosslinks is found to be M(c) = 2300.

EQUILIBRIUM BINDING MODEL OF BILE SALT-MEDIATED CHIRAL MICELLAR ELECTROKINETIC CAPILLARY CHROMATOGRAPHY

Optimum concentration of bile salts in chiral separations depends on both the aggregation properties of the surfactant and the stability of the analyte‒micelle complexes. An equilibrium model is proposed in which these two effects are treated separately. First the aggregation constants should be determined under the experimental conditions of the chiral MEKC analysis. With these data, the equilibrium concentrations of bile salt aggregates can be calculated at any total surfactant concentration. Using the Offord equation to approximate the mobilities of the enantiomer‒bile salt complexes, a model function has been derived to fit the experimental mobilities. The method yields the binding constants of the enantiomers to each aggregate present. Those species are assumed to be important in the chiral recognition process, which have significantly different stability constants for the enantiomers. The method is demonstrated by the chiral separation of R‒ and S‒1,1′‒binaphthyl‒2,2′‒diyl hydrogen phosphate with sodium taurodeoxycholate. Based on the calculated binding constants, tetrameric aggregates are assumed to be the discriminating species, while no significant difference in enantiomer binding to dimers was found.

NMR, CD and UV spectroscopic studies reveal uncommon binding modes of dapoxetine to native cyclodextrins

Complex formation between the selective serotonin reuptake inhibitor drug (S)-dapoxetine (Dpx) and β-, γ-, and methylated γ-cyclodextrins (CyDs) was studied by complementary experimental techniques. Phase solubility studies indicated 1 : 1 stoichiometry for all CyDs, with β-CyD being the most effective to enhance the solubility of and form the most stable complex with Dpx. 1H NMR titrations and Jobs method of continuous variation were also employed to corroborate the stoichiometry and association constants with γ- and β-CyDs. The simultaneous evaluation of titration datasets of several Dpx and CyD protons revealed for both CyDs that the best fit was achieved when the coexistence of Dpx·CyD and 2Dpx·CyD complexes were assumed. This finding was rather unanticipated considering the smaller cavity size of β-CyD, which inspired further investigations. The UV hypochromism and circular dichroism spectroscopic data suggested the inclusion of two Dpx molecules into the CyD cavity (albeit to a less extent with β-CyD). This conclusion was fully supported by 2D ROESY and 1D NOESY NMR spectra, observing NOEs between the inner methine protons of CyDs and both the phenyl and naphthyl ring protons of Dpx. Additional molecular dynamics calculations identified the energetically most favoured species for both the Dpx·CyD and 2Dpx·CyD complexes.

Solution-state NMR spectroscopy of famotidine revisited: spectral assignment, protonation sites, and their structural consequences

AbstractMultinuclear one (1D-) and two-dimensional (2D) nuclear magnetic resonance (NMR) spectroscopic investigations of famotidine, the most potent and widely used histamine H2-receptor antagonist, were carried out in dimethyl sulfoxide-d6 (DMSO-d6) and water. Previous NMR assignments were either incomplete or full assignment was based only on 1D spectra and quantum-chemical calculations. Our work revealed several literature misassignments of the 1H, 13C, and 15N NMR signals and clarified the acid–base properties of the compound at the site-specific level. The erroneous assignment of Baranska et al. (J. Mol. Struct. 2001, 563) probably originates from an incorrect hypothesis about the major conformation of famotidine in DMSO-d6. A folded conformation similar to that observed in the solid-state was also assumed in solution, stabilized by an intramolecular hydrogen bond involving one of the sulphonamide NH2 protons and the thiazole nitrogen. Our detailed 1D and 2D NMR experiments enabled complete ab initio1H, 13C, and 15N assignments and disproved the existence of the sulphonamide NH hydrogen bond in the major conformer. Rather, the molecule is predominantly present in an extended conformation in DMSO-d6. The aqueous acid–base properties of famotidine were studied by 1D 1H- and 2D 1H/13C heteronuclear multiple-bond correlation (HMBC) NMR-pH titrations. The experiments identified its basic centers including a new protonation step at highly acidic conditions, which was also confirmed by titrations and quantum-chemical calculations on a model compound, 2-[4-(sulfanylmethyl)-1,3-thiazol-2-yl]guanidine. Famotidine is now proved to have four protonation steps in the following basicity order: the sulfonamidate anion protonates at pH = 11.3, followed by the protonation of the guanidine group at pH = 6.8, whereas, in strong acidic solutions, two overlapping protonation processes occur involving the amidine and thiazole moieties. FigureCorrect amidine assignments and protonation sites of famotidine

Capillary electrophoresis separation of vinpocetine and related compounds: prediction of electrophoretic mobilities in partly aqueous media.

Offords equation, a relationship between electrophoretic mobility and charge, size and shape of peptides, has been extended to quantitate the electrophoretic mobility of vinca alkaloids. Partly aqueous protonation constants and the derived theoretical mobilities have been proven to be able to predict experimental electrophoretic mobilies. In practice, seven vincamine derivatives of very low water‐solubility were separated by capillary electrophoresis. Buffer total concentration, apparent pH and methanol content, the three most important parameters of the running buffer, were used in triangular resolution mapping to characterize separation. Even though electrophoresis is well known to slow down in partly aqueous media, under our optimized circumstances a baseline separation was achieved within 8 min in each case.

Determination of the aggregation constants of bile salts by capillary electrophoresis

SummaryCapillary electrophoresis has been investigated as a novel experimental method for determination of the aggregation constants of surfactants. The tendency of sodium cholate and sodium taurodeoxycholate to associate was studied in phosphate buffers of pH 8.0 and pH 7.0, respectively. Stepwise aggregation equilibria of bile salt monomers has been described in terms of massbalance equations. The Offord equation was used to model the electrophoretic mobility of the bile salt associates, and the experimental mobility values could be fitted to the model. Interestingly, only even-membered aggregates-dimers and tetramers-besides the monomers were proposed from the results of the curve-fitting for both bile salts. The aggregation constants calculated were (in molar units): cholate logKA2=1.37, logKA4=4.98 taurodeoxycholate logKA2=1.68, logKA4=6.46. From these values, more pronounced aggregation of taurodeoxycholate starting at lower concentrations has been deduced, supporting the back-to-back association model of bile salts.

Capillary electrophoretic separation of mono- and dimaltosyl-β-cyclodextrins and determination of the stability constants of their benzoate complexes

SummaryCapillary zone electrophoresis method has been developed for the separation (and determination) of monoand dimaltosyl-β-cyclodextrins using indirect UV absorbance detection. The separation is based on inclusion complex formation with benzoate ion which is used as background electrolyte and absorbance provider simultaneously. Good resolution of cyclodextrin mixtures can be achieved optimizing the pH and the benzoic acid concentration in the background electrolyte. Capillary zone electrophoresis method has also been developed for the determination of formation constants of inclusion complexes.