Biomedical subjects

Roman Kaliszan

Publications and source records attributed to Roman Kaliszan.

At least 19 recordsLinked to original sources

PubMed2006

Simultaneous determination of pKa and lipophilicity by gradient RP HPLC.

High-performance methods of testing of drug candidates for properties of pharmacokinetics and pharmacodynamics importance, in particular lipophilicity and acidity, are necessary to overcome innovation stagnation in the pharmaceutical industry. Reversed-phase high-performance liquid chromatography (RP HPLC) might be a unique tool for the determination of both pKa and the apparent (pH-dependent) partition coefficient, applicable in high-throughput analysis of multicomponent mixtures, e.g., samples originating from automated synthesis. In this work, the pH/organic modifier gradient RP HPLC is presented as a means of simultaneous determination of an analyte's acidity and lipophilicity. The approach consists of retention measurements in a series of methanol gradient runs differing in pH range and duration of the gradient. Two different models of the influence of pH on retention in organic modifier gradient RP HPLC are compared regarding the quality of the simultaneously determined lipophilicity and dissociation constants. Advantages of the proposed approach over currently employed procedures are that it can be applied to compound mixtures, it requires only minute amounts of substances, and pKa values can be determined in the range 3-10 units and lipophilicity in the range 0-7 units. Verification of the reliability of the parameters determined by the new method was demonstrated on a series of 93 acidic and basic drug analytes.

Chemical Phenomena↗

PubMed2006

Evaluation of the silanol-suppressing potency of ionic liquids.

Recently, increasing attention has been paid to the use of ionic liquids for high-performance liquid chromatography (HPLC) and capillary electrophoresis. In the present study, the silanol-suppressing potency of ionic liquids was evaluated by HPLC using the two-retention site model proposed previously by Nahum and Horváth (J. Chromatogr. 1981, 203, 53-63). The binding constant, KA, in that approach has been demonstrated to reliably reflect the ability of the ionic liquids to block the silanols of the silica support material of the stationary phase. The determinations were carried out for ionic liquids of the 1-alkyl-3-methylimidazolium group with the use of a series of basic drugs as the test analytes. Comparison of ionic liquids with standard mobile phase additives such as triethylamine showed the former to possess advantages as silanol suppressors in HPLC. The main advantage of the method is that it provides a simple and fast determination of the silanol complex stability, which allowed comparison of the suppressing efficiency of several ionic liquids.

Chromatography, High Pressure Liquid↗

PubMed2005

Separation of nicotinic acid and its structural isomers using 1-ethyl-3-methylimidazolium ionic liquid as a buffer additive by capillary electrophoresis.

The growing interest in application of ionic liquids (ILs) in analytical chemistry has been observed. The aim of presented investigation was to verify whether ILs would be a suitable modifier of the background electrolyte (BGE) for pharmaceutical analysis of the closely related drug analogues. The study demonstrates the use of 1-ethyl-3-methylimidazolium tetrafluoroborate (1E-3MI-TFB) ionic liquid as modifiers in the separation of nicotinic acid and its structural isomers by capillary electrophoresis. Dependences of the ionic liquid concentration in a BGE on the separation parameters like migration time, resolution factor and width at peak's baseline have been compared. The separation mechanism involves the free imidazolium ions, which can interact with inner surface of the capillary wall. Increased 1E-3MI-TFB concentration to 150 mmol/L caused decrease of migration times of analytes, improve peaks shape and increase of separation performances. At this ionic liquid concentration in a BGE resolution factor between nicotinic and isonicotinic acids increased to 1.86. The proposed CE separation procedure is highly reproducible and can be applied in qualitative and quantitative analysis of carboxylic acids.

Buffers↗

PubMed2005

Human red blood cells targeted metabolome analysis of glycolysis cycle metabolites by capillary electrophoresis using an indirect photometric detection method.

A capillary electrophoretic (CE) method with UV indirect spectrophotometric detection for determination of the main compounds of glycolysis in human erythrocytes has been elaborated. Blood samples for electrophoretic analysis were prepared by isolation of erythrocytes, lysis of the cells by heating in double-distilled water and subsequent ultrafiltration with a centrifuge equipped with filter devices: Mr cut off 5000. Using 20 mM 2,6-pyridinedicarboxylic acid (PDC) as a highly UV absorbing carrier electrolyte and 4 mM cetyltrimetylammonium bromide (CTAB), at the resulting pH 12.3, nine of the glycolysis intermediates were separated and characterized. The repeatability and linearity of the method was assessed with percent relative standard deviation (%R.S.D.) for migration time ranging from 0.3 to 1.9% and correlation coefficients of 0.991-0.999 for the studied concentration range. Limits of detection (LOD) for the analyzed metabolites were in the range of 6.25 x 10(-6) to 5.0 x 10(-5) M. The optimized CE method was used to compare metabolome content of red blood cells of 22 healthy volunteers. Mean metabolite concentrations in erythrocytes ranged from 49.6 microM for fructose-6-phosphate (F-6-P) to 3.1 mM for 2,3-diphospho-D-glyceric acid (2,3-DPG). The method can be readily applied in clinical, pathophysiological and epidemiological studies.

2,3-Diphosphoglycerate↗

PubMed2005

Verification of the exponential model of body temperature decrease after death in pigs.

The authors have conducted a systematic study in pigs to verify the models of post-mortem body temperature decrease currently employed in forensic medicine. Twenty-four hour automatic temperature recordings were performed in four body sites starting 1.25 h after pig killing in an industrial slaughterhouse under typical environmental conditions (19.5-22.5 degrees C). The animals had been randomly selected under a regular manufacturing process. The temperature decrease time plots drawn starting 75 min after death for the eyeball, the orbit soft tissues, the rectum and muscle tissue were found to fit the single-exponential thermodynamic model originally proposed by H. Rainy in 1868. In view of the actual intersubject variability, the addition of a second exponential term to the model was demonstrated to be statistically insignificant. Therefore, the two-exponential model for death time estimation frequently recommended in the forensic medicine literature, even if theoretically substantiated for individual test cases, provides no advantage as regards the reliability of estimation in an actual case. The improvement of the precision of time of death estimation by the reconstruction of an individual curve on the basis of two dead body temperature measurements taken 1 h apart or taken continuously for a longer time (about 4 h), has also been proved incorrect. It was demonstrated that the reported increase of precision of time of death estimation due to use of a multiexponential model, with individual exponential terms to account for the cooling rate of the specific body sites separately, is artifactual. The results of this study support the use of the eyeball and/or the orbit soft tissues as temperature measuring sites at times shortly after death. A single-exponential model applied to the eyeball cooling has been shown to provide a very precise estimation of the time of death up to approximately 13 h after death. For the period thereafter, a better estimation of the time of death is obtained from temperature data collected from the muscles or the rectum.

Animals↗

PubMed2005

Comparative characteristics of HPLC columns based on quantitative structure-retention relationships (QSRR) and hydrophobic-subtraction model.

The study was aimed at quantitative comparison of retention properties of modern stationary phases for reversed-phase HPLC. Three approaches, the calculated logarithm of octanol/water partition coefficient (clogP)-based model, the molecular modeling descriptors-based model and the hydrophobic-subtraction model, were compared and discussed. Gradient retention time, tR, of a series of test analytes was a dependent variable in the quantitative structure-retention relationship (QSRR) equations describing retention in terms of analytes' structure descriptors. The QSRRs derived were used to characterize in quantitative manner the specific retention properties of nine representative reversed-phase HPLC. Either the theoretically calculated logarithm of octanol/water partition coefficient, or the structural descriptors from molecular modeling were employed to quantitatively characterize the structure of the analytes. The three molecular modeling-derived structural descriptors considered were: the total dipole moment, the electron excess charge of the most negatively charged atom and the water-accessible molecular surface area. In addition to the above standard QSRR approaches, a recently developed parameterization of reversed-phase column selectivity based on the hydrophobic-subtraction model of Snyder et al. [L.R. Snyder, J.W. Dolan, J.W. Carr, The hydrophobic-subtraction model of reversed-phase column selectivity, J. Chromatogr. A 1060 (2004) 77] was considered. According to the hydrophobic-subtraction model, reversed-phase columns are characterized by five selectivity parameters derived from the linear solvation energy relationships (LSER) theory. Values of these parameters are available for more than 300 different columns. It has been demonstrated that the clogP-based model, the molecular modeling descriptors-based model and the hydrophobic-subtraction model provide generally similar classification of the HPLC columns studied. Some differences in column classification by the three approaches considered are discussed in terms of specific properties of individual stationary phases. All the approaches allow a quantitative, although multidimensional, characteristic of HPLC columns, however, the nonempirical QSRR-based approach is simpler and require less labor.

Chromatography, High Pressure Liquid↗

PubMed2005

Combined pH/organic solvent gradient HPLC in analysis of forensic material.

A combined pH/organic solvent linear gradient mode in high performance liquid chromatography (HPLC) is presented as a new approach to determination of low concentrations of ionogenic analytes in biological material. The approach consists in simultaneous development of linear gradients of pH and organic modifier in the mobile phase. Advantages of the method are illustrated in postmortem analysis of opipramol in material from suicide victims. Very narrow peaks without tailing were obtained and several times lower limits of analyte quantitation were achieved using ultraviolet detection as compared to a standard isocratic method. The double gradient HPLC method seems to be especially valuable in case of ionogenic analytes dispersed in complex biological matrices. That is due to a high selectivity of the double gradient method and the lack of peak tailing, which is commonly observed for basic analytes chromatographed at isocratic conditions.

Chromatography, High Pressure Liquid↗

PubMed2005

Theoretical opportunities and actual limitations of pH gradient HPLC.

In a series of reports published recently by our laboratory comprehensive theory and experimental conditions were established for reversed-phase high-performance liquid chromatography (RP HPLC) employing the programmed pH gradient of mobile phase. A procedure was developed providing, rapidly and conveniently, the acidity (pK(a)) of weak acids and bases and their lipophilicity (hydrophobicity) log k(w). The basis of the double-gradient RP HPLC, employing simultaneous gradients of organic modifier content and mobile phase pH, was also elaborated. The fundamentals of the approach are presented briefly and systematically and its advantages and limitations are discussed. It is demonstrated that the newly introduced pH gradient method increases the analytical versatility of RP HPLC and our understanding of its physicochemical basis.

Chromatography, High Pressure Liquid↗

PubMed2005

pH/organic solvent double-gradient reversed-phase HPLC.

A new reversed-phase high-performance liquid chromatographic (RP HPLC) procedure has been theoretically and experimentally established. The approach consists of the simultaneous development of a gradient of pH and of the organic modifier in the mobile phase. The proposed theoretical model of the pH/organic solvent double-gradient RP HPLC allows determination of both pK(a) and the lipophilicity parameter of the ionized and the nonionized form of the analyte and prediction of the retention times at specific separation conditions as well as bandwidth for all analytes. The model provides a rational basis for optimization of separation of ionizable analytes at any given chromatographic mode and analysis conditions. In addition, in the case of pH/organic solvent double-gradient RP HPLC, a compression of analyte peak and its reduced tailing can be expected.

Chromatography, High Pressure Liquid↗

PubMed2005

Behavior of peptides and computer-assisted optimization of peptides separations in a normal-phase thin-layer chromatography system with and without the addition of ionic liquid in the eluent.

The addition of an ionic liquid into the mobile phase appeared to be useful in optimization of chromatographic separation of peptides. Different behavior of peptides in thin-layer chromatography (TLC) was observed after addition of 1-ethyl-3-methylimidazolium tetra fluoroborate to the eluent in comparison to the system without the ionic liquid. Nonlinear dependence of the retention coefficient, R(M), of peptides on the volume percentage of acetonitrile in the eluent was found in normal-phase TLC with and without immidazolium tetra fluoroborate in the mobile phase. In general, R(M) increased with increasing concentration of acetonitrile. In TLC systems without the ionic liquid, R(M) can be described well with a quadratic function. On the other hand, in a TLC system with an ionic liquid as the additive to the mobile phase, the retention behavior is better described with a third-degree polynomial function. The potential usefulness of ionic liquids for optimization of separation of peptides was demonstrated. Optimization of the separation conditions was supported by a commercially available computer program.

Acetonitriles↗

PubMed2005

Prediction of high-performance liquid chromatography retention of peptides with the use of quantitative structure-retention relationships.

Quantitative structure retention relationships (QSRR) were derived allowing prediction of reversed-phase high-performance liquid chromatography (HPLC) retention of peptides. To quantitatively characterize the structure of a peptide, and then to predict its gradient retention time under given HPLC conditions, the following descriptors are employed: logarithm of the sum of retention times of the amino acids composing the peptide, log Sum(AA), logarithm of Van der Waals volume of the peptide, log VDW(Vol), and logarithm of its calculated n-octanol-water partition coefficient, clog P. The first descriptor is based on a set of empirical data for 20 natural amino acids. The next two descriptors are easily calculated from a structural formula. The predicted gradient retention times are in excellent agreement with the experimental data, determined for a structurally diversified series of 101 peptides. The QSRR equation obtained predicts in a convenient and reliable manner the retention times for any peptide in a once characterized HPLC system.

Amino Acid Sequence↗

PubMed2004

pH gradient high-performance liquid chromatography: theory and applications.

pH gradient high-performance liquid chromatography (HPLC) is a method of reversed-phase high-performance liquid chromatography suitable for ionogenic substances. It consists in programmed increase during the chromatographic process of the eluting strength of eluent with respect to the analytes separated. On the analogy of the conventional organic modifier gradient reversed-phase HPLC, in the pH gradient approach the eluting strength of the mobile phase increases due to its changing pH: increasing in case of acids or decreasing in case of bases. At the same time the content of organic modifier remains constant. A theory of the pH gradient HPLC has been elaborated. The resulting mathematical model is easily manageable. Its ability to predict changes in retention and separation of analytes following the changes in chromatographic conditions is demonstrated. The pH gradient method is uniquely suitable to determine pKa values of analytes. An equation is presented allowing to calculate pKa values basing on appropriate retention data. The effects on pKa are discussed of the concentration of methanol in the mobile phase. The RP HPLC-derived pKa data correlate to the reference pKa values (w(w)pKa) but are not identical. That may be explained by the effects on the chromatographically determined pKa of the specific interactions of analytes with stationary phases. The proposed pH gradient RP HPLC procedure offers a fast and convenient means to get comparable acidity parameters for larger series of compounds, like drug candidates, also when the analytes are available only in minute amounts and/or as complex mixtures.

Chromatography, High Pressure Liquid↗

PubMed2004

Determination of pKa by pH gradient reversed-phase HPLC.

pH gradient reversed-phase HPLC consists of a programmed increase during the chromatographic run of the eluting power of the mobile phase with regard to ionizable analytes. On the analogy of the conventional organic modifier gradient RP HPLC, in the pH gradient mode, the eluting strength of the mobile phase increases due to its increasing (with acid analytes) or decreasing (with basic analytes) pH, whereas the content of organic modifier is kept constant. We have shown previously that the pH gradient separations are technically possible using standard chromatographic equipment. Here we demonstrate that the method is uniquely suitable to determine pK(a) values of analytes. A strict theoretical model is proposed to determine pK(a) values based on the retention data from a pH gradient RP HPLC run. The pK(a) data so obtained are discussed in relation to the concentration of methanol in the mobile phase, the type of stationary phase, and the duration of the gradient. The pK(a) values determined by the pH gradient method are related to the respective data obtained conventionally in a series of isocratic experiments. A close similarity of the two types of chromatographically determined pK(a) data is demonstrated. The HPLC-derived pK(a) parameters correlate to the literature pK(a) values determined by titrations in water. The chromatographically derived and the reference pK(a) values are not identical, however. That is probably due to the effects on the chromatographic pK(a) of the specific sites of interactions with analytes on the surfaces of the HPLC stationary phases. Nonetheless, the proposed pH gradient HPLC method may supply in a fast and convenient manner comparable acidity parameters for larger series of drug candidates, including those available in only minute amounts, without need of their purification, and also when the compounds are provided as complex mixtures, like those produced by combinatorial chemistry.

Journal Article↗

PubMed2004

Suppression of deleterious effects of free silanols in liquid chromatography by imidazolium tetrafluoroborate ionic liquids.

Silica-based stationary phases are commonly used in liquid chromatography, but their surface acidity causes known problems, especially when separating basic compounds. Deleterious effects of free silanols are not fully removed by standard prevention procedures consisting in adding alkylamines or other amino quenchers to the eluents. We found that ionic liquids of the imidazolium tetrafluoroborate class, added to mobile phases at concentrations of 0.5-1.5% (v/v), blocked silanols and provided excellent thin-layer chromatographic separations of strongly basic drugs which were otherwise not eluted, even with neat acetonitrile as the mobile phase. The silanol suppressing potency of imidazolium tetrafluoroborates was demonstrated to markedly exceed that of the standard mobile phase additives, like triethylamine, dimethyloctylamine and ammonia. The proposed new mobile phase additives were also demonstrated to provide reliable lipophilicity parameters of base drug analytes as determined by gradient mode of high-performance liquid chromatography. By applying the readily available and environmentally friendly imidazolium tetrafluoroborate ionic liquids, simple and efficient means of improvement of liquid chromatographic analysis of organic bases were elaborated.

Borates↗

PubMed2004

Artificial neural network analysis for evaluation of peptide MS/MS spectra in proteomics.

The aim of the work was to explore usefulness of artificial neural network (ANN) analysis for the evaluation of proteomics data. The analysis was applied to the data generated by the widely used protein identification program Sequest, completed with several structural parameters readily calculated from peptide molecular formulas. Proteins from yeast cells were identified based on the MS/MS spectra of peptides. The constructed ANN was demonstrated to classify automatically as either "good" or "bad" the peptide MS/MS spectra otherwise classified manually. An appropriately trained ANN proves to be a high-throughput tool facilitating examination of Sequest's results. ANNs are recommended as a means of automatic processing of large amounts of MS/MS data, which normally must be considered in the analysis of complex mixtures of proteins in proteomics.

Artificial Intelligence↗

PubMed2004

pH gradient reversed-phase HPLC.

pH gradient HPLC is reported, which is a new original mode of reversed-phase high-performance liquid chromatography applicable to ionogenic analytes. The method consists of programmed increase during the chromatographic run of the eluting strength of the mobile phase with respect to the acid/base analytes separated. Unlike the well-established conventional gradient HPLC, where the eluting power of the mobile phase is increased with time due to the increasing content of organic modifier, in the pH gradient HPLC that is realized by linearly increasing (in the case of acids) or decreasing (in the case of bases) the pH of the eluent of a fixed organic modifier content, thus providing functional increase in the degree of analyte dissociation and, hence, a decrease in its retention. The pH gradient mode has typical features of gradient HPLC, such as reduced peak width and minimized peak-tailing due to peak compression, which is especially advantageous in the case of organic base analytes. It may be of special value for separation of those analytes which are susceptible to the higher concentrations of organic solvents, as many bioanalytes are. A theory of the pH gradient HPLC has been elaborated, and its full mathematical formalistic is presented step by step in a comprehensive manner. Although fundamental relationships at the basis of pH gradient HPLC are more complex than in the case of the organic gradient variant, the resulting mathematical model is easily manageable. Its applicability to predict changes in retention and separation of test mixtures of analytes accompanying the changes in chromatographic conditions has been demonstrated experimentally in both gradient and isocratic HPLC. The proposed model supplies a rational basis for modifications of eluent pH aimed at optimization of separations and for convenient assessment of chromatographically relevant physicochemical parameters of analytes, such as pK(a).

Journal Article↗

PubMed2004

Artificial neural networks for prediction of antibacterial activity in series of imidazole derivatives.

Artificial neural networks (ANNs) have been applied for the quantitative structure-activity relationships (QSAR) studies of antibacterial activity against Escherichia coli, Serratia marcescens, Proteus vulgaris, Klebsiella pneumoniae and Pseudomonas aeruginosa of a large series of new imidazole derivatives. Antibacterial activity against individual bacteria, expressed as logarithm of reciprocal of the minimal inhibitory concentrations, log 1/MIC, has been related to a number of physicochemical and structural parameters of the imidazole derivatives investigated. Molecular descriptors of agents were obtained by quantum-chemical calculations combined with molecular modelling and from respective structure fragment reference data (e.g., log P). A high correlation resulted between the predicted from ANN model antibacterial activity, log 1/MIC(ANN), and that from biological experiments, log 1/MIC(exp), both for the data used in learning and in the testing sets of imidazoles. Correlation coefficient, R, depending on the type of bacteria and structural subset of analysed imidazole compounds, varies from 0.875 to 0.969. The applicability of ANNs has been demonstrated for the prediction of pharmacological potency of new imidazole derivatives based on their structural descriptors generated exclusively by calculation chemistry.

Anti-Bacterial Agents↗

PubMed2004

High-throughput evaluation of lipophilicity and acidity by new gradient HPLC methods.

There is a need for fast testing of drug candidates for properties of pharmacokinetics and pharmacodynamics importance, in particular lipophilicity and acidity. These two parameters can conveniently be estimated by gradient reversed-phase HPLC. Appropriate conventional organic solvent gradient and the new pH gradient HPLC procedures are presented. The chromatographic parameter of lipophilicity, log kw, can be determined from two organic solvent gradient runs instead of 6-8 runs necessary in the standard isocratic (polycratic) approach. The newly introduced pH gradient reversed-phase HPLC consists in a programmed increase during the chromatographic run of the eluting power of the mobile phase with regards to ionizable analytes. The eluting strength of the mobile phase increases due to its increasing (in case of acidic analytes) or decreasing (basic analytes) pH, whereas the content of organic modifier remains constant. It has been theoretically and experimentally demonstrated that the pKa and log kw values can be evaluated based on retention data from a pH gradient run, combined with appropriate data from two organic solvent gradient runs. The gradient HPLC-derived log kw parameters correlate well with analogous parameters determined isocratically as well as with reference lipophilicity parameter log P (logarithm of n-octanol/water partition coefficient). Also, the HPLC-derived pKa parameters correlate to the literature pKa values (w(w)pKa), conventionally determined by titrations in water. The approach described allows rapid and high-throughput assessment of log kw and pKa for large series of drugs candidates, also when the analytes are available in a form of mixture, e.g. produced by combinatorial synthesis.

Chemical Phenomena↗