Jiří Klimeš

The role of van der Waals forces in water adsorption on metals

The interaction of water molecules with metal surfaces is typically weak and as a result van der Waals (vdW) forces can be expected to be of importance. Here we account for the systematic poor treatment of vdW forces in most popular density functional theory exchange-correlation functionals by applying accurate non-local vdW density functionals. We have computed the adsorption of a variety of exemplar systems including water monomer adsorption on Al(111), Cu(111), Cu(110), Ru(0001), Rh(111), Pd(111), Ag(111), Pt(111), and unreconstructed Au(111), and small clusters (up to 6 waters) on Cu(110). We show that non-local correlations contribute substantially to the water-metal bond in all systems, whilst water-water bonding is much less affected by non-local correlations. Interestingly non-local correlations contribute more to the adsorption of water on the reactive transition metal substrates than they do on the noble metals. The relative stability, adsorption sites, and adsorption geometries of competing water adstructures rarely differ when comparing results obtained with semi-local functionals and the non-local vdW density functionals, which explains the previous success of semi-local functionals in characterizing adsorbed water structures on a number of metal surfaces.

Investigation of the stability of aromatic hydrazones in plasma and related biological material

Novel aromatic hydrazones derived from pyridoxal isonicotinoyl hydrazone (PIH) are interesting compounds from the viewpoint of their pharmacodynamic activity. However, they were recently shown to suffer from relatively short biological half-lives. The purpose of the present study was to investigate the stability of novel aroylhydrazones in plasma and related biological media in order to reveal its potential involvement in the pharmacokinetics of these drugs. Three different aroylhydrazones (pyridoxal isonicotinoyl hydrazone - PIH, salicylaldehyde isonicotinoyl hydrazone - SIH and pyridoxal 2-chlorobenzoyl hydrazone - o-108) were incubated in plasma from different species, plasma ultrafiltrate, bovine serum albumin, RPMI cell medium and phosphate buffer saline (PBS) at 37 degrees C. Stability of these compounds was determined using precise, selective and validated HPLC methods. Although the aroylhydrazones were relatively stable in PBS, they underwent rapid degradation in plasma. Plasma proteins as well as low molecular weight components were involved in this matter. Furthermore, the products of hydrazone bond splitting revealed in this study were also found in the chromatograms from pharmacokinetic experiments. In the light of short biological half-lives determined in vivo, these in vitro findings strongly suggest that hydrolysis of investigated aromatic hydrazones in plasma could have a significant impact on their pharmacokinetics. Furthermore, these results also suggest that plasma stability of other novel drug candidates containing the hydrazone bond deserves to be considered.

HPLC-DAD and MS/MS analysis of novel drug candidates from the group of aromatic hydrazones revealing the presence of geometric isomers.

Salicylaldehyde isonicotinoyl hydrazone (SIH) is an iron-chelating aromatic hydrazone with promising pharmacological properties. However, it suffers from relatively short biological half-life. Hence, two novel derivates of SIH, HAP-INH and HPP-INH were synthesized in order to overcome this pharmacokinetic drawback. The aim of the present study was to employ HPLC-DAD and HPLC-MS/MS methods to investigate the identity of the putative impurities of these newly prepared substances, which are being formed in aqueous environment. At first, it was shown that their retention times as well as UV spectra did not correspond to any expected synthetic precursor, by-product or degradation product. HPLC-DAD analysis confirmed purity of peaks and revealed close but not identical UV spectra of putative impurities and corresponding hydrazones. The subsequent HPLC-MS/MS analyses using ESI and the ion trap mass analyzer showed the identical molecular ions (in both modes) as well as their fragmentation, which implicated presence of geometric isomers. This suggestion was further supported by the NMR analyses. Since the Z/E isomers can have different biological activities, results of this study might be of great importance for further development of the aroylhydrazones as novel drug candidates as well as from the theoretical point of view.

Photochemical stability of nimesulide.

HPLC and TLC methods for monitoring of the photochemical stability of nimesulide are presented. Solution of nimesulide sodium salt was exposed to the light of wavelengths 254 nm. The presence of degradation products (2-phenoxy4-nitroaniline and methanesulfonic acid) was observed. In the exposed sample, 2-phenoxy4-nitroaniline was detected by HPLC analysis and sulfonic acid was detected by TLC analysis. An isocratic HPLC chromatographic condition was described for determination of nimesulide in a presence of its degradation product. The sample was analysed on Separon SGX, C(18), 250 x 4.6 i.d. 7 microm analytical column. The mobile phase was consisted of a mixture of acetonitrile and ammonium phosphate (pH 7.9; 0.02 M) (35:65 v/v). UV detector was performed at 245 nm. Propylparaben was employed as an internal standard. Standard area response was linear respect to concentration of nimesulide over range 150-500 microg/ml. As a validation of the method, the accuracy and between-day precision were done. The detection limit of 2-phenoxy4-nitroaniline was 0.12 microg/ml. The solvent system for TLC analysis was consisted of ethylacetate and cyclohexane (45:55), the samples were plotted on silica gel UV-254 nm. UV lamp (254 nm) and the chemical detection were used.

HPLC evaluation of diclofenac in transdermal therapeutic preparations

High-performance liquid chromatography (HPLC) was selected for analytical evaluation of sodium diclofenac in original transdermal therapeutic preparations containing adjuvant substances (capsaicin, hyoscyamine). After isolation from laminated adhesive patches, diclofenac was analysed on columns with reversed phase, using the mobile phase ethanol and phosphate buffer (pH 6.5) with an addition of tetrabutylammonium iodide and detection at 284 nm. Not only the total amount of diclofenac in the patch was evaluated, but HPLC methodology was also employed to select a suitable acceptor medium for permeation experiments. In patches manufactured in the tested series, HPLC was also employed to examine the release of diclofenac and its in vitro permeation through the human skin.

The retention behaviour of polar compounds on zirconia based stationary phases under hydrophilic interaction liquid chromatography conditions.

The most separations in HILIC mode are performed on silica-based supports. Nevertheless, recently published results have indicated that the metal oxides stationary phases also possess the ability to interact with hydrophilic compounds under HILIC conditions. This paper primarily describes the retention behaviour of model hydrophilic analytes (4-aminobenzene sulfonic acid, 4-aminobenzoic acid, 4-hydroxybenzoic acid, 3,4-diaminobenzoic acid, 3-aminophenol and 3-nitrophenol) on the polybutadine modified zirconia in HILIC. The results were simultaneously compared with a bare zirconia and a silica-based HILIC phase. The mobile phase strength, pH and the column temperature were systematically modified to assess their impact on the retention of model compounds. It was found that the retention of our model hydrophilic analytes on both zirconia phases was mainly governed by adsorption while on the silica-based HILIC phase partitioning was primarily involved. The ability of ligand-exchange interactions of zirconia surface with a carboxylic moiety influenced substantially the response of carboxylic acids on the elevated temperature as well as to the change of the mobile phase pH in contrast to the silica phase. However, no or negligible ligand-exchange interactions were observed for sulfanilic acid. The results of this study clearly demonstrated the ability of modified zirconia phase to retain polar acidic compounds under HILIC conditions, which might substantially enlarge the application area of the zirconia-based stationary phases.

Hydrophilic interaction liquid chromatography in the separation of a moderately lipophilic drug from its highly polar metabolites--the cardioprotectant dexrazoxane as a model case.

This paper presents a systematic study of the retention behavior of a model bisdioxopiperazine drug, dexrazoxane (DEX) and its three polar metabolites (two single open-ring intermediates-B and C and an EDTA-like active compound ADR-925) on different stationary phases intended for hydrophilic interaction liquid chromatography (HILIC). The main aim was to estimate advantages and limitations of HILIC in the simultaneous analysis of a moderately lipophilic parent drug and its highly polar metabolites, including positional isomers, under MS compatible conditions. The study involved two bare silica columns (Ascentic Express HILIC, Atlantis HILIC) and two stationary phases with distinct zwitterionic properties (Obelisc N and ZIC HILIC). The chromatographic conditions (mobile phase strength and pH, column temperature) were systematically modified to assess their impact on retention and separation of the studied compounds. It was found that the bare silica phases were unable to separate the positional isomers (intermediates B and C), whereas both columns with zwitterionic properties (Obelisc N and ZIC HILIC) were able to separate these structurally very similar compounds. However, only ZIC HILIC phase allowed appropriate separation of DEX and all its metabolites to a base line within a single run. A mobile phase composed of a mixture of ammonium formate (0.5 mM) and acetonitrile (25:75, v/v) was suggested as optimal for the simultaneous analysis of DEX and its metabolites on ZIC HILIC. Thereafter, HILIC-LC-MS analysis of DEX and all its metabolites was performed for the first time to obtain basic data about the applicability of the suggested chromatographic conditions. Hence, this study demonstrates that HILIC could be a viable solution for the challenging analysis of moderately polar parent drug along with its highly polar metabolites including the ability to separate structurally very similar compounds, such as positional isomers.

Determination of lipophilicity of novel potential antituberculotic agents using HPLC on monolithic stationary phase and theoretical calculations

The HPLC analyses on the monolithic stationary phase were employed for rapid determination of lipophilicity of the two sets of newly synthesized potential antituberculotic agents. The analyses utilized the mixture of methanol and phosphate buffer (pH 7.4) as a mobile phase and a flow rate of 4mL/min. Monolithic stationary phase enabled to significantly reduce the time of analyses, achieve appropriate peak shapes for all tested compounds as well as the separation of positional isomers. Furthermore, the theoretical lipophilic parameters (logP) for all compounds were calculated employing the chemical programs (e.g., ACD/logP, HyperChem, miLogP, AlogP, KOWWIN and COSMOFrag, etc.). The experimental data (logk) and calculated logP values were compared by linear regression analysis. The highest correlation for both series was obtained for KOWWIN and miLogP programs. However, capability of particular chemical software to precisely predict lipophilicity of a compound is structurally dependent. Thus the predictive power of the selected program should be verified using experimental method. The results of this study documented that experimental determination of lipophilicity using HPLC on monolithic stationary phase is practical and reasonable for this purpose.

Combination of molecular modeling and quantitative structure-activity relationship analysis in the study of antimycobacterial activity of pyridine derivatives

A set of 4-benzylsulfanyl derivatives of pyridine-2-carbonitriles and pyridine-2-carbothioamides, previously tested for their antimycobacterial activity, were analysed by quantitative structure-activity relationship (QSAR) techniques, using some physicochemical and quantum-chemical parameters. The resulting QSAR revealed that the activity increases with electron withdrawing substituents in the benzyl moiety of studied compounds. HOMO orbitals can play an important role in the description of the mechanism of interactions at the molecular level. Additionally, the results of multiple linear regression indicate the differences between Mycobacterium tuberculosis and M. avium. The hydrophobicity of studied compounds is important for activity against M. avium.

HPLC methods for determination of two novel thiosemicarbazone anti-cancer drugs (N4mT and Dp44mT) in plasma and their application to in vitro plasma stability of these agents.

The aim of this study was to develop and validate HPLC methods for the determination in plasma of two novel thiosemicarbazone anti-tumour drugs developed in our laboratories (Dp44mT and N4mT). The appropriate separations were achieved using a HS F5 HPLC column with the mobile phase composed of a mixture of either acetate buffer/EDTA or EDTA and acetonitrile (62:38 and 50:50, v/v, respectively). The plasma samples were pretreated with SPE (phenyl and C18, respectively). Furthermore, these methods were successfully applied to in vitro plasma stability experiments. The investigation has clearly shown that both thiosemicarbazones are markedly more stable in plasma than their aroylhydrazone forerunners.