Supercritical fluid extraction of amino acids from birch (Betula pendula Roth) leaves and their liquid chromatographic determination with fluorimetric detection

A method for supercritical fluid extraction (SFE) of amino acids was adapted and optimal experimental conditions were selected for a matrix consisting of dry leaves. The matrix-dependent SFE method uses a mixture of MeOH-H(2)O-acetonitrile (10:10:1 v/v/v) as a modifier (0.5 mL in situ, 300 muL on-line) at 70 degrees C and 40 MPa and no HCl is needed as an entrainer. The amino acids were quantified using high-performance liquid chromatography with fluorimetric detection (HPLC/FLD) after gradient elution on Zorbax Eclipse AAA columns (4.6x150 mm, 3.5 mum) with aqueous Na(2)HPO(4 )buffer of pH 7.8 and ACN-MeOH-water as a mobile phase. In comparison with Soxhlet extraction, SFE gave higher recovery and selectivity, but it required longer extraction time (90 min) and it was more labor-intensive (clean-up step after the pre-concentration). Both methods should be used separately or in combination according to the matrix, number of samples, and levels of ballast compounds.     

Application of infrared spectroscopy to the assessment of authenticity of tea

Diffusion reflectance fourier transform infrared spectroscopy in the mid-IR region was used to assess the authenticity of tea varieties. The differences between the spectra of 12 different tea varieties (seven black, two green, three semifermented grades) were sufficient to allow their discrimination by the soft independent modelling of class analogy classification method or linear discriminant analysis, despite a significant heterogeneity of the samples as revealed by variance analysis.     

Determination of a series of quinolones in pig plasma using solid-phase extraction and liquid chromatography coupled with mass spectrometric detection Application to pharmacokinetic studies

A simple and sensitive liquid chromatography (LC) method was developed for the simultaneous determination of eight quinolones in pig plasma samples. The following two methods of detection were used: ultraviolet (UV) and mass spectrometry with electrospray ionization (ESI/MS). Sample preparation consisted of solid-phase extraction (SPE) on Strata X cartridges prior to the analysis by LC/UV or LC/ESI/MS. The recovery, linearity, limit of detection (LOD) and limit of quantification (LOQ), precision and accuracy of the method were evaluated using spiked pig plasma samples. The suitability of the method for pharmacokinetic studies was evaluated by determining the concentrations of enrofloxacin (ENR) and ciprofloxacin (CIP) also in pig plasma, after administration of 200mg of enrofloxacin per kilogram of fodder during 5 consecutive days.     

Separation and determination of terbinafine and its four impurities of similar structure using simple RP-HPLC method

A novel reversed-phase HPLC method for the simultaneous determination of active component terbinafine, its one impurity 1-methylaminomethylnaphtalene and three degradation products, β-terbinafine, Z-terbinafine and 4-methyl-terbinafine occurring in pharmaceutical formulations after long-term stability tests, was developed and validated using propylparaben as an internal standard.The chromatographic separation was performed on a NUCLEOSIL 100-5-CN column, mobile phase for separation of all compounds consisted of a mixture of tetrahydrofurane, acetonitrile and citrate buffer pH 4.50 (10:20:70, v/v/v). The analysis time was less than 32 min at flow-rate of 0.8 ml min⁻¹. UV detection was performed at 226 nm. The method was validated and system suitability parameters were investigated. Method robustness and short-term standard solution stability were verified. Limits of detection for terbinafine degradation products/impurity were from 0.023 to 0.098 μg ml⁻¹, limits of quantitation were from 0.078 to 0.327 μg ml⁻¹. The method was applicable for routine determination of terbinafine and all its found impurities of similar structure with sufficient selectivity, precision and accuracy.     

226Ra translocation from soil to selected vegetation in the Crucea (Romania) uranium mining area

A two year field survey of ²²⁶Ra translocation from soil to selected spontaneous plants growing in the Crucea (Romania) uranium mining area is presented. Since Crucea is a large and relatively highly contaminated area, an easy and cheap way for a desired lowering of radiopollution represents phytoremediation using plentifully occurring plant species. The ²²⁶Ra translocation from soil was affected by analyzed species, collecting sites and vegetation periods. The specific ²²⁶Ra activities of the roots were considerably higher than the aboveground parts confirming the root tropism of radioisotopes. Mentha piperita was found to be highly effective in phytoremediation, especially in autumn.     

Isolation of polymerase chain reaction-ready bacterial DNA from Lake Baikal sediments by carboxyl-functionalised magnetic polymer microspheres

Carboxyl group-containing magnetic nonporous poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate) (P(HEMA-co-EDMA)) microspheres were used for the isolation of polymerase chain reaction (PCR)-ready DNA from samples of Baikal sediments. DNA was isolated using the phenol extraction method or the Soil DNA Isolation Kit. The occurrence of false-negative results in PCR caused by the presence of extracellular inhibitors in DNA samples was solved using solid phase reversible DNA immobilisation. PCR-ready DNA was reversibly adsorbed to the microspheres in the presence of 8.0% (w/v) poly(ethylene glycol) (PEG 6000) and 2.0M sodium chloride concentrations. The adsorbed DNA was released from the microspheres in a low ionic strength TE buffer. The quality of isolated DNA was checked by PCR amplification.     

Possibilities of column coupling electrophoresis provided with a fiber-based diode array detection in enantioselective analysis of drugs in pharmaceutical and clinical samples

The present work illustrated possibilities of column coupling electrophoresis combined with ionizable chiral selector and diode array detection (DAD) for the enantioselective analysis of trace drugs (pheniramine and its analogs) in pharmaceutical and clinical samples. Isotachophoresis (ITP), on-line coupled with capillary zone electrophoresis (CZE), served as an ideal injection technique (high sample load capacity, narrow and sharp drugs zones) of on-line pretreated samples (preseparation, purification and preconcentration of drugs) for the CZE stage. Enhanced (enantio)separation selectivity of CZE with ionizable chiral selector (carboxyethyl-beta-cyclodextrin recognized between drugs enantiomers on one hand as well as between drugs and sample matrix constituents on the other hand) enabled to obtain pure zones of the drugs enantiomers, suitable for their detection and quantitation. DAD in comparison with single wavelength UV detection enhanced value of analytical information verifying purity of drugs enantiomers zones (indicating interferents with different spectra to those of drugs). Obtained results indicated pure zones of interest confirming effective ITP-CZE (enantio)separation process. Distinguishing the trace analytes signals superposed on the baseline noise was provided with sufficient reliability (for this purpose the background correction and smoothing procedure had to be applied to the raw DAD spectra). The proposed ITP-CZE-DAD methods were characterized by favorable performance parameters (sensitivity, linearity, precision, recovery, accuracy, robustness, selectivity) and successfully applied for (i) enantiomeric purity testing of dexbrompheniramine in commercial pharmaceutical tablets and (ii) enantioselective metabolic study of pheniramine in human urine.     

Facile synthesis and relaxation properties of novel bispolyazamacrocyclic Gd3+ complexes: an attempt towards calcium-sensitive MRI contrast agents

Three novel GdDO3A-type bismacrocyclic complexes, conjugated to Ca (2+) chelating moieties like ethylenediaminetetraacetic acid and diethylenetriamine pentaacetic acid bisamides, were synthesized as potential "smart" magnetic resonance imaging contrast agents. Their sensitivity toward Ca (2+) was studied by relaxometric titrations. A maximum relaxivity increase of 15, 6, and 32% was observed upon Ca (2+) binding for Gd 2L (1), Gd 2L (2), and Gd 2L (3), respectively (L (1) = N, N-bis{1-[{[({1-[1,4,7-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-10-yl]eth-2-yl}amino)carbonyl]methyl}-(carboxymethyl)amino]eth-2-yl}aminoacetic acid; L (2) = N, N-bis[1-({[({alpha-[1,4,7-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-10-yl]- p-tolylamino}carbonyl)methyl]-(carboxymethyl)}amino)eth-2-yl]aminoacetic acid; L (3) = 1,2-bis[{[({1-[1,4,7-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-10-yl]eth-2-yl}amino)carbonyl]methyl}(carboxymethyl)amino]ethane). The apparent association constants are log K A = 3.6 +/- 0.1 for Gd 2L (1) and log K A = 3.4 +/- 0.1 for Gd 2L (3). For the interaction between Mg (2+) and Gd 2L (1), log K A = 2.7 +/- 0.1 has been determined, while no relaxivity change was detected with Gd 2L (3). Luminescence lifetime measurements on the Eu (3+) complexes in the absence of Ca (2+) gave hydration numbers of q = 0.9 (Eu 2L (1)), 0.7 (Eu 2L (2)), and 1.3 (Eu 2L (3)). The parameters influencing proton relaxivity of the Gd (3+) complexes were assessed by a combined nuclear magnetic relaxation dispersion (NMRD) and (17)O NMR study. Water exchange is relatively slow on Gd 2L (1) and Gd 2L (2) ( k ex (298) = 0.5 and 0.8 x 10 (6) s (-1)), while it is faster on Gd 2L (3) (k ex (298) = 80 x 10 (6) s (-1)); in any case, it is not sensitive to the presence of Ca (2+). The rotational correlation time, tau R (298), differs for the three complexes and reflects their rigidity. Due to the benzene linker, the Gd 2L (2) complex is remarkably rigid, with a correspondingly high relaxivity despite the low hydration number ( r 1 = 10.2 mM (-1)s (-1) at 60 MHz, 298 K). On the basis of all available experimental data from luminescence, (17)O NMR, and NMRD studies on the Eu (3+) and Gd (3+) complexes of L (1) and L (3) in the absence and in the presence of Ca (2+), we conclude that the relaxivity increase observed upon Ca (2+) addition can be mainly ascribed to the increase in the hydration number, and, to a smaller extent, to the Ca (2+)-induced rigidification of the complex.     

Separation of phenolic acids by capillary electrophoresis with indirect contactless conductometric detection

A new method for the electrophoretic separation of nine phenolic acids (derivatives of benzoic and cinnamic acids) with contactless conductometric detection is presented. Based on theoretical calculations, in which the mobility of the electrolyte co- and counterions and mobility of analytes are taken into consideration, the electrolyte composition and detection mode was selected. This approach was found to be especially valuable for optimization of the electrolyte composition for the separation of analytes having medium mobility. Indirect conductometric detection mode was superior to the direct mode as predicted theoretically. The best performance was achieved with 150 mM 2-amino-2-methylpropanol electrolyte at pH 11.6. The separation was carried out in a counter-electroosmotic mode and completed in less than 6 min. The LODs achieved were about 2.3-3.3 microM and could be further improved to 0.12-0.17 microM by using a sample stacking procedure. The method compares well to the UV-Vis detection.     

Negative electrospray, ion trap multistage mass spectrometry of synthetic fragments of the O-PS of Vibrio cholerae O:1

Saccharides (mono through hexasaccharides) that mimic the terminal epitopes of O-antigens of Vibrio cholerae O:1, serotypes Ogawa and Inaba, were studied by electrospray ion trap (ESI IT) mass spectrometry (MS) in the negative mode. Anionized adducts are the characteristic ions formed by the capture of H(3)O(2)(-) under the condition of ESI MS analysis. The reactive species are produced by reaction of hydroxyl anions with the molecule of water. Thus the [M + H(3)O(2)](-) have the highest m/z value in the ESI IT negative mass spectra. After dissociation of adducts by loss of 2H(2)O the [M-H](-) ions are produced. The fragmentation pathways were confirmed by multistage measurements (MS(n)). The predominant pathway of fragmentation of the mono- and oligomers is the elimination of a molecule of alpha- hydroxy--gammabutyrolactone from the 4-(3-deoxy-L-glycero-tetronamido) group. The other characteristic pathway occurs by shortening the length of oligosaccharides. In this way, conversion of the Ogawa to Inaba fragments takes place under the conditions of measurement. Negative ESI MS/MS provided sufficient information about molecular mass, the number of saccharide residues, basic structure of saccharides, about the tetronamide part of the compounds investigated and allowed Ogawa and Inaba serotypes to be distinguished.