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Journal of Biochemical and Biophysical Methods (v.69, #1-2)
Screening and optimization of the derivatization of polar herbicides with trimethylanilinium hydroxide for GC-MS analysis by A. Ranz; E. Lankmayr (pp. 3-14).
In the present study, a derivatization method for the determination of acidic herbicides has been investigated. This procedure involves the methylation with the quaternary ammonium salt trimethylanilinium hydroxide (TMAH) directly in the gas chromatographic auto-sampler vial for analysis by gas chromatography combined with mass spectrometry. The derivatization reaction has been screened for influential factors and statistically significant parameters. The identified factors, reaction time, temperature and hold-up time were optimized by a complete factorial response surface design and optimal reaction conditions were generated. Finally, the optimized methylation procedure was compared to different alkylation methods and obtained results demonstrated the applicability of derivatization with trimethylanilinium hydroxide. Acidic herbicides used in the study consist of several families of compounds like derivatives of acetic acid (2,4-D and 2,4,5-T), butanoic acid (MCPB), benzoic acid (chloramben, dicamba), phenol (dinoseb and dinoterb), propanoic acid (mecoprop) and other miscellaneous acids such as pyridinecarboxlyic acid (picloram). A reliably working, rapid method for the preparation of methyl compounds is generated with respect to automation for routine analysis.
Keywords: Acidic herbicides; Derivatization; Gas chromatography; Optimization; Screening; Trimethylanilinium hydroxide
Screening and optimization of derivatization in heating block for the determination of aliphatic aldehydes by HPLC by A. Stafiej; K. Pyrzynska; A. Ranz; E. Lankmayr (pp. 15-24).
For the study of the derivatization behavior of aliphatic C1–C10 aldehydes with 2,4-dinitrophenylhydrazine (DNPH) in a heating block a gradient elution HPLC method for separation and determination by UV detection at 360nm was applied. The influence of time, temperature, excess of reagent and stirring onto the reaction yields were examined for conducting the reaction in a thermostated heating block. First, the derivatization procedure had been screened by experiments according to a complete factorial design in order to evaluate the statistically significant variables. Those parameters were used to establish the optimum conditions for the reaction by means of a Box-Behnken design.
Keywords: Abbreviations; DNPH; dinitrophenylhydrazine; HPLC; high performance liquid chromatographyHPLC; Aldehydes; 2,4-Dinitrophenylhydrazine; Optimization; Hexanal
Analysis of amino acids and carbohydrates in green coffee by Michael Murkovic; Karin Derler (pp. 25-32).
The analysis of carbohydrates and amino acids in green coffee is of the utmost importance since these two classes of compounds act as precursors of the Maillard reaction during which the colour and aroma are formed. During the course of the Maillard reaction potentially harmful substances like acrylamide or 5-hydroxymethyl-furfural accrue as well. The carbohydrates were analysed by anion-exchange chromatography with pulsed amperometric detection and the amino acids by reversed phase chromatography after derivatization with 6-amino-quinolyl- N-hydroxysuccinimidyl carbamate and fluorescence detection. Both methods had to be optimized to obtain a sufficient resolution of the analytes for identification and quantification. Sucrose is the dominant carbohydrate in green coffee with a concentration of up to 90 mg/g (mean=73 mg/g) in arabica beans and significantly lower amounts in robusta beans (mean=45 mg/g). Alanine is the amino acid with the highest concentration (mean=1200 μg/g) followed by asparagine (mean=680 μg/g) in robusta and 800 μg/g and 360 μg/g in arabica respectively. In general, the concentration of amino acids is higher in robusta than in arabica.
Keywords: Green coffee; Carbohydrates; Amino acids
Chiral separation of T3 enantiomers using stereoselective antibodies as a selector in micro-HPLC by Julia Koidl; Heike Hödl; Martin G. Schmid; Marlene Konrad; Sabine Petschauer; Gerhard M. Kostner; Gerald Gübitz (pp. 33-42).
This work deals with the application of stereoselective antibodies againstl-T3 as a tailor-made chiral selector in micro-HPLC. The separations were performed in microbore columns using commercially available anti-l-T3 antibodies chemically bonded to 5 μm silica gel. The enantiomers of T3 were baseline separated under mild continuous isocratic elution conditions using 10 mM phosphate buffer, pH 7.4. Thed-enantiomer eluted with the void volume, while thel-enantiomer was retained by the antibody phase and eluted second. An indirect competitive and non-competitive enzyme linked immunosorbent assay (ELISA) was used for testing the stereoselectivity of anti-l-T3 antibodies.
Keywords: Abbreviations; PBS; Phosphate-buffered saline.Micro-HPLC; Thyroid hormones; Stereoselective antibodies; Enantiomer separation; Enzyme-linked immunosorbent assay
A simple and rapid ion-pair HPLC method for simultaneous quantitation of 4-nitrophenol and its glucuronide and sulfate conjugates by Attila Almási; Emil Fischer; Pál Perjési (pp. 43-50).
Because of its simple and well characterized metabolic profile, 4-nitrophenol is widely used as a model substrate to investigate the influence of drug therapy, disease, nutrient deficiencies and other physiologically altered conditions on conjugative drug metabolism in animal studies. For simultaneous determination of 4-nitrophenol (PNP), 4-nitrophenyl-β-d-glucuronide (PNP-G) and 4-nitrophenyl-sulfate (PNP-S) in samples generated in rat small intestine luminal perfusion experiments, an ion-pair HPLC assay coupled with UV detection was set up. The RP-HPLC separation was achieved with a methanol–water mixture (50:50, v/v) containing 0.01 M tetrabutyl-ammonium-bromide with UV detection of the analytes at 290 nm. The isocratic system was operated at ambient temperature and required less than 7 min of chromatographic time. The method provided good enough within-day precision, between-day precision and linearity in the target concentration ranges of 6–1200 μM (PNP) and 2.5–100 μM (PNP-G and PNP-S). The instrumental limit of quantification for PNP-G and PNP-S was found to be 2.7 μM and 2.1 μM, respectively. The assay was applied for determination of PNP, PNP-G and PNP-S in rat small intestine perfusates.
Keywords: 4-nitrophenol; 4-nitrophenyl-glucuronide; 4-nitrophenyl-sulfate; HPLC assay; Ion-pair chromatography; Intestinal metabolism
HPLC-RIA analysis of the ectopic cortisol production in a cancerous pancreas tumor by M. Szécsi; I. Tóth; J. Gardi; M. Vecsernyés; J. Németh; J. Julesz (pp. 51-55).
Steroidal pathophysiology of a malignant, ACTH-producing pancreas tumor was investigated via HPLC-RIA determinations of intratissular concentrations of eleven main steroid hormones. The tumor specimen underwent extraction procedure with ethyl acetate and the extract was purified on a C18 minicolumn. Steroids were isolated by HPLC (C18-silica reversed phase stationary phase and methanol–water eluent system) and quantified by specific RIAs. Cortisol content of the tumor specimen was 15,700 pmol/g, the further steroid hormones were found in much lower concentrations (<1.5–28 pmol/g). The extremely high cortisol concentration in the tissue witnesses the synthesis of the main glucocorticoid steroid in the ACTH-producing pancreas tumor and suggests a stimulating paracrine effect of ACTH on cortisol production. The present data verify that the determination of intratissular steroid concentrations by HPLC-RIA methods may identify even the most peculiar hormone sources and the hormone profiles facilitate studying pathophysiology of ectopic endocrine tumors.
Keywords: Steroid isolation and purification; High performance liquid chromatography; Radioimmunoassay; Hypercortisolism; Ectopic ACTH syndrome
Effect of antifungal agents on protein composition of Candida albicans studied by capillary electrophoresis and chip technology by Ildikó Kustos; Adrienn Nyul; Tamás Lóránd; Béla Kocsis; Ferenc Kilár (pp. 57-65).
In the present study protein profile of a Candida albicans strain had been examined by chip technology and conventional capillary electrophoresis (CE). Profiles could be characterised by the presence of ten dominating protein peaks. These proteins could be distinguished by both techniques, but their quantity showed significant differences in the electropherograms obtained by CE and chip method. Changes in the protein profile were induced by administration of different antifungal agents. Fluconazole and amphotericin B treatment was able to induce similar changes in the pattern, appearance of a 40-kDa protein and up-regulation of a 60-kDa protein was observed by chip technology. Increase in the quantity of these proteins under stress effect (antifungal treatment) might refer to their stress function in the fungal cell. Treatment of C. albicans cells with MK 94 (fused cyclic Mannich ketone) antifungal compound induced not only the previously mentioned changes, but further specific alterations, appearance of a 19-kDa protein and up-regulation of the low molecular weight proteins. This might refer to the different mode of action of this agent on the fungal cells. Conventional capillary electrophoresis was suitable to detect the appearance of the 19-kDa peak, and up-regulation of the 60Â kDa protein, but the other changes could not be detected by this technique. Shorter running time, more effective and baseline separation of proteins refer to the advantages of microchip-based method in the analysis of complex biological samples.
Keywords: Abbreviations; C. albicans; Candida albicans; Hsps; heat shock proteins; MIC; minimum inhibitory concentration; MK; Mannich ketone.Microchip electrophoresis; Capillary electrophoresis; Antifungal drugs; Candida albicans; Stress proteins; Fused cyclic Mannich ketone
Enantioseparation of glycyl-dipeptides by CEC using particle-loaded monoliths prepared by ring-opening metathesis polymerization (ROMP) by Christina Gatschelhofer; Martin G. Schmid; Karin Schreiner; Thomas R. Pieber; Frank M. Sinner; Gerald Gübitz (pp. 67-77).
Novel particle-loaded monolithic capillary electrochromatography (CEC) phases for chiral separations were prepared via ring-opening metathesis polymerization (ROMP) within the confines of fused silica columns with 200 μm i.d. using norborn-2-ene (NBE), 1,4,4a,5,8,8a-hexahydro-1,4,5,8, exo,endo-dimethanonaphthalene (DMN-H6) as monomers, 2-propanol and toluene as porogens, RuCl2(PCy3)2(CHPh) as initiator and silica-based particles containing the chiral selector. By suspending silica particles bearing the chiral selector in the polymerization mixture, particle-based monoliths are easily prepared. This approach has several advantages compared to particle-based separation media: (i) the concept of particle-based monoliths is broadly applicable, as any silica-based chiral phase can be used; (ii) they are inexpensive to prepare; and (iii) the manufacturing process is very simple, no sophisticated packing procedures or the preparation of end frits are required. To show the usefulness of this concept for chiral CEC, the chiral separation performance of particle-loaded CEC monoliths bearing teicoplanin aglycone, chemically bonded to 3 μm silica gel, was investigated for a set of glycyl-dipeptides. Particle-loaded ROMP CEC monoliths showed good separation performance for glycyl-dipeptides.
Keywords: Chiral separation; Particle-loaded monoliths; Capillary electrochromatography; Monolithic phases; ROMP; Glycyl dipeptides
Determination of glucose metabolites in stored erythrocytes and in erythrocytes from patients with thalassemia by analytical isotachophoresis by Zyrafete Kuçi; Jürgen Hins; Selim Kuçi; Susanne Renner; Dirk Flottmann; Gernot Bruchelt (pp. 79-87).
Glycolysis is for some cells, such as erythrocytes, neutrophil granulocytes and many cancer cells, the only or most important source of energy (ATP) production. Based on previous studies we developed an isotachophoretic (ITP) method which allows, in principle, the simultaneous determination of all metabolites of glycolysis. Since glucose metabolites are small anions, mobility of some of them may overlap in isotachophoresis and, therefore, partial mixed zones are generated. By variation of the leading/terminating system, however, it is possible to separate the compounds of interest. In this communication, we describe a method for analysis of glucose metabolites in erythrocytes from healthy donors during storage in blood bags, and from patients with thalassemia, with special respect to intracellular 2,3 bisphosphoglycerate, lactate and ATP/ADP. The well known characteristic changes of glycolysis in erythrocytes during blood storage and in erythrocytes from thalassemia patients, which are often analysed by separate enzymatic assays, could be confirmed with this isotachophoretic procedure. The method is currently adapted for analysis of glycolysis in neutrophil granulocytes and cancer cells which requires some modifications of sample preparation and performance of the isotachophoretic analysis.
Keywords: Isotachophoresis; Glycolysis; Erythrocytes; 2,3 bisphosphoglycerate; Thalassemia; Erythrocyte storage
New model substrates for enzymes hydrolysing polyethyleneterephthalate and polyamide fibres by Sonja Heumann; Anita Eberl; Herbert Pobeheim; Stefan Liebminger; Gudrun Fischer-Colbrie; Eva Almansa; Artur Cavaco-Paulo; Georg M. Gübitz (pp. 89-99).
Recently the potential of enzymes for surface hydrophilisation and/or functionalisation of polyethyleneterephthalate (PET) and polyamide (PA) has been discovered. However, there was no correlation between enzyme class/activity (e.g. esterase, lipase, cutinase) and surface hydrolysis of these polymers and consequently no simple assay to estimate this capability. Enzymes active on the model substrates bis (benzoyloxyethyl) terephthalate and adipic acid bishexyl-amide, were also capable of increasing the hydrophilicity of PET and PA. When dosed at the identical activity on 4-nitrophenyl butyrate, only enzymes from Thermobifida fusca, Aspergillus sp., Beauveria sp. and commercial enzymes (TEXAZYME PES sp5 and Lipase PS) increased the hydrophilicity of PET fibres while other esterases and lipases did not show any effect. Activity on PET correlated with the activity on the model substrate. Hydrophilicity of fibres was greatly improved based on increases in rising height of up to 4.3Â cm and the relative decrease of water absorption time between control and sample of the water was up to 76%. Similarly, enzymes increasing the hydrophilicity of PA fibres such as from Nocardia sp., Beauveria sp. and F. solani hydrolysed the model substrate; however, there was no common enzyme activity (e.g. protease, esterase, amidase) which could be attributed to all these enzymes.
Keywords: Polyamide; Polyethyleneterphthalate; Enzyme; Model substrate
Determination of phylloquinone and cholecalciferol encapsulated in granulates formed by melt extrusion by E. Petritz; T. Tritthart; R. Wintersteiger (pp. 101-112).
Vitamin K1 (phylloquinone) and vitamin D3 (cholecalciferol) play a dominant role in bone metabolism. Both vitamins are sensitive to ultraviolet radiation, oxygen and other environmental influences. For this reason a special extrusion technology was developed, that enables an encapsulation of these sensitive substances in a matrix of carbohydrates and hydrogenated carbohydrates. To exclude decomposition products possibly originating under process conditions quantitative analysis was carried out by HPLC/UV using a modified method based on United States Pharmacopoeia. Under the used chromatographic conditions it has to be possible to separate cis-phylloquinone, trans-phylloquinone and phylloquinone 2,3-oxide, as well as pre-cholecalciferol, cis-cholecalciferol and trans-cholecalciferol. A silica column as stationary phase and a mixture of n-hexane and 1-amyl alcohol as mobile phase were used for quantification. UV detection ensued at 254 nm. A linear relationship between peak area and concentration was found over almost two orders of magnitude for cis-phylloquinone, trans-phylloquinone and cholecalciferol. The detection limits (S/N 3) on column were 0.1 μg for phylloquinone and 0.4 μg for cholecalciferol. Analytical results showed that the vitamins were encapsulated sufficiently in the used carbohydrate matrix and that they were protected against environmental influences. After granulation process all of the samples tested met the pharmacopoeial requirements.
Keywords: Phylloquinone; Cholecalciferol; HPLC; Solvent free granulation
Determination of the thermodynamic parameters of the complex formation between malvidin-3-O-glucoside and polyphenols. Copigmentation effect in red wines by Sándor Kunsági-Máté; Kornélia Szabó; Martin P. Nikfardjam; László Kollár (pp. 113-119).
The thermodynamics of the molecular association process between the malvidin-3-O-glucoside and a series of polyphenol derivatives (called ‘copigmentation’ in food chemistry) were studied in aqueous media. The Gibbs free energy, enthalpy and entropy values were determined by the fluorometric method. A combination of the Job's method with the van't Hoff theory was applied for data evaluation. The results show the exothermic character of the copigmentation process. The change of the enthalpy seems to be the same in every complexation step. However, the decreasing of the entropy term is higher at higher stoichiometries. As a result, the Gibbs free energy changes and, thus, the complex stability decreases quickly with increasing stoichiometry. Quantum-chemical investigation reveals the complexity of molecular interactions between malvidin and polyphenols, which is preferably based on π–π and OH–π interaction moderated by repulsive Coulomb-type interactions.
Keywords: Polyphenols; Copigmentation; π–π interaction
Electrochemical behavior of ferrocene in ionic liquid media by LÃvia Nagy; Gergely Gyetvai; László Kollár; Géza Nagy (pp. 121-132).
Chemistry and applicability of ionic liquids (IL), – organic salts with low melting point – are in the focus of interest today. The ILs with melting point below room temperature are expected to be good solvents. Their applicability in organic synthetic work, in separation processes as well as in electrochemistry is very promising. In the work reported here the voltammetric behavior of ferrocene in 1-butyl-3-methylimidazolium hexafluorophosphate (BMIM+ PF6−) ionic liquid has been investigated. Conventional size and micro platinum and carbon electrodes were employed in CV and in chronoamperometric measurements. Karl Fischer method was used for the determination of water content of the solvent.Voltammetric measurements without addition of background electrolyte could be carried out in (BMIM+ PF6−) ionic liquid. A broad potential window could be used. Concentration dependence of the electrochemically determined diffusion coefficient of the ferrocene was observed.
Keywords: Ionic liquids; 1-butyl-3-methylimidazolium hexafluorophosphate; Ferrocene; Voltammetry; Cyclic voltammetry; Platinum microelectrode; Diffusion coefficient measurement
Periodically interrupted amperometry. A way of improving analytical performance of membrane coated electrodes by Livia Nagy; Nikoletta Kálmán; Géza Nagy (pp. 133-141).
Amperometry is a powerful voltammetric measuring method. Its application is specially advantageous when used in combination with a separation step or with some other sample treatment method providing selectivity. The selectivity is often achieved by coating the amperometric working electrode surface with a membrane of special character. Size exclusion membrane, immobilized enzyme containing reaction layer, protecting dialysis membrane, perm selective ion exchange film etc can be mentioned here. In conventional amperometry the measuring potential is continuously applied, therefore in case of membrane coated electrodes the electrode process depletes the diffusion layer. In this work the performance of a new periodically interrupted amperometric (PIA) measuring program has been investigated in case of glucose enzyme sensor. The measuring program allowing time for reloading the diffusion layer provided higher current and therefore improved sensitivity and lower limit of detection.
Keywords: Pulsed amperometry; Glucose enzyme sensor; Sensitivity
Comparison of the effects of selected chalcones, dihydrochalcones and some cyclic flavonoids on mitochondrial outer membrane determined by fluorescence spectroscopy by VladimÃra TomeÄ?ková; Juraj Guzy; Jaroslav KuÅ¡nÃr; Krisztina Fodor; Mária Mareková; Zenóbia Chavková; Pál Perjési (pp. 143-150).
The effect on mitochondrial outer membrane of 4-hydroxychalcone (1), the cyclic chalcone analogues E-2-(4′-hydroxybenzylidene)-1-indanone (2a) and E-2-(4′-hydroxybenzylidene)-1-tetralone (2b), the dihydrochalcones phloretin (3a) and phloridzin (3b), the flavanones naringenin (4a) and naringin (4b), and the flavonol quercetin (5) was investigated by fluorescence spectroscopy. Excitation and emission fluorescence spectra of each flavonoid and synthetic analogue were recorded in respiration medium containing 1mM succinate. Initial interaction of the compounds with the outer mitochondrial membrane was investigated by recording their fluorescence polarization in the presence of rat liver mitochondria. Most of the compounds displayed an elevated fluorescence polarization on mixing with mitochondria at the zero time point. During the investigated 20min period the initial fluorescence polarization values remained constant (1,2a), or a gradual depression of the measured polarization values could be observed (2b,3a,4b,5). In the case of naringenin (4a), however, similar to the previously investigated seven-membered cyclic chalcone analogue E-2-(4´-methoxybenzylidene)-1-benzosuberone, a slight, continuous increase of fluorescence polarization could be detected during the 20min experiment. Phloridzin (3b) showed an increased fluorescence polarization in first 10min, which was slightly depressed by the 20min time point.
Keywords: Chalcones; Dihydrochalcones; Flavanones; Quercetin; Mitochondrial outer membrane; Fluorescence polarization
Three generations of α,γ-diaminobutyric acid modified poly(propyleneimine) dendrimers and their cisplatin-type platinum complexes by E. Bellis; L. Hajba; B. Kovács; K. Sándor; L. Kollár; G. Kokotos (pp. 151-161).
Three generations of α,γ-diaminobutyric acid modified poly(propyleneimine) dendrimers [DAB(AM) n, n=4, 8, 16] containing 4, 8, 16 free amino groups were coupled with Boc-protected α,γ-diaminobutyric acid (DABA) moieties in high yields. These modified dendrimers were deprotected and the chiral dendritic amines with 8, 16 and 32 amino groups on the surface were isolated in excellent yields. Dendrimers with cisplatin moieties at the periphery were obtained in the reaction of the free amine dendrimers and potassium tetrachloroplatinate(II). The highly insoluble complexes were isolated as hydrates and characterized by means of IR, TGA and elemental analysis.
Keywords: Dendrimer; Cisplatin; Platinum; Infrared spectroscopy
Analysis of the vibrational spectra of new OH-containing E-4-arylmethylene-3-isochromanones and 3-arylcoumarins by Gábor Keresztury �; Sándor Holly; Viktória Komlósi; Krisztina István; Tamás Lóránd (pp. 163-177).
A combined experimental and theoretical approach is presented to structural characterization of fairly large, newly synthesized organic molecules in order to enhance the effectiveness of their instrumental analysis by vibrational spectroscopy. The method consists of measurement of FT-IR and Raman spectra of the reaction products and subsequent ab initio or DFT quantum mechanical calculations (prediction) of the vibrational spectra for any anticipated structural varieties of the synthesized molecules. Comparison of the measured and computed frequencies as well as the observed and simulated spectra is performed to resolve any uncertainties in identifying the reaction products. Vibrational frequency and normal mode calculations based on scaled quantum mechanical (SQM) force fields performed at the DFT/B3LYP/6-31G� level of theory are demonstrated to provide a wealth of information that have been used in this work to ascertain the molecular structure, probable conformation and H-bond properties of three new isochromanone or coumarin derivatives, namely: 3-([2′-hydroxymethyl]-phenyl)-coumarin (1), E-4-(3′-hydroxyphenylmethylene)-3-isochromanone (2), and 2-[(2′-hydroxymethyl)phenyl]-3 H-naphto[2,1-b]pyran-3-one (3).
Keywords: Vibrational spectra; SQM force field; DFT calculations; Isomerism; Isochromanones; Arylcoumarin conformers; Hydrogen bonding
Reactions of 3-isochromanone with aromatic aldehydes—microwave assisted condensations performed on solid basic inorganic supports by András Vass; András Földesi; Tamás Lóránd (pp. 179-187).
An improved Knoevenagel condensation of 3-isochromanone and aromatic aldehydes can be achieved by microwave irradiation on solid supports in the presence of various catalysts. This synthetic method offers some major advantages, especially the possibility to change the ratio of E/ Z isomers.
Keywords: Abbreviations; TLC; thin-layer chromatography; MW; microwave heating; Term.; conventional heatingIsochromane; Knoevenagel condensation; Microwave; Validation; TLC
Comparative study of integrating cavity absorption meters by János Erostyák; Tamás Jávorfi; Andrea Buzády; K. Razi Naqvi; GyoË?zoË? Garab (pp. 189-196).
Integrating spheres (IS) are widely used for recording spectra of scattering samples by placing the specimen inside or outside the sphere. An unusual application of integrating spheres has been also demonstrated earlier where the liquid sample completely filled the spherical cavity; such a device is often called an integrating cavity absorption meter (ICAM). In the present work, integrating cavities with different coatings are compared. The spheres were made of glass, covered by metal or white paint, and their surfaces were prepared for diffuse or specular reflectance. The spheres were evaluated by recording kinetic traces following a short light pulse with the aid of time-correlated single photon counting (TCSPC), and by recording steady-state spectra through single-photon counting (SPC) detection. The relative efficiencies of the spheres were determined by comparing the steady-state spectra. Possible reasons for differences in the performance are discussed.
Keywords: Integrating sphere; Absorption; Scattering; Corrected spectra
Investigation on drug dissolution and particle characteristics of pellets related to manufacturing process variables of high-shear granulation by Attila Dévay; Klára Mayer; Szilárd Pál; István Antal (pp. 197-205).
There is a growing interest for multiparticulate solid dosage forms such as pellets, because of their several advantages over tablets during drug therapy. It is essential to investigate the drug dissolution process which can be influenced by the composition and manufacturing process technology, too. This study was performed applying experimental design in order to evaluate the effects of independent process variables during high-shear pelletisation, taking the impeller speed ( x1) and granulation binder flow rate ( x2) as factors into consideration. Theophylline containing pellet formulation was prepared using a matrix consisted of ethylcellulose, microcrystalline cellulose and lactose. Dissolution profiles were modeled by the Weibull function to evaluate the power of process variables. Both process variables were powerful to influence the particle agglomeration. A linear regression was found between the particle size and the diffuse reflectance values after the Kubelka–Munk transformation. Differences in the diffuse reflectance spectra of pellet samples related to particle size offer a fast instrumental method for the in-process control.
Keywords: High-shear pelletisation; Dissolution profile; Particle size; NIR diffuse reflectance spectra; Experimental design
Determination of uranine tracer dye from underground water of Mecsek Hill, Hungary by Andrea Buzády; János Erostyák; Gábor Paál (pp. 207-214).
The underground water system of Mecsek Hill, Hungary was studied using spectrofluorometry. A quantitative dye-tracing technique was used to map the connections between ground water recharge points and the springs and wells. Excitation–emission matrices were used to establish optimal spectroscopic parameters for spectral measurements. Only an EEM gives full spectrofluorimetric information about the sample and makes possible the determination of the optimal wavelength parameters. The uranine concentration was determined by measuring the synchronous excitation spectra of water samples. It was found that under the 4 μg/L uranine concentration, no special chemical treatment was necessary if the pH values remained between 7 and 8. However, above this concentration, the solution should be diluted to avoid spectral distortion from the increasing absorbance of the sample. Long-term concentration measurements were conducted based on spectrofluorometry.
Keywords: Uranine; Tracer dye; Ground-water contamination; Synchronous excitation spectrum; Excitation–emission matrix
Analysis of 3-aminopropionamide: A potential precursor of acrylamide by Kristina Bagdonaite; Gunilla Viklund; Kerstin Skog; Michael Murkovic (pp. 215-221).
An analytical method for the analysis of 3-aminopropionamide (3-APA) based on derivatization with dansyl chloride and liquid chromatography/fluorescence detection was developed. We have analysed 3-APA formation in raw potatoes, grown and stored under different condition, green and roasted coffee beans and in freeze dried mixtures of asparagine with sucrose and glucose in molar ratio of 1:0.5, 1:1, and 1:1.5. In potatoes the 3-APA content varied depending on the potato variety. We detected 3-APA in potatoes up to 14 μg/g fresh weight. In the model experiment glucose had a stronger capacity to form 3-APA. The substance was formed at temperatures as low as 130 °C. However, in the model experiment with sucrose 3-APA was formed not below 150 °C. In heated mixtures with increasing molar ratio of sucrose at 170 °C we noticed a decrease of 3-APA and in the same mixtures at 150 °C we observed an increase of 3-APA. In coffee 3-APA was not formed, neither in green nor in roasted beans.
Keywords: 3-Aminopropionamide; Dansyl chloride; Acrylamide; Coffee; Potato
Effect of perchlorate on the fluorescence of protonated 2,2′-bipyridine by Klára Szabó; Nándor Marek (pp. 223-226).
Interaction between the monocation of an aromatic amine, 2,2′-bipyridine and perchlorate anion was studied. The protonation process by perchloric acid has been followed by the changes in the UV absorption spectra and in the fluorescence emission. Addition of lithium perchlorate with increasing amount results in a significant increase in the fluorescence of 2,2′-bipyridine (the bipyridyl cation). This increase in fluorescence indicates that the rotation in the protonated monocation is hindered in the presence of perchlorate. We assume that this hindrance is due to the formation of the BpH+·ClO4− ion-pair.
Keywords: Protonated 2,2′-bipyridine; Perchlorate anion; Ion-pair inhibited rotation; Fluorescence
The application of in vivo microiontophoresis for the investigation of mast cell–neuron interactions in the rat brain by Zsombor Kőszegi; Péter Kovács; Márta Wilhelm; Tamás Atlasz; Norbert Babai; Veronika Kállai; István Hernádi (pp. 227-231).
Although mast cells are immune cells of hematopoietic origin, they can be found in parts of the central nervous system of many mammalian species. In the rat brain they are located in the thalamic region. Their function is not defined yet, although they are mostly known to secrete several chemicals, which may influence the surrounding neurons. There are no in vivo electrophysiological data available on the possible effects of brain mast cells on neurons. In this study, we used a combined method of microiontophoresis and extracellular single unit recording to simultaneously activate mast cells and record neuronal action potentials. Four-barrelled micropipettes were used for recording neuronal activity and for microiontophoretic application of mast cell degranulator Compound 48/80 (C48/80). Spike sorting routines were performed on-line and off-line to ensure that data were always recorded from a single neuron. C48/80 did not modify the firing rate of cortical neurons (no mast cells are found there), however, it caused excitation ( n=16/37, 43%), or inhibition ( n=9/37, 24%) in thalamic neurons possibly due to mast cell activation. Further investigations will clarify the biochemical nature of changes in neural excitability due to mast cell degranulation in the mammalian brain.
Keywords: Electrophysiology; Mast cells; Microiontophoresis; Thalamus
The application of surface plethysmography for heart rate variability analysis after GSM radiofrequency exposure by Tamás Atlasz; Lóránd Kellényi; Péter Kovács; Norbert Babai; György Thuróczy; László Hejjel; István Hernádi (pp. 233-236).
The aim of the present study was to test whether the electromagnetic field emitted by standard GSM mobile phones results in changes in heart rate (HR) and heart rate variability (HRV) of 35 healthy young male and female subjects. Two parallel signals, electrocardiogram and infrared surface plethysmogram were recorded and compared to test their validity for the analysis. Plethysmographic recording is proved to be a fast and reliable method for HRV measurements. In the radiofrequency (RF) exposure study, there was no significant difference in the values of HR and HRV between the RF and the sham groups. Our preliminary study demonstrates that, in our experimental conditions, RF fields emitted by cellular phones do not cause observable effects on the regulation of heart rate of healthy, young adults.
Keywords: RF fields; Infrared surface plethysmography; Heart rate variability; Human