RESUMEN
A force field has been previously designed for a dodecasaccharide chain of chondroitin-6-sulfate (C6S) and has proved to yield valuable data going from basic conformational properties to a more detailed H-bonding network. This force field is further used here to unravel the interaction of C6S with its pathological counterpart in low density lipoprotein (LDL) particles. In particular, well-selected peptide fragment p2 (residues 3359-3377) also identified as the principal proteoglycan binding site (PPBS) of the major protein in LDL, apolipoproteinB-100 (apoB-100), was chosen. We study here the interaction between C6S and p2. The role of arginine and lysine, positively charged amino acids of p2, in the crucial interaction of C6S with LDL is highlighted. The secondary structure of p2 is shown to affect the efficiency of the interaction, as the α-helical structure of p2 allows optimal interaction with C6S also in dynamic conditions. One point mutation in p2 appeared to affect consequently p2-C6S interaction.
Asunto(s)
Apolipoproteína B-100/química , Sulfatos de Condroitina/química , Simulación de Dinámica Molecular , Secuencia de Aminoácidos , Apolipoproteína B-100/metabolismo , Sitios de Unión , Enlace de Hidrógeno , Datos de Secuencia MolecularRESUMEN
Field-flow fractionation (FFF) is one of the most versatile separation techniques in the field of analytical separation sciences, capable of separating macromolecules in the range 10³-10¹5 g mol⻹ and/or particles with 1 nm-100 µm in diameter. The most universal and most frequently used FFF technique, flow FFF, includes three types of techniques, namely symmetrical flow FFF, hollow fiber flow FFF, and asymmetrical flow FFF which is most established variant among them. This review provides a brief look at the theoretical background of analyte retention and separation efficiency in FFF, followed by a comprehensive overview of the current status of asymmetrical flow FFF with selected applications in the field of biopolymers and bioparticles.
Asunto(s)
Biopolímeros/química , Fraccionamiento de Campo-Flujo/métodos , Biopolímeros/aislamiento & purificación , Liposomas/química , Modelos Químicos , Tamaño de la Partícula , Virión/químicaRESUMEN
Chondroitin-6-sulfate (C6S) is a glycosaminoglycan (GAG) constituent in the extracellular matrix, which participates actively in crucial biological processes, as well as in various pathological conditions, such as atherosclerosis and cancer. Molecular interactions involving the C6S chain are therefore of considerable interest. A computational model for atomistic simulation was built. This work describes the design and validation of a force field for a C6S dodecasaccharide chain. The results of an extensive molecular dynamics simulation performed with the new force field provide a novel insight into the structure and dynamics of the C6S chain. The intramolecular H-bonds in the disaccharide linkage region are suggested to play a major role in determining the chain structural dynamics. Moreover, the unravelling of an additional H-bond involving the sulfate groups in C6S is interesting as changes in sulfation have been claimed to be an important factor in several diseases. The force field will prove useful for future studies of crucial interactions between C6S and various nanoassemblies. It can also be used as a basis for modeling of other GAGs.
Asunto(s)
Sulfatos de Condroitina/química , Simulación de Dinámica Molecular , Teoría CuánticaRESUMEN
Pressurised hot water extraction-microporous membrane liquid-liquid extraction was coupled on-line with gas chromatography-mass spectrometry (PHWE-MMLLE-GC-MS) for the analysis of pesticides in grapes. MMLLE serves as a trapping step after PHWE. Water from PHWE is directed to the donor side of the membrane unit and the analytes are extracted to the acceptor solution on the other side. The role of MMLLE is to clean and concentrate the extract before on-line transfer to the GC via a sample loop and an on-column interface using partially concurrent solvent evaporation. The extraction conditions were investigated, and then the quantitative features such as linearity, limit of quantification (LOQ), extraction yield and enrichment factors. LOQs in the range 0.3-1.8 microg kg(-1) were achieved. Procymidone and tetradifon were found in the skins of the grapes. The results were in good agreement with those obtained by liquid-solid and ultrasonic extractions.
Asunto(s)
Contaminación de Alimentos/análisis , Residuos de Plaguicidas/análisis , Vitis , Autoanálisis , Cromatografía de Gases y Espectrometría de Masas/métodos , HumanosRESUMEN
Microporous membrane liquid-liquid extraction (MMLLE) was coupled on-line with gas chromatography for the determination of pesticides in wine. The MMLLE-GC provided to be efficient and selective and the method was linear, repeatable and sensitive. The limits of detection ranged from 0.05 to 2.3 microg/l and the limits of quantification were 0.2-7.5 microg/l for all the analytes using FID as detector. With MS detection LODs in the range 0.03-0.4 and LOQs of 0.3-3.5 microg/l were achieved. The method was applied to the determination of pesticides in several red wines of different origin.
Asunto(s)
Cromatografía de Gases/métodos , Plaguicidas/análisis , Vino/análisisRESUMEN
Pressurised hot water extraction (PHWE) was coupled on-line with microporous membrane liquid-liquid extraction (MMLLE) and gas chromatography (GC) in the analysis of polycyclic aromatic hydrocarbon (PAH) compounds in soil. The MMLLE serves as a trapping device after the PHWE. Water from PHWE is directed to the donor side of the membrane unit and the analytes are extracted to the acceptor solution on the other side of the membrane. The role of MMLLE is to clean and concentrate the extract, which is then transferred on-line to the GC via a sample loop and an on-column interface using partially concurrent solvent evaporation. Separate optimisation of MMLLE and simulations of the PHWE-MMLLE connection were carried out before the actual on-line coupling. After optimisation of the whole on-line system, the efficiencies of the PHWE-MMLLE-GC and PHWE-solid-phase trap extractions were compared. The PHWE-MMLLE-GC method allowed on-line analysis of soil samples. The method was linear, with limits of detection in the range 0.05-0.13 ng and limits of quantification 0.65-1.66 microg g(-1). Comparison of the results with those obtained by other techniques confirmed the good performance.
Asunto(s)
Fraccionamiento Químico/métodos , Hidrocarburos Policíclicos Aromáticos/análisis , Contaminantes del Suelo/análisis , Fraccionamiento Químico/instrumentación , Cromatografía de Gases , Diseño de Equipo , Sistemas en Línea , Sensibilidad y Especificidad , Suelo/normasRESUMEN
A sample pretreatment method based on microporous membrane liquid-liquid extraction (MMLLE) was developed for the subsequent gas chromatographic determination of pesticides in wine. MMLLE provided efficient and selective extraction with enrichment factors in the range 3-13. The gas chromatographic separation was carried out using on-column injection and flame ionization detection. The method was linear, repeatable and sensitive. The limits of quantification were better than 0.006 mg/L for all the analytes except for iprodione (0.37 mg/L). The method was applied to the determination of pesticides in several red wines of different origin.
Asunto(s)
Cromatografía de Gases/métodos , Membranas Artificiales , Residuos de Plaguicidas/análisis , Vino/análisis , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
The resolution of the analytes was predicted from their known pKa* values and actual mobilities in nonaqueous methanolic solutions according to theory taking longitudinal diffusion as the only cause for peak dispersion. This leads to an equation of the resolution as a function of the pH*, as both selectivity and efficiency are dependent on the pH of the buffer. The experimentally obtained resolution values were in acceptable agreement with the predicted theoretical ones in most cases. This was demonstrated for substituted benzoic acids as analytes. The pKa* values needed for the calculation of the resolution were derived from the pH* dependence of the effective mobility. The pH* scale in methanol was based on conventional pKa* values of acetic acid and chloroacetic acid taken from the literature.
Asunto(s)
Electroforesis Capilar/métodos , Metanol , Solventes , Benzoatos/aislamiento & purificación , Modelos Teóricos , Relación Estructura-ActividadRESUMEN
Coated capillaries can be advantageous in many capillary electrophoretic applications where nonaqueous background electrolytes are used. In the present work, a new dynamic polymer coating (poly(glycidylmethacrylate-co-N-vinylpyrrolidone)) for methanol-based background electrolytes is introduced. The magnitude and stability of electroosmotic flow was investigated with coated capillaries at pH* values of 3, 7.8, and 10.4 in methanol. At pH* 7.8 and 10.4 the electroosmotic flow was negligible and repeatable. On the other hand, at pH* 3 a weak, unstable electroosmotic flow was observed, due to a change in the conformation of the polymer under acidic conditions. The dynamically coated capillaries were successfully applied to the separations of cationic drugs, phenols, and benzoic acids. The synthesis and characterization of the polymer are described in detail.
Asunto(s)
Materiales Biocompatibles Revestidos , Codeína/análogos & derivados , Electroforesis Capilar/métodos , Concentración de Iones de Hidrógeno , Alprenolol/aislamiento & purificación , Materiales Biocompatibles Revestidos/química , Codeína/aislamiento & purificación , Electrólitos , Electroforesis Capilar/instrumentación , Efedrina/aislamiento & purificación , Luz , Espectroscopía de Resonancia Magnética , Metanfetamina/aislamiento & purificación , Metanol , Propranolol/aislamiento & purificación , Dispersión de RadiaciónRESUMEN
This study describes three ways to couple micellar electrokinetic chromatography (MEKC) on-line with electrospray ionization mass spectrometry (ESI-MS) for the analysis of N-methylcarbamate pesticides. The methods involved the use of a partial filling (PF) technique under basic conditions and the use of reverse migrating micelles (RMMs) under acidic and basic conditions. The use of RMMs in basic electrolyte solutions required coated capillaries with low electroosmotic flows, and capillaries coated with anionic poly(sodium 2-acrylamide-2-methylpropanesulfonate) were selected for the purpose. Before the on-line MEKC-ESI-MS coupling, the MEKC and MS conditions were separately optimized under off-line conditions. The methods were compared in terms of detection limits and the stability of the electrospray process. The PF method offered good separation but poorer stability of the electrospray relative to the other methods. A more stable electrospray performance was obtained with use of RMMs in acidic electrolyte solutions, but some of the analytes were protonated and could not be detected due to the increase in their retention factors. However, with the use of anionic polymer-coated capillaries and RMMs at pH 8.5, all analytes were successfully separated. The high-salt stacking method was applied to improve the sensitivity of MEKC-ESI-MS and the detection limits were in the range of 0.04-2.0 microg/ml.
Asunto(s)
Carbamatos/química , Cromatografía Capilar Electrocinética Micelar/métodos , Insecticidas/química , Micelas , Espectrometría de Masa por Ionización de Electrospray/métodos , Sensibilidad y EspecificidadRESUMEN
The mobilities of cationic analytes in organic solvents and water are compared, and the reasons for differences in the mobilities are discussed in detail. Actual mobilities (at background electrolyte concentration 10 mmol/l) of anilinium ions were determined by capillary zone electrophoresis in water, methanol, acetonitrile and mixtures of methanol and acetonitrile (in volume ratios 1:1, 1:3 and 3:1). The actual mobilities correlated with the viscosity of the organic solvent: the products of actual mobility and viscosity were constant within 7%. However, these products were significantly larger in water. Larger products of mobility and viscosity in water were also found for unsubstituted anilinium when the absolute mobility (at zero ionic strength) was taken into consideration. Thus, ion-solvent interactions must be responsible for the seemingly high mobility in water compared with that in organic solvents. This finding can be explained by the effect of the ion on the water structure. Based on equilibrium constant for ion-pair formation given in the literature, about 20% of the main background electrolyte constituent (tetrapropylammonium perchlorate) is associated at 10 mmol/l concentration in acetonitrile. Comparison of the plot of the measured mobilities of the analytes vs. the square root of the corrected ionic strength of the background electrolyte in acetonitrile with the prediction based on the Debye-Hückel-Onsager theory showed the measured mobilities deviate negatively from the theoretical line. This is apparently due to ion pairing, which takes place for the analytes as well.
Asunto(s)
Acetonitrilos/química , Cationes/aislamiento & purificación , Electroforesis Capilar/métodos , Metanol/química , Concentración OsmolarRESUMEN
4-Chloro-3-methylphenol (c = 2.0 mM), representing a model pollutant, was oxidized in pressurized hot water and in supercritical water in a continuous flow system. Potassium persulfate was used as oxidant in concentrations of 8.0 and 40.0 mM. Contact times (reaction times) were 3-59 s, temperatures 110-390 degrees C, and pressures 235-310 bar. A wide temperature range was tested to determine the range over which potassium persulfate can be used effectively. Good oxidation efficiencies for 4-chloro-3-methylphenol were obtained at both oxidant concentrations and with short contact times at temperatures clearly underthe critical temperature of water; total organic carbon content of the effluent was low under optimized conditions. Corrosion, measured as nickel and chromium concentrations of the effluent, was more severe at oxidant concentration of 40.0 mM. Sulfate was present in the effluent in high concentrations. Sulfate is the limiting factor in the use of potassium persulfate in wastewater treatment and requires further water treatment.
Asunto(s)
Oxidantes/química , Fenoles/química , Compuestos de Potasio/química , Sulfatos/química , Contaminantes Químicos del Agua/análisis , Purificación del Agua/métodos , Oxidación-Reducción , TemperaturaRESUMEN
A fast and easy method was sought for determination of the iridoid glycosides catalpol, ketologanin, verbenalin, loganin, 8-epi-loganic acid, geniposidic acid and 10-cinnamoyl catalpol in plant samples. The method involved micellar electrokinetic capillary chromatography (MEKC) coupled on-line to mass spectrometry. The partial filling technique and electrospray ionization were used. Seven iridoid glycosides could be separated with use of MEKC under basic conditions. However, 8-epi-loganic acid and geniposidic acid could not be detected simultaneously with the five neutral iridoid glycosides by mass spectrometry. Therefore, only the neutral iridoid glycosides were screened from plant samples. Catalpol, verbenalin, loganin and possibly 10-cinnamoyl catalpol were found in an examination of seven plant species in the genera Plantago, Veronica, Melampyrum, Succisa, and Valeriana. Aucubin, which was not included in the sample mixture used in method development because of overlapping with catalpol in MEKC, was also detected. The limits of detection for the iridoid glycosides, both at the UV and at the mass spectrometer, are given.
Asunto(s)
Cromatografía Capilar Electrocinética Micelar/métodos , Glucósidos/análisis , Espectrometría de Masas/métodos , Plantas Medicinales/química , Piranos/análisis , Glucósidos/química , Concentración de Iones de Hidrógeno , Glucósidos Iridoides , Glicósidos Iridoides , Iridoides , Estructura Molecular , Peso Molecular , Plantago/química , Piranos/química , Scrophulariaceae/química , Especificidad de la Especie , Espectrofotometría Ultravioleta , Valerianaceae/químicaRESUMEN
The influence of high electric field strength on the separation of basic analytes in non-aqueous alcohol background electrolyte (BGE) solutions was investigated. Increasing the separation voltage in capillary electrophoresis (CE) may be advantageous if the conductivity of the BGE solution is low enough to allow fast separations without excessive Joule heating or band broadening. The voltage range tested was 20-60 kV with methanol and ethanol, and 25-60 kV with propanol and butanol as solvent for BGE. The resulting electric field strengths ranged from 660 V cm(-1) to 2000 V cm(-1). Experiments were made with a special laboratory constructed CE instrument. The separation efficiency vs. voltage curve was found to vary with the alcohol BGE solution. The increase in voltage decreased the separation efficiency in the case of methanol BGE solution, but with the other BGEs a clear efficiency maximum was obtained above 30 kV. The highest separation efficiencies were achieved with propanol BGE solution, where the efficiency maximum was reached at 45 kV. However, reasonable efficiency was achieved even at 60 kV. The extent of Joule heating was determined by calculating the temperature inside the capillary and the observed plate heights were interpreted in terms of the Van Deemter equation. The decrease in the separation efficiency with higher voltage was attributed mainly to Joule heating in the case of methanol and ethanol BGE solution and to the analyte adsorption on the capillary wall with propanol and butanol BGE solutions.
Asunto(s)
Electroforesis Capilar/métodos , Electricidad , TemperaturaRESUMEN
Liposomes made of mixtures of zwitterionic and anionic lipids were investigated by means of capillary electrophoresis and dynamic light scattering. The influence of the molar lipid ratio and of the buffers, used in the running electrolyte solution, on the physical characteristics of the liposomes were investigated. Data on effective electrophoretic mobilities, total charges as well as sizes of the liposomes are given. In addition, examples on the use of liposomes as carriers in electrokinetic capillary electrophoresis for the separation of benzene derivatives, steroids, and phenols are shown.
Asunto(s)
Liposomas/química , Electroforesis Capilar , Liposomas/análisisRESUMEN
The electrophoretically relevant properties of monoacidic 21 bases (including common drugs) containing aliphatic or aromatic amino groups were determined in methanol as solvent. These properties are the actual mobilities (that of the fully ionised weak bases), and their pKa values. Actual mobilities were measured in acidic methanolic solutions containing perchloric acid. The ionisation constants of the amines were derived from the dependence of the ionic mobilities on the pH of the background electrolyte solution. The pH scale in methanol was established from acids with known conventional pK*a values in this solvent used as buffers, avoiding thus further adjustment with a pH sensitive electrode that might bias the scale. Actual mobilities in methanol were found larger than in water, and do not correlate well with the solvent's viscosity. The pK*a values of the cation acids, HB-, the corresponding form of the base, B, are higher in methanol, whereas a less pronounced shift was found than for neutral acids of type HA. The mean increase (compared to pure aqueous solution) for aliphatic ammonium type analytes is 1.8, for substituted anilinium 1.1, and for aromatic ammonium from pyridinium type 0.5 units. The interpretation of this shift was undertaken with the concept of the medium effect on the particles involved in the acid-base equilibrium: the proton, the molecular base, B, and the cation HB+.
Asunto(s)
Electroforesis Capilar/métodos , Metanol/química , Solventes/química , Concentración de Iones de Hidrógeno , Espectrofotometría UltravioletaRESUMEN
An online multidimensional liquid chromatographic system was developed for online clean-up, derivatisation and separation of biogenic amines in wines. The system consists of a cation-exchange precolumn, a derivatisation coil and an analytical column, and a column-switching valve. The entire system can be easily automated. The method proved to be quantitative and sensitive. Limits of detection were below 0.05 mg l(-1) for all amines and linearity was preserved over the tested range (0.05-15 mg l(-1)). The method was applied to the analysis of red wines of different origin.
Asunto(s)
Aminas Biogénicas/análisis , Vino/análisis , Cromatografía Liquida/métodosRESUMEN
A simple method with minimal manual sample preparation was developed for the analysis of phenols in pyrolysis oils. Sample pre-treatment was done by gel permeation chromatography (GPC), where the high-molecular-mass lignins were separated from the phenols. Multidimensional liquid chromatography (LC-LC) was used in the analysis of the phenolic fraction. The pre-column was used for sample clean-up and pre-fractionation before introduction of the phenolic fraction to the analytical column. The repeatability and linearity of the total GPC and LC-LC methods were excellent. The results were in accordance with the reference method in which the sample pre-treatment was done by precipitating the lignins with water, and the phenols were extracted with toluene and analysed by GC-MS.
Asunto(s)
Cromatografía en Gel/métodos , Cromatografía Liquida/métodos , Petróleo/análisis , Fenoles/análisis , Reproducibilidad de los ResultadosRESUMEN
The benefits of non-aqueous capillary electrophoresis have been described in a number of recent publications. The wide selection of organic solvents, with their very different physicochemical properties, broadens our scope to manipulate separation selectivity. The lower currents present in non-aqueous solvents allow the use of high electric field strengths and wide bore capillaries, the latter in turn allowing larger sample load. In many cases detection sensitivity can also be enhanced. The potential of non-aqueous capillary electrophoresis is discussed throughout the paper, and the feasibility of capillary electrophoresis under non-aqueous media is demonstrated with reference to several applications.
Asunto(s)
Electroforesis Capilar/métodos , Análisis Espectral/métodosRESUMEN
Liposomes are small membrane-enclosed vesicles composed of either natural or synthetic lipids. Their size can be adjusted on a wide scale and they can be made with well-defined compositions. While liposomes have been extensively used as model biomembranes they have also gained a considerable degree of attention as carriers for drugs as well as for genetic material. The physical properties of liposomes are critically dependent on their chemical composition. In this study liposomes were applied as pseudostationary phases in electrokinetic capillary chromatography. Various negatively charged liposomes, consisting of mixtures of zwitterionic and anionic lipids, were investigated. Major emphasis was put on clarifying the effects of the total lipid concentration, the lipid molar ratio, the lipid head group, and the buffer on the capillary electrophoretic separation of neutral analytes. In addition, the influence of the physical state of the membrane, ie., gel vs. fluid, on the separation was investigated. Corticosteroids were applied as model analytes.