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Biomedical subjects

W T Kok

Publications and source records attributed to W T Kok.

At least 19 recordsLinked to original sources
PubMed2001

Retention behaviour of amylopectins in asymmetrical flow field-flow fractionation studied by multi-angle light scattering detection.

Asymmetrical flow field-flow fractionation (FFF) with multi-angle light scattering (MALS) detection was applied for the fractionation of amylopectins from four different sources. Samples originated from genetically modified potatoes and waxy maize. Amylopectins were dissolved in a 1 mol l(-1) sodium hydroxide solution or water. With an injected mass of 0.2 microg, well below overloading conditions, a decrease of the apparent hydrodynamic radius with increasing inlet flow-rate was observed. Moreover, a decrease of the radius of gyration with increasing elution volume was recorded by the MALS detector. Steric/hyperlayer effects are a feasible explanation for this behaviour. The observed radius of gyration at the steric inversion point was in the order of 0.3 microm, which is smaller than the theoretically calculated inversion point. Apparently, the amylopectin behave as macromolecules with a larger hydrodynamic radius than expected on basis of their radius of gyration and are subjected to significant lift forces. The results were confirmed by four fractionations with varying flow-rates but constant ratio of cross to outlet-flow. In contrast to the normal mode operation, the retention of the amylopectins depended strongly on the applied flow-rates and was close to that of a much smaller 10 kDa dextran. Apparent molar masses in the order of between 10(7) and 10(9) g mol(-1) were obtained. The results are contrasted with enzymatically degraded and oxidised starch samples that were fractionated in the normal mode.

Amylopectin↗
PubMed2001

Determination of the molecular mass distribution of synthetic polymers by size-exclusion electrochromatography.

The performance of size-exclusion electrochromatography (SEEC) for the mass distribution analysis of synthetic polymers was studied and compared to conventional, pressure-driven size-exclusion chromatography (SEC). Electroosmotic flow control, within-day, day-to-day and column-to-column repeatability were determined for SEEC with respect to retention and separation efficiency. It was shown that by using the retention ratio instead of the migration time, the precision of the mass distribution calculations is sufficiently high, and that similar distributions were obtained for a sample analyzed by pressure-driven SEC and by SEEC. Furthermore, hexafluoroisopropanol was demonstrated to be a new and potent solvent for SEEC. It was used for the separation of narrow polymethylmethacrylate standards and several commercially important polymers such as polycarbonate, polycaprolactam and poly(ethylene terephthalate), using UV detection in the deep UV region (195-230 nm).

Chromatography, Gel↗
PubMed2001

Effects of pore flow on the separation efficiency in capillary electrochromatography with porous particles.

The effect of pore flow on the separation efficiency of capillary electrochromatography (CEC) has been studied using columns packed with particles with different pore sizes. A previously developed model was used to predict the (relative) pore flow velocity in these columns under various experimental conditions. Equations are derived describing the effect of pore flow on peak broadening in CEC. The theory has been compared with practice in the reversed-phase CEC separation of various polyaromatic hydrocarbons. It is shown, by theory and experimentally, that the mass-transfer resistance contribution to peak dispersion can be effectively eliminated when using porous particles with a high (> or =50 nm) average pore diameter. Moreover, at high pore-to-interstitial flow ratios the flow inhomogeneity contribution (the A term in the plate height equation) is also shown to decrease. Under optimal conditions, a reduced plate height of 0.3 for the nonretained compound could be obtained. It is argued that fully perfusive porous particles can be a more efficient separation medium in CEC than nonporous particles.

Journal Article↗
PubMed2001

Size-exclusion electrochromatography with controlled pore flow.

In size-exclusion electrochromatography (SEEC) there exists an optimum in pore-to-interstitial flow ratio with respect to the resolution. Two methods for finding and controlling the optimal pore-to-interstitial flow ratio in SEEC have been studied: (i) varying the ionic strength of the mobile phase and (ii) the application of a hydrodynamic flow in addition to the electrco-driven flow. Both methodologies have been evaluated in terms of efficiency and applicability with columns packed with silica particles containing pores of either 10 or 50 nm in diameter, and with different ionic strength mobile phases. Using the first method with the 10-nm pore particles, the flow ratio could be adjusted within an appropriate range. However, with the wide-pore (50 nm) particles it appeared that the pore-to-interstitial flow ratio was too high at all conditions tested to obtain proper selectivity. In the second approach, the desired pore flow was generated by the electric field and the pore-to-interstitial flow ratio could then be adjusted by an applied pressure over the column. This method was applicable with both particle types studied. The application of a (low) voltage gradient in addition to a pressure-driven flow resulted in a sharply improved separation efficiency as a result of a strongly improved mass transfer due to intra-particle electroosmotic flow. When optimized, pressurized SEEC generates identical separation efficiencies for polystyrene standards as are obtained with pure SEEC, while the reduction in selectivity, in comparison to pressure-driven SEC, is kept minimal.

Chromatography, Gel↗
PubMed2001

Influence of the carrier composition on thermal field-flow fractionation for the characterisation of sub-micron polystyrene latex particles.

A study on the influence of the carrier composition in a ThFFF system on the retention and thermal diffusion of sub-micron polystyrene latex particles has been carried out. Various factors that may influence retention were studied. These include: the type of electrolyte and surfactant, their respective concentrations, and the addition of an organic modifier. Particle retention is highly sensitive to small changes in the carrier composition. It is demonstrated that under the conditions applied, secondary effects, such as particle-wall and particle-particle interactions, are negligible. Addition of surfactants is required to minimise particle-wall interactions. Generally, retention increases at higher electrolyte concentration. Furthermore, the addition of acetonitrile (ACN) to an aqueous carrier leads also to an increased retention. The type of surfactant as well as its concentration is of influence on the retention time. The three surfactants that were studied, i.e., sodium dodecyl sulfate, Brij 35 and cetyltrimethylammonium bromide, showed significant differences in particle retention behaviour. The observed differences in retention in the carriers can be attributed to actual changes in thermal diffusion. D(T) appears to be mainly determined by the interaction between the particle's surface and the carrier liquid, and is therefore highly sensitive to changes in the chemical composition of the particle surface and the carrier. Strong differences in size selectivity were found for different carrier compositions. This allows a relatively easy optimisation of the separation. On the other hand, it complicates the size and composition analysis of particles.

Chemical Fractionation↗
PubMed2000

Determination of aromatic amines in water samples by capillary electrophoresis with electrochemical and fluorescence detection.

Two capillary electrophoresis methods have been compared for the determination of aniline derivatives in environmental water samples. With the first method the anilines were separated as cations by free zone electrophoresis at low pH, and detected by amperometry. For this, the separation capillary was connected through a palladium field decoupler to an electrochemical detection cell which had been modified to match the volume scale of the separation. Most anilines tested, except chlorinated compounds, could be detected with full sensitivity at a detection potential of +0.7 V. Detection limits with this detection scheme were on a low microg/l level. The alternative method involved the derivatization of the anilines with fluorescamine, the separation of the derivatives formed by micellar electrokinetic chromatography, and fluorescence detection. For detection a lamp-based, fibre optics instrument was used. Detection limits with fluorimetry were comparable with those obtained with amperometric detection (in the order of 1 microg/l). Still, this method was preferred since it gave a higher separation efficiency and shorter analysis times (approximately 4 min). The most important argument, however, was its higher reliability and ease-of-handling. Preliminary experiments with water samples collected in areas where pollution with anilines may be expected showed that the method is highly specific, with few interferences showing up in the electropherograms.

Amines↗
PubMed2000

Modelling of the pore flow in capillary electrochromatography.

Pore flow in capillary electrochromatography (CEC) on porous silica particles has been investigated. To that end the migration behaviour of narrow polystyrene (PS) standards dissolved in di-methylformamide (DMF) with lithium chloride in 1 and 10 mmol/l concentration has been measured. These data have been compared to theoretical predictions. The latter were based on a model comprising cylindrical pores of varying diameter as measured experimentally by porosimetry, while the flow in each set of pores was calculated with the expression given by Rice and Whitehead. A reasonable to good agreement between experimental and predicted data was observed, provided it was assumed that pores of differing diameter occur in series. It was found that the flow in pores with a nominal size of 100 A can be considerable compared to the interstitial flow, especially at 10 mmol/l ionic strength. It is concluded that pore flow within porous particles in CEC, of great importance for improved efficiency in both interactive and exclusion type CEC, can be predicted fairly reliably by means of the Rice and Whitehead expression.

Dimethylformamide↗
PubMed2000

Application of micellar electrokinetic chromatography for the separation of retinoids.

The applicability of micellar electrokinetic chromatography (MEKC) using sodium dodecylsulphate (SDS) as pseudo-stationary phase for the separation of five retinoids (retinol, retinal, retinyl acetate, retinyl palmitate, retinoic acid), was investigated. The effects of the acetonitrile content, the SDS concentration, the pH and the addition of Brij 35 to the background electrolyte on the migration behaviour of the retinoids were determined. It was found that the effective mobilities of retinol, retinal and retinyl acetate could be easily regulated through the ACN content and the SDS concentration of the BGE. The electrophoretic behaviour of the very hydrophobic retinyl palmitate was abnormal. Under various conditions this compound showed up as a late, very sharp peak. A strong indication was found that the retinyl palmitate forms a stable, charged complex with SDS during sample preparation. The mobility of the retinyl palmitate peak could be regulated, independently from the other peaks, through the Brij concentration of the BGE. Using a running buffer consisting of Tris buffer (pH 8), 20 mmol l(-1) SDS, 1 mmol l(-1) Brij 35 and 35% (v/v) acetonitrile, a complete separation of the five retinoids could be realised in less than 20 min.

Acetonitriles↗
PubMed1999

Capillary electrochromatography with macroporous particles.

The performance of macro-porous particles in capillary electrochromatography is studied. Three reversed-phase stationary phases with pore diameters between 500 A and 4000 A have been tested for separation efficiency and mobile phase velocity. With these stationary phases, a large portion of the total flow appears to be through the pores of particles, thereby increasing the separation efficiency through a further decrease of the flow inhomogeneity and through enhancement of the mass transfer kinetics. The effects of pore size and mobile phase composition on the plate height and mobile phase velocity have been studied. With increasing buffer concentrations and larger pore diameters, higher mobile phase velocities and higher separation efficiencies have been obtained. Columns packed with 7 microns particles containing pores with a diameter of 4000 A generated up to 430,000 theoretical plates/m for retained compounds. Reduced plate heights as low as 0.34 have been observed, clearly demonstrating that a significant portion of the flow is through the pores. For the particles containing 4000 A pores no minimum was observed in the H-u plot up to linear velocities of 3.3 mm/s, suggesting that the separation efficiency is dominated by axial diffusion. On relatively long (72 cm) columns, efficiencies of up to 230,000 theoretical plates/column have been obtained under non-optimal running conditions. On short (8.3 cm) columns fast separations could be performed with approximately 15,000 theoretical plates generated in less than 30 s.

Chromatography↗
PubMed1998

Post-column derivatization for fluorescence and chemiluminescence detection in capillary electrophoresis.

Instrumental developments and applications of post-column derivatization for fluorescence and chemiluminescence detection in capillary electrophoresis (CE) are reviewed. Various systems to merge the reagent solution with the separation medium have been developed, including coaxial capillary reactors, gap reactors and free solution or end-column systems. For all reactor types the geometry of the system, as well as the method to propel the reaction mixture (by pressure or by voltage) appeared to be critical to preserve the separation efficiency. Plate numbers of over 100,000 could be realised with different reactors. The strict requirements on the rate of post-column derivatization reactions to be applied in CE limit the number of different reagents that have been used. For fluorescence detection, with laser or lamps as the excitation source, so far mainly o-phthalaldehyde and its naphthalene analogue have been used as reagent. Derivatization systems that are based on complexation reactions also showed good promise for application in CE. Detection limits could be obtained that were comparable to those obtained after pre-column derivatization. Various reagents for chemiluminescence detection (e.g. the luminol and peroxyoxalate systems) have been studied. The often complicated chemistry involved made application of these reagents in CE even more difficult. Results obtained so far, in terms of sensitivity, have not been up to expectation, with detection limits usually in the order of micromol l(-1).

Chemistry Techniques, Analytical↗
PubMed1998

Postcolumn derivatization of peptides with fluorescamine in capillary electrophoresis.

Fluorescamine is used as a postcolumn derivatization reagent for fluorescence detection detection of peptides after separation by capillary electrophoresis. The problems resulting from the use of an organic solvent have been solved by introducing LiC1O4 and 5% water into the postcolumn derivatization reagent. The reaction rate and detection sensitivity of amino acids and small peptides observed with fluorescamine and OPA were compared. Fluorescamine gives much higher sensitivity than o-phthaldialdehyde (OPA) for small peptides, with detection limits for the selected peptides and amino acids below 0.1 mumol 1-1. Under optimized experimental conditions, the method has a good reproducibility and separation efficiency for peptides. The method was applied for the analysis of the protein tryptic digests. Only submicromolar concentrations of proteins were required.

Amino Acids↗
PubMed1997

Determination of catecholamines and related compounds by capillary electrophoresis with postcolumn terbium complexation and sensitized luminescence detection.

A method has been developed for the determination of catecholamines and related compounds (DA, E, NE, DOPA, DOPAC, DOPAG) in urine samples. The compounds of interest were separated by capillary electrophoresis with a borate buffer as background electrolyte. Detection was based on the sensitized luminescence of terbium ions. Using a homemade postcolumn reactor, a basic solution containing a stoichiometric mixture of terbium(III) chloride and EDTA was added postcolumn to the separation buffer. The ternary catechol-EDTA-terbium complexes formed in the reaction mixture showed a strong luminescence with excitation and emission maxima at 300 and 545 nm, respectively. By optimization of the experimental conditions, zone broadening could be restrained and plate numbers up to 130,000 appeared to be possible. Detection limits found were in the order of 10(-7) mol L-1. The catecholic compounds could be determined in urine samples after a standard cleanup and preconcentration procedure.

Catecholamines↗
PubMed1996

Magnesium fractions in serum of healthy individuals and CAPD patients, measured by an ion-selective electrode and ultrafiltration.

OBJECTIVES: Validation and comparison of a magnesium ion-selective electrode (ISE) with a cation-exchange resin technique, followed by determination of all magnesium fractions in serum of healthy volunteers and continuous ambulatory peritoneal dialysis (CAPD) patients. DESIGN AND METHODS: The analytical aspect has been studied by measuring the influence of complexing agents on the fraction ionized magnesium (friMg2+). A theoretical approximation of friMg2+, based on mass equilibria and complexation constants, was calculated and compared with the measurements. RESULTS: ISE measurements showed good agreement with theory. Reference values of the ionized, protein-bound, and complexed magnesium fractions were (mean +/- SD) 0.65 +/- 0.04, 0.27 +/- 0.04, and 0.08 +/- 0.03, respectively. Fractions obtained in the CAPD group were 0.62 +/- 0.04, 0.22 +/- 0.05, and 0.16 +/- 0.05, respectively, and differed significantly from the values of the reference population. CONCLUSIONS: All known serum magnesium parameters can be established by a combination of ultrafiltration, atomic absorption spectrometry, and ISE measurements. Unknown complexing compounds most probably account for the increased fraction of complexed magnesium in the serum of CAPD patients.

Adult↗
PubMed1994

Simultaneous determination of urinary creatinine, calcium and other inorganic cations by capillary zone electrophoresis with indirect ultraviolet detection.

A method for the rapid analysis of calcium and creatinine in urine has been developed using capillary zone electrophoresis without sample pretreatment other than dilution. Cations of NH3, K, Na, Mg, Ca, Li, Ba and creatinine in human urine samples could be separated within 7 min. Background electrolyte (BGE) was composed of 5 mmol/l pyridine (chromophore), 3.6 mmol/l tartaric acid and 2 mmol/l 18-crown-6, having a pH of 4.05. An indirect UV detection mode was employed at 255 nm. Efforts were made to eliminate the interaction between cations and proteins and to minimize the electromigrative dispersion (EMD) for the analytes of interest by selecting a suitable BGE and sample dilution solution. Quantitative analyses were performed for calcium and creatinine. The calibration plots showed good linearity over the concentration range of interest to clinical analysis. Data on recoveries and reproducibilities are also reported. Results for urine samples (previously collected and frozen) from a variety of healthy and pathological individuals were in good agreement with those obtained by the Technicon SMA II calcium and creatinine methods.

Ammonia↗
PubMed1994

Derivatization reactions for the determination of aflatoxins by liquid chromatography with fluorescence detection.

Various derivatization methods for the fluorometric detection of aflatoxins after separation by HPLC are reviewed. In normal-phase chromatography the sensitivity for aflatoxins B1 and B2 was improved by using special mobile phases or a flow cell packed with silica-gel particles. In the nowadays more popular reversed-phase methods, the fluorescence intensity of B1 and G1 can be increased by precolumn derivatization with trifluoroacetic acid or by postcolumn derivatization with iodine or bromine. Optimum conditions for the reactions are discussed. In terms of sensitivity, the three derivatization schemes give similar results. The methods are compared with respect to experimental convenience, selectivity, reproducibility and suitability for automation.

Aflatoxins↗
PubMed1994

Analytical evaluation of Kone Microlyte determination of ionized magnesium.

We performed an analytical evaluation of a commercially available instrument for determining ionized magnesium through use of a neutral carrier, liquid-membrane-based ion-selective electrode. Reproducibility (CV 2-4%), linearity (0.30-2.50 mmol/L), lower limit of detection (0.30 mmol/L), and absence of interference from Ca2+ indicate adequate performance for measuring ionized magnesium in plasma or serum samples in the normal to high-concentration range. Sodium in excess of 150 mmol/L caused a negative bias, which can be explained by ionic strength-induced changes in activity coefficients. The use of heparin as an anticoagulant must be restricted to concentrations < 15 units/mL because of the binding of magnesium to heparin. The mean +/- SD concentration of ionized magnesium and its fraction of total magnesium in 76 healthy volunteers were 0.56 +/- 0.05 mmol/L and 0.65 +/- 0.04, respectively.

Adolescent↗
PubMed1990

Selective determination of trace levels of phenol in river water using electrochemical concentration modulation correlation chromatography.

Electrochemical concentration modulation (ECM) was used as a sample introduction technique in the correlation chromatographic (CC) trace determination of phenol in water. The linearity and sensitivity of the method were tested and detection limits were calculated. The selectivity of the technique was confirmed by comparison with loop injection experiments using several detection methods. In preliminary experiments is was found that ECM-CC, in combination with fluorescence detection, is selective and sensitive enough to be used for the monitoring of the phenol concentration in river water at the draining points for drinking water production.

Chromatography↗