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Gerhardus J de Jong

Publications and source records attributed to Gerhardus J de Jong.

At least 19 recordsLinked to original sources
PubMed2006

Direct sample injection for capillary electrophoretic determination of organic acids in cerebrospinal fluid.

Organic acids in cerebrospinal fluid (CSF) are potential diagnostic markers for neurological diseases and metabolic disorders. A capillary electrophoretic (CE) method for the direct analysis, i.e., without any sample preparation, of six organic acids in CSF was developed. A capillary coating consisting of a triple layer of charged polymers (polybrene-dextran sulfate-polybrene) was used in combination with a negative separation voltage, providing fast and efficient analysis of acidic compounds. Separation conditions, such as background electrolyte (BGE) concentration and pH were optimized, and the influence of albumin and sodium chloride was systematically studied using a set of test compounds. With injection volumes of ca. 44 nL, plate numbers of up to ca. 150,000 were obtained with a BGE of 200 mM sodium phosphate (pH 6.0). It appeared that high sodium chloride concentrations in the sample hardly affected the peak width and shape of the organic acids, most probably due to transient isotachophoresis effects occurring in the sample zone. Adverse effects of CSF proteins, which frequently compromise the CE performance, could be effectively minimized by the triple layer coating in combination with rinses of 0.1 M hydrochloric acid. Overall, the developed CE system allowed direct injections of CSF samples, yielding good separation efficiencies and stable migration times (RSDs<2%) for organic acids. Validation of the method with artificial and real CSF samples showed good linear responses (r>0.99), and LODs for the organic acids were in the range of 2-8 microg/mL when applying UV detection. RSDs for migration times and peak areas were <2% and <7%, respectively. The applicability of the CE system is shown for the determination of organic acids in CSF samples.

3-Hydroxybutyric Acid↗
PubMed2006

Capillary electrophoresis-mass spectrometry of proteins at medium pH using bilayer-coated capillaries.

The feasibility of using noncovalently bilayer-coated capillaries for capillary electrophoresis-mass spectrometry (CE-MS) of acidic proteins was investigated using background electrolytes (BGEs) of medium pH. The capillary was coated by successively rinsing the capillary with solutions of the oppositely charged polymers polybrene (PB) and poly(vinyl sulfonic acid) (PVS). Volatile BGEs containing ammonium formate and/or N-methyl morpholine were tested at pH 7.5 and 8.5. Overall, these BGEs provided relatively fast protein separations (analysis times of ca. 12 min) and showed high efficiencies (70,000-300,000 plates) when the ionic strength was sufficiently high. Migration-time reproducibilities were very favorable with RSDs of less than 1.0%. Infusion experiments showed satisfactory MS responses for studied proteins dissolved in ammonium formate (pH 8.5), however, high concentrations of N-methyl morpholine appeared to seriously suppress the MS protein signals. Evaluation of the CE-MS system was performed by analyzing a mixture of intact proteins yielding efficient separations and good-quality mass spectra. CE-MS analysis of a reconstituted formulation of the biopharmaceutical recombinant human growth hormone (rhGH) which was stored for a prolonged time, revealed one degradation product which was provisionally identified as desamido rhGH. Based on the MS responses the amount of degradation was estimated to be ca. 25%.

Animals↗
PubMed2006

Coupling of sequential injection analysis and capillary electrophoresis - Laser-induced fluorescence via a valve interface for on-line derivatization and analysis of amino acids and peptides.

The on-line coupling of sequential injection analysis (SIA) and capillary electrophoresis (CE) via an in-line injection valve is presented. The SIA system is used for automated derivatization of amino acids and peptides. Dichlorotriazinylaminofluorescein serves as the derivatization agent, thus enabling sensitive laser-induced fluorescence detection of the derivatized analytes. The SIA procedure includes the following steps: (a) introduction of reagent and sample zones in a holding coil, (b) sample and reagent mixing in a reaction coil, (c) stop-flow step for increase of the reaction time, and (d) delivery of derivatized sample into the loop of the micro-valve interface. A small portion of the analyte zone is introduced electrokinetically in the separation capillary via the valve interface and CE analysis is performed. Factors affecting the CE separation, such as pH, the borate and sodium dodecyl sulphate concentration of the background electrolyte have been optimized. The derivatization conditions have been studied to obtain a high reaction yield in a relative short time. The transfer of a part of the reaction plug into the loop of the valve interface has been optimized. Using des-Tyr(1)-[Met]-enkephalinamide as test compound, it is demonstrated that after automated derivatization, on-line electrophoretic analysis could be achieved. Glycine has been selected as the internal standard in order to correct for variations in reaction time and filling of the injection loop. For the enkephalin, good reproducibility (RSD<4.5% calculated by the ratio of the peak areas) and linearity (0.5-5 microg mL(-1), R(2)>or=0.994) are obtained with a detection limit of 30 ng mL(-1) (S/N=3).

Amino Acids↗
PubMed2006

Micellar electrokinetic chromatography-electrospray ionization mass spectrometry for the identification of drug impurities.

Previously, we have presented a system hyphenating continuous micellar electrokinetic chromatography (MEKC) with electrospray ionization mass spectrometry (ESI-MS). Here we evaluate this technique for its applicability in impurity profiling of drugs using galantamine and ipratropium as test samples. A background electrolyte (BGE) of 10mM sodium phosphate (pH 7.5), 12.5-15% acetonitrile and 20mM sodium dodecylsulfate (SDS) was used for the MEKC-MS analysis of a galantamine sample containing a number of related impurities, and a heat-treated solution of ipratropium containing a number of unknown degradation products. MEKC provided efficient separation of all sample constituents. Despite the presence of non-volatile BGEs, all impurities in the galantamine sample could be detected by ESI-MS in their respective extracted ion traces (XICs) with a detection sensitivity in the sub-microg/ml range (full-scan mode). MS/MS detection provided useful product spectra allowing the structural characterization of the respective galantamine impurities. With the MEKC-MS/MS system, two degradation products could be revealed and identified in the heat-stressed ipratropium sample. The presented method shows good potential for the detection and structure elucidation of minor impurities in drug substances.

Chromatography, Micellar Electrokinetic Capillary↗
PubMed2006

Fast LC separation of a myoglobin digest: a case study using monolithic and particulate RP 18 silica capillary columns.

A method was developed for the fast separation of a myoglobin digest using a monolithic RP 18 silica capillary column of 100 microm I.D. The results were compared with those obtained with a particulate RP 18 silica capillary column of 100 microm I.D. at a flow-rate between 0.6 and 1.2 microl/min. The digest was analyzed at the monolithic column at a flow-rate up to 2.8 microl/min. This high flow-rate could not be applied to the particulate column due to the high back-pressure. When the starting composition of the gradient was changed from 0 to 20% and a gradient steepness of 16%/min was used, the analysis time was less than 4 min. A positive Mascot identification score of 115 was achieved for the MS-MS data. When a lower gradient steepness was employed, the chromatographic resolution and the peak capacity did not increase for most compounds. The intraday repeatability for the retention time of the monolithic column was better than 1.5% at 2.8 microl/min and even less than 0.5% using a flow-rate of 0.6 or 1.0 microl/min. For the particulate column, it was between 0.5 and 1.4% for a flow-rate of 0.6 microl/min, probably due to the high column back-pressure. The interday reproducibility for the retention time of the monolithic column was less than 0.9% using a flow-rate of 1.0 microl/min.

Animals↗
PubMed2006

On-line coupling of size exclusion chromatography and capillary electrophoresis via solid-phase extraction and a Tee-split interface.

An on-line size exclusion chromatography (SEC)-solid-phase extraction (SPE)-capillary electrophoresis (CE) system using a Tee-split interface has been developed for the analysis of peptides in biological fluids. The SEC column fractionates the sample by molecular size and the low-molecular-weight fraction, which contains the peptides, is directed to a C(18) SPE microcolumn, where the peptides are trapped and concentrated. The SPE column is desorbed with 425 nL acetonitrile and the effluent is sent to the Tee-split interface, which hydrodynamically splits (1:40) the flow and, thus, allows appropriate injection of analytes into the CE system. The performance of the system is investigated by the analysis of enkephalins in cerebrospinal fluid (CSF). It is demonstrated that the SEC step efficiently removes potentially interfering proteins, permitting reproducible SPE and CE. The total system provides efficient separations of the enkephalins with plate numbers up to 100,000. Concentration limits of detection (S/N = 3) for the peptides are about 100 ng/mL for injection of 20 microL spiked CSF samples. Plots of enkephalin peak areas versus concentration showed good linearity over the 0.25-10 microg/mL range (R2 > or = 0.985). Repeatability of migration time and peak area was within 2% and 10% R.S.D., respectively.

Chromatography, Gel↗
PubMed2006

Interlaboratory study of a NACE method for the determination of R-timolol content in S-timolol maleate: assessment of uncertainty.

Analyses of statistical variance were applied to evaluate the precision and practicality of a CD-based NACE assay for R-timolol after enantiomeric separation of R- and S-timolol. Data were collected in an interlaboratory study by 11 participating laboratories located in Europe and North America. General qualitative method performance was examined using suitability descriptors (i.e. resolution, selectivity, migration times and S/N), while precision was determined by quantification of variances in the determination of R-timolol at four different impurity levels in S-timolol maleate samples. The interlaboratory trials were designed in accordance with the ISO guideline 5725-2. This allowed estimating for each sample, the different variances, i.e. between-laboratory (s2(Laboratories)), between-day (s2(Days)) and between-replicate (s2(Replicates)). The variances of repeatability (s2r) and reproducibility (s2R) were then calculated. The estimated uncertainty, derived from the precision estimates, seems to be concentration-dependent above a given threshold. This example of R-timolol illustrates how a laboratory can evaluate uncertainty in general.

Adrenergic beta-Antagonists↗
PubMed2006

Efficient and highly reproducible capillary electrophoresis-mass spectrometry of peptides using Polybrene-poly(vinyl sulfonate)-coated capillaries.

The potential of capillaries noncovalently coated with a bilayer of oppositely charged polymers for the analysis of peptides by CE-MS was investigated. Bilayer coatings were produced by subsequently rinsing fused-silica capillaries with a solution of Polybrene (PB) and poly(vinyl sulfonate) (PVS). The PB-PVS coating showed to be fully compatible with MS detection causing no ionization suppression or background signals. The bilayer coating provided a considerable EOF at low pH, thereby facilitating the fast separation of peptides using a BGE of formic acid (pH 2.5). Under optimized CE-MS conditions, for enkephalin peptides high separation efficiencies were obtained with plate numbers in the range of 300,000-500,000. It is demonstrated that both the cancellation of the hydrodynamic capillary flow induced by the nebulizer gas and a sufficiently high-data acquisition rate are crucial for achieving these efficiencies. The overall performance of the CE-MS system using PB-PVS-coated capillaries was evaluated by the analysis of a tryptic digest of cytochrome c. The system provided an efficient separation of the peptide mixture, which could be effectively monitored by MS/MS detection allowing identification of at least 13 peptides within a time interval of 1.5 min. In addition, the PB-PVS coating proved to be very consistent yielding stable CE-MS patterns with highly favorable migration time reproducibilities (RSDs < 1% over a 3-day period).

Electrophoresis, Capillary↗
PubMed2005

On-line coupling of cyclodextrin mediated nonaqueous capillary electrophoresis to mass spectrometry for the determination of salbutamol enantiomers in urine.

The usefulness of the on-line coupling of nonaqueous capillary electrophoresis (NACE) with electrospray ionization (ESI) mass spectrometry (MS) using heptakis(2,3-di-O-acetyl-6-O-sulfo)-beta-cyclodextrin (HDAS-beta-CD) was demonstrated for the enantioselective determination of low concentrations of salbutamol in human urine. After optimization of several parameters, such as sheath-liquid composition and flow rate, nebulizing gas pressure, CE counter-pressure and position of the CE capillary outlet, a limit of quantification of 18 and 20 ng/ml was obtained for salbutamol enantiomers. Moreover, the relative standard deviation values for repeatability at a concentration of 30 ng/ml were below 7% for both enantiomers. Typical regression lines obtained after application of a simple linear regression model revealed a good relationship between peak area and analyte concentration (with 0.9988 and 0.9966 as coefficients of determination). This paper proposes an easy to use and sensitive NACE-MS method to determine enantiomers of a basic chiral drug in biological fluids preceded by solid-phase extraction as sample cleanup.

Adrenergic beta-Agonists↗
PubMed2005

Atmospheric pressure photoionization for enhanced compatibility in on-line micellar electrokinetic chromatography-mass spectrometry.

Atmospheric pressure photoionization (APPI) is presented as a novel means for the combination of micellar electrokinetic chromatography (MEKC) and mass spectrometry (MS). The on-line coupling is achieved using an adapted sheath flow interface installed on an orthogonal APPI source. Acetone or toluene is added as dopant to the sheath liquid to enhance analyte photoionization. It is demonstrated that with APPI signal suppression and interferences by the surfactant sodium dodecyl sulfate (SDS) and nonvolatile buffers can be circumvented. This implies that MEKC conditions can be selected independently from MS detection. Moreover, it is shown that both polar and apolar compounds can be photoionized, thereby also facilitating the analysis of compounds that are not amenable to electrospray ionization. Consequently, the MEKC-APPI-MS system can provide effective separation and detection of compounds of diverse character in one run using background electrolytes containing up to 50 mM SDS. Concentration limits of detection derived from extracted-ion traces (full scan mode) of test compounds were approximately 1 microg/mL, and the detection sensitivity remained unaffected during 1 day of continuous use. Overall, the system features are very favorable for applications such as drug impurity profiling as is illustrated by the analysis of mebeverine and related compounds (both charged and neutral) at the 0.25% (w/w) level.

Atmospheric Pressure↗
PubMed2005

Noncovalently bilayer-coated capillaries for efficient and reproducible analysis of proteins by capillary electrophoresis.

The suitability of noncovalently bilayer-coated capillaries for the analysis of proteins by capillary electrophoresis (CE) at medium pH was investigated. Fused-silica capillaries were coated simply by successively flushing with a polybrene (PB) and a poly(vinyl sulfonate) (PVS) solution. A protein test mixture was used to evaluate the performance of the coated capillaries. Comparisons with bare fused-silica capillaries were made. Several background electrolytes (BGEs) were tested in combination with the PB-PVS coating, showing that optimum performance was obtained for the proteins using high BGE concentrations. With a 300 mM Tris phosphate buffer (pH 7.0), good plate numbers (150,000-300,000), symmetrical peaks, and favorable migration-time repeatabilities (RSDs below 0.8%) were obtained for the proteins. Using bare fused-silica capillaries, the protein peaks were significantly broadened and the migration-time RSDs often exceeded 5%. It is concluded that the PB-PVS coating effectively minimizes adverse protein adsorption and provides a very stable electroosmotic flow (EOF). We also investigated the potential of a commercially available bilayer coating (CEofix) for protein analysis. It is demonstrated that with this coating, good plate numbers and peak symmetries for proteins can be achieved when the CEofix BGE ("accelerator") is replaced by a common BGE such as sodium or Tris phosphate. Apparently, the negatively charged polymer present in the "accelerator" interacts with the proteins causing band broadening. The utility of the bilayer coatings is further illustrated by the separation of proteins such as interferon-alpha 2b, myoglobin and carbonic anhydrase, by the analysis of a degraded insulin sample in time, and by the profiling of the glycoprotein ovalbumin. In addition, it is demonstrated that even in the presence of concentrations of human serum albumin in the sample of up to 60 mg/mL, the PB-PVS coating still provides reproducible protein separations of good performance.

Coated Materials, Biocompatible↗
PubMed2005

On-line multidimensional liquid chromatography and capillary electrophoresis systems for peptides and proteins.

Peptides and proteins are gaining increasing attention in biosciences and, consequently, in analysis. This overview highlights the different approaches to couple on-line various separation techniques for the determination of proteins and peptides. The first section discusses the liquid chromatography (LC)-LC coupling, the second one reviews the on-line LC-capillary electrophoresis (CE) coupled systems and the third section summarizes the strategies for on-line CE-CE. The advantages, disadvantages, most relevant difficulties and particular systems for on-line coupling are discussed. Special attention is paid to the interface between the two dimensions. Applications are summarized in tables and a few typical examples are discussed. Many multidimensional separation methods are available, and it is demonstrated that peptide and protein mapping, or quantitation of proteins or peptides in various samples (aqueous solutions, cells, plasma) require different coupled systems. For mapping a semi-quantitative detection is often sufficient, while comprehensiveness is very important. For quantitation of a certain peptide or protein at a low concentration level a validated method should be used, while a heart-cut transport of the first dimension to the second one can offer sufficient selectivity. The combination with mass spectrometry as part of the total system is stressed and illustrated.

Chromatography, Liquid↗
PubMed2005

On-line capillary electrophoresis-mass spectrometry using dopant-assisted atmospheric pressure photoionization: setup and system performance.

The on-line coupling of capillary electrophoresis (CE) and mass spectrometry (MS) via atmospheric pressure photoionization (APPI) is demonstrated. To achieve CE-APPI-MS, an adapted coaxial sheath-flow interface was combined with an ion-trap mass spectrometer equipped with an APPI source originally designed for liquid chromatography-MS. Effective photoionization of test compounds was accomplished after optimization of several interface and MS parameters, and of the composition and flow rate of the sheath liquid. Further enhancement of the ionization efficiency could be achieved by adding a dopant, such as acetone or toluene, to the sheath liquid to aid indirect ionization. Acetone significantly increased the ionization of the polar test compounds by proton transfer, while toluene was more useful for the enhanced formation of molecular ions from nonpolar compounds. The effect of several common CE background electrolytes (BGEs) on the APPI-MS response of the analytes was also studied. It appeared that in contrast with electrospray ionization, nonvolatile BGEs do not cause suppression of analyte signals using APPI. Therefore, in CE-APPI-MS, a variety of buffers can be chosen, which obviously is a great advantage during method development. Remarkably, also sodium dodecyl sulfate (SDS) did not affect the photoionization of the test compounds, indicating a strong potential of APPI for the on-line coupling of micellar electrokinetic chromatography (MEKC) and MS.

Atmospheric Pressure↗
PubMed2005

Evaluation of the sensitivity of miniaturized liquid chromatography-electrospray ionization-mass spectrometry for pharmaceutical analysis.

LC-ESI-MS is applied frequently in pharmaceutical analysis. The sample amount is generally not restricted, however with LC-ESI-MS, a lack of sensitivity may still be observed with standard-bore LC columns in isocratic mode. Therefore, it was investigated whether increased sensitivity could be achieved by using miniaturized LC-ESI-MS. Seven columns ranging from 0.1 to 4.6 mm ID were tested using several instrument setups. For proper comparison, a sensitivity gain factor (SGF) was introduced. The SGF expresses the extra sensitivity that may be obtained on top of the normal increase of peak concentration, which can be expected when the column ID is reduced. Desogestrel, mirtazapine, and sugammadex sodium were used as test compounds. For desogestrel and sugammadex sodium, the SGF increased up to a factor of 5-13 when the column ID was reduced, indicating enhanced ionization efficiencies at lower flow rates. Optimum sensitivity was found for the 0.3 mm column coupled in combination with a microinjection valve and a dedicated low flow rate interface. For mirtazapine, no increase of SGF was observed when the column ID was decreased. Apparently, the ionization efficiency of this compound is not affected by the flow rate and the spray quality.

Chromatography, High Pressure Liquid↗
PubMed2004

Chromatographic preconcentration coupled on-line to capillary electrophoresis via a tee-split interface.

Solid-phase extraction (SPE) and capillary electrophoresis (CE) are on-line coupled via a Tee-split interface, which provides hydrodynamic injection of the SPE eluate by flow splitting. The interface allows sample preconcentration independently from the CE separation and prevents sample matrix and washing solvents from entering the separation capillary. The effect of the Tee-split interface on the CE efficiency was examined using enkephalin peptides as model compounds. Most favorable plate numbers were obtained using a split ratio of 1:40. Breakthrough volume, desorption efficiency and elution volume for the C18 micro SPE column (5 mm x 0.5 mm i.d.) were found to be 750 microL, 65% and 1 microL, respectively. The performance of the complete system was demonstrated by the preconcentration and separation of an enkephalin mixture. Plate numbers up to 120,000 were obtained using a sample volume of 250 microL and a split ratio of 1:40. Enkephalin peak areas were linear (R2 = 0.996) over the 10-1000 ng/mL range. UV absorbance concentration limits of detection (SIN = 3) were about 5 ng/mL. For 250 microL injections of 100 ng/mL, the relative standard deviation (n = 5) of peak area was lower than 10%.

Chromatography, Liquid↗
PubMed2004

Chromatographic preconcentration coupled to capillary electrophoresis via an in-line injection valve.

A preconcentration-capillary electrophoresis (CE) system using a small precolumn in combination with an in-line injection valve is presented. The advantage of the present design is the ability to perform the sample preconcentration fully independently from the CE separation and to prevent sample matrix and washing solvents from entering the CE capillary. With a micro injection valve, sample could be effectively introduced into the CE system in an in-line fashion without seriously affecting the CE separation efficiency. Breakthrough volume, desorption efficiency, and elution volume for the C18 microcolumn (5 x 0.5 mm i.d.) were established, yielding values of 750 microL, 70%, and 0.9-1.1 microL, respectively, using enkephalin peptides. The time between the start of the desorption of the analytes from the precolumn and the injection into the CE system was also studied in order to achieve optimal sensitivity and separation efficiency. The performance of the complete system was demonstrated by the preconcentration and separation of an enkephalin mixture. Using a sample volume of 250 microL and a CE injection voltage of -15 kV for 12 s, linearity was observed over 2 orders of magnitude, and detection limits (S/N = 3) were in the 5-10 ng/mL range. A 1000-fold sensitivity enhancement is obtained using this setup, as compared to a regular CE setup. For 100 ng/mL samples, repeatabilities (RSDs) of migration time and peak area were 1.2 and 11%, respectively.

Chromatography↗