R.A.M. van der Hoeven

Liquid chromatography-mass spectrometry with on-line solid-phase extraction by a restricted-access C18 precolumn for direct plasma and urine injection.

In this paper, the on-line coupling of solid-phase extraction, based on a restricted-access support with liquid chromatography-mass spectrometry (LC-MS), for the analysis of biological samples is described. The system was tested with cortisol and prednisolone for plasma analysis and arachidonic acid for urine analysis. A precolumn packed with a 25-micron C18 alkyl-diol support is used for direct plasma or urine injection. Using column-switching techniques, the analytes enriched on the precolumn are eluted to the analytical column without transfer loss. An on-line heart-cut technique was employed and only the analyte-containing fraction eluting from the LC column is directed to the MS to protect the LC-MS interface and ion-source from contamination. The whole system is operated in a parallel mode, that is, sample pre-treatment and LC-MS analysis are performed simultaneously to provide the shortest possible analysis time. The only off-line sample pre-treatment step required was centrifugation to remove particulate matter. With the fully automated system, total analysis times of 5 and 9.5 min were achieved for cortisol in serum and arachidonic acid in urine, respectively. Cortisol and related compounds were quantitatively recovered from plasma with a detection limit for prednisolone (direct injection of 100 microliters on restricted-access precolumn) of 2 ng/ml.

Microdialysis introduction high‐performance anion‐exchange chromatography/ionspray mass spectrometry for monitoring of on‐line desalted carbohydrate hydrolysates

A system for the determination of carbohydrates based on on-line microdialysis introduction high-performance anion-exchange chromatography integrated pulsed electrochemical detection/mass spectrometry (MS) using ionspray as the ionization technique, with on-line desalting, is presented. This coupling of techniques utilizes a cationexchange membrane desalting device (CEMDD) which permitted the exchange of sodium ions with hydronium ions when sodium ion concentrations up to 600 mM were used to separate and elute oligosaccharides. The e†ective on-line desalting by the CEMDD of the effluent before its introduction into the mass spectrometer was achieved by the electrolysis of water with a 500 mA current. The use of the CEMDD in combination with ionspray eliminated the need for a booster pump and also the regenerating water improved the sensitivity in comparison with the use of sulphuric acid as oligosaccharides could be detected down to 3 l gm lo1. The system was developed and used to monitor cationized oligosaccharides based on their singly [M + Na ]e and doubly sodiated [(M + 2Na)/2 ]2e molecules during the hydrolysis of wheat starch. The MS/MS properties of maltose, trehalose and sucrose were used in combination with retention times to identify these disaccharides based on their dissociation patterns, which are characteristic of the type of glycosidic linkage. 1998 by John Wiley & Sons, Ltd.

Coupled column chromatography—mass spectrometry : Thermospray liquid chromatography—Mass spectrometric and liquid chromatography—tandem mass spectrometric analysis of metoprolol enantiomers in plasma using phase-system switching

Abstract The applicability of phase-system switching using thermospray tandem mass spectrometry is demonstrated for the bioanalysis of the enantiomers of the betablocker metoprolol. Independent optimization of the chromatography, using an α1-acid glycoprotein chiral stationar phase, and the mass spectrometric detection system is realized. By utilizing peak compression, 10 ng (37 pmol) of each enantiomer is easily detected in standard solutions. In plasma samples, with use of tandem mass spectrometry for additional selectivity, levels of 61 ng/ml (230 nmol/l) can be measured by using [2H6]metoprolol as internal standard.

Some aspects of peak broadening in particle-beam liquid chromatography-mass spectrometry

Abstract The particle-beam interface has recently been introduced for coupling liquid chromatography and mass spectrometry. The coupling of a particle-beam interface to a Finnigan MAT TSQ-70 triple quadrupole instrument is described. A compound-dependent peak broadening in the interface has been observed. In this paper various experiments are described to investigate some of the sources of peak broadening. For this purpose, the transfer efficiency of the particle-beam interface has been measured, and the potential of volatility-enhancing derivatization procedures has been explored. The detection of 40 pg of the pentafluorobenzyl derivative of palmitic acid is demonstrated.

Characterization of sugar oligomers by on-line high-performance anion-exchange chromatography-thermospray mass spectrometry.

Abstract The on-line coupling of high-performance anion-exchange chromatography and thermospray mass spectrometry via an anion-micromembrane suppressor and a booster pump is described. The system was applied to the analysis of homologous series of oligosaccharides. Among others, the mass spectrometric detection of β-1,4-xylose oligomers up to degree of polymerization (DP) 25 is demonstrated. Further, the system was used in the analysis of more complex oligosaccharide samples, containing mixed oligomers of hexoses, pentoses and uronic acids. In such samples oligomers up to DP 10 can be tested. The potential use of this approach in the characterization of oligosaccharides obtained from (enzymic) degradation of plant cell wall oligosaccharides is discussed.

Development of optimization strategies in thermospray liquid chromatography—mass spectrometry

Abstract In order to optimize the sensitivity and reproducibility of thermospray liquid chromatography—mass spectrometry in (bio)analytical applications, some of the experimental parameters that influence thermospray buffer ionization have been investigated systematically. Attention was paid to the vaporizer temperature, which is especially important in the analysis of thermolabile compounds, the ammonium acetate concentration, the methanol content and the repeller potential. General optimization strategies for theirmospray buffer ionization have been developed. The usefulness of extensive optimization is discussed for qualitative and quantitative analysis. In quantitative target compound analysis optimization on the analyte is necessary. In qualitative analysis, however, usually unknown compounds have to be analysed and no parent compound is available for optimization purposes.

Electrospray mass spectrometry of neutral and acidic oligosaccharides: methylated cyclodextrins and identification of unknowns derived from fruit material.

Abstract A qualitative study of the characteristics in the mass spectrometric analysis of various neutral and acidic oligosaccharides using electrospray ionization is reported. Experiments were performed in both positive- and negative-ion modes. In the positive-ion mode molecular mass information for neutral non-derivatized oligosaccharides could be obtained up to Mr 4000. The method was applied to the determination of the degree of methylation of methylated β-cyclodextrins and the identification of unknown oligosaccharides enzymatically derived from pear and apple fruit material.

Automated analysis of mitomycin C in body fluids by high-performance liquid chromatography with on-line sample pre-treatment

A fully automated liquid chromatographic system for the bioanalysis of mitomycin C has been described. The isolation of the analyte from the biological matrix (plasma, ascites and urine) is performed using a continuous-flow system equipped with a dialysis membrane in order to remove proteins. The samples are concentrated on a reversed-phase pre-column and subsequently introduced on to a reversed-phase analytical column by applying column-switching techniques. The drug is detected by absorbance measurements at 360 nm. Using the described system up to 100 samples a day can be analysed with determination limits of the order of 1 ng/ml, with a linear dynamic range of at least three decades for plasma and urine samples. The procedure was applied to pharmacokinetic studies of ovarian cancer patients treated intraperitoneally with mitomycin C.

Recent progress in high-performance anion-exchange chromatography—thermospray mass spectrometry of oligosaccharides

Abstract The on-line combination of high-performance anion-exchange chromatography and mass spectrometry via a thermospray interface has proved to be a powerful tool in the characterization of sugar oligomers obtained by enzymatic digestion of plant cell wall polysaccharides. The potential of the method can be improved by the use of a new column material, CarboPac PA100, which requires lower sodium acetate concentrations for the elution of large sugar oligomers. Further, the application of multiple-ion detection optimizes the information obtained from the analysis by improving both the signal-to-noise ratio and the conservation of the chromatographic resolution. Negative-ion instead of positive-ion detection results in significantly better signals, especially for the larger sugar oligomers.

Application of microdialysis for on-line coupling of capillary isoelectric focusing with electrospray mass spectrometry on a magnetic sector instrument

The on-line coupling of capillary isoelectric focusing (CIEF) with electrospray mass spectrometry (MS) by means of microdialysis (MD) for application to three carbonic anhydrase isozymes is described. The dialysis device consisted of a donor and an acceptor compartment that were separated by a flat membrane, and served to clear the CIEF effluent that was directed to the mass spectrometer from ampholytes that were necessary for the establishment of a pH gradient in the IEF capillary. In addition to single-ion monitoring and full-scan experiments, a position- and time-resolved ion counting (PATRIC) detector was used in the static array mode. For CIEF/MD/MS of unknown proteins the minimum amount of analyte required for the analysis was estimated to be in the mid-femtomole range using full-scan MS data acquisition.