On-line coupling of solid-phase extraction with mass spectrometry for the analysis of biological samples. I. Determination of clenbuterol in urine.

The potential of the direct coupling of solid-phase extraction (SPE) with mass spectrometry (MS) for the analysis of biological samples is demonstrated. For SPE a cartridge exchanger is used and the eluate is directly introduced into the mass spectrometer. This system has been investigated for the determination of clenbuterol in urine. With mixed-mode cartridges, a considerable ion suppression has been obtained. The mass spectrum at the elution time of clenbuterol is dominated by that of creatinine and adduct formation of clenbuterol and creatinine has been observed. The whole procedure including injection of 1 ml urine, washing and desorption has been developed with cartridges containing 8-microm C18-bonded silica. If only a single MS step is used, the selectivity and, therefore, the sensitivity are insufficient. The detection limit is about 100 ng/ml. However, with atmospheric pressure chemical ionisation and the tandem MS mode the detection limit has been decreased to about 2 ng/ml and the ion suppression is only about 10%. For the electrospray ionisation the detection limit is about 10-times higher and the ion suppression is less favourable. The repeatability for the SPE-MS-MS procedure was 6.5% at 10 ng/ml (n=5) and the difference between the response factors at 10 ng/ml and 100 ng/ml was only 2.5%. The MS behaviour of clenbuterol and the matrix under the present conditions is discussed.

Evaluation of the characteristics of the differential amperometric detector in combination with anion-exchange chromatography, using L-ascorbic acid as test compound.

The hydrodynamic characteristics of the thin-layer flow cell of the differential amperometric detector were deduced theoretically and confirmed by experiment. The response of the differential amperometric detector was compared with that of a fixed-wavelength UV detector in the analysis of L-ascorbic acid in combination with anion-exchange chromatography.

Optimization of the reversed-phase high-performance liquid chromatographic separation of synthetic estrogenic and progestogenic steroids using the multi-criteria decision making method.

The optimization of the reversed-phase high-performance liquid chromatographic separation of a mixture of ethynylestradiol, desogestrel and three related compounds is described. A procedure is used that allows the prediction of the capacity factors of each individual synthetic steroid, depending on the mobile phase composition. Therefore, complete chromatograms can be predicted and evaluated with the multi-criteria decision making method.

On-fiber derivatization for direct immersion solid-phase microextraction. Part II. Acylation of amphetamine with pentafluorobenzoyl chloride for urine analysis.

On-fiber derivatization was used for solid-phase microextraction (SPME) in order to increase the detectability and extractability of drugs in biological samples. Amphetamine, which was used as a model compound, was derivatized with pentafluorobenzoyl chloride (PFBCl) and subjected to gas chromatography with electron capture or mass spectrometric detection. Extraction was performed by direct immersion of a 100 microm polydimethylsiloxane-coated fiber into buffered human urine. On-fiber derivatization was performed either after or simultaneously with extraction. The former procedure gave cleaner chromatograms but the latter turned out to be superior with respect to linearity and repeatability. For the on-fiber derivatization of amphetamine an excess of reagent is required. Because a considerable part of the PFBCl loaded on to the fiber is used up by reaction with matrix compounds and water, a reagent loading time of 5 min was needed to obtain a linear range (r = 0.9756) from 250 pg mL(-1) to 15 ng mL(-1). Due to an interfering matrix compound, the limit of detection was also found to be dependent on the reagent loading time, i.e., the limit of detection for a PFBCl loading time of 5 min is 250 pg mL(-1) whereas that for a 1 min loading time it is 100 pg mL(-1). The relative standard deviation (n = 7) of the method was about 11% at an amphetamine concentration of 1 ng mL(-1). The applicability of the method for the determination of drugs in biological samples is shown.