Lars B. Nilsson

Simultaneous determination of dopamine, DOPAC and homovanillic acid. Direct injection of supernatants from brain tissue homogenates in a liquid chromatography--electrochemical detection system.

A simple method based on high-performance liquid column chromatography with electro-chemical detection is described for the simultaneous determination of dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in discrete brain regions of rats. The supernatant of a tissue homogenate is injected directly onto a liquid chromatograph, thus omitting the commonly adopted adsorption step. Of the four different supports tested Nucleosil C15 (5 micron) was found superior with respect to chromatographic performance. The effects of pH, methanol and the ion-pairing agent hexyl sulfate on the retention were studied. The mobile phase used in the final studies consisted of citrate buffer pH 4.25-methanol (92:8, v/v) containing hexyl sulfate (1.7 . 10(-3) M). Standard curves of dopamine, DOPAC and HVA were found linear up to about 600 pmol per injection for each compound. The precisions of the chromatographic step were (Srel. %): 0.72% (dopamine), 1.26% (DOPAC) and 2.69% (HVA).

Straight-Phase Ion-Pair Chromatography of Zimelidine and Similar Divalent Amines Part I Bioanalysis

Abstract Methods are presented for the quantitative determination of ZIMELIDINE, a new antidepressant drug, and its active metabolite norZIMELIDINE in biological material (whole blood, plasma, urine and rat brain). The extraction is optimized regarding recoveries and blank chromatograms and the compounds are separated by high performance ion-pair liquid chromagraphy with perchlorate as counter ion in the stationary phase. Internal standards are chlorpheniramine and the geometrical isomer to norzimelidine. The precision for determinations in plasma ranges 2 - 7% (CV) for the concentrations 100 - 5 ng/ml, and the detection limits are 150 pg/ml but can be lowered about five times by using larger sample volumes. The selectivity against metabolites is investigated and the use of the method in routine is discussed. The isolation and identification of the primary amine metabolite by collecting the peak for subsequent GC-MS-analysis is demonstrated.

Determination of remoxipride in plasma and urine by reversed-phase column liquid chromatography

A reversed-phase column liquid chromatographic method for the determination of remoxipride, a novel antipsychotic drug, in biological fluids is described. A simple one-step extraction is used followed by liquid chromatography on a 3-microns octadecylsilica column and ultraviolet absorbance detection. The method is accurate and precise for clinical remoxipride levels in both plasma and urine. For situations where a higher sensitivity is necessary a two-step extraction and a modified mobile phase are used. With this modification plasma concentrations down to 2 nM can be determined with acceptable precision.

Optimization of a peak compression system for a remoxipride metabolite (FLA 797) and its application to bioanalysis

A peak compression system is optimized for FLA797 (I), a phenolic tertiary amine and a metabolite to the antipsychotic drug remoxipride. An application is described where this effect is used to give a 6-7-fold improvement of the quantification limit in an assay of I in plasma. The liquid chromatographic system is constructed so that the injection of I dissolved in a solution of a competing amine gives a very high and narrow analyte peak with an apparent efficiency of 1.5 x 10(6) plates/m. When the levels of I in plasma are determined, an internal standard, giving a normal isocratic peak, is added to the plasma sample before the extraction. Concentrations of I down to 0.5-1.0 nM can be determined with reasonable precision. FLA908, another phenolic remoxipride metabolite and a regioisomer to I, eluting as a normal isocratic peak, can be determined simultaneously although only at concentrations higher than 10-15 nM.

Straight-phase ion-pair chromatography of zimelidine and similar divalent amines : II. The chromatographic system

Abstract The properties of a straight-phase ion-pair liquid chromatographic system are described. The system is based on perchlorate as the anion component in a strongly acidic stationary phase with methylene chloride-n-butanol as the mobile phase, and has been used for the separation of zimelidine, a divalent hydrophobic amine, and related compounds. Batch distribution data for some of the amines as bases and as 1 + 1 and 1 + 2 ion pairs with perchlorate are presented and used to calculate capacity ratios, which were found to be in good agreement with the experimental chromatographic data. It is concluded that the retention mechanism is based mainly on liquid—liquid distribution and that selectivity factors can be calculated from batch extraction constants. The ion-pair equilibria were found to include an association of a 1 + 2 ion pair in the aqueous phase and also dissociation of the 1 + 1 ion pair in the organic phase. The relationship between chemical structure and selectivity is discussed, and it is emphasized that it is complicated because of the possible existence of two kinds of ion pairs with the divalent amines. The baseline separation of four compounds that are both geometric and bromo-positional isomers demonstrate the excellent selectivity of the system in practice. The capacity ratios increase both with increasing flow-rates and at very low flow-rates, but with maintained selectivities, and possible reasons are discussed. The effects of the injection of large sample volumes (up to 500 μl) on chromatographic efficiencies and resolutions are demonstrated, and linear relationships between the standard deviation (σ) of the dispersion and the injected volume were obtained.

Reversed-phase ion-pair liquid chromatographic method for the determination of low concentrations of haloperidol in plasma

A sensitive liquid chromatographic method for the determination of haloperidol in plasma is described. The efficient and simple extraction procedure, followed by reversed-phase ion-pair liquid chromatography on a 3-micron octadecylsilica column and UV absorbance detection, makes it possible to determine concentrations down to 0.5 nmol/l with acceptable precision. In a pharmacokinetic study, in which 5 mg of haloperidol were given orally, the plasma levels were followed for 48 h.

High sensitivity determination of the remoxipride hydroquinone metabolite NCQ-344 in plasma by coupled column reversed-phase liquid chromatography and electrochemical detection.

An HPLC method for the determination of NCQ-344, a remoxipride metabolite with a hydroquinone structure, in human plasma is described. Special precautions for the sampling were necessary as the compound rapidly decomposes. An efficient clean-up of the plasma samples was necessary to make use of the inherent sensitivity of the electrochemical detector. This was accomplished by a fast and simple liquid-liquid extraction at pH 7.05 combined with further cleaning on-line by using a short cyanopropyl column as the first column in a column switching system. A heart-cut from the cyanopropyl column containing the NCQ-344 fraction was then injected onto the analytical octadecyl silica column and NCQ-344 was detected at an oxidation potential of 0.70 V. The absolute recovery was > 95% and concentrations down to 0.10 nM could be determined with acceptable precision. The NCQ-344 levels in a limited number of samples from patients given remoxipride were found to be between 0.10 and 1 nM. The remoxipride concentrations in the same samples were 5,000-20,000 nM.

Regulation of peak compression effects for substituted benzamides in reversed-phase liquid chromatography

Abstract The possibilities of regulating peak compression effects, i.e. , of making the analyte peak coelute with the system peak, were investigated for substituted benzamides. The changes in capacity factor for the system peak relative to the retention of the analyte were studied by varying the composition of the mobile phase. The parameters useful for altering the capacity factor ratio were found to be the ionic strength of the phosphate buffer and to some extent the pH, whereas the amount of acetonitrile and the concentration of the amine modifier gave negligible effects. Variation of the amount of silanol groups on the solid phase could also be used as an efficient means for regulation of the capacity factor ratio.

Determination of dextropropoxyphene and norpropoxyphene in plasma by high-performance liquid chromatography

A simple and sensitive procedure for the routine assay of the analgesic drug dextropropoxyphene and its main metabolite, norpropoxyphene, in plasma is described. After liquid-liquid extraction from alkalinized plasma and back-extraction into a small volume of an acidic aqueous phase, the aqueous phase was injected into a column packed with 3-microns octadecylsilica particles. Ultraviolet absorbance detection at 210 nm was used. Concentrations down to 2 nM could be determined for both compounds; at this level, the intra-assay coefficient of variation was 5%.