Gas chromatographic determination of the interconversion energy barrier for dialkyl 2,3-pentadienedioate enantiomers.

The enantiomers of dialkyl 2,3-pentadienedioate undergo interconversion during gas chromatographic separation on chiral stationary phases. In this paper the on-column apparent interconversion kinetic and thermodynamic activation data were determined for dimethyl, diethyl, propylbutyl and dibutyl 2,3-pentadienedioate enantiomers by gas chromatographic separation of the racemic mixtures on a capillary column containing a polydimethylsiloxane stationary phase coupled to 2,3-di-O-methyl-6-O-tertbutyldimethylsilyl-beta-cyclodextrin. A deconvolution method was used to determine the individual enantiomer peak areas and retention times that are needed to calculate the interconversion rate constants and the energy barriers. The apparent rate constants and interconversion energy barriers decrease slightly with an increase in the alkyl chain length of the dialkyl 2,3-pentadienedioate esters. The optimum conformation of the dialkyl 2,3-pentadienedioate molecules, their separation selectivity factors and apparent interconversion enthalpy and entropy data changes with the alkyl chain length. The dependence of the apparent interconversion energy barrier (deltaG(app)(a-->b), deltaG(app)(b-->a)) on temperature was used to determine the apparent activation enthalpy (deltaH(app)(a-->b), deltaH(app)(b-->a)) and apparent entropy (deltaS(app)(a-->b), deltaS(app)(a-->b)) (where a denotes the first and b second eluted enantiomer). The comparison of the activation enthalpy and entropy (deltaS(app)(a-->b), deltaS(app)(a-->b)) indicated that the interconversion of dialkyl 2,3-pentadienedioate enantiomers on the HP-5+Chiraldex B-DM column series is an entropy driven process at 160 degrees C. Data obtained for dimethyl 2,3-pentadienedioate enantiomers on the HP-5+Chiraldex B-DM column series at 120 degrees C (deltaG(app)(a-->b) = 123.3 and deltaG(app)(b-->a) = 124.4 kJ mol(-1)) corresponds (at the 95% confidence interval) with the value of deltaG(#) = 128+/-1 kJ mol(-1) found at this temperature by gas chromatography using a two-dimensional stop flow technique on an empty capillary column [V. Schurig, F. Keller, S. Reich, M. Fluck, Tetrahedron: Asymmetry 8 (1997) 3475].

Determination of fendiline and gallopamil by capillary isotachophoresis.

Capillary isotachophoresis was applied for the determination of fendiline and gallopamil--calcium antagonists--in serum. The cationic electrolyte system containing Na+ with acetic acid as a counter constituent was used as a leading electrolyte with the pH 4.7 and the terminating electrolyte was beta-alanine. Most of the proteins were precipitated with methanol, ethanol and dimethylketone. The lowest limits of quantitation were obtained for the pretreatment of serum with methanol. The recoveries of both compounds varied from 91.3 to 97.5%. The relative standard deviations varied from 0.6 to 7.7%.

Isotachophoretic determination of naproxen in the presence of its metabolite in human serum.

An isotachophoretic method with conductivity detection was developed to determine naproxen in the presence of its metabolite 6-O-desmethylnaproxen in human serum. The leading electrolyte contained 10 mM hydrochloric acid, beta-alanine, pH 4.0 and 0.1% methylhydroxypropylcellulose. The terminating electrolyte was 10 mM 2-(N-morpholino)ethanesulfonic acid-tris(hydroxymethyl)aminomethane, pH 6.9, containing 20% (v/v) of ethanol. Naproxen was determined in serum supernatant after simple deproteination of the sample with ethanol. The isotachophoretic results were compared with those obtained by synchronous fluorescence spectrometry.

Determination of ibuprofen and naproxen in tablets.

Ibuprofen and naproxen have been quantified in tablets by capillary isotachophoresis. Hydrochloric acid (10 mmol/l) adjusted with creatinine to pH 5.0 plus 0.1% polyvinylpyrrolidone was used as the leading electrolyte and 10 mmol/l 4-morpholineethanesulfonic acid as the terminating electrolyte. Linearity was observed from 40.0 to 200.0 mg/l of ibuprofen (naproxen), with a coefficient of determination (r2) of 0.999. Good quantitation was obtained in short analysis time. The isotachophoretic results were compared with those obtained by the fluorescence spectrometry. Experimental parameters for ibuprofen were: lambdaEX=224 nm and lambdaEM=290 nm. Experimental parameters for naproxen were: lambdaEX=230 nm and lambdaEM=355 nm. The calibration plot was found to be linear in the range 0.4-2.4 mg/l for ibuprofen and 5.0-20.0 microg/l for naproxen. The minimal sample pretreatment and relatively low running cost make isotachophoresis a good alternative to existing methods.

On the calculation of Gibbs energy corresponding to enantioselective interactions at a direct HRGC separation of enantiomers.

A novel procedure is proposed for the calculation of Gibbs energy corresponding to enantiospecific interactions of 2-(2, 4-dinitrophenoxy)-, 2-phenoxy-, and 2-halogen-n-pentane enantiomers with a beta-cyclodextrin (ChirasilDex) stationary phase under gas chromatographic conditions. This energy is calculated from retention data as a difference between the Gibbs energy of an enantiomer and its corresponding achiral congener. The procedure for the determination of 2-(2,4-dinitrophenoxy)-, 2-phenoxy- and 2-halogen- n-pentane achiral congener retention data is discussed in detail.

SPE-HPLC determination of catecholamines using an affinity principle.

Solid-phase extraction (SPE) with the affinity chromatography principle was applied for high performance liquid chromatography (HPLC) determination of catecholamines in patients urine samples. Electrochemical amperometric detection (ED) was used for all HPLC analyses and both analytical and microbore columns were tested for optimal chromatographic resolution of all analyzed catecholamines. Capacity ratio k' and detection limits were evaluated for all columns used. Precolumn affinity chromatographic procedure of catecholamines with diphenylboronic acid (DPBA) in alkaline medium improved their retention on different commercial SPE reversed-phase cartridges. After clean-up and preconcentration step the complex catecholamine-diphenylboronic acid was degraded by acidic pH and eluted from SPE cartridge. After SPE affinity step catecholamines were analyzed by HPLC-ED. The optimal elution solutions was chosen also as suitable SPE cartridges. Extraction recoveries of catecholamines were 89.6-93.3% with relative standard deviation RSD = 3.8-4.3%. Total analysis time was 30 min including 15 min for the preseparation procedure.

Quantitative assay of verapamil in drugs and serum by capillary isotachophoresis.

Experimental conditions for a simple capillary isotachophoresis (ITP) with conductometric detection have been created for the determination of verapamil in drugs and serum. The limit of detection (LD) was 80 ng.cm-3 for standard verapamil solution. Under optimal experimental conditions and instrumental comparison, the LD of verapamil in serum was 420 ng.cm-3 without sample pretreatment. To lower the LD of verapamil in serum a liquid-liquid extraction was used. The high recovery (more than 94%) facilitated the verapamil ITP analysis at the therapeutic concentration.

HPLC-ED determination of catecholamines and their metabolites in urine.

High performance liquid chromatography (HPLC) with electrochemical detection (ED) was applied for the monitoring of catecholamines (epinephrine - E, norepinephrine - NE, dopamine - DA) and their main metabolites (homovanillic acid - HVA, vanillylmandelic acid - VMA) in urine samples of patients with the pheochromocytoma diagnosis. Both amperometric and colorimetric detectors were tested for the clinical applications and the results were compared. The optimization of separation conditions was worked out, especially the effects of mobile phase compositions (pH, ionic strength, organic modifier and ion-pair agent concentrations). Dependences of capacity ratio values k' on all optimized components of the mobile phases were evaluated. Chromatographic conditions were optimized for the suitable chromatographic resolution (Rij > 1.25). Extraction recoveries for liquid-liquid and solid-phase extractions have been worked-out. Detection limits for catecholamines in urine were in the range of 0.3-0.6 ng/ml and 10-20 ng/ml for HVA, VMA using coulometric detector.