Milos Novotný

Laser fluorimetry for capillary column liquid chromatography: high-sensitivity detection of derivatized biological compounds

A high-sensitivity, low-dead-volume laser-induced fluorescence detector has been designed for use in capillary column liquid chromatography. This detector exhibits femtogram detection limits and a linear dynamic range spanning five orders of magnitude. Derivatization schemes were utilized to add a fluorescent moiety to biologically important compounds such that their excitation maxima corresponded well with the output wavelength of the laser. Sample chromatograms are presented for picogram amounts of standard bile acid and steroid hormone derivatives. In addition, a high-efficiency separation of picogram amounts of the solvolyzable steroid hormones from human serum is illustrated.

Fractionation and capillary gas chromatographic—mass spectrometric characterization of the neutral components in marijuana and tobacco smoke condensates

Abstract Polar and non-polar “neutral” constituents of the smoke condensates of Mexican marijuana and standard tobacco were resolved by capillary gas chromatography and structurally characterized through mass spectrometry. Comparisons of the two materials reveal substantial qualitative and quantitative differences. The cannabinoid-like substances partition into both the non-polar and polar fraction. In total, over 130 “neutral” smoke components have been characterized.

Optimization and evaluation of packed capillary columns for high-performance liquid chromatography

Abstract Packed capillary columns were systematically optimized for adsorption high-performance liquid chromatography, and their performance was evaluated using conventional plate height vs . linear velocity curves and the Knox separation impedance. Column performance was improved by decreasing the inner diameter, nominal particle size, and mean pore size of the silica adsorbent. Irregularly shaped silica particles gave consistently higher efficiencies than spherical materials. The performance of packed capillary columns, as measured by the Knox separation impedance, is currently equal to or exceeds that of conventional or microbore packed columns.

Micro-column high-performance liquid chromatography and flame-based detection principles

Abstract Novel detectors are described for micro-column high-performance liquid chromatography; they are based on the well-known principles of flame photometric and thermionic detection. These devices exhibit good selectivity and sensitivity for organophosphorus compounds in both aqueous and selected organic solvents. The extremely small volume of these flame-based detectors and their enhanced response at low flow-rates make them particularly attractive for capillary-column high-performance liquid chromatography.

Small-volume electrochemical detector for microcolumn liquid chromatography

A submicroliter electrochemical detector for liquid chromatography has been designed, using pressure-annealed pyrolytic graphite technology. The analytical performance of this detector was studied in connection with a reversed-phase packed microcapillary column at very low flow-rates. Although the miniaturized version of the electrochemical detector is less sensitive, a direct analysis of a number of urinary metabolites in 0.1--1.0 microliters samples is feasible.

Application of pattern recognition and feature extraction techniques to volatile constituent metabolic profiles obtained by capillary gas chromatography.

The applicability of threshold logic units, a form of nonparametric pattern recognition, to the processing of metabolic profile data obtained by high-efficiency glass capillary column gas chromatography has been investigated. The test data included profiles of the volatile constituents of urine from normal individuals and from individuals with diabetes mellitus. A feature extraction algorithm allowed for dimensionality reduction and indicated the constituents most important in the normal versus pathological distinction. With an optimum number of dimensions, a normal versus pathological prediction rate of 93.75% was achieved. Gas chromatography-mass spectrometry was utilized to identify important profile constituents.

Automated high-resolution gas chromatographic system for recording and evaluation of metabolic profiles

A fully automated high-resolution gas chromatographic system is described that incorporates repetitive sampling of volatile biological samples onto a glass capillary column, and a simultaneous flame ionization and nitrogen-sensitive detection prior to data acquisition and computer handling. Reliable headspace sampling procedure and reproducible column characteristics that are essential to automation have been studied. The application of this system to the reproducible chromatographic recording of human urinary volatile constituents is shown.

Fundamentals of column performance in supercritical fluid chromatography

Abstract The principles governing chromatography with a supercritical mobile phase are examined in light of measurements made with open-tubular columns. Plate height vs. linear velocity curves obtained for uncoated columns having different radii confirm earlier theoretical predictions. These measurements and their implications for both practical separations and physico-chemical measurements in supercritical fluid chromatography are discussed. Consideration is also given to other parameters such as selection of phase systems, film thickness and operating pressure.

Light-scattering detection in liquid chromatography

Abstract A novel detection system for liquid chromatography is described which consists of post-column solute precipitation and light-scattering measurement of the formed precipitate. Its efficiency is demonstrated by the sensitive detection of non-polar lipids of biological origin separated by reversed-phase chromatography. A linear relationship between the square root of the detector response and sample concentration was found for several lipid solutes. Detection sensitivity below 10 −6 g has been achieved with the present system. Some response-affecting factors were also investigated.

Preparation and chromatographic evaluation of chemically bonded ion-exchange stationary phases : II. Weak and strong cation exchangers

Abstract Surface reactions of chlorodimethyl [4-(4-chloromethylphenyl)butyl] silane and chlorodimethyl (4-phenylbutyl) silane with small-particle silica, followed by simple chemical modifications, lead to the preparation of weak and strong cation-exchange packing materials for high-performance liquid chromatography. Retention of various solutes on such bonded phases was investigated under different conditions of pH, ionic strength, and mobile-phase composition. Results of these studies are compared with those previously obtained on classical cation-exchange materials under comparable operating conditions.