Wiktor Kemula

Alternating voltage polarographic detection for high-performance liquid chromatography and its evaluation for the analysis of bile acids

Abstract The first attempt to apply alternating voltage (a.v.) polarography for detection in high-performance liquid chromatography is presented. The performance of a flow-through polarographic detector in the a.v. polarographic mode is illustrated with the separation of a mixture of bile acids. Generally, the retention time of a substance is found to be proportional to its extent of adsorption at the electrode at the applied potential. Thus, the more the substance is retained, the easier is the detection of its peak. The observed noise level is close to 0.024 μF −1 . The linear dynamic range for cholic acid determination extends from 1.25 nmol to over 50 nmol with a correlation coefficient of 0.0998. The sensitivity is close to 8·10 4 sec 1 μF −1 mol −1 and the detector response index is 0.98. The standard deviation of the peak area from nine consecutive injections was 4.5%. The detection limit was close to 1 nmol per injection, almost one order of magnitude larger than that for a UV detector operated at 210 nm, but still one order of magnitude lower than that for a refractive index detector.

Separation mechanism and determination of acidic compounds by ion-exclusion liquid chromatography with electrokinetic detection☆

It was found that in ion-exclusion chromatography both strong acids (pKa 6.5) were eluted unseparated, the first at the beginning and the latter at the end of the elution. The retention volumes of the remaining acids (2.5 < pKa < 6.5) were found to be proportional to their pKa values. This kind of behaviour is described in terms of appropriate equations. The dead and inner volumes of the chromatographic column were determined from the observed dependence of retention volumes on the pKa values of the analysed acids. The monitoring of the acids was performed using electrokinetic detection. The detectabilities of the acids were found to be inversely proportional to their dissociation constants.

Polarographic detection for high-performance liquid chromatography using a flow-through detector

A new arrangement of a flow-through polarographic detector has been designed with a rapidly dropping mercury electrode. The detector is especially usefull for reversed-phase high-performance liquid chromatography (HPLC). It has been shown that the properties of the detector are similar to those of a UV detector operated at 254 nm. An electroactive substance in the eluate can be determined at flow-rates not exceeding 6 ml min−1. An upwards direction of flow of the solution facilitates the automatic removal of gas bubbles, which are swept to the detector with the column effluent. The volume of solution necessary to wash out 95% of the detection passage filled with another solution is about 0.1 ml, which is approximately equal to the volume when using a UV (254 nm) detector with a geometrical volume of the detection passage of 8 μl. The detector sensitivity with injection of cadmium nitrate solution is (3.6 ± 0.4)·10−4 A 1 mol−1, and the linear dynamic range is 5·10−4–5·10−2M cadmium nitrate with 5-μl injections. The selection of a suitably short drop time at a given flow-rate enables one to eliminate current signal disturbances on the recorded curves. Thus the current is diffusion or convective-diffusion controlled, irrespective of the presence or absence of turbulence. The application of the detector to HPLC is illustrated with the separation of a mixture of testosteroids.

Investigation of the electrochemical generation and the visible spectra of free radical anions of the mono-nitro-naphthalenes and nitromesitylene, in dimethylformamide

Abstract As a result of certain polarographic investigations, the free radical anions of α- and β-nitronaphthalenes and nitromesitylene were generated by electrolysis in a 0.2 N solution of NaNO 3 in dimethylformamide. The free radical anions of the mononitronaphthalenes were green in solution and those of nitromesitylene pinkish-red. The visible spectra of the free radical anions were determined, using their polarographic oxidation current as a measure of their concentration. The interpretation of the spectra of the free radical anions and nitrobenzene free radical anion, and their correlation with the u.v. spectra of the nitro-compounds has been discussed. It has been suggested that in the free radical anions of the aromatic nitro-compounds, the unpaired electron occupies a low-energy orbital of the nitro-group. The visible absorption bands of the free radical anions correspond to transitions of the unpaired electron from this orbital to higher molecular orbitals.

Microdetermination of chloride in water

Abstract An indirect colorimetric method for the determination of chloride has been developed. This method is based on the decrease in the absorption of the mercury-diphenylcarbazone complex caused by chloride ions. The influence of pH and of temperature on the determination have been thoroughly investigated and experimental conditions derived which make possible the determination of 10 to 60 μg of chloride in the presence of an excess of copper, nickel, cobalt, zinc, cadmium and lead, which are removed by chloroform extraction as diphenylcarbazonates.

Electrochemical behaviour of nitrosobenzene in dimethylformamide

Abstract Nitrosobenzene is reduced in a solution of 0.2 N NaNO3 in dimethytformamide in two polarographic waves. In the first reduction wave the free radical anions of nitrosobenzene form primarily. The free radical anions are unstable and react with formation of azoxybenzene as a final product. This was ascertained by means of controlled potential electrolysis at the potential of the first reduction wave of azoxybenzene. Nitrobenzene is formed during simultaneous electrochemical reduction of oxygen and nitrosobenzene in a solution of 0.2 N NaNO3 in dimethylformamide. This was ascertained by means of electrolysis and also by oscillopolarography.

Extra-column effects in polarographic versus UV detection in high-performance liquid chromatography

Abstract A comparison of extra-column effects of chromatographic peaks in high-performance liquid chromatography recorded using a flow-through polarographic detector and a typical UV (254 nm) detector is presented. The diameter of the detection channel of the polarographic detector was 2 mm and the volume of the detection channel of the UV detector was 9.2 μl. When extra-column effects due to injection, tubing and the electronic system were considered, it was found that within experimental error the values of the peak variances are comparable for UV and polarographic detection and equal to (2.7 ± 1.0)·10 3 μl 2 .

Chromato-polarographic analysis of mononitroethyl-benzene mixtures

Abstract A method of partition and determination of o-, m- and p-nitroethylbenzenes with clathrates as fillers of the chromatographic column is described. The clathrate used was of composition Ni(γ-picoline)4(SCN)2γ-picoline. The degree of filling of clatlirate used was 0.4–0.7. The aqueous moving phase contained 2 M NH4SCN, 0.3 M γ-picoline and 40–60 vol.% organic solvent. The applicability of the solvents: acetone, acetonitrile, ethanol, methanol, formaimide and dimethylformtimide, was checked; acetone and dimethylformamide were found best. Quantitative partition and evaluation of all three isomers was achieved on a 2.5-cm column. The recovery of the eluted individuals was complete. The samples used weighed only 0.05–0.3 mg. The relative error of evaluated components was not greater than ±5%. The clathrate filler can be used many times.

Electrokinetic Detection in Reversed Phase High Performance Liquid Chromatography Part I. Volatile Fatty Acids

Abstract Conditions of electrokinetic detection are elaborated for volatile fatty acids (acetic, propionic, isobutyric and valeric) in reversed phase high performance liquid chromatography, HPLC. A simple, open tube capillary electrokinetic detector was constructed. The working unit of the detector was a capillary made of polytetra-fluoroethylene, PTFE, or stainless-steel. The output signal of the detector was the streaming potential of the capillary which was measured against earth. When chro-matograms were developed in non-buffered polar solution of mobile phase, the retention volume V R, of acids increased with the increase of concentration of acids in the sample. The detectability of the detector with PTFE capillary used as a working unit was of the order of 10−12 mole for a 5 μl injected sample and the reproducibility was 5% (relative standard deviation, R. S. D., for ten consecutive injections). The linear dynamic range was close to two orders of magnitude of the concentrations of acids.

New model of the hanging mercury drop electrode and its application in aqueous and non-aqueous media

ZusammenfassungEine neu konstruierte Quecksilbertropfelektrode wird vorgestellt, die sich durch besonders gute Reproduzierbarkeit und Stabilität der Tropfenoberfläche auszeichnet. Ein Doppelkonusverschluß gewährleistet die Dichtigkeit des Hg-Reservoirs. Die Abweichungen der Oberflächengröße überschreiten nicht 1%. Die Elektrode wurde erfolgreich in Wasser sowie acht gebräuchlichen reinen organischen Lösungsmitteln getestet, auch im negativen Potentialbereich.SummaryA new design of the hanging mercury drop electrode is described, offering a particularly good reproducibility and stability of the drop surface. The tightness of the mercury reservoir is obtained by a double-cone seal system. The deviation of the electrode surface does not exceed 1%.The electrode was successfully tested in water and also in eight commonly used organic solvents and shows stability even in negative potential range.