Mirko Deml

Electrically controlled electrofocusing of ampholytes between two zones of modified electrolyte with two different values of pH

Abstract A method is described for the separation and concentration of ampholytes from their mixtures with other ionic species. In a quadrupole electromigration column, two zones of different pHs are created by using controlled flows of the solvolytic ions (H + and OH − ) from appropriate electrode chambers at opposite sides of the column. Thus, a time-variable interface is created between two zones with a sharp change of pH. The position of the interface in the column, the direction and velocity of its movement and the difference in pH across the interface-pH gap can be adjusted by electric currents. This arrested interface is reasonably stable with time and has the following separation properties: ampholytes with p I values between the pHs of both zones are focused into a zone at interface; and other types of ampholytes and other weak or strong ions are not trapped at the interface. The basic properties of the above system are described and experiments showing the effects of the type of sample (ampholyte, weak ion), time, the concentration of primary electrolyte and the additives changing the viscosity, solubility or p I of ampholyte are given. The method proposed offers the following advantages: the ampholytes in the sample may be concentrated several hundred-fold; the focused zones have sharp boundaries (zones 0.1 mm in length were prepared) and high concentrations of the trapped species; the zone of a trapped ampholyte contains the ampholyte proper and simple ions of primary electrolyte (KCI) only; and the zones can be shifted to any selected position in the column (potentially to the location of a detector cell or a collection device).

Use of complex formation equilibria in the analytical isotachophoresis of strong electrolyte ions : Separation of halides and sulphates

Abstract The possibility of effecting isotachophoretic separations by means of complex formation equilibria between a counter ion and the components being separated was investigated. This procedure was shown to be of potential significance mainly for the separation of ionic species of strong electrolytes, and the separation of sulphates and halides was examined. The use of Cd(II) as a counter ion and NO3− as a leading ion is suggested. The migration of sulphate and nitrate zones is considered theoretically, and data calculated for actual operating conditions are verified experimentally.

Instrumentation for high-speed isotachophoresis

Abstract The design and operating parameters of the individual elements of an isotachophoregraph were examined from the viewpoint of rapid quantitative analysis. An arrangement comprising an inlet port, separation column and detection cell was constructed, in which dead spaces were eliminated and the capillary and detection cell were efficiently cooled. This arrangement permits the time of isotachophoretic analysis from the instant of sample introduction to be shortened to 3-5 min, the development of the isotachophoregram proper taking 20-40 sec. The basis of the arrangement is a monolithic block of Perspex that comprises both electrode compartments, control stopcocks of the sampling device and the capillary. The latter is formed by a flat groove in the block, closed with a PTFE foil overlapping the groove and compressed on to the block with a thermostatted metallic plate. The detection cell is formed by part of the capillary and consists of two platinum contacts that extend into the groove in the monolithic block. The detector functions by sensing the electric gradient of the migrating zone. Recording was effected by means of a home-made voltmeter of high input resistance, with the detector input insulated electrically from the output of the recorder. The high-voltage source employed was also home made and yielded a stabilized current of up to 400 μA at 16 kV.

High-speed isotachophoresis: current supply and detection system

Abstract A stabilized d.c. power supply for high-speed analytical isotachophoresis, providing a stepless constant current adjustable up to 400 μA at a voltage up to 16 kV, and a detection system for the direct measurement of potential gradients in isotachophoretic zones are described. The design enables any of the power supply output terminals to be earthed. Hence the detection cell can be kept at a relatively low above-earth potential when analyzing both anions and cations. It has been verified that the ionic species separated can be identified by determining the relative ratios of the potential gradients in their zones and by comparing these experimental data with the inverse values of the relative ratios of the tabulated mobilities. The relative deviations found did not exceed 2% for a model mixture of inorganic anions.

Electric sample splitter for capillary zone electrophoresis

Abstract A method for reproducible sampling of small quantities of sample for high-performance capillary electrophoretic separations was developed. The method is based on the principle of the splitter. The sample migrates electrophoretically in two electrical circuits and the splitting ratio is given by the ratio of the corresponding electric currents. Very small aliquots of the sample in the form of short zones (pulses) can be introduced corresponding to direct sampling of nanolitre volumes.

Large sample volume preseparation for trace analysis in isotachophoresis

An isotachophoretic device for the analysis of trace components in a sample with the efficient pre-separation and elimination of bulk components has been suggested and realized. A large volume of the sample in question is separated in a rectangular wide-bore channel packed with a suspension of a granulated polyacrylamide gel and analyzed in free electrolytes in a narrow-bore tube. Trace components, the concentrations of which are 10(-6) mol/l, can be analyzed in 60-70 min applying ca. 1 ml of the sample even in the presence of bulk excess of a major component the concentration of which is by 5 orders of magnitude higher.

Concept of the effective mobility of the hydrogen ion and its use in cationic isotachophoresis

Abstract The concept of the effective mobility of the terminating H+ ion for cationic isotachophoresis in an acidic medium is introduced and its application to the selection of the electrolyte system for a required separation is discussed. Based on the theoretical description of the migrating reaction boundary, a relationship is derived for the calculation of the effective mobility of the H+ ion. Further, a rule is defined for the selection of a terminating cation such that the loss of the control of the migration of H+ ions through the isotachophoretic system is avoided. The method for the calculation and the rule mentioned above have been verified experimentally for model systems.

Migration behaviour of the hydrogen ion and its role in isotachophoresis of cations

Abstract A theoretical and experimental study has been made of the behaviour of hydrogen ions under the conditions of cationic isotachophoresis. It is shown that, for successful cationic isotachophoresis, the electrolyte conditions should be such that H+ migrates in an enforced isotachophoretic manner as terminator and its front boundary shows self-sharpening. A mathematical description of the migration of H+ as the terminating zone is elaborated, giving the criteria decisive for the character of the terminating zone front boundary depending on the composition of the leading electrolyte and on the properties of the cationic substances separated (strong and weak bases). It has been found theoretically that, for a given set of cations to be separated and counter anion in the leading electrolyte, the concentration of the leading electrolyte must be higher than a certain critical value to obtain stable isotachophoretic migration. Model experiments with different counter-anion systems (acetic acid—acetatate and formic acid—formate) at various concentrations have given good agreement with the theory. Illustrative examples of cationic isotachophoresis of some amino acids and dipeptides are described. The model described enables both the prediction of migration behaviour of cationic substances and the selection of suitable electrolyte systems in order to obtain stable isotachophoretic migration of these substances.

Analytical isotachophoresis : The concept of the separation capacity

Abstract The concept of separation capacity in isotachophoresis has been investigated in order that the analytical separation possibilities may be predicted for given working conditions. A relationship has been derived between the amount and composition of a binary mixture under isotachophoretic separation and the amount of electricity which has to be passed through the system to obtain complete separation of the mixture. By using this relationship, values of the separation capacity have been calculated for three two-component mixtures analyzed by means of two isotachophoretic instruments and verified experimentally.

Generation of operational electrolytes for isotachophoresis and capillary zone electrophoresis in a three-pole column

Abstract A new method has been developed for controlling the composition of the operational electrolytes directly in the separation capillary in isotachophoresis or capillary zone electrophoresis. The method is based on feeding the capillary with two different suitable ionic species from two separate electrode chambers by simultaneous electromigration. The composition and pH of the electrolyte in the separation capillary is thus controlled by setting the ratio of two electric currents. The theory has been developed and verified experimentally to predict both the electrolyte composition in the separation capillary and the time necessary to change this composition in the required way. Some of the possible ionic matrices realizable in the three-pole arrangement have been studied experimentally and used in isotachophoretic experiments. The technique described does not require moving parts in the instrumentation and provides the possibility to make very fine changes of pH in the capillary in a reproducible and easy way. The procedure itself is feasible for automation.