Stanislawa Koronkiewicz

Programmable chronopotentiometry as a tool for the study of electroporation and resealing of pores in bilayer lipid membranes.

This paper presents the application of chronopotentiometry in the study of membrane electroporation. Chronopotentiometry with a programmable current intensity was used. The experiments were performed on planar bilayer phosphatidylcholine and cholesterol membranes formed by the Mueller-Rudin method. It was demonstrated that a constant-intensity current flow through the bilayer membranes generated voltage fluctuations during electroporation. These fluctuations (following an increase and decrease in membrane conductance) were interpreted as a result of the opening and closing of pores in membrane structures. The decrease in membrane potential to zero did not cause the pore to close immediately. The pore was maintained for about 200 s. The closing of the pore and recovery of the continuous structure of the membrane proceeded not only when the membrane potential equalled zero, but also at membrane potentials up to several tens of millivolts. The fluctuations of the pore were possible at values of membrane potential in the order of at least 100 mV. The size of the pore changed slightly and it closed after some time below this potential value.

A novel direct-injection photometric detector integrated with solenoid pulse-pump flow system.

A novel flow photometric detector based on paired emitter-detector diodes (PEDD), coupled with solenoid pulse micro-pumps is presented. The photometric detection chamber also plays a role of the reaction chamber. Both solutions, a sample and a reagent, are injected by solenoid micro-pumps directly and simultaneously into the chamber in countercurrent. This method ensures fast and effective mixing of the injected solutions. A good accuracy and precision of the injected volumes assure the pulse micro-pumps. This method, in comparison with other flow methods, does not require the application of the reaction coil. Thanks to this, the dispersion of the sample is minimized. The presented flow network is easy to control, miniaturize and exhibits a very low consumption of reagents and the sample. Two chemical systems were chosen and presented as models of photometric reactions: the first - Fe(III) with thiocyanate, and the second - Cr(VI) with 1,5-diphenylcarbazide. To obtain the highest repeatability, the total volume of the solutions should be smaller than the volume of the reaction-detection chamber. In this case, the whole coloured product remains in the chamber. The use of the proposed direct-injection PEDD detector considerably simplifies analytical procedures. The shape of analytical signals and their potential applications have been discussed.

Changes of structural and dynamic properties of model lipid membranes induced by α-tocopherol: implication to the membrane stabilization under external electric field

The effects of alpha-tocopherol on electric properties of bilayer lipid membranes were investigated. Planar bilayer membranes formed by the Mueller-Rudin method were used. Voltammetric and chronopotentiometric measurements were performed using a four-electrode potentiostat-galvanostat. It was demonstrated that registration of membrane capacitance, resistance, and voltammetric characteristics provided information about the change in the structure and permeability of bilayer lipid membranes. The results suggested that incorporation of alpha-tocopherol into lipid membrane destabilized its structure and facilitated the electrogeneration of pores. The possible role of observed changes in physiological functions of alpha-tocopherol was discussed.

Application of direct-injection detector integrated with the multi-pumping flow system to photometric stop-flow determination of total iron

A novel direct-injection detector (DID) integrated with multi-pumping flow system (MPFS) for the photometric determination of iron is proposed. Paired emitter-detector diodes have been used as a photometric detection system. The sample and reagent were injected using appropriate solenoid pulse micro-pumps directly into the detection chamber where effective mixing occured. The use of proposed stop-flow detector considerably simplified the analytical procedure. The potassium thiocyanate has been chosen as a chromogenic reagent for photometric Fe(III) detection. The total volume of reagent and sample/standard solutions involved in the detection process was adjusted to the volume of the reaction-detection chamber. Calibration graph was found to be linear in the range up to 10mgL(-1). The detection limit (3s(b)/S) was 0.15mgL(-1). The repeatability (R.S.D.), calculated from 10 analyses of sample containing 5mgL(-1) Fe(III), was 1.5% and the sample throughput 180 determinations per hour. The consumption of sample and reagent was 20μL each with the waste generation at the level of 0.24mL. The applicability of the proposed method to the determination of total iron in groundwater samples has been proved. The analytical parameters are compared to those obtained exploiting the MPFS system with typical configuration containing a confluence point and reaction coil.

An automatic chemiluminescence method based on the multi-pumping flow system coupled with the fluidized reactor and direct-injection detector: Determination of uric acid in saliva samples

A novel approach for the automatic chemiluminescence (CL) analysis of the complex samples is proposed. A multi-pumping flow system was successfully combined with fluidized reactor and direct-injection CL detector. The possibility of the approach was demonstrated on the determination of uric acid in saliva samples. Uric acid is clinically important analyte and its determination in biological fluids is related to human organism dysfunctions, such as gout. For the first time, the fast luminol - N-bromosuccinimide (NBS) reaction in an alkaline medium was used for the CL determination of uric acid in saliva samples. The CL intensity is greatly quenched in the presence of the analyte. The method includes on-line separation of uric acid from the saliva samples based on fluidized beds strategy using anion-exchange resin Dowex® 2×8 followed by the elution and CL determination using a direct-injection CL detector. The stroke pulsations of the solenoid micro-pumps provided the floating of the anion exchange resin in a sample phase and uric acid separation from the sample matrix into a sample pre-treatment block of flow system. To obtain efficient elution of analyte an eluent circulation was applied. Under the optimal conditions, the detector response for uric acid was linear in the logarithmic concentration ranges from 6·10-6 to 1·10-3molL-1. The limit of detection, calculated from a blank test based on 3σ, was 2·10-6molL-1. Fluidized bed strategy allows us to exclude saliva matrices influence on the luminol-NBS CL reaction, which results in improved selectivity. The applicability of the method developed is demonstrated with the help of real sample analysis. The obtained results are confirmed by reference HPLC-UV method.

Direct-injection chemiluminescence detector. Properties and potential applications in flow analysis.

We present a novel chemiluminescence detector, with a cone-shaped detection chamber where the analytical reaction takes place. The sample and appropriate reagents are injected directly into the chamber in countercurrent using solenoid-operated pulse micro-pumps. The proposed detector allows for fast measurement of the chemiluminescence signal in stop-flow conditions from the moment of reagents mixing. To evaluate potential applications of the detector the Fenton-like reaction with a luminol-H2O2 system and several transition metal ions (Co(2+), Cu(2+), Cr(3+), Fe(3+)) as a catalyst were investigated. The results demonstrate suitability of the proposed detector for quantitative analysis and for investigations of reaction kinetics, particularly rapid reactions. A multi-pumping flow system was designed and optimized. The developed methodology demonstrated that the shape of the analytical signals strongly depends on the type and concentration of the metal ions. The application of the detector in quantitative analysis was assessed for determination of Fe(III). The direct-injection chemiluminescence detector allows for a sensitive and repeatable (R.S.D. 2%) determination. The intensity of chemiluminescence increased linearly in the range from about 0.5 to 10 mg L(-1) Fe(III) with the detection limit of 0.025 mg L(-1). The time of analysis depended mainly on reaction kinetics. It is possible to achieve the high sampling rate of 144 samples per hour.

A novel pulsed xenon flashlamp photoreactor and its potential applications in flow analysis with chemiluminescence detection

The pulsed xenon flashlamp as a source of UV-light in a flow analytical system is proposed. Milliseconds long, high energy pulses were applied to a suspension of photocatalyst (TiO2) in order to generate free radicals capable of participating in a luminol chemiluminescence reaction. Three commercially available TiO2 powders of different crystal structures and sizes were selected to prepare suspensions in an H2O2 solution or in ultrapure water. The preliminary results indicate that the induction of chemiluminescence signals using the proposed photoreactor is possible. The intensity of the chemiluminescence signals depends on the number of flashes applied and the composition of the irradiated suspension. The photoreactor described is compact, small in size and capable of saving reagents and energy. Additionally, the system allows us to obtain chemiluminescence signals with shorter irradiation time when compared to configurations employing classical low-pressure mercury lamps. The detailed characteristics of a flashlamp photoreactor and its potential applications in flow systems are discussed.

A chemiluminescence method for screening of fluoroquinolones in milk samples based on a multi-pumping flow system

An automated and miniaturized chemiluminescence method for screening of fluoroquinolones in milk samples was proposed. The method was based on magnetic dispersive micro-solid phase extraction of analytes followed by the chemiluminescence determination of the total fluoroquinolones content using a multi-pumping flow system. In the developed method, Zr-Fe-C magnetic nanoparticles were used as an efficient sorbent for separation of fluoroquinolones from sample matrix. The chemiluminescence intensity obtained for Ce4+-SO32--Tb3+ chemiluminescence system was greatly increased in the presence of the analytes. Under the optimal conditions, the detector response for fluoroquinolones was linear in the concentration ranges from 5·10-9 to 1·10-6 mol L-1 with respect to fleroxacin. The limit of detection, calculated from a blank test based on 3σ, was 2·10-9 mol L-1 with respect to fleroxacin. The presented method demonstrated to be a good tool for available and cost-effective point-of-need screening fluoroquinolones in milk samples.