Paweł Knihnicki

Tyrosinase-based biosensor for determination of bisphenol A in a flow-batch system

A tyrosinase-based amperometric biosensor is proposed for determination of bisphenol A (BPA) in a flow-batch monosegmented sequential injection system. The enzyme was entrapped in a sol-gel TiO2 matrix modified with multi-walled carbon nanotubes (MWCNTs), polycationic polymer poly(diallyldimethylammonium chloride), (PDDA) and Nafion. Morphology of TYR/TiO2/MWCNTs/PDDA/Nafion matrix composite was studied via scanning electron microscopy (SEM). Electrochemical behavior of the developed biosensor towards bisphenol A was examined and analytical characteristics were assessed with respect to linear range, biosensor sensitivity, limit of detection, long term stability, repeatability and reproducibility. Linear range of biosensor response was found between 0.28 and 45.05 µM with high sensitivity of 3263 µA mM(-1) cm(-2) and detection limit 0.066 µM. The approach was successfully employed for determination of BPA in natural samples.

Analytical methodologies for the determination of benzodiazepines in biological samples.

Benzodiazepine drugs belong to important and most widely used medicaments. They demonstrate such therapeutic properties as anxiolytic, sedative, somnifacient, anticonvulsant, diastolic and muscle relaxant effects. However, despite the fact that benzodiazepines possess high therapeutic index and are considered to be relatively safe, their use can be dangerous when: (1) co-administered with alcohol, (2) co-administered with other medicaments like sedatives, antidepressants, neuroleptics or morphine like substances, (3) driving under their influence, (4) using benzodiazepines non-therapeutically as drugs of abuse or in drug-facilitated crimes. For these reasons benzodiazepines are still studied and determined in a variety of biological materials. In this article, sample preparation techniques which have been applied in analysis of benzodiazepine drugs in biological samples have been reviewed and presented. The next part of the article is focused on a review of analytical methods which have been employed for pharmacological, toxicological or forensic study of this group of drugs in the biological matrices. The review was preceded by a description of the physicochemical properties of the selected benzodiazepines and two, very often coexisting in the same analyzed samples, sedative-hypnotic drugs.

Development of microextraction by packed sorbent for toxicological analysis of tricyclic antidepressant drugs in human oral fluid.

The aim of this study was to apply microextraction by packed sorbent (MEPS) to the isolation of six tricyclic antidepressants (TCADs): nordoxepin, doxepin, desipramine, nortriptyline, imipramine, and amitriptyline from human oral fluid. Samples were collected from healthy volunteers via free spillage from the oral cavity to disposable test tubes. A method of oral fluid sample pretreatment was developed and optimized in terms of suitability for MEPS extraction and removing of interfering agents (protein, food debris, or air bubbles). Moreover, it was short and simple to perform with limited sample consumption (150μL). Extracts were analysed by UHPLC-MS. The MEPS/UHPLC-MS method was validated at three concentration levels (2.00, 4.00 and 8.00ng/mL) of all analytes in the range 1.25-10.0ng/mL. The following parameters were determined: limit of detection, limit of quantification, precision, and accuracy. For all tested concentration levels, the intra- and inter-day repeatability did not exceeded 8.1% and 12.2%, respectively. Gained LOQ value, 0.50ng/mL, made the MEPS/UHPLC-MS method to be a useful tool in clinical and forensic laboratories, which was demonstrated on the basis of analysis of real samples.

Novel electroanalytical method based on the electrostriction phenomenon and its application to determination of Cr(VI) by the flow injection technique

The electrostriction phenomenon is observed in membranes of thickness in the nanometer range, e.g. bilayer lipid membranes or self-assembled monolayers. Strong electrical field of 105-106V/cm in intensity appears when applying the potential lower than 1V to these membranes. Electrostatic forces change the dimensions of a dielectric, hence the membrane compression and decrease in thickness are observed. As a result, increase in the membrane capacitance is recorded. The presented work covers development and application of a new analytical method based on the innovative capacitance-to-frequency conversion method and flow technique. Construction of the novel flow manifold designed for realization of the whole analytical procedure of chromium(VI) determination with the use of capacitance measurements and its operating rules are shown. The main element of the system is a specially designed measurement cell for realization of chromium determination using a three-electrode system. Au electrode with a self-assembled monolayer of thiols grafted with functional groups selectively interacting with the analyte is employed as a dielectric layer. Ag/AgCl electrode and Pt one are used as a reference and auxiliary electrode, respectively. Accuracy, repeatability and reproducibility of the proposed analytical procedure were tested in determination of Cr(VI) in synthetic solutions and environmental water matrices spiked with the analyte. Benefits and drawbacks of the developed manifold were critically discussed.

Critical approach to flow injection gradient titration as a calibration method.

An attempt was made to demonstrate that flow injection gradient titration should not be considered as classical titration but rather as the indirect calibration method. This was shown experimentally by exploiting indirect spectrophotometric determination of chlorite ions in the presence of iron(II) in an acid environment with measurement of absorbance for the coloured Fe(II)/o-phenanthroline complex at 512nm. In such an analytical system, the peaks of a cut-off profile were obtained with two characteristic points corresponding to equivalent amounts of analyte and reagent, and peak widths were used as the analytical signal. The only difference between the approach presented and the classical flow injection titration procedure was not fundamental but of a technical nature and consisted in the fact that a sample loop of relatively great volume instead of a mixing chamber was installed into the flow injection system. The analytical performance of the developed calibration procedure is also presented and discussed.

Application of polypyrrole nanowires for the development of a tyrosinase biosensor

Polypyrrole nanowires (PPyNWs) were fabricated and examined as a structural component of amperometric biosensor matrix. An enzyme, tyrosinase (TYR), was immobilized onto PPyNWs using glutaraldehyde (GA). Matrix composite morphology was investigated using scanning electron microscopy. Electrochemical behavior of the prepared PPyNWs/GA/TYR biosensor towards catechol was studied and the assessment of its analytical characteristics was carried out taking into account linear range, sensitivity, repeatability, reproducibility and operational stability.

Flow manifold for chemical H-point standard addition method implemented to electrochemical analysis based on the capacitance measurements

The paper presents a novel analytical method for electrochemical potassium determination using gold electrodes with self-assembled thiol monolayers (SATMs) deposited on its surface. The whole analytical procedure was carried out in a dedicated flow manifold using capacitance detection mode. The terminal functional groups on the surface of the dielectric monolayer interacts selectively with the analyte, changing the thickness of the layer depending on the amount of the analyte and the applied voltage, resulting in a change in the registered dielectric capacitance. New calibration approach based on the Chemical H-point Standard Addition Method (C-HPSAM), allowing both specific (proportional) and unspecific (constant) interference effects caused by sodium ions to be corrected, was applied. The calibration method is based on the registration of the calibration graphs by the standard addition method (SAM) three times in such different chemical conditions, which are able to differentiate the slope of each graph. The C-HPSAM was applied for the first time in electrochemical analysis. As a chemical parameter differentiating the sensitivity of the calibration graphs, the concentration of ionic strength stabilizer (ethylenediamine) was used. In order to improve the analytical procedure, to make it faster and automated, the dedicated flow system was applied. The constructed flow system was composed of several modules individually dedicated to the appropriate step of the whole analytical procedure: electrochemical cleaning of work electrode surface, adsorption of SATMs and analytical calibration. The calibration curves were obtained in the range of 0.1-0.9 mmol L-1 with good linearity (R2 = 0.996 ± 0.001) and the LOD and LOQ of 28.6 and 85.8 μmol L-1, respectively. The proposed method was employed for potassium determination in highly mineralized water, juice and pharmaceutical samples without any special pretreatment.