Electrochemical uranyl biosensor with DNA oligonucleotides as receptor layer.

The feasibility of using gold electrodes modified with short-chain ssDNA oligonucleotides for determination of uranyl cation is examined. Interaction between UO(2)(2+) and proposed recognition layer was studied by means of voltammetric and quartz crystal microbalance measurements. It was postulated that ssDNA recognition layer functions via strong binding of UO(2)(2+) to phosphate DNA backbone. The methylene blue was used as a redox marker for analytical signal generation. Biosensor response was based on the difference in electrochemical signal before and after subjecting it to sample containing uranyl ion. The lower detection limit of 30 nmol L(-1) for UO(2)(2+) was observed for a sample incubation time of 60 min. Proposed ssDNA-modified electrodes demonstrated good selectivity towards UO(2)(2+) against common metal cations, with only Pb(2+) and Ca(2+) showing considerable interfering effect.

Capillary electrophoretic separation of nanoparticles.

In the present work, CdSe nanocrystals (NCs) synthesized with a trioctylphosphine surface passivation layer were modified using amphiphilic molecules to form a surface bilayer capable of providing stable NCs aqueous solutions. Such modified nanocrystals were used as a test solute in order to analyze new electrophoretic phenomena, by applying a micellar plug as a separation tool for discriminating nanocrystals between micellar and micelle-free zones during electrophoresis. The distribution of NCs between both zones depended on the affinity of nanocrystals towards the micellar zone, and this relies on the kind of surface ligands attached to the NCs, as well as electrophoretic conditions applied. In this case, the NCs that migrated within a micellar zone can be focused using a preconcentration mechanism. By modifying electrophoretic conditions, NCs were forced to migrate outside the micellar zone in the form of a typical CZE peak. In this situation, a two-order difference in separation efficiencies, in terms of theoretical plates, was observed between focused NCs (N ~ 10(7)) and a typical CZE peak for NCs (N ~ 10(5)). By applying the amino-functionalized NCs the preconcentration of NCs, using a micellar plug, was examined, with the conclusion that preconcentration efficiency, in terms of the enhancement factor for peak height (SEF(height)) can be, at least 20. The distribution effect was applied to separate CdSe/ZnS NCs encapsulated in silica, as well as surface-modified with DNA, which allows the estimation of the yield of conjugation of biologically active molecules to a particle surface.

Molecular diagnostic of toxigenic Corynebacterium diphtheriae strain by DNA sensor potentially suitable for electrochemical point-of-care diagnostic.

The presented study is focused on the development of electrochemical genosensor for detection of tox gene fragment of toxigenic Corynebacterium diphtheriae strain. Together with our previous studies it fulfils the whole procedure for fast and accurate diagnostic of diphtheria at its early stage of infection with the use of electrochemical methods. The developed DNA sensor potentially can be used in more sophisticated portable device. After the electrochemical stem-loop probe structure optimization the conditions for real asymmetric PCR (aPCR) product detection were selected. As was shown it was crucial to optimize the magnesium and organic solvent concentrations in detection buffer. Under optimal conditions it was possible to selectively detect as low as 20.8 nM of complementary stand in 5 min or 0.5 nM in 30 min with sensitivity of 12.81 and 0.24 1⋅µM-1 respectively. The unspecific biosensor response was elucidated with the use of new electrode blocking agent, diethyldithiocarbamate. Its application in electrochemical genosensors lead to significant higher current values and the biosensor response even in conditions with magnesium ion depletion. The developed biosensor selectivity was examined using samples containing genetic material originated from a number of non-target bacterial species which potentially can be present in the human upper respiratory tract.

Capillary electrophoresis study on segment/segment system and its role in characterization of nanoparticles.

Capillary electrophoresis (CE) coupled with contactless conductivity detector (C4D) was applied to characterize the migration of micellar segments under BGE/segment/BGE system, where a segment is a micellar phase surrounded by BGE. The transformation (non-ionic, ionic) → mixed micelles under the system was explained in terms of the features of the micellar phase discussed already in the literature. In the second part of the work, based on established phenomena for BGE/segment/BGE system, the BGE/segment/segment/BGE system was used to analyze the re-equilibration of two micellar phases (SDS│(TX-100/SDS)) during a run. This aspect was deeper analyzed with the conclusion that re-equilibration in the present system induces decomposition of the phase of mixed micelles by the phase of ionic micelles and this is a mechanism able to generate the boundary conditions. Established features for BGE/segment/segment/BGE system were further implemented towards preconcentration of nanoparticles and systematic studies show that the system in the configuration (SDS│(TX-100/SDS)) generates a trap state for nanoparticles. Due to this the preconcentration of nanoparticles, expressed in the terms of an enhancement factor (SEFheight), was found to reach level ca. 60.

Development of quantitative structure-activity relationships for interpretation of the migration behavior of neutral platinum(II) complexes in microemulsion electrokinetic chromatography.

Metal-ligand complexes present an important group of analytes in capillary electrophoresis (CE) utilized as an analytical tool for metal ion analysis, metal speciation studies, and characterization of metal complexes of pharmaceutical or industrial importance. In the majority of CE techniques but not yet in microemulsion electrokinetic chromatography (MEEKC), the migration behavior of metal complexes has been well described using the formalism of quantitative structure-activity relationships (QSARs). To fill this gap, a number of migration models, operating with the fundamental properties of analytes as molecular descriptors, were derived and tested for platinum(II) complexes with cyclohexanediamine and its alkylsubstituted derivatives as variable ligand. It was demonstrated that the complexes under examination are consistent with respect to the experimental versus calculated values of n-octanol-water partition coefficients (log P). Meaningful two-parametric regressions between the retention factors and the analyte structural descriptors, such as log P, molecular volumes or surface areas and dipole moments, were obtained. In addition, statistically significant correlations between k' of the platinum(II) complexes and their theoretical binding energies onto a hydrophobic surface were established. The validity of the QSAR modeling approach allowed the authors to postulate that it can be applied to the investigation of binding phenomena for neutral metal complexes to a microdroplet pseudostationary phase.

Chelates of cobalt(III) and iron(II) with 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol as test probes for the characterization of chromatographic effects on a reversed-phase liquid chromatography stationary phase.

The chromatographic effects on a reversed-phase liquid chromatography (RPLC) phase were established with the use of chelates system: cobalt(III) and iron(II) with 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol as the test probes. Both chelates have the same octahedral structure, M:L = 1:2, the former chelate is cationic and can be used to probe the ion-exchange phenomena on a RPLC phase, whereas the latter is not charged and can be used as reference molecule with respect to the charged one. Based on well-established LC phenomena referred in the literature, the suitability of the chelate system for examining some chromatographic effects was tested. It was concluded that the proposed test method is sensitive towards the ion-exchange phenomena on the LC phase and the hydrogen bonding between the solvent eluent and the LC phase. In addition, chromatographic effects due to the presence of ultrasonic field or due to the presence of aromatic amines in the eluent were observed with the help of the proposed test method. Based on molecular computation, the properties of chelates were compared with properties of quaternary amines, the probes most frequently used for testing LC phases, and the possible differences in an interaction of the mentioned compounds with a surface of a RPLC phase were indicated.

Capillary electrophoresis study on segment/segment system for segments based on phase of mixed micelles and its role in transport of particles between the two segments.

Capillary electrophoresis coupled with contactless conductivity detector was applied to characterize BGE/segment/segment/BGE and BGE/segment/electrolyte/segment/BGE systems, where segment is the phase of mixed micelles migrating surrounded by BGE and composition of the first segment≠second segment. It was established that both systems are subject of evolution during electrophoretic run induced by different electrophoretic mobilities of segments and the phenomenon that generates the evolution is exchange of micelles between the two segments. This leads to segments re-equilibration during a run, which generates sub-zones from the two segments in the form of a cumulative zone or two isolated zones, depending on the injection scheme applied. Further analysis based on the system BGE/segment/electrolyte/segment/BGE shows that electrolyte solution between segments can act as a spacer to isolate the two micellar segments, and thereby to control the exchange of micelles between the two segments. Established features for both systems were further implemented towards characterization of the transport of nanocrystals (NCs) between two segments using CE/UV-vis technique and two examples were discussed: (i) on-line coating of NCs with surfactants and (ii) distribution of NCs between segments. The former aspect was found to be useful to discuss the state of particle in micellar media, whereas the latter shows system ability for the transport of NCs from the first segment or BGE based sample to the second segment, controlled by the electrolyte characteristics. It was concluded that transport of micelles and NCs is the subject of the same phenomena since basic electrolyte characteristics, i.e. length and concentration, act in the same way. This means that NCs in these systems can play the role of pseudomicelles, which mimic behaviour of micelles. Definitely, the tools established in the present work can be used to examine dynamic phenomena for pseudophase during electrophoresis and for NCs migrating in the presence of pseudophase in various configurations.

Capillary electrophoresis study on phase of mixed micelles and its role in transport phenomena of particles.

In the present work comprehensive studies on electrophoretic effects induced by a phase of mixed micelles, that migrates surrounded with background electrolyte (BGE) and is denoted as the BGE/segment of mixed micelles/BGE system, were undertaken using capillary electrophoresis coupled with contactless conductivity or UV-vis detector. It was established that mixed micelles under electrophoresis are subject of evolution in terms of mobility, peak area and presence of sub-zones enforced by the composition of micellar phase, segment length and applied voltage. Established features allowed us to explain the electrophoretic behavior of nanoparticles in the system BGE/sample containing nanocrystals/segment of mixed micelles/BGE and it was postulated that a pseudomicellar state of nanoparticles can be useful term in analyzing the migration phenomena of nanoparticles within micellar environment. In contrast to the previous works, where transport of nanocrystals (NCs) within micellar segment or between two micellar segments was analyzed, the present work is focused on the transport of NCs from sample of NCs dispersed in BGE to phase of mixed micelles, i.e., to rear boundary between micellar zone and BGE. Based on these results, systematic studies on transport efficiency for nanoparticles in the system BGE/sample containing nanocrystals/segment of mixed micelles/BGE show that the system assures efficient transport of nanoparticles from BGE based sample to micellar phase and their efficient preconcentration at the micellar segment/BGE rear boundary.

Comparative study on coating CdSe nanocrystals with surfactants.

We have synthesized CdSe nanocrystals (NCs) in sizes from 2.2 to 5.1 nm passivated with hydrophobic trioctylphosphine oxide (TOPO) in combination trioctylphosphine (TOP) or tributylphosphine (TBP) to obtain particles of the type CdSe/TOPO/TOP or CdSe/TOPO/TBP. These NCs were then dispersed in aqueous solution of ionic or non-ionic surfactants (such as stearate, oleic acid, Tween) using a biphase (water and chloroform or hexane) transfer method. It is found that both the structure of the surfactant and the native surface of the ligand govern the coating of the NCs with surfactants. More specifically, the hydrophobicity-hydrophilicity balance of the surfactant regulates the coating efficacy, thereby transferring the NC from the organic to the aqueous phase. The type of ligand on the NCs and the kind of coating surfactant also affect photoluminescence (PL). The ratio of PL and absorbance unit (defined as PL per 0.1 AU) was implemented as a tool to monitor changes in PL intensity and wavelength as a function of size, coatings and surface defects. Finally, the distribution of CdSe nanocrystals between pseudophases in cloud point extraction was discussed based on experimental results. It was concluded that the size of CdSe nanocrystal present in an appropriate pseudophase is correlated with the way in which the non-ionic surfactant coats CdSe nanocrystals. FigureCoating of CdSe semiconductor nanocrystals with surfactants impacts nanocrystals' spectral features. Absorbance of first exciton absorption band was used to estimate ability of surfactant to disperse CdSe nanocrystals. Photoluminescence (PL) intensity and position of PL band were analysed in terms of nanocrystal's surface phenomena via surfactants applied for coating.

Capillary zone electrophoresis of quantum dots dispersed in mixed micelles: new evidence of the concentration effect.

CZE was applied to characterizing a nanoparticle/micellar system, in which CdSe nanoparticles dispersed in a mixture of non-ionic and ionic surfactants are introduced into a capillary to form the sample zone that migrates being sandwiched by the background electrolyte (BGE). Parameters that affect the migration of the surface-modified CdSe nanoparticles and conditions under which they are focused within the micellar zone (or released from it into bulk BGE) were explored, including the sample composition, sample plug length, and applied voltage. The observed migration behavior was analyzed within the framework of a concept of formation of a mixed pseudomicellar system, according to which a nanoparticle coated with an ionic surfactant is treated as a micelle-like entity. It was found out that the nanoparticles are subject to transformation that results in building-up a mixed pseudomicellar system (with regular micelles), with a consequence of exhibiting distinctive migration phenomena. Particularly observed and brought into focus is the event of focusing of the nanoparticles.

Characterization of CdSe nanocrystals coated with amphiphiles. A capillary electrophoresis study.

We have synthesized CdSe nanocrystals (NCs) possessing a trioctylphosphine surface passivation layer and modified with amphiphilic molecules to form a surface bilayer. The NCs covered with single amphiphiles are not stable in aqueous solution, but a mixed amphiphilic system is shown to provide stability in solution over several months. The solutions of the modified NCs were characterized by UV-Vis absorbance, photoluminescence, and transmission electron microscopy. An electrophoretic study revealed two operational modes. The first relies on the enrichment of NCs using a micellar plug as a tool. The accumulation of NCs at the plug-electrolyte buffer interface results in a sharp peak. By controlling the electrophoretic conditions, nanocrystals were forced to exit a micellar plug into an electrolyte buffer. We conclude that a system consisting of modified nanocrystals and a micellar plug can act as a mixed pseudomicellar system, where modified nanocrystals play the role of pseudomicelles.FigureElectrophoretic focusing of amphiphile coated CdSe nanocrystals using a micellar plug. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00604-011-0727-8) contains supplementary material, which is available to authorized users.

Interactions of tumour-targeting nanoparticles with proteins: potential of using capillary electrophoresis as a direct probe.

Metal-based nanoscale particles possess unique optoelectronic or magnetic properties that make them highly promising as imaging agents in cancer therapy research. The fate of nanoparticles in vivo and particularly, the delivery to tumours are closely related to their interactions with plasma proteins. Furthermore, proteins can be used to modify the nanoparticle surface in order to facilitate active targeting to tumours. Therefore, there is an ongoing need for new and more capable analytical methodologies to characterize the protein-nanoparticle binding. Due to the small-sample volume requirement, high degree of resolution and, most importantly, mild, species-friendly separation conditions, capillary electrophoresis (CE) is gaining increasing popularity in the analysis of protein-nanoparticle interaction. This perspective article highlights the potential of CE in studying reactions associated with protein-mediated transformations of nanoparticles, with the focus on quantum dots, gold and iron oxide nanoparticles. Different ways by which CE can be applied to such monitoring are summarized and critically assessed using a representative coverage of recent publications.

An advanced application of the quantitative structure-activity relationship concept in electrokinetic chromatography of metal complexes.

The relevance of the quantitative structure-activity relationship (QSAR) principle in MEKC and microemulsion EKC (MEEKC) of metal-ligand complexes was evaluated for a better understanding of analyte migration mechanism. A series of gallium chelates were applied as test solutes with available experimental migration data in order to reveal the molecular properties that govern the separation. The QSAR models operating with n-octanol-water partition coefficients or van der Waals volumes were found to be valid for estimation of the retention factors (log k') of neutral compounds when using only an aqueous MEEKC electrolyte. On the other hand, consistent approximations of log k' for both uncharged and charged complexes in either EKC mode (and also with hydro-organic BGEs) were achievable with two-parametric QSARs in which the dipole moment is additionally incorporated as a structural descriptor, reflecting the electrostatic solute-pseudostationary phase interaction. The theoretical analysis of significant molecular parameters in MEKC systems, in which the micellar BGE is modified with an organic solvent, confirmed that concomitant consideration of hydrophobic, electrostatic, and solvation factors is essential for explaining the migration behavior of neutral metal complexes.

Reversed-phase liquid chromatographic simultaneous determination of iron(III) and iron(II) as complexes with 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol Determination of iron(III) and iron(II) in water samples and ultrasound field effect on distribution of iron(III) and iron(II) in micellar solution.

The formation of Fe(III) and Fe(II) chelates with pyridylazo and thiazolylazo reagents was examined. Optimum conditions for the formation of Fe(III) and Fe(II) chelates with 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol (5-Br-PADAP) were in detail evaluated. The LC method for simultaneous separation of Fe(III) and Fe(II) ions as 5-Br-PADAP chelates was evaluated using the PEEK column with C18 e.c. stationary phase and acetonitrile+water (90:10, v/v) eluent containing the 1x10(-3) mol l(-1) C(12)H(25)SO(3)Na, the ion-pairing reagent, pH 3.4-3.6. The simultaneous determination of 20-500 mug l(-1) Fe(II) ions (detection at 555 nm) and 20-500 mug l(-1) Fe(III) ions (detection at 585 nm) as 5-Br-PADAP chelates (for both ions, detection limit, 18 mug l(-1) for 20 mul loop) was established. The chromatographic method was applied to the water analysis. Although the present method is able to determine both Fe(III) and Fe(II) ions, the Fe(III) ion was not detected in all water samples. The Fe(II) was detected only in fresh gathered oligocene water at the level of 135 mug l(-1). The present method was used to the investigation of the distribution of Fe(III)/Fe(II) ions in aqueous and micellar solutions after action of external, ultrasonic field.

Simultaneous determination of zirconium and hafnium as ternary complexes with 5-Br-PADAP and fluoride using solid-phase extraction and reversed-phase liquid chromatography.

Solid-phase extraction (SPE) along with reversed-phase liquid chromatography (RP-LC) was used for the simultaneous determination of Zr(IV) and Hf(IV) by means of their ternary chelates with fluoride and 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol (5-Br-PADAP). The conditions of SPE sorption were examined in detail: type of SPE column, volume of the sample, volume of the eluent, concentrations of metal ions, fluoride salt, chromogenic reagent, organic phase, and pH. It was established that the sorption of Zr(IV) and Hf(IV), as their ternary chelates, on SPE Zorbax SPE C18 (EC) cartridge was the most efficient, when the sample containing metal ion (Zr(IV), Hf(IV), both, up to 2 mug), 5-Br-PADAP 1.5x10(-4), NaF 7.5x10(-5) mol l(-1), methanol 40%, pH 4.5+/-1 was applied for the SPE sorption. The chelates were discarded from SPE cartridge using acetonitrile/water (99.75+0.25, v/v) eluent containing 3.8x10(-4) mol l(-1) sodium fluoride and subsequently separated by RP-LC method. The RP-LC separation of both chelates was optimized and Zorbax SB-C18 analytical LC column along with acetonitrile/water (65+35, v/v) eluent containing the 1.5x10(-4) mol l(-1) sodium fluoride was used. The established SPE/LC conditions allow Zr(IV) 0.08-2.0 mug and Hf(IV) 0.04-2.0 mug determination in a sample volume up to 150 ml. The detection limits, 0.03 mug Hf(IV) and 0.05 mug Zr(IV), were obtained. Recoveries, (94+/-2)% for Hf(IV) chelate and (106+/-2)% for Zr(IV) chelate were obtained, when 1 mug of Zr(IV) and Hf(IV) ions were determined by the present SPE/LC method from the sample volume of 100 ml. The established, pre-concentration SPE conditions, along with the LC separation and determination allow the assay of Zr(IV) and Hf(IV) in complicated matrix materials. The present SPE/LC method was applied to the determination of Zr(IV) and Hf(IV) in tap water and reference geological material (rock, NCS DC 73303; certified content: Zr, 27.7x10(-3)% (w/w) and Hf, 6.5x10(-4)% (w/w)).

Platinum metallodrug-protein binding studies by capillary electrophoresis-inductively coupled plasma-mass spectrometry: characterization of interactions between Pt(II) complexes and human serum albumin.

Characterizing how platinum metallocomplexes bind to human serum albumin (HSA) is essential in evaluating anticancer drug candidates. Using cisplatin as a reference complex, the application of capillary electrophoresis (CE) to reliably assess drug/HSA interactions was validated. Since this complex is small compared to the size of the protein, the binding response could only be recognized when applying CE coupled to a (platinum) metal-specific mode of detection, namely inductively coupled plasma-mass spectrometry (ICP-MS). This coupling allowed for confirmation of a specific affinity of cisplatin and novel Pt complexes to HSA, measurement of the kinetics of binding reactions, and determination of the number of drug molecules attached to the protein. As the cisplatin/HSA molar ratio increased, the reaction rate became faster with a maximum on the kinetic curve appearing at about 50 h of incubation at 20 times excess of cisplatin. The reaction was characterized as a pseudo-first order reaction with the rate constant k = 0.003 min(-1) at 37 degrees C. When incubated with a 20-fold excess of cisplatin, HSA bound up to 10 mol of Pt per mol of the protein. This is indicative for a strong metal-protein coordination occurring at several HSA sites other than the only protein cysteine residue. Structural analogs of cisplatin, bearing aminoalcohol ligands, showed comparable protein binding reactivity and stoichiometry but a common equilibrium was not reached even after one week of incubation. Also apparent was a two-step mechanism of the binding reaction. Results demonstrated the suitability of CE-ICP-MS as a rapid assay for high-throughput studying of drug/HSA interactions.