para-Phenylenediamine Dimer as a Redox-Active Guest for Supramolecular Systems.

Redox-active components are highly valuable in the construction of molecular devices. We combined two p-phenylenediamines (p-PDA) with a biphenyl (BiPhe) unit to prepare a supramolecular guest 4 consisting of three binding sites for cucurbit[7/8]uril (CBn) and/or cyclodextrins (CD). Supramolecular properties of 4 were investigated using NMR, UV-vis, mass spectrometry and isothermal titration calorimetry. Our analysis revealed that 4 forms higher-order host-guest complexes, wherein a CD unit occupies the central BiPhe site, secured by two CBn units at the terminal p-PDA sites. Additionally, 1 : 1 complexes with α-CD and ß-CD, a 1 : 2 complex with γ-CD and 2 : 1 complexes with CB7 and CB8 were identified. Through UV-vis and cyclic voltammetry, redox processes leading to the formation of a stable, deep blue dication diradical of 4 are elucidated. Furthermore, it is demonstrated that CB7 selectively protects oxidised 4 from reduction in the presence of a reducing agent. The supramolecular and redox properties of the structural motif represented by 4 render it an interesting candidate for the construction of supramolecular devices.

Materials interacting with inorganic selenium from the perspective of electrochemical sensing.

Inorganic selenium, the most common form of harmful selenium in the environment, can be determined using electrochemical sensors, which are compact, fast, reliable and easy-to-operate devices. Despite progress in this area, there is still significant room for developing high-performance selenium electrochemical sensors. To achieve this, one should take into account (i) the electrochemical process that selenium undergoes on the electrode; (ii) the valence state of selenium species in the sample and (iii) modification of the sensor surface by a material with high affinity to selenium. The goal of this review is to provide a knowledge base for these issues. After the Introduction section, mechanisms and principles of the electrochemical reduction of selenium are introduced, followed by a section introducing the modification of electrodes with materials interacting with selenium and a section dedicated to speciation methods, including the reduction of non-detectable Se(VI) to detectable Se(IV). In the following sections, the main types of materials (metallic, polymers, hybrid (nano)materials…) interacting with inorganic selenium (mostly absorbents) are reviewed to show the diversity of properties that may be endowed to sensors if the materials were to be used for the modification of electrodes. These features for the main material categories are outlined in the conclusion section, where it is stated that the engineered polymers may be the most promising modifiers.

Simplified synthesis of silver nanoparticles on graphene oxide and their applications in electrocatalysis.

In this work the possibility of synthesizing in situ silver nanoparticles (AgNPs) on graphene oxide (GO) surfaces without commonly used additional reducing or alkalizing agents or increased temperature was investigated. Using diverse microscopic (atomic force microscopy, transmission electron microscopy) and spectroscopic methods, it was proved that very small AgNPs were formed on GO by simple incubation for 2 h in a mixture of GO dispersion and AgNO3. The prepared nanomaterial (GO_Ag) was also assessed using electrochemical methods, and it exhibited electrochemical behavior similar to the GO_Ag nanomaterial prepared with a help of citric acid as a reducing agent. Furthermore, it was found that (i) the electrochemical reduction of the GO_Ag on the electrode surface decreased the voltammetric response even though this step increased the surface conductivity and (ii) GO_Ag can be employed for the sensing of chlorides with a detection limit of 79 µM and a linear range of up to 10 mM. It could also provide an electrochemical response toward the chloroacetanilide herbicide metazachlor. Hence, the reducing capabilities of GO were proved to be applicable for in situ synthesis of metal nanoparticles with the highest possible simplification, and the as-prepared nanomaterials could be employed for fabrication of different electrochemical sensors.

Modulation of wettability, gradient and adhesion on self-assembled monolayer by counterion exchange and pH.

In this study, two quaternary ammonium salts derived from l-lipoic acid were applied for self-assembled monolayers formation on rough structured gold surface. The derivatives differ in functionality since one possesses simple quaternary ammonium group whereas the other one is carboxybetaine ester containing quaternary ammonium group with pH hydrolysable ester group as a pendant. The response of surface wettability to ion exchange between Cl- and perfluorooctanoate, kinetics and gradient wettability were examined by water contact angle measurement and confirmed by X-ray photoelectron spectroscopy. Furthermore, adhesion forces related to applied counterion on the entire surface and after hydrolysis were investigated by atomic force microscopy measurement at nanometer scales. A dramatic change in wettability upon counterion exchange from superhydrophilic for Cl- to very or superhydrophobic for perfluorooctanoate in a repeatable manner was observed for both derivatives. Kinetics of counterion exchanges revealed faster hydration of simple quaternary derivate. The wettability gradient could be designed from superhydrophobic to superhydrophilic either in a reversible manner by simple immersion of the modified surface in a counterion solution modulated by ionic strength or in an irreversible manner for carboxybetaine ester derivate by time-controlled hydrolysis to charge balanced carboxybetaine.

Electrochemical performance of Ti3C2Tx MXene in aqueous media: towards ultrasensitive H2O2 sensing.

An extensive characterization of pristine and oxidized Ti3C2Tx (T: =O, -OH, -F) MXene showed that exposure of MXene to an anodic potential in the aqueous solution oxidizes the nanomaterial forming TiO2 layer or TiO2 domains with subsequent TiO2 dissolution by F- ions, making the resulting nanomaterial less electrochemically active compared to the pristine Ti3C2Tx. The Ti3C2Tx could be thus applied for electrochemical reactions in a cathodic potential window i.e. for ultrasensitive detection of H2O2 down to nM level with a response time of approx. 10 s. The manuscript also shows electrochemical behavior of Ti3C2Tx modified electrode towards oxidation of NADH and towards oxygen reduction reactions.

pH-Switchable Interaction of a Carboxybetaine Ester-Based SAM with DNA and Gold Nanoparticles.

We describe a self-assembled monolayer (SAM) on a gold surface with a carboxybetaine ester functionality to control the interaction between DNA and gold nanoparticles via pH. The negatively charged phosphate backbone of DNA interacts with and adsorbs to the positively charged carboxybetaine esters on the SAM. DNA release can be achieved by the hydrolysis of carboxybetaine ester (CBE) to a zwitterionic carboxybetaine state. Furthermore, the adsorption of negatively charged citrate-capped gold nanoparticles to a SAM-modified plain gold surface can be controlled by the pH. The SAM based on carboxybetaine ester allows for the homogeneous adsorption of particles, whereas the SAM after hydrolysis at high pH repels AuNP adsorption. The antifouling surface properties of the surface modified with carboxybetaine were investigated with protein samples.

Progress in biocatalysis with immobilized viable whole cells: systems development, reaction engineering and applications.

Viable microbial cells are important biocatalysts in the production of fine chemicals and biofuels, in environmental applications and also in emerging applications such as biosensors or medicine. Their increasing significance is driven mainly by the intensive development of high performance recombinant strains supplying multienzyme cascade reaction pathways, and by advances in preservation of the native state and stability of whole-cell biocatalysts throughout their application. In many cases, the stability and performance of whole-cell biocatalysts can be highly improved by controlled immobilization techniques. This review summarizes the current progress in the development of immobilized whole-cell biocatalysts, the immobilization methods as well as in the bioreaction engineering aspects and economical aspects of their biocatalytic applications.

Immobilization of bilirubin oxidase on graphene oxide flakes with different negative charge density for oxygen reduction. The effect of GO charge density on enzyme coverage, electron transfer rate and current density.

Previously we showed that an effective bilirubin oxidase (BOD)-based biocathode using graphene oxide (GO) could be prepared in 2 steps: 1. electrostatic adsorption of BOD on GO; 2. electrochemical reduction of the BOD-GO composite to form a BOD-ErGO (electrochemically reduced GO) film on the electrode. In order to identify an optimal charge density of GO for BOD-ErGO composite preparation, several GO fractions differing in an average flake size and ζ-potential were prepared using centrifugation and consequently employed for BOD-ErGO biocathode preparation. A simple way to express surface charge density of these particular GO nanosheets was developed. The values obtained were then correlated with biocatalytic and electrochemical parameters of the prepared biocathodes, i.e. electrocatalytically active BOD surface coverage (Γ), heterogeneous electron transfer rate (kS) and a maximum biocatalytic current density. The highest bioelectrocatalytic current density of (597±25)µAcm-2 and the highest Γ of (23.6±0.9)pmolcm-2 were obtained on BOD-GO composite having the same moderate negative charge density, but the highest kS of (79.4±4.6)s-1 was observed on BOD-GO composite having different negative charge density. This study is a solid foundation for others to consider the influence of a charge density of GO on direct bioelectrochemistry/bioelectrocatalysis of other redox enzymes applicable for construction of biosensors, bioanodes, biocathodes or biofuel cells.

Composite films prepared from agricultural by-products.

In our study we used holocelluloses from sugar beet and bagasse for film preparation. Films from sugar beet holocellulose have better mechanical properties than from bagasse holocellulose. By subsequent carboxymethylation of bagasse holocellulose, films with better properties were produced. Specimens prepared from combined sugar beet and bagasse carboxymethylated holocellulose had the best mechanical properties. The results could be explained by the ratios of cellulose, arabinan, polygalacturonan and xylan content in individual films, based on the elemental analysis data. The use of microwaves to prepare holocellulose film speed up the process, but negatively affected the mechanical properties. Lignin content of the sugar beet holocellulose and bagasse samples was low and did not affect the mechanical properties. Both types of agricultural by-products could be used for preparation of composite film with high strength and stiffness suitable for broad range of applications.

Nanotechnology in Glycomics: Applications in Diagnostics, Therapy, Imaging, and Separation Processes.

This review comprehensively covers the most recent achievements (from 2013) in the successful integration of nanomaterials in the field of glycomics. The first part of the paper addresses the beneficial properties of nanomaterials for the construction of biosensors, bioanalytical devices, and protocols for the detection of various analytes, including viruses and whole cells, together with their key characteristics. The second part of the review focuses on the application of nanomaterials integrated with glycans for various biomedical applications, that is, vaccines against viral and bacterial infections and cancer cells, as therapeutic agents, for in vivo imaging and nuclear magnetic resonance imaging, and for selective drug delivery. The final part of the review describes various ways in which glycan enrichment can be effectively done using nanomaterials, molecularly imprinted polymers with polymer thickness controlled at the nanoscale, with a subsequent analysis of glycans by mass spectrometry. A short section describing an active glycoprofiling by microengines (microrockets) is covered as well.

Simple, Reversible, and Fast Modulation in Superwettability, Gradient, and Adsorption by Counterion Exchange on Self-Assembled Monolayer.

A simple fabrication method for preparation of surfaces able to switch from superhydrophobic to superhydrophilic state in a reversible and fast way is described. A self-assembled monolayer (SAM) consisting of quaternary ammonium group with aliphatic tail bearing terminal thiol functionality was created on gold nano/microstructured and gold planar surfaces, respectively. A rough nano/microstructured surface was prepared by galvanic reaction on a silicon wafer. The reversible counterion exchange on the rough surface resulted in a switchable contact angle between <5° and 151°. The prewetted rough surface with Cl(-) as a counterion possesses a superoleophobic underwater character. The kinetics of counterion exchanges suggests a long hydration process and strong electron ion pairing between quaternary ammonium group and perfluorooctanoate counterion. Moreover, a wettability gradient from superhydrophobic to superhydrophilic can be formed on the modified rough gold surface in a robust and simple way by passive incubation of the substrate in a counterion solution and controlled by ionic strength. Furthermore, adsorption of gold nanoparticles to modified plain gold surface can be controlled to a high extent by counterions present on the SAM layer.

Biophysical properties of carboxymethyl derivatives of mannan and dextran.

Mannan from Candida albicans, dextran from Leuconostoc spp. and their carboxymethyl (CM)-derivatives were tested on antioxidant and thrombolytic activities. As antioxidant tests, protection of liposomes against OH radicals and reducing power assay were used. Dextran and mannan protected liposomes in dose-dependent manner. Carboxymethylation significantly increased antioxidant properties of both CM-derivatives up to concentration of 10mg/mL, higher concentrations did not change the protection of liposomes. The reducing power of CM-mannan (DS 0.92) was significantly lower (P<0.05) than underivatized mannan. No reductive activity was found for dextran and CM-dextran. All CM-derivatives demonstrated statistically significant increasing activity compared with underivatized polysaccharides. The highest thrombolytic activity was found using CM-mannan (DS 0.92). The clot lysis here amounted to 68.78 ± 6.52% compared with 0.9% NaCl control (18.3 ± 6.3%). Three-dimensional surface profiles of mannan, dextran, and their CM-derivatives were compared by atomic force microscopy.

Carboxybetaine Modified Interface for Electrochemical Glycoprofiling of Antibodies Isolated from Human Serum.

Impedimetric lectin biosensors capable of recognizing two different carbohydrates (galactose and sialic acid) in glycans attached to antibodies isolated from human serum were prepared. The first step entailed the modification of a gold surface by a self-assembled monolayer (SAM) deposited from a solution containing a carboxybetaine-terminated thiol applied to the subsequent covalent immobilization of lectins and to resist nonspecific protein adsorption. In the next step, Sambucus nigra agglutinin (SNA) or Ricinus communis agglutinin (RCA) was covalently attached to the SAM, and the whole process of building a bioreceptive layer was optimized and characterized using a diverse range of techniques including electrochemical impedance spectroscopy, cyclic voltammetry, quartz crystal microbalance, contact angle measurements, zeta-potential assays, X-ray photoelectron spectroscopy, and atomic force microscopy. In addition, the application of the SNA-based lectin biosensor in the glycoprofiling of antibodies isolated from the human sera of healthy individuals and of patients suffering from rheumatoid arthritis (RA) was successfully validated using an SNA-based lectin microarray. The results showed that the SNA lectin, in particular, is capable of discriminating between the antibodies isolated from healthy individuals and those from RA patients based on changes in the amount of sialic acid present in the antibodies. In addition, the results obtained by the application of RCA and SNA biosensors indicate that the abundance of galactose and sialic acid in antibodies isolated from healthy individuals is age-related.

Graphene as a signal amplifier for preparation of ultrasensitive electrochemical biosensors.

Early diagnostics of diseases performed with minimal money and time consumption has become achievable due to recent advances in development of biosensors. These devices use biorecognition elements for selective interaction with an analyte and signal readout is obtained via different types of transducers. Operational characteristics of biosensors have been reported to improve substantially, when a diverse range of nanomaterials was employed. This review presents construction of electrochemical biosensors based on graphene, atomically thin 2D carbon crystals, which is currently intensively studied nanomaterial. The most attractive directions of graphene applications in biosensor preparation are discussed here including novel detection and amplification schemes exploiting graphene's unique electrochemical, physical and chemical properties. The future of graphene-based biosensors is most likely bright, but there is still a lot of work to do to fulfill high expectations.

Glycoprofiling as a novel tool in serological assays of systemic sclerosis: a comparative study with three bioanalytical methods.

Systemic sclerosis (SSc) is an autoimmune disease seriously affecting patient's quality of life. The heterogeneity of the disease also means that identification and subsequent validation of biomarkers of the disease is quite challenging. A fully validated single biomarker for diagnosis, prognosis, disease activity and assessment of response to therapy is not yet available. The main aim of this study was to apply an alternative assay protocol to the immunoassay-based analysis of this disease by employment of sialic acid recognizing lectin Sambucus nigra agglutinin (SNA) to glycoprofile serum samples. To our best knowledge this is the first study describing direct lectin-based glycoprofiling of serum SSc samples. Three different analytical methods for glycoprofiling of serum samples relying on application of lectins are compared here from a bioanalytical point of view including traditional ELISA-like lectin-based method (ELLA), novel fluorescent lectin microarrays and ultrasensitive impedimetric lectin biosensors. Results obtained by all three bioanalytical methods consistently showed differences in the level of sialic acid present on glycoproteins, when serum from healthy people was compared to the one from patients having SSc. Thus, analysis of sialic acid content in human serum could be of a diagnostic value for future detection of SSc, but further work is needed to enhance selectivity of assays for example by glycoprofiling of a fraction of human serum enriched in antibodies for individual diagnostics.

Cationization of heparin for film applications.

Trimethylammonium-2-hydroxypropyl-(TMAHP) spacer was introduced into heparin (H) and the prepared films were characterized by elemental analysis, NMR, SEC-MALS, TG/DTG/DTA, AFM and mechanical tester. When quaternized at the ratio of H/NaOH/alkylating agent/H2O=0.1-1/0-2/0.1-1/50-500 mmol, H was substituted at A6 and A3 positions. The formation of double-substituted structures by substitution of free hydroxyl group of the previously introduced TMAHP substituent is evident. In the absence of NaOH (H/GTMAC/H2O=1:1:500) the most drastic decrease of M(n) to 8.639 kg/mol and M(w)/M(n) at 1.48 was observed in comparison to H (Mn=9532 g/mol with M(w)/M(n)=1.38). The film mechanical properties were better on H (E=4030 MPa; σ(b)=65 MPa; ɛ(b)=4.6%) than on quaternized specimens (E=2500-3340 MPa; σ(b)=25-40 MPa; ɛ(b)=1.7-1.8%). The AFM images did not prove relation between mechanical properties and surface shape.

The pH dependence of the cathodic peak potential of the active sites in bilirubin oxidase.

This is the first study showing pH dependence of three distinct redox sites within bilirubin oxidase (BOD) adsorbed on a nanocomposite modified electrode. The 1st redox centre with the highest redox potential Ec(1st)=404 mV vs. Ag/AgCl (614 mV vs. NHE at pH7.0) exhibited pH dependence with a slope -dEc(1st)/dpH=66(±3) mV under a non-turnover process. The 2nd redox centre with a potential Ec(2nd)=228 mV vs. Ag/AgCl (438 mV vs. NHE at pH7.0) was not dependent on pH in the absence and presence of O2. Finally, the 3rd redox site with a redox potential Ec(3rd)=92 mV vs. Ag/AgCl (302 mV vs. NHE at pH7.0) exhibited pH dependence for a cathodic process with -dEc(3rd)/dpH=70(±6) mV and for anodic process with -dEa(3rd)/dpH=73(±2) mV, respectively. Moreover, two break points for dependence of Ec(1st) or Ec(3rd) on pH were observed for the 1st (T1) site and the 3rd site assigned to involvement of two acidic amino acids (Asp105 and Glu463). A diagram of a potential difference between cathodic peaks of BOD as a dependence on pH is shown. The results obtained can be of interest for construction of biofuel cells based on BOD such as for generation of a low level of electricity from body fluids.

Analysis of ethanol in fermentation samples by a robust nanocomposite-based microbial biosensor.

A robust microbial biosensor was constructed from a bionanocomposite prepared by a direct mixing of bacterial cells of Gluconobacter oxydans and carbon nanotubes with ferricyanide employed as a mediator for enhanced sensitivity of ethanol oxidation. A successful integration of the device into flow injection analysis mode of operation provided a high sensitivity of detection of (74 ± 2.7) µA mM(-1) cm(-2), a low detection limit of 5 µM and a linear range from 10 µM up to 1 mM. A short response time of the biosensor allowed a sample throughput of 67 h(-1) at 0.3 ml min(-1). The biosensor exhibited high operational stability with a decrease in the biosensor response of 1.7% during 43 h of continuous operation. The device was used to analyse ethanol in fermentation samples with a good agreement with a HPLC method.

A hyaluronic acid dispersed carbon nanotube electrode used for a mediatorless NADH sensing and biosensing.

A biocompatible nanocomposite consisting of single-walled carbon nanotubes (CNTs) dispersed in a hyaluronic acid (HA) was investigated as a sensing platform for a mediatorless electrochemical detection of NADH. The device was characterised by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) and extensively by electrochemistry. CNT-HA bionanocomposite showed more reversible electrochemistry, higher short-term stability of NADH sensing and higher selectivity of NADH detection compared to frequently used CNT-CHI (chitosan) modified GCE. Finally the performance of the sensor modified by CNT-HA was tested in a batch and flow injection analysis (FIA) mode of operation with basic characteristics revealed. The NADH sensor exhibits a good long-term operational stability (95% of the original sensitivity after 22 h of continuous operation). Subsequently a d-sorbitol biosensor based on such a nanoscale built interface was prepared and characterised with a d-sorbitol dehydrogenase used as a biocatalyst.