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Biomedical subjects

U Wollenberger

Publications and source records attributed to U Wollenberger.

At least 19 recordsLinked to original sources
PubMed2002

GDH biosensor based off-line capillary immunoassay for alkylphenols and their ethoxylates.

The application of a quinoprotein glucose dehydrogenase modified thick-film sensor as label detector in a capillary immunoassay (CIA) for xenoestrogens is presented. The detection of the alkylphenols and their ethoxylates is based on the competition between the analyte and tracer molecules for the binding sites of anti-alkylphenol ethoxylate antibodies. This assay is performed off-line in small disposable PVC capillaries coated with immobilized antibodies. This format allows the combination of the assay with a small portable device potentially useful for on-site environmental monitoring. Beside high amplification the utilization of beta-galactosidase as enzyme label allows the direct combination with a GDH biosensor at optimal pH conditions. The bioelectrocatalytic properties of this biosensor offer an additional amplification and thus allow a very sensitive quantification of 4-aminophenol, generated by the beta-galactosidase. Detection limits of the analytes in the microg/l range were obtained, while other phenolics and surfactants showed no or very little cross reactivity.

Biosensing Techniques↗
PubMed2002

Detection of Escherichia coli in water by culture-based amperometric and luminometric methods.

The application of amperometric biosensor- and chemiluminiscence based methods for rapid detection of viable E. coli in water has been investigated. An amplification of the amperometric signal by a factor of 4 was obtained when the cellobiose dehydrogenase (CDH) biosensor was used instead of a plain graphite electrode for detection of b-galactosidase (b-GAL) activity at 22.5 degrees C. A linear correlation was demonstrated for detection time (DT) vs. initial concentrations (logarithmic units) of E. coli IT1 and E. coli in environmental samples, respectively, by use of the CDH biosensor or a chemiluminometric technique. The study has shown that an E. coli concentration > or = 10(4) cfu/100 mL in environmental samples was determined by the CDH biosensor within one working day. However, further reduction of the DT can be obtained, e.g. by increasing the signal amplification factor using other biosensors.

Biosensing Techniques↗
PubMed2001

Quinoprotein glucose dehydrogenase modified thick-film electrodes for the amperometric detection of phenolic compounds in flow injection analysis.

The use of thick-film electrodes as basic transducers for highly sensitive amperometric biosensors using PQQ (pyrroloquinoline quinone) dependent glucose dehydrogenase (GDH) with short response times is described. The enzyme is embedded in a polyurethane matrix on top of a platinum based thick film electrode and its ability to reduce oxidized phenolic compounds is exploited. The electrochemical amplification is based on the oxidation of the analyte on the surface of the electrode followed by its enzymatic reduction. Different parameters of the glucose dehydrogenase electrode system using dopamine as a model analyte were optimized, e.g., membrane thickness, pH value, buffer system, flow rate and storage conditions. Using optimized parameters the sensitivity and detection limits for various phenolic compounds were evaluated. The comparison of electrodes from the identical as well as from different batches shows the ability to produce a number of well reproducible sensors showing remarkably small differences with respect to parameters as sensitivity, response times and measuring range.

Acinetobacter calcoaceticus↗
PubMed2001

Research and development in biosensors.

Progress in biosensors has mainly been made by the improvement of the biological components and the implementation of microsystem technologies. Enzymes are still the most appropriate recognition elements because they combine high chemical specificity and inherent biocatalytic signal amplification. A breakthrough has been achieved in the application of membrane-integrated receptor systems for analyte recognition and signal transduction in biosensors. Sensor integration of RNA aptamers has been initiated, and the performance of fully synthetic molecularly imprinted polymers has been improved.

Antibodies↗
PubMed2000

Clay-bridged electron transfer between cytochrome p450(cam) and electrode.

We demonstrate a very fast heterogeneous redox reaction of substrate-free cytochrome P450(cam) on a glassy carbon electrode modified with sodium montmorillonite. The linear relationship of the peak current in the cyclic voltammogram with the scan rate indicates a reversible one-electron transfer surface process. The electron transfer rate is in the range from 5 to 152 s(-1) with scan rates from 0.4 to 12 V/s, respectively. These values are comparable to rates reported for the natural electron transfer from putidaredoxin to P450(cam). The formal potential of adsorbed P450(cam) is -139 mV (vs NHE) and therefore positively shifted by 164 mV compared to the potential of substrate-free P450(cam) in solution. UV-VIS and FTIR spectra do not indicate an influence of the clay colloidal particles on the heme and the secondary structure of P450(cam) in solution. However, P450(cam) adsorbed on the surface of the clay-modified electrode may undergo partial dehydration resulting in the shift of the formal potential.

Bentonite↗
PubMed2000

Electrochemical investigation of cellobiose oxidation by cellobiose dehydrogenase in the presence of cytochrome c as mediator.

An important aspect of the cytochrome c electrochemistry is the possibility of coupling the 'heterogeneous reactions' with other redox enzymes. Cellobiose dehydrogenase, a 89170 Da glycoprotein that contains both FAD and a b-type haem as prosthetic groups, donates electrons to a number of acceptors, including cytochrome c. While haem b is surrounded mainly by acidic amino acids, cytochrome c displays positive charged lysine groups around the haem site. Thus a fast reaction between both proteins is explicable. In the presence of cellobiose, a catalytic current was observed, owing to the interaction of cellobiose dehydrogenase with electrostatically adsorbed cytochrome c. Adsorption of cytochrome c provides a technological model surface for vectorial electron transfer.

Carbohydrate Dehydrogenases↗
PubMed1997

Catecholamine detection using enzymatic amplification.

Different amplification sensors based on the substrate recycling principle were investigated with respect to their applicability to catecholamine detection. In the bioelectrocatalytic approach, glassy carbon electrodes were modified by laccase or a PQQ-dependent glucose dehydrogenase. Substrate recycling occurs and the detection limit is in the lower nanomolar concentration range (e.g. 10 nM dopamine and 1 nM noradrenaline for the laccase- and glucose dehydrogenase-modified electrodes, respectively). Combinations of glucose dehydrogenase with laccase or tyrosinase were investigated as bienzymatic probes. Among the systems we studied, the laccase/glucose dehydrogenase sensor is the most sensitive (detection limit: 0.5 nM adrenaline). The selectivities of the different sensor systems are discussed. Application of the laccase/glucose dehydrogenase electrode in different media (i.e. brain homogenate, heart effluate) was successfully shown. For samples with high concentrations of interfering substances (uric and ascorbic acid), the interferences can be effectively removed using enzymatic methods.

Biosensing Techniques↗
PubMed1997

Phenol-oxidizing enzymes: mechanisms and applications in biosensors.

Phenolic compounds are widely distributed in nature. Enzymes which catalyze their oxidation are monophenol monooxygenases, such as tyrosinases and laccases, and peroxidases. Their metabolic role includes the decomposition of natural complex aromatic polymers as well as polymerization of the oxidation products and the degradation of xenobiotics. Their catalytic properties and broad availability gained impact on the development of biosenors for both environmentally important pollutants and clinically relevant metabolites. Mechanisms for the phenol-oxidizine enzymes tyrosinases, laccases, and peroxidases are reviewed and some examples for their use in the construction of phenol selective biosenors are given.

Biosensing Techniques↗
PubMed1997

Enzymatic substrate recycling electrodes.

A weak chemical signal might result in a large response when biochemically amplified. Enzymatic recycling of the analyte is one of the biochemical ways of providing an effective increase in biosensor sensitivity by several orders of magnitude. The enhancement of sensitivity is provided by consecutive consumption and generation of the analyte on the sensor surface. The principle of enzymatic substrate regeneration using bioelectrocatalysis and coupled enzymes is shortly reviewed and illustrated with some recent developments of biosensors for catecholamines, and its potential for electrochemical immunoassays is outlined.

Biosensing Techniques↗
PubMed1997

Ultrasensitive bienzyme sensor for adrenaline.

A biosensor consisting of an analyte-recycling two-enzyme system using laccase (Coriolus hirsutus) and PQQ-dependent glucose dehydrogenase in combination with the electrochemical detection of oxygen depletion at a platinum electrode was used for adrenaline determination in the nano- and subnanomolar concentration range. Measurements were performed in a flow cell providing excellent baseline stability and fast recovery of the sensor. Improved design of the polymer matrix resulted in a lower detection limit of 200 pmol/l for adrenaline. The sensor has successfully been applied to the analysis of adrenaline in effluate of isolated rabbit hearts.

Animals↗
PubMed1993

Enhancing biosensor performance using multienzyme systems.

Enhancing the performance of biosensors, in terms of increasing the range of analytes that may be detected, and the sensitivity and specificity of the detection event, would improve the prospects for commercializing this technology. Coupling the catalytic activities of several enzymes is one approach being used to address these issues. Sequences of enzymes, where ligand binding triggers the activation of enzymes, or where biocatalytic pre-concentration of intermediates permits augmentation of the signal, may be used. In addition, enzymatic recycling of the analyte can be used to increase the sensitivity by several orders of magnitude.

Animals↗
PubMed1993

Enzyme activation for activator and enzyme activity measurement.

A new sensing principle of enzyme activation is demonstrated for the determination of glycogen phosphorylase b and its allosteric effector AMP. As the indicator of the phosphorylase catalysed glycogen phosphorolysis, glucose-1-phosphate formation has been detected with an enzyme sequence comprising coentrapped alkaline phosphatase, mutarotase and glucose oxidase on a hydrogen peroxide indicating electrode. The optimized three-enzyme sensor was useful for the determination of 0.005-0.2 U.ml-1 glycogen phosphorylase a and b. A biosensor for AMP and inorganic phosphate has been developed by coupling glycogen entrapped phosphorylases to the three-enzyme indicator membrane. The measurement of AMP is based on the modulation of the phosphorylase b catalysed glycogen phosphorylating activity. The proposed sensor responds to AMP between 5 and 150 microM. The calibration graph of the reagentless phosphate sensor is linear between 0.05 and 1 mM.

Adenosine Monophosphate↗
PubMed1992

A lysine dehydrogenase-based electrode for biosensing of L-lysine.

An amperometric biosensor for L-lysine based on the recently isolated enzyme lysine dehydrogenase is described. Immobilization of the enzyme onto a platinum electrode is achieved via entrapment within a gelatin support on a cellulose membrane. Anodic detection (at 0.4 V vs. Ag/AgCl) is facilitated by the presence of a redox-mediating ferricyanide ion. The effect of experimental variables such as pH, enzyme loading, applied potential, cofactor and mediator concentrations were evaluated in order to optimize the analytical performance. A detection limit of 7 x 10(-8) M, and linearity up to 7 x 10(-4) M are reported. The fast response permits adaptation for flow injection operation with good precision (RSD = 1.9%) and high sample throughout (40 samples per hour). The high specificity offered by this new enzyme is indicated by the lack of interference by other L-amino acids, alcohols or carbohydrates.

Amino Acid Oxidoreductases↗
PubMed1991

Enzyme sensor-FIA-system for on-line monitoring of glucose, lactate and glutamine in animal cell cultures.

Enzyme sensors for glucose, lactate and glutamine were connected via flow-injection analysis (FIA) devices to two different bioprocesses. They were used for on-line process control of perfused bioreactor systems containing mammalian cell lines producing a monoclonal antibody and recombinant interleukin-2. The biosensor system gives direct access to important process data which can be used as control parameters for long term cell cultivation systems.

Animals↗
PubMed1991

Second generation biosensors.

Enzyme-membrane electrodes using glucose oxidase in combination with peroxide detection dominate in the field of laboratory analyzers for diluted samples. Using the same indication principle, extremely fast responding glucose sensors have been fabricated by covering thin metal electrodes with a porous enzyme layer. In the second generation auxiliary enzymes and/or co-reactants are coimmobilized with the analyte converting enzyme in order to improve the analytical quality and to simplify the performance. Following this line oxidizable interferences are suppressed by using a glucose oxidase/peroxidase complex which communicates with the electrode at a low working potential. Furthermore, fluctuations of pH or buffer capacity are ineffective when using a glucose oxidase/peroxidase layer covered fluoride FET in the potentiometric glucose determination. Enzymatic recycling of the analyte and/or accumulation of intermediates increase the sensitivity by several orders of magnitude. Inclusion of NAD bound to PEG in the glucose dehydrogenase layer allows a reagentless glucose measurement.

Biosensing Techniques↗