TY - JOUR A1 - Beissenhirtz, Moritz Karl T1 - Electrochemical quartz crystal microbalance studies on cytochrome c/polyelectrode multilayer assemblies on gold electrodes N2 - Polyelectrolyte multilayer assemblies containing proteins are of interest for applications such as sensors, bioreactors, and bioelectronics. A multilayer electrode was built up by the layer-by-layer strategy consisting of alternating layers of cytochrome c and poly(aniline sulfonic acid). The electrode showed a linear increase of redox active protein with the number of deposited layers. The principle of electrode preparation was transferred from needle electrodes to planar surfaces in order to further the understanding of electron transfer through the layer assembly by means of electrochemical quartz crystal microbalance studies. The deposition process was followed on-line by detection of the frequency shift of the crystals and was found to be rather fast (minutes). The total mass deposited was found to correlate well with the electrochemical response of the immobilized cyt.c. Furthermore, the influence of the polyelectrolyte was investigated by addition of PSS to the PASA solution. The strong interaction of the former polyelectrolyte seemed to hinder the electron transfer although a multilayer formation was proved. Dilution of the protein solution with redox inactive apo-cyt.c led to a strong decrease of the voltammetric signal, well beyond the percentage of apo-cyt.c inside the assembly. Thus, arguments for an electron transfer via protein-protein interaction were found Y1 - 2005 ER - TY - JOUR A1 - Beissenhirtz, Moritz Karl A1 - Kwan, R. C. H. A1 - Ko, K. M. A1 - Renneberg, Reinhard A1 - Scheller, Frieder W. A1 - Lisdat, Fred T1 - Comparing in vitro electrochemical measurement of superoxide scavenging activity with an in vivo assessment of antioxidant potential in Chinese tonifying herbs N2 - The in vitro superoxide scavenging activity (as determined by electrochemical measurement) and the in vivo antioxidant potential (as determined by a mouse model of carbon tetrachloride (CCl4) hepatotoxicity) of methanolic extracts prepared from 10 Chinese tonifying herbs were compared. Electrochemical measurement using a cytochrome c (Cyt. c) sensor showed that all of the tested herbal extracts exhibited a medium superoxide scavenging activity of different potency, as indicated by their IC50 values. The in vivo measurement demonstrated that 80% of the herbal extracts displayed in vivo antioxidant potential, as assessed by the percentage of protection of the activity of plasma alanine aminotransferases and the hepatic glutathione regeneration capacity under CCl4-intoxicated condition. Although the in vitro antioxidant activity did not correlate quantitatively with the in vivo antioxidant potential, for 8 out of 10 samples a similar tendency was found. The rapid amperometric assessment of antioxidant potential by Cyt. c sensor may offer a convenient and direct method for screening as well as the quality control of herbal products. Copyright (C) 2004 John Wiley Sons, Ltd Y1 - 2004 ER - TY - JOUR A1 - Beissenhirtz, Moritz Karl A1 - Scheller, Frieder W. A1 - Lisdat, Fred T1 - A superoxide sensor based on a multilayer cytochrome c electrode N2 - A novel multilayer cytochrome c electrode for the quantification of superoxide radical concentrations is introduced. The electrode consists of alternating layers of cytochrome c and poly(aniline(sulfonic acid)) on a gold wire electrode. The formation of multilayer structures was proven by SPR experiments. Assemblies with 2-15 protein layers showed electrochemical communication with the gold electrode. For every additional layer, a substantial increase in electrochemically active cytochrome c (cyt. c) was found. For electrodes of more than 10 layers, the increase was more than 1 order of magnitude as compared to monolayer electrode systems. Thermodynamic and kinetic parameters of the electrodes were characterized. The mechanism of electron transfer within the multilayer assembly was studied, with results suggesting a protein-protein electron-transfer model. Electrodes of 2-15 layers were applied to the in vitro quantification of enzymatically generated superoxide, showing superior sensitivity as compared to a monolayer-based sensor. An electrode with 6 cyt. c/PASA layers showed the highest sensitivity of the systems studied, giving an increase in sensitivity of half an order of magnitude versus the that of the monolayer electrode. The stability of the system was optimized using thermal treatment, resulting in no loss in sensor signal or protein loading after 10 successive measurements or 2 days of storage Y1 - 2004 SN - 0003-2700 ER - TY - JOUR A1 - Beissenhirtz, Moritz Karl A1 - Scheller, Frieder W. A1 - Lisdat, Fred T1 - Immobilized cytochrome c sensor in organic / aqueous media for the characterization of hydrophilic and hydrophobic antioxidants Y1 - 2003 ER - TY - JOUR A1 - Beissenhirtz, Moritz Karl A1 - Scheller, Frieder W. A1 - Stöcklein, Walter F. M. A1 - Kurth, D. A1 - Möhwald, Helmuth A1 - Lisdat, Fred T1 - Electroactive cytochrome c multilayers within a polyelectrolyte assembly Y1 - 2004 ER - TY - JOUR A1 - Beissenhirtz, Moritz Karl A1 - Scheller, Frieder W. A1 - Viezzoli, Maria Silvia A1 - Lisdat, Fred T1 - Engineered superoxide dismutase monomers for superoxide biosensor applications N2 - Because of its high reaction rate and specificity, the enzyme superoxide dismutase (SOD) offers great potential for the sensitive quantification of superoxide radicals in electrochemical biosensors. In this work, monomeric mutants of human Cu,Zn-SOD were engineered to contain one or two additional cysteine residues, which could be used to bind the protein to gold surfaces, thus making the use of promotor molecules unnecessary. Six mutants were successfully designed, expressed, and purified. All mutants bound directly to unmodified gold surfaces via the sulfur of the cysteine residues and showed a quasireversible, direct electron transfer to the electrode. Thermodynamic and kinetic parameters of the electron transfer were characterized and showed only slight variations between the individual mutants. For one of the mutants, the interaction with the superoxide radical was studied in more detail. For both partial reactions of the dismutation, an interaction between protein and radical could be shown. In an amperometric biosensorial approach, the SOD-mutant electrode was successfully applied for the detection of superoxide radicals. In the oxidation region, the electrode surpassed the sensitivity of the commonly used cytochrome c electrodes by similar to 1 order of magnitude while not being limited by interferences, but the electrode did not fully reach the sensitivity of dimeric Cu,Zn-SOD immobilized on MPA-modified gold Y1 - 2006 UR - http://pubs.acs.org/journal/ancham U6 - https://doi.org/10.1021/Ac051465g SN - 0003-2700 ER - TY - JOUR A1 - Büttemeyer, R. A1 - Philipp, A. W A1 - Schlenzka, L. A1 - Mall, J. W. A1 - Beissenhirtz, Moritz Karl A1 - Lisdat, Fred T1 - Epigallocatechin gallate can significantly decrease free oxygen radicals in the reperfusion insury in vivo Y1 - 2003 ER - TY - JOUR A1 - Kapp, A. A1 - Beissenhirtz, Moritz Karl A1 - Geyer, F. A1 - Scheller, F. A1 - Viezzoli, Maria Silvia A1 - Lisdat, Fred T1 - Electrochemical and sensorial behavior of SOD mutants immobilized on gold electrodes in aqueous/organic solvent mixtures JF - Electroanalysis : an international journal devoted to fundamental and practical aspects of electroanalysis N2 - A cysteine mutant of a monomeric human Cu, Zn-SOD (Glycine 61, Serine 142) has been immobilized directly on gold electrodes using the thiol groups introduced. The electrochemical behavior of the surface confined protein was studied in mixtures of aqueous buffer and DMSO up to an organic solvent content of 60%. The formal potential was found to be rather independent of the DMSO content. However, half peak width increased and the redoxactive amount clearly decreased with raising DMSO content. In addition, the kinetics of the heterogeneous electron transfer became slower; but still a quasireversible electrochemical conversion of the mutant SOD was feasible. Thus, the electrodes were applied for sensorial superoxide detection. At a potential of +220 mV vs. Ag/AgCl advantage was taken of the partial oxidation reaction of the enzyme. A defined superoxide signal was obtained in solutions up to 40% DMSO. The sensitivity of the mutant electrodes decreased linearly with the organic solvent content in solution but was still higher compared to conventional cyt.c based sensors. At DMSO concentrations higher than 40% no sensor response was detected. KW - SOD KW - mutants KW - gold electrodes KW - DMSO KW - electrochemistry Y1 - 2006 U6 - https://doi.org/10.1002/elan.200603620 SN - 1040-0397 VL - 18 SP - 1909 EP - 1915 PB - Wiley CY - Weinheim ER - TY - JOUR A1 - Kapp, Andreas A1 - Beissenhirtz, Moritz Karl A1 - Geyer, F. A1 - Scheller, Frieder W. A1 - Viezzoli, Maria Silvia A1 - Lisdat, Fred T1 - Electrochemical and sensorial behaviour of SOD mutants immobilized on gold electrodes in aqueous / organic solvent mixtures Y1 - 2006 UR - http://www3.interscience.wiley.com/cgi-bin/jhome/26571/ U6 - https://doi.org/10.1002/elan.200603620 SN - 1040-0397 ER - TY - JOUR A1 - Krylov, Andrey V. A1 - Beissenhirtz, Moritz Karl A1 - Adamzig, Holger A1 - Scheller, Frieder W. A1 - Lisdat, Fred T1 - Thick-film electrodes for measurement of superoxide and hydrogen peroxide based on direct protein-electrode contacts N2 - Cytochrome c was immobilized on screen-printed thick-film gold electrodes by a self-assembly approach using mixed monolayers of mercaptoundecanoic acid and mercaptoundecanol. Cyclic voltammetry revealed quasi-reversible electrochemical behavior of the covalently fixed protein with a formal potential of +10 mV vs. Ag/AgCl. Polarized at +150 mV vs. Ag/AgCl the electrode was found to be sensitive to superoxide radicals in the range 300-1200 nmol L-1. Compared with metal needle electrodes sensitivity and reproducibility could be improved and combined with the easiness of preparation. This allows the fabrication of disposable sensors for nanomolar superoxide concentrations. By changing the electrode potential the sensor can be switched from response to superoxide radicals to hydrogen peroxide-another reactive oxygen species. H2O2 sensitivity can be provided in the range 10-1000 mumol L-1 which makes the electrode suitable for oxidative stress studies Y1 - 2004 ER -