Fachbereich Medizintechnik und Technomathematik
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Das Diskussionspapier beschreibt einen Prozess an der FH Aachen zur Entwicklung und Implementierung eines Self-Assessment-Tools für Studiengänge. Dieser Prozess zielte darauf ab, die Relevanz der Themen Digitalisierung, Internationalisierung und Nachhaltigkeit in Studiengängen zu stärken. Durch Workshops und kollaborative Entwicklung mit Studiendekan:innen entstand ein Fragebogen, der zur Reflexion und strategischen Weiterentwicklung der Studiengänge dient.
An acetoin biosensor based on a capacitive electrolyte–insulator–semiconductor (EIS) structure modified with the enzyme acetoin reductase, also known as butane-2,3-diol dehydrogenase (Bacillus clausii DSM 8716ᵀ), is applied for acetoin detection in beer, red wine, and fermentation broth samples for the first time. The EIS sensor consists of an Al/p-Si/SiO₂/Ta₂O₅ layer structure with immobilized acetoin reductase on top of the Ta₂O₅ transducer layer by means of crosslinking via glutaraldehyde. The unmodified and enzyme-modified sensors are electrochemically characterized by means of leakage current, capacitance–voltage, and constant capacitance methods, respectively.
The enantioselective synthesis of α-hydroxy ketones and vicinal diols is an intriguing field because of the broad applicability of these molecules. Although, butandiol dehydrogenases are known to play a key role in the production of 2,3-butandiol, their potential as biocatalysts is still not well studied. Here, we investigate the biocatalytic properties of the meso-butanediol dehydrogenase from Bacillus licheniformis DSM 13T (BlBDH). The encoding gene was cloned with an N-terminal StrepII-tag and recombinantly overexpressed in E. coli. BlBDH is highly active towards several non-physiological diketones and α-hydroxyketones with varying aliphatic chain lengths or even containing phenyl moieties. By adjusting the reaction parameters in biotransformations the formation of either the α-hydroxyketone intermediate or the diol can be controlled.
α-hydroxy ketones (HK) and 1,2-diols are important building blocks for fine chemical synthesis. Here, we describe the R-selective 2,3-butanediol dehydrogenase from B. clausii DSM 8716ᵀ (BcBDH) that belongs to the metal-dependent medium chain dehydrogenases/reductases family (MDR) and catalyzes the selective asymmetric reduction of prochiral 1,2-diketones to the corresponding HK and, in some cases, the reduction of the same to the corresponding 1,2-diols. Aliphatic diketones, like 2,3-pentanedione, 2,3-hexanedione, 5-methyl-2,3-hexanedione, 3,4-hexanedione and 2,3-heptanedione are well transformed. In addition, surprisingly alkyl phenyl dicarbonyls, like 2-hydroxy-1-phenylpropan-1-one and phenylglyoxal are accepted, whereas their derivatives with two phenyl groups are not substrates. Supplementation of Mn²⁺ (1 mM) increases BcBDH's activity in biotransformations. Furthermore, the biocatalytic reduction of 5-methyl-2,3-hexanedione to mainly 5-methyl-3-hydroxy-2-hexanone with only small amounts of 5-methyl-2-hydroxy-3-hexanone within an enzyme membrane reactor is demonstrated.
Monitoring of organic acids (OA) and volatile fatty acids (VFA) is crucial for the control of anaerobic digestion. In case of unstable process conditions, an accumulation of these intermediates occurs. In the present work, two different enzyme-based biosensor arrays are combined and presented for facile electrochemical determination of several process-relevant analytes. Each biosensor utilizes a platinum sensor chip (14 × 14 mm²) with five individual working electrodes. The OA biosensor enables simultaneous measurement of ethanol, formate, d- and l-lactate, based on a bi-enzymatic detection principle. The second VFA biosensor provides an amperometric platform for quantification of acetate and propionate, mediated by oxidation of hydrogen peroxide. The cross-sensitivity of both biosensors toward potential interferents, typically present in fermentation samples, was investigated. The potential for practical application in complex media was successfully demonstrated in spiked sludge samples collected from three different biogas plants. Thereby, the results obtained by both of the biosensors were in good agreement to the applied reference measurements by photometry and gas chromatography, respectively. The proposed hybrid biosensor system was also used for long-term monitoring of a lab-scale biogas reactor (0.01 m³) for a period of 2 months. In combination with typically monitored parameters, such as gas quality, pH and FOS/TAC (volatile organic acids/total anorganic carbonate), the amperometric measurements of OA and VFA concentration could enhance the understanding of ongoing fermentation processes.
Multi-analyte biosensors may offer the opportunity to perform cost-effective and rapid analysis with reduced sample volume, as compared to electrochemical biosensing of each analyte individually. This work describes the development of an enzyme-based biosensor system for multi-parametric determination of four different organic acids. The biosensor array comprises five working electrodes for simultaneous sensing of ethanol, formate, d-lactate, and l-lactate, and an integrated counter electrode. Storage stability of the biosensor was evaluated under different conditions (stored at +4 °C in buffer solution and dry at −21 °C, +4 °C, and room temperature) over a period of 140 days. After repeated and regular application, the individual sensing electrodes exhibited the best stability when stored at −21 °C. Furthermore, measurements in silage samples (maize and sugarcane silage) were conducted with the portable biosensor system. Comparison with a conventional photometric technique demonstrated successful employment for rapid monitoring of complex media.
A capacitive electrolyte-insulator-semiconductor (EIS) field-effect biosensor for acetoin detection has been presented for the first time. The EIS sensor consists of a layer structure of Al/p-Si/SiO₂/Ta₂O₅/enzyme acetoin reductase. The enzyme, also referred to as butane-2,3-diol dehydrogenase from B. clausii DSM 8716T, has been recently characterized. The enzyme catalyzes the (R)-specific reduction of racemic acetoin to (R,R)- and meso-butane-2,3-diol, respectively. Two different enzyme immobilization strategies (cross-linking by using glutaraldehyde and adsorption) have been studied. Typical biosensor parameters such as optimal pH working range, sensitivity, hysteresis, linear concentration range and long-term stability have been examined by means of constant-capacitance (ConCap) mode measurements. Furthermore, preliminary experiments have been successfully carried out for the detection of acetoin in diluted white wine samples.
Reconstructive surgery and tissue replacements like ureters or bladders reconstruction have been recently studied, taking into account growth and remodelling of cells since living cells are capable of growing, adapting, remodelling or degrading and restoring in order to deform and respond to stimuli. Hence, shapes of ureters or bladders and their microstructure change during growth and these changes strongly depend on external stimuli such as training. We present the mechanical stimulation of smooth muscle cells in a tubular fibrin-PVDFA scaffold and the modelling of the growth of tissue by stimuli. To this end, mechanotransduction was performed with a kyphoplasty balloon catheter that was guided through the lumen of the tubular structure. The bursting pressure was examined to compare the stability of the incubated tissue constructs. The results showed the significant changes on tissues with training by increasing the burst pressure as a characteristic mechanical property and the smooth muscle cells were more oriented with uniformly higher density. Besides, the computational growth models also exhibited the accurate tendencies of growth of the cells under different external stimuli. Such models may lead to design standards for the better layered tissue structure in reconstructing of tubular organs characterized as composite materials such as intestines, ureters and arteries.
The immobilization of NAD+-dependent dehydrogenases, in combination with a diaphorase, enables the facile development of multiparametric sensing devices. In this work, an amperometric biosensor array for simultaneous determination of ethanol, formate, d- and l-lactate is presented. Enzyme immobilization on platinum thin-film electrodes was realized by chemical cross-linking with glutaraldehyde. The optimization of the sensor performance was investigated with regard to enzyme loading, glutaraldehyde concentration, pH, cofactor concentration and temperature. Under optimal working conditions (potassium phosphate buffer with pH 7.5, 2.5 mmol L-1 NAD+, 2.0 mmol L-1 ferricyanide, 25 °C and 0.4% glutaraldehyde) the linear working range and sensitivity of the four sensor elements was improved. Simultaneous and cross-talk free measurements of four different metabolic parameters were performed successfully. The reliable analytical performance of the biosensor array was demonstrated by application in a clarified sample of inoculum sludge. Thereby, a promising approach for on-site monitoring of fermentation processes is provided.
Three amperometric biosensors have been developed for the detection of L-malic acid, fumaric acid, and L -aspartic acid, all based on the combination of a malate-specific dehydrogenase (MDH, EC 1.1.1.37) and diaphorase (DIA, EC 1.8.1.4). The stepwise expansion of the malate platform with the enzymes fumarate hydratase (FH, EC 4.2.1.2) and aspartate ammonia-lyase (ASPA, EC 4.3.1.1) resulted in multi-enzyme reaction cascades and, thus, augmentation of the substrate spectrum of the sensors. Electrochemical measurements were carried out in presence of the cofactor β-nicotinamide adenine dinucleotide (NAD+) and the redox mediator hexacyanoferrate (III) (HCFIII). The amperometric detection is mediated by oxidation of hexacyanoferrate (II) (HCFII) at an applied potential of + 0.3 V vs. Ag/AgCl. For each biosensor, optimum working conditions were defined by adjustment of cofactor concentrations, buffer pH, and immobilization procedure. Under these improved conditions, amperometric responses were linear up to 3.0 mM for L-malate and fumarate, respectively, with a corresponding sensitivity of 0.7 μA mM−1 (L-malate biosensor) and 0.4 μA mM−1 (fumarate biosensor). The L-aspartate detection system displayed a linear range of 1.0–10.0 mM with a sensitivity of 0.09 μA mM−1. The sensor characteristics suggest that the developed platform provides a promising method for the detection and differentiation of the three substrates.
The gene encoding a putative (R,R)-butane-2,3-diol dehydrogenase (bdhA) from Bacillus clausii DSM 8716T was isolated, sequenced and expressed in Escherichia coli. The amino acid sequence of the encoded protein is only distantly related to previously studied enzymes (identity 33–43%) and exhibited some uncharted peculiarities. An N-terminally StrepII-tagged enzyme variant was purified and initially characterized. The isolated enzyme catalyzed the (R)-specific oxidation of (R,R)- and meso-butane-2,3-diol to (R)- and (S)-acetoin with specific activities of 12 U/mg and 23 U/mg, respectively. Likewise, racemic acetoin was reduced with a specific activity of up to 115 U/mg yielding a mixture of (R,R)- and meso-butane-2,3-diol, while the enzyme reduced butane-2,3-dione (Vmax 74 U/mg) solely to (R,R)-butane-2,3-diol via (R)-acetoin. For these reactions only activity with the co-substrates NADH/NAD+ was observed. The enzyme accepted a selection of vicinal diketones, α-hydroxy ketones and vicinal diols as alternative substrates. Although the physiological function of the enzyme in B. clausii remains elusive, the data presented herein clearly demonstrates that the encoded enzyme is a genuine (R,R)-butane-2,3-diol dehydrogenase with potential for applications in biocatalysis and sensor development.
An amperometric biosensor using a substrate recycling principle was realized for the detection of low adrenaline concentrations (1 nM) by measurements in phosphate buffer and Ringer’s solution at pH 6.5 and pH 7.4, respectively. In proof-of-concept experiments, a Boolean logic-gate principle has been applied to develop a digital adrenaline biosensor based on an enzyme AND logic gate. The obtained results demonstrate that the developed digital biosensor is capable for a rapid qualitative determination of the presence/absence of adrenaline in a YES/NO statement. Such digital biosensor could be used in clinical diagnostics for the control of a correct insertion of a catheter in the adrenal veins during adrenal venous-sampling procedure.
Abstractauthoren Graphene oxide (GO) nanoparticles were incorporated in temperature-sensitive Poly(N-isopropylacrylamide) (PNIPAAm) hydrogels. The nanoparticles increase the light absorption and convert light energy into heat efficiently. Thus, the hydrogels with GO can be stimulated spatially resolved by illumination as it was demonstrated by IR thermography. The temporal progression of the temperature maximum was detected for different concentrations of GO within the polymer network. Furthermore, the compatibility of PNIPAAm hydrogels with GO and cell cultures was investigated. For this purpose, culture medium was incubated with hydrogels containing GO and the viability and morphology of chinese hamster ovary (CHO) cells was examined after several days of culturing in presence of this medium.
Light-stimulated hydrogel actuators with incorporated graphene oxide for microfluidic applications
(2015)
An amperometric enzyme biosensor has been applied for the detection of adrenaline. The adrenaline biosensor has been prepared by modification of an oxygen electrode with the enzyme laccase that operates at a broad pH range between pH 3.5 to pH 8. The enzyme molecules were immobilized via cross-linking with glutaraldehyde. The sensitivity of the developed adrenaline biosensor in different pH buffer solutions has been studied.
A new microfluidic assembly method for semiconductor-based biosensors using 3D-printing technologies was proposed for a rapid and cost-efficient design of new sensor systems. The microfluidic unit is designed and printed by a 3D-printer in just a few hours and assembled on a light-addressable potentiometric sensor (LAPS) chip using a photo resin. The cell growth curves obtained from culturing cells within microfluidics-based LAPS systems were compared with cell growth curves in cell culture flasks to examine biocompatibility of the 3D-printed chips. Furthermore, an optimal cell culturing within microfluidics-based LAPS chips was achieved by adjusting the fetal calf serum concentrations of the cell culture medium, an important factor for the cell proliferation.