Dilek Odaci

Biosensing approach for glutathione detection using glutathione reductase and sulfhydryl oxidase bienzymatic system

Chitosan membrane with glutathione reductase and sulfhydryl oxidase (SOX) was subsequently integrated onto the surface of spectrographic graphite rods for obtaining a glutathione biosensor. The working principle was based on the monitoring of O(2) consumption that correlates the concentration of glutathione during the enzymatic reaction. A linear relationship between sensor response and concentration was obtained between 0.5 and 2.0 mM for oxidized glutathione (GSSG), and 0.2-1.0 mM for reduced glutathione (GSH) in the presence of 2 microM nicotinamide adenine dinucleotide phosphate (NADPH) under the optimum working conditions. Also, reduced/oxidized glutathione were separated by HPLC and utility of bienzymatic system was investigated as an electrochemical detector for the analysis of these compounds. All data were given as a comparison of two systems: biosensor and diode array detector (DAD).

A microbial biosensor based on bacterial cells immobilized on chitosan matrix

A bio-electrochemical system consisting of Gluconobacter oxydans DSM 2343 cells as a biological material and carbon nanotube (CNT)-free and CNT-modified chitosan as immobilizing matrices has been developed. The measurement was based on the respiratory activity of the cells estimated by the oxygen consumption at -0.7 V (versus the Ag|AgCl reference electrode) due to the metabolic activity in the presence of substrates. The system was calibrated and dependence of signal amplitude on the measuring conditions and cell amount was studied as well as the substrate specificity, pH, temperature and working potential. The biosensors (CNT-modified and unmodified) were demonstrated for the quantification of glucose in the range of 0.05-1.0 mM, at 30 degrees C and pH 7.0 with the 40 s of response time. The linear relationships between sensor response (y; microA/cm(2)) and substrate concentration (x; mM) were defined by the equations of y=1.160x+0.151 (R(2)=0.990) and y=1.261x+0.197 (R(2)=0.982), respectively. All other data were also given as comparison of two systems one with CNT-modified and CNT-free.

Determination of phenolic acids using Trametes versicolor laccase

Two biosensors based on Trametes versicolor laccase (TvL) were developed for the determination of phenolic compounds. Commercial oxygen electrode and ferrocene-modified screen-printed graphite electrodes were used for preparation of laccase biosensors. The systems were calibrated for three phenolic acids. Linearity was obtained in the concentration range 0.1-1.0muM caffeic acid, 0.05-0.2muM ferulic acid, 2.0-14.0muM syringic acid for laccase immobilised on a commercial oxygen electrode and 2.0-30.0muM caffeic acid, 2.0-10.0muM ferulic acid, 4.0-30.0muM syringic acid for laccase immobilised on ferrocene-modified screen-printed electrodes. Furthermore, optimal pH, temperature and thermal stability studies were performed with the commercial oxygen electrode. Both electrodes were used for determination of a class of phenolic acids, achieving a cheap and fast tool and an easy to be used procedure for screening real samples of human plasma.

Alcohol biosensing by polyamidoamine (PAMAM)/cysteamine/alcohol oxidase-modified gold electrode.

A highly stable and sensitive amperometric alcohol biosensor was developed by immobilizing alcohol oxidase (AOX) through Polyamidoamine (PAMAM) dendrimers on a cysteamine‐modified gold electrode surface. Ethanol determination is based on the consumption of dissolved oxygen content due to the enzymatic reaction. The decrease in oxygen level was monitored at −0.7 V vs. Ag/AgCl and correlated with ethanol concentration. Optimization of variables affecting the system was performed. The optimized ethanol biosensor showed a wide linearity from 0.025 to 1.0 mM with 100 s response time and detection limit of (LOD) 0.016 mM. In the characterization studies, besides linearity some parameters such as operational and storage stability, reproducibility, repeatability, and substrate specificity were studied in detail. Stability studies showed a good preservation of the bioanalytical properties of the sensor, 67% of its initial sensitivity was kept after 1 month storage at 4°C. The analytical characteristics of the system were also evaluated for alcohol determination in flow injection analysis (FIA) mode. Finally, proposed biosensor was applied for ethanol analysis in various alcoholic beverage as well as offline monitoring of alcohol production through the yeast cultivation.

Electrochemical polymerization of 1-(4-nitrophenyl)-2,5-di(2-thienyl)-1 H-pyrrole as a novel immobilization platform for microbial sensing.

Two types of bacterial biosensor were constructed by immobilization of Gluconobacter oxydans and Pseudomonas fluorescens cells on graphite electrodes modified with the conducting polymer; poly(1-(4-nitrophenyl)-2,5-di(2-thienyl)-1 H-pyrrole) [SNS(NO(2))]. The measurement was based on the respiratory activity of cells estimated by the oxygen consumption at -0.7 V due to the metabolic activity in the presence of substrate. As well as analytical characterization, the linear detection ranges, effects of electropolymerization time, pH and cell amount were examined by using glucose as the substrate. The linear relationships were observed in the range of 0.25-4.0 mM and 0.2-1.0 mM for G. oxydans and P. fluorescens based sensors, respectively.

In situ synthesis of biomolecule encapsulated gold-cross-linked poly(ethylene glycol) nanocomposite as biosensing platform: a model study.

In situ synthesis of poly(ethylene glycol) (PEG) hydrogels containing gold nanoparticles (AuNPs) and glucose oxidase (GOx) enzyme by photo-induced electron transfer process was reported here and applied in electrochemical glucose biosensing as the model system. Newly designed bionanocomposite matrix by simple one-step fabrication offered a good contact between the active site of the enzyme and AuNPs inside the network that caused the promotion in the electron transfer properties that was evidenced by cyclic voltammetry as well as higher amperometric biosensing responses in comparing with response signals obtained from the matrix without AuNPs. As well as some parameters important in the optimization studies such as optimum pH, enzyme loading and AuNP amount, the analytical characteristics of the biosensor (AuNP/GOx) were examined by the monitoring of chronoamperometric response due to the oxygen consumption through the enzymatic reaction at -0.7 V under optimized conditions at sodium acetate buffer (50 mM, pH 4.0) and the linear graph was obtained in the range of 0.1-1.0 mM glucose. The detection limit (LOD) of the biosensor was calculated as 0.06 mM by using the signal to noise ratio of 3. Moreover, the presence of AuNPs was visualized by TEM. Finally, the biosensor was applied for glucose analysis for some beverages and obtained data were compared with HPLC as the reference method to test the possible matrix effect due to the nature of the samples.

Maltose biosensing based on co-immobilization of α-glucosidase and pyranose oxidase

A new bi-enzymatic system was designed by co-immobilization of alpha-glucosidase (AG) and pyranose oxidase (PyOx) for maltose analysis. The immobilization was carried out by cross-linking enzyme mixture, chitosan (CHIT) and carbon nanotube (CNT) via glutaraldehyde. The structure of biosensor including enzyme, CHIT, glutaraldehyde and CNT amount together with operational conditions like pH, temperature and applied potential were optimized. Then analytical characterization was performed. A fast linear response of the biosensor was observed for maltose in the concentration range from 0.25 to 2.0 mM at 35 degrees C and pH 6.0. The effect of CNT addition into the immobilization matrix was also investigated. The linear relationships between sensor response (y; microA/cm(2)) and substrate concentration (x; mM) were defined by the equations of y=0.844x+0.029 (R(2)=0.999) and y=0.882x+0.0625 (R(2)=0.996) for AG/PyOx/CHIT and AG/PyOx/CHIT-CNT biosensors, respectively. All other data were also given as comparison of two systems one with CNT-modified and CNT-free. Finally, for the sample application, maltose was analyzed in beer samples. As a result, it has been found that; complex matrix of natural beer samples had no influence on the biosensing response. Also the results were in good agreement with those obtained by spectrophotometric measurements.

Effects of mediators on the laccase biosensor response in paracetamol detection

An enzyme electrode suitable for paracetamol detection was developed by immobilizing laccase on a dissolved‐oxygen probe surface. The immobilization procedure was achieved by means of gelatin, which was then cross‐linked with glutaraldehyde. The measurement was based on the detection of oxygen consumption in relation to analyte oxidation. The optimum experimental conditions for the biosensor were investigated and the system was calibrated for paracetamol. Also the effects of three different mediators, namely HBT (1‐hydroxybenzotriazole), VLA [violuric acid (5‐isonitrosobarbituric acid)] and TEMPO (2,2′,6,6′‐tetramethylpiperidine‐N‐oxyl radical) were tested for the biosensors response. As a result, it was observed that HBT has a remarkable effect on the signal by providing more oxygen consumption during the enzymatic reaction. A linear relationship between sensor responses and analyte concentrations was obtained over the concentration range 2.0–15.0 μM, whereas, in the presence of the mediator HBT, this range became 0.5–3.0 μM.

Pseudomonas putida Based Amperometric Biosensors for 2,4-D Detection

Abstract Amperometric biosensors using Pseudomonas putida cells as a bioelement were developed for 2,4-dichloro phenoxy acetic acid (2,4-D). After the adaptation process of Pseudomonas putida to 2,4-D, cells were immobilized onto the screen printed graphite electrodes (SPG) as well as Clark oxygen probe by gelatin and glutaraldehyde. Optimum pH, temperature, and stability of the biosensor were investigated. Substrate specificities for various phenolic compounds were also searched. In repeatability studies, variation coefficients and standard deviations for both type of systems were calculated; SPG and Clark electrodes were calculated and results are given as a comparison of two systems. Finally, the biosensors were applied to 2,4-D determination in a real herbicide sample.

Use of Eggshell Membrane as an Immobilization Platform in Microbial Sensing

Abstract A new microbial biosensor was developed by immobilizing Pseudomonas fluorescens cells on eggshell membrane via physical adsorbtion. Bacteria-modified eggshell membrane was fixed tightly onto the surface of a carbon paste electrode (CPE) with a silicone rubber o-ring to construct a microbial biosensor. The measurements were based on the respiratory activity of the cells caused by oxygen consumption during bacterial metabolism. A mediated biosensor was also developed by ferrocene. As well as the response characteristics, stabilities and substrate specificities were investigated. Data were given as the comparison of mediated and unmediated systems.