Erol Akyilmaz

A biosensor based on urate oxidase–peroxidase coupled enzyme system for uric acid determination in urine

A new amperometric biosensor based on urate oxidase-peroxidase coupled enzyme system for the specific and selective determination of uric acid in urine was developed. Commercially available urate oxidase and peroxidase were immobilized with gelatin by using glutaraldehyde and fixed on a pretreated teflon membrane. The method is based on generation of H(2)O(2) from urine uric acid by urate oxidase and its consuming by peroxidase and then measurement of the decreasing of dissolved oxygen concentration by the biosensor. The biosensor response depends linearly on uric acid concentration between 0.1 and 0.5 muM. In the optimization studies of the biosensor, phosphate buffer (pH 7.5; 50 mM) and 35 degrees C were obtained as the optimum working conditions. In addition, the most suitable enzyme activities were found as 64.9x10(-3) U cm(-2) for urate oxidase and 512.7 U cm(-2) for peroxidase. And also some characteristic studies of the biosensor such as reproducibility, substrate specificity and storage stability were carried out.

A new enzyme electrode based on ascorbate oxidase immobilized in gelatin for specific determination of l-ascorbic acid

A biosensor for the specific determination of l-ascorbic acid in fruit juices and vitamin C tablets was developed using ascorbate oxidase (EC 1.10.3.3) from cucumber (Cucumis sativus L.) in combination with a dissolved oxygen probe. Ascorbate oxidase immobilized with gelatin using glutaraldehyde and fixed on pretreated teflon membrane served as an enzyme electrode. The phosphate buffer (50 mM, pH 7.5) and 35 degrees C were established as providing the optimum conditions. The biosensor response depends linearly on l-ascorbic acid concentration between 5.0x10(-5) and 1.2x10(-3) M with a response time 45 s. The biosensor is stable for more than 2 months, while more than 200 assays were performed. The results obtained for fruit juices and tablets were compared with DCIP (2,6 dichlorophenolindophenol) method.

A mushroom (Agaricus bisporus) tissue homogenate based alcohol oxidase electrode for alcohol determination in serum

To construct homogenized tissue based biosensors by using plant tissue materials is a relatively new development in the biosensor technology. In this study, a homogenized mushroom (Agaricus bisporus) tissue based electrode in which alcohol oxidase activity was developed by immobilizing with gelatin and cross-linking agent glutaraldehyde at dissolved oxygen probe for determination of ethyl alcohol. The electrode response depends linearly on ethyl alcohol concentration between 0.2 and 20 mM. In the optimization studies, phosphate buffer (pH 7.5, 50 mM) and 35 degrees C were obtained as the optimum conditions. By using the electrode prepared we have made approximately 60 measurements in 10-h period, and response time of the electrode is 2 min for each measurement.

MWCNT–cysteamine–Nafion modified gold electrode based on myoglobin for determination of hydrogen peroxide and nitrite

In this work, a novel amperometric biosensor of hydrogen peroxide (H2O2) was developed based on the immobilization of myoglobin (Mb) on the surface of the multi-walled carbon nanotube (MWCNT) -Nafion-cysteamine (CA) modified gold electrode (Au) and its electrocatalytic activity was used for the determination of nitrite (NO2(-)). In the optimization studies, the best MWCNT and myoglobin amount were investigated. It was discovered at the experiments for the optimization of the working conditions that the buffer at this study as 50.0mM, pH7.0 phosphate buffer (PBS) and working temperature as 30°C for the H2O2 biosensor. It was determined at the characterization studies on the biosensor that linear results are obtained between the ranges of 0.1μM to 70.0μM for H2O2 concentration and 1-250μM for NO2(-). The reproducibility of the biosensor was determined both H2O2 and nitrite. From the experiments, average value, standard deviation (SD) and coefficients of variation (CV%) were calculated to be 10.02±0.43μM, and 4.29% for 10.0μM H2O2 (n=6) and 52.0±2.1μM, and 3.89% for 50.0μM nitrite (n=8), respectively. At the same time the sample was analyzed for NO2(-) in drinking and mineral waters.

Preparation and optimization of a bienzymic biosensor based on self-assembled monolayer modified gold electrode for alcohol and glucose detection

The aim of this project was to develop a bienzymic biosensor, which was based on co-immobilization of alcohol oxidase and glucose oxidase on the same electrode by formation of self-assembled monolayer (SAM) for selective determination of ethanol and glucose. In the biosensor construction the enzymes and the mediator, tetrathiafulvalene (TTF), were immobilized with cross-linking agents glutaraldehyde and cysteamine by forming a self-assembled monolayer (SAM) on a gold disc electrode. Amounts of ethanol and glucose were amperometrically detected by monitoring current values at reduction potential of TTF(+), 0.1V. Decreases in biosensor responses were linearly related to glucose concentrations between 0.1 and 1.0 mM and ethanol concentrations between 1.0 and 10 mM. Limits of detection of the biosensor for ethanol and glucose were calculated to be 0.75 and 0.03 mM, respectively. In the optimization studies of the biosensor some parameters such as optimum pH, optimum temperature, enzyme amount, effect of TTF concentration and duration of SAM formation were investigated.

Design of a multiwalled carbon nanotube–Nafion–cysteamine modified tyrosinase biosensor and its adaptation of dopamine determination

In this work, a multiwalled carbon nanotube (MWCNT)-Nafion-cysteamine (CA) modified tyrosinase biosensor brings a new and original perspective to biosensor technology intended for the development of dopamine determination. Dopamine measurements were done at 0.2V with the amperometric method by the developed biosensor system. In addition, in this study dopamine determination was carried out by using the differential pulse voltammetry method between potentials of 0.4 and -0.15 V. In the optimization studies of the biosensor, some parameters such as optimal pH, optimal temperature, optimal enzyme amount, and effect of MWCNT concentration were investigated. Afterward, in the characterization studies, some parameters such as linearity and reproducibility were determined. In the reproducibility experiment, an average value of 1.026 μM, a standard deviation of ±0.03975, and a coefficient of variation of 3.8% were determined for a 1-μM dopamine concentration (n=15). Determination of dopamine was carried out in drug samples by the developed biosensor.

Voltammetric determination of epinephrine by White rot fungi (Phanerochaete chrysosporium ME446) cells based microbial biosensor

The lyophilized biomass of White rot fungi (Phanerochaete chrysosporium ME446) was immobilized in gelatine using glutaraldehyde crosslinking agent on a Pt working electrode. The fungal cells retained their laccase activity under entrapped state. The immobilized cells were used as a source of laccase to develop amperometric epinephrine biosensor. The catalytic action of the laccase in the biosensor released an epinephrinequinone as a result of redox activity, thereby causing an increase in the current. The optimal working conditions of the biosensor were carried out at pH 4.5 (50 mM acetate buffer containing 100 mM K(3)Fe(CN)(6)), and 20°C. The sensor response was linear over a range of 5-100 μM epinephrine. The detection limit of the biosensor was found to be 1.04 μM. In the optimization and characterization studies of the microbial biosensor some parameters such as effect of fungi and gelatine amount, percentage of glutaraldehyde on the biosensor response and substrate specificity were carried out. In the application studies of the biosensor, sensitive determination of epinephrine in pharmaceutical ampules was investigated.

Development of a catalase based biosensor for alcohol determination in beer samples

An amperometric biosensor based on catalase enzyme for alcohol determination was developed. To construct the biosensor catalase was immobilized by using gelatin and glutaraldehyde on a Clark type dissolved oxygen (DO) probe covered with a teflon membrane which is sensitive for oxygen. The working principle of the biosensor depends on two reactions, which one is related to another, catalyzed by catalase enzyme. In the first reaction catalase catalyzes the degradation of hydrogen peroxide and oxygen is produced and also a steady-state DO concentration occurs in a few minutes. When ethanol added to the medium catalase catalyzes the degradation of both hydrogen peroxide and ethanol and this results in a new steady-state DO concentration. Difference for first and the last steady-state DO concentration occurred in the interval surface of DO probe membrane, which related to ethanol concentration, are detected by the biosensor. The biosensor response depends linearly on ethanol concentration between 0.05 and 1.0 mM with a detection limit of 0.05 mM and a response time of 3 min. In the optimization studies of the biosensor phosphate buffer (pH 7.0; 50 mM) and 35 degrees C were established as providing the optimum working conditions. In the characterization studies of the biosensor some parameters such as reproducibility, substrate specificity, operational and storage stability were carried out. Finally, by using the biosensor developed and enzimatic-spectrophotometric method alcohol concentration of some alcoholic drinks were determined and results were compared.

Do copper ions activate tyrosinase enzyme? A biosensor model for the solution

Some metal ions play a cofactor role for the activity of tyrosinase enzyme and one of them is copper ion. In this study an amperometric biosensor was developed in order to investigate the effect of the copper ions on the activity of tyrosinase enzyme. In the construction of the biosensor tyrosinase enzyme was immobilized on a Clark-type dissolved oxygen probe which was covered with a oxygen sensitive teflon membrane, by using a chemical covalent immobilization method based on gelatine and bifunctional reagent, glutaraldehyde. The principle of the measurement was based on the determination of the differentiation of dissolved oxygen level in the enzymatic reaction catalyzed by tyrosinase in the absence and the presence of copper ions. Differences between the dissolved oxygen concentrations were related to copper ion concentration which was added in to the reaction medium. The biosensor response depends linearly on copper ion concentration between 2.5-20.0microM with a response time 1min. The detection limit of the biosensor is 0.95microM. In the optimization studies of the biosensor, the most suitable amounts of tyrosinase, gelatin and glutaraldehyde ratio were determined to be 69.0U/cm(-2), 4.21mg/cm(-2), and 2.5%, respectively. In the optimization studies of the biosensor, phosphate buffer (pH 7.0 ,50mM) and 30 degrees C were detected to be working conditions. For the characterization of the biosensor some parameters such as reproducibility, thermal and pH stability were carried out.

An inhibition type amperometric biosensor based on tyrosinase enzyme for fluoride determination

In this study, a new biosensor based on the inhibition of tyrosinase for the determination of fluoride is described. To construct the biosensor tyrosinase was immobilized by using gelatine and cross-linking agent glutaraldehyde on a Clark type dissolved oxygen (DO) probe covered with a teflon membrane which is sensitive for oxygen. The phosphate buffer (50mM, pH 7.0) at 30 degrees C were established as providing the optimum working conditions. The method is based on the measurement of the decreasing of dissolved oxygen level of the interval surface that related to fluoride concentration added into reaction medium in the presence of catechol. Inhibitor effect of fluoride results in decrease in dissolved oxygen concentration. The biosensor response depends linearly on fluoride concentration between 1.0 and 20 microM with a response time of 3 min. In the characterization studies of the biosensor some parameters such as reproducibility, substrate specificity and storage stability were carried out. From the experiments, the average value (x), Standard deviation (S.D) and coefficient of variation (C.V %) were found as 10.5 microM, +/-0.57 microM, 5.43%, respectively for 10 microM fluoride standard.