Fulya Ekiz Kanik

A novel promising biomolecule immobilization matrix: Synthesis of functional benzimidazole containing conducting polymer and its biosensor applications

In order to construct a robust covalent binding between biomolecule and immobilization platform in biosensor preparation, a novel functional monomer 4-(4,7-di(thiophen-2-yl)-1H-benzo[d]imidazol-2-yl)benzaldehyde (BIBA) was designed and successfully synthesized. After electropolymerization of this monomer, electrochemical and spectroelectrochemical properties were investigated in detail. To fabricate the desired biosensor, glucose oxidase (GOx) was immobilized as a model enzyme on the polymer coated graphite electrode with the help of glutaraldehyde (GA). During the immobilization step, an imine bond was formed between the free amino groups of enzyme and aldehyde group of polymer. The surface characterization and morphology were investigated to confirm bioconjugation by X-ray photoelectron spectroscopy (XPS) and transmission electron microscopy (TEM) at each step of biosensor fabrication. The optimized biosensor shows good linearity between 0.02mM and 1.20mM and a low limit of detection (LOD) of 2.29μM. Kinetic parameters Km(app) and Imax were determined as 0.94mM and 10.91μA, respectively. The biosensor was tested for human blood serum samples.

Development of a novel biosensor based on a conducting polymer.

A new type of amperometric cholesterol biosensor was fabricated to improve the biosensor characteristics such as sensitivity and reliability. For this purpose, a novel immobilization matrix 2-(4-fluorophenyl)-4,7-di(thiophene-2-yl)-1H-benzo[d]imidazole (BIPF) was electrochemically deposited on a graphite electrode and used as a matrix for the immobilization of cholesterol oxidase (ChOx). Due to strong π-π stacking of aromatic groups in the structures of polymer backbone and enzyme molecule, one can easily achieve a sensitive and reliable biosensor without using any membrane or covalent bond formation between the enzyme molecules and polymer surface. Moreover, through pendant fluorine group of the polymer, H-bond formation between with enzyme molecules and polymer was generated. Cholesterol was used as the substrate and amperometric response was measured in correlation with cholesterol amount, at -0.7 V vs. Ag/AgCl in phosphate buffer (pH 7.0). Consequently, optimum conditions for this constructed biosensor were determined. K(M)app, I(max), LOD and sensitivity values were investigated and calculated as 4.0 nM, 2.27 µA, 0.404 µM and 1.47 mA/mM cm(2), respectively. A novel and accurate cholesterol biosensor was developed for the determination of total cholesterol in food samples.

Development of an efficient immobilization matrix based on a conducting polymer and functionalized multiwall carbon nanotubes: synthesis and its application to ethanol biosensors

Material modification is one of the hot topics recently. Hereby a novel functional monomer, 2-(4-nitrophenyl)-4,7-di(thiophen-2-yl)-1H-benzo[d]imidazole (BIPN), was synthesized for matrix generation through electrochemical polymerization. Its conducting polymer was successfully used for the biolayer construction in the biosensor preparation. The electrochemical and morphological properties were improved by the introduction of carboxylic acid functionalized multiwall carbon nanotubes (f-MWCNTs). Carboxylic acid functionalization of MWCNTs was carried out via acid treatment. The electrode surface was modified with the polymer and f-MWCNTs during electropolymerization to achieve a perfect immobilization matrix for alcohol oxidase. In order to prepare a new alcohol biosensor, alcohol oxidase (AOx) was immobilized onto the modified electrode. The modified electrode was characterized by scanning electron microscopy (SEM), X-ray photoelectron microscopy (XPS) and Fourier transform infrared (FTIR) spectroscopy techniques. Electrochemical responses of the enzyme electrodes were monitored at -0.7 V vs. Ag reference electrode by monitoring oxygen consumption in the presence of ethanol. Kinetic parameters, operational and storage stabilities were investigated. K, Imax, LOD and sensitivity were calculated as 16.946 mM, 3.31 μA, 0.806 mM and 476 μA mM-1 cm-2, respectively. Finally, this biosensor was applied to estimate the alcohol content in various beverages successfully.

An amperometric acetylcholine biosensor based on a conducting polymer.

An amperometric acetylcholine biosensor was prepared by the generation of the conducting polymer poly(4-(2,5-di(thiophen-2-yl)-1H-pyrrol-1-yl)benzenamine) (poly(SNS-NH2)) on graphite electrodes. For pesticide detection, the enzymes acetylcholinesterase (AChE) and choline oxidase (ChO) were co-immobilized onto the conducting polymer poly(SNS-NH2) films using covalent binding technique. Electrochemical polymerization was carried out using a three-electrode cell configuration via cyclic voltammetry. Characterization of resulting acetylcholine biosensor was done in terms of optimum pH, enzyme loading, range of linear response and shelf-life. Linear range was 0.12-10mM and shelf-life 4 weeks. Sensitivity was calculated as 2.19μAmM(-1)cm(-2). The designed biosensor was tested for the determination of paraoxon-ethyl in spiked tap water samples. The results were compared with a conventional quantification method using HPLC-DAD. Linear correlation of the quantification results with both methods (R(2)=0.998) was obtained.

A novel functional conducting polymer: synthesis and application to biomolecule immobilization

A recently synthesized conducting polymer poly(TBT6–NH2); poly(6-(4,7-di(thiophen-2-yl)-2H-benzo[d][1,2,3]triazol-2-yl)hexan-1-amine) was utilized as a matrix for biomolecule immobilization. After successful electrochemical deposition the polymer poly(TBT6–NH2) on the graphite electrodes, immobilization of choline oxidase (ChO) was carried out. Due to the free amino functional groups of the polymeric structure, ChO molecules were successfully immobilized onto the polymer surface via covalent binding. For this, glutaraldehyde (GA) was used as crosslinker and bifunctional agent. Hence, a robust binding between the support and the protein molecules was achieved. Scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy (XPS) were used to monitor the surface morphologies of both the polymer and the bioactive layer and to confirm the binding of the protein. Amperometric measurements were recorded by monitoring oxygen consumption in the presence of choline as the substrate at −0.7 V. The optimized biosensor showed a very good linearity between 0.1 and 10 mM with a 7 s response time and a detection limit (LOD) of 16.8 μM to choline. Also, kinetic parameters, operational and storage stabilities were determined. Finally, designed system was applied for pesticide detection.

A sepiolite modified conducting polymer based biosensor.

A conducting polymer modified with sepiolite was utilized in the construction of a highly sensitive and fast amperometric cholesterol biosensor. In this study a monomer; (10,13-bis(2,3-dihydrothieno[3,4-b][1,4]dioxin-5-yl)dibenzo[a,c]phenazine (PHED)) was synthesized and then its polymer was coated on a graphite electrode by electropolymerization to obtain a matrix for enzyme immobilization. Cholesterol oxidase was immobilized onto polymer coated electrode by adsorption technique. Sepiolite was introduced for a successful immobilization of the cholesterol oxidase. Immobilized enzyme kinetic parameters (KM(app), Imax) were evaluated by Michaelis-Menten kinetics and calculated as 0.031 mM and 6.06 μA, respectively. LOD and sensitivity were estimated as 0.36 μM and 1.64 mA/mMcm(2). Characterization of designed biosensor was done to examine the effect of various factors such as enzyme amount, optimum pH and shelf-life. A novel accurate and inexpensive cholesterol biosensor was developed for the determination of total cholesterol in food samples.

A novel DAD type and folic acid conjugated fluorescent monomer as a targeting probe for imaging of folate receptor overexpressed cells

We describe a modification and post‐functionalization technique for a donor–acceptor–donor type monomer; 6‐(4,7‐bis(2,3‐dihydrothieno[3,4‐b][1,4]dioxin‐5‐yl)‐2H‐benzo[d][1,2, 3]triazol‐2‐yl)hexan‐1‐amine. Folic acid was attached to the fluorescent structure. The conjugation was confirmed via NMR and Fourier transform infrared analyses. Cytotoxicity was investigated and the comparison of association of targeted monomeric structures in tumor cells was monitored via fluorescence microscopy.

Preparation and Characterization of a Novel Solid-Phase Microextraction Material for Application to the Determination of Pesticides

ABSTRACT The preparation and characterization of a novel solid-phase microextraction fiber is reported with application to the determination of pesticides in fruit juice. The fiber was fabricated by electrochemically coating a stainless steel wire with a thin polymeric film of 4-(2,5-di(thiophen-2-yl)-1H-pyrrol-1-yl) benzenamine. The procedure was initiated in 10 mL of acetonitrile containing 5.5 mg of monomer, 0.1 mol NaClO4, and 0.1 mol LiClO4 by cycling the potential between −0.5 and 1.2 V with a scan rate of 100 mV/sec. The morphology of the fiber surface was examined by scanning electron microscopy and its stability was characterized by thermal gravimetric analysis. The fiber was exposed to headspace extraction of bromopropylate, chlorpyrifos, lambda-cyhalothrin, penconazole, and procymidone prior to the analysis by gas chromatography with an electron capture detector. Operational parameters affecting the extraction efficiency, adsorption and desorption times and temperature, and stirring rate were screened using a Plackett-Burman Design. Emerging parameters were further optimized via Central Composite Design that were 20 min at 64°C for adsorption and 4.4 min at 250°C for desorption. Solution parameters were optimized to be 5.0 mL of sample in pH 2.0 Britton-Robinson buffer containing 0.1 mg/L NaCl to promote the volatilization of the analytes. The limits of detection were at the ng/mL level for the pesticides. The fiber was used as a selective and sensitive tool for the trace determination of these pesticides in grape juice.