Masoud Khayyami

Development of an amperometric biosensor based on acetylcholine esterase covalently bound to a new support material.

A new type of amperometric biosensor based on immobilised acetylcholine esterase was designed and constructed. The enzyme was immobilised on a flow-through working electrode, which was prepared from reticulated vitreous carbon (RVC) or from a composite material consisting of RVC and superporous agarose. The sensor was operated in FIA mode using acetylthiocholine as a substrate. The sensor responded to inhibitors such as paraoxon-10(-9) mol was detected by the sensor in a non-optimised configuration. The practical lifetime of the sensor was at least 1 month.

Development of a simple detector for microbial metabolism, based on a polypyrrole dc resistometric device

Abstract A simple device for the detection of metabolic effects was constructed based on a conducting polymer (polypyrrole) coated with an agarose layer containing immobilised yeast cells. A galvanostatic potentiometric method was used to measure the resistance of the sensor, thus minimising interferences from fluctuations in solution conductivity. The resulting biosensor was used as a simple indicator of yeast metabolism. It responded within 20 minutes to increases in the glucose concentration by decreased resistance (40% per 5 g/l). When the metabolism was inhibited by a toxic substance the response disappeared. The device could be reconditioned by washing off the agarose/cell layer with hot water, followed by the application of a new agarose layer with fresh cells. Because the manufacture of the device is simple and the measuring principle is uncomplicated, it is specially suited for mass production for quick field tests.

Ferrocene-mediated thermal biosensor

A biosensor that combines electrochemistry and calorimetry was investigated. Using this sensor, glucose measurements were performed by employing glucose oxidase together with ferrocene as electron-transfer mediator. The electrochemical reaction was accomplished by applying a voltage between a platinum column (working electrode), in contact with a crushed reticulated vitreous carbon (RVC) matrix onto which glucose oxidase was immobilized, and platinum wires (counter electrodes) located at the inlet and outlet of the column. For detection, the thermal signal generated by the redox reaction was measured as opposed to measuring the electrochemical signal. By using this method, a linear range of glucose concentration up to 20 mmol l–1 was achieved.

Coulometric determination of NAD+ and NADH in normal and cancer cells using LDH, RVC and a polymer mediator.

An electrochemical method for the measurement of NAD(+) and NADH in normal and cancer tissues using flow injection analysis (FIA) is reported. Reticulated vitreous carbon (RVC) electrodes with entrapped l-lactate dehydrogenase (LDH) and a new redox polymer containing covalently bound toluidine blue O (TBO) were employed for this purpose. Both NAD(+) and NADH were estimated coulometrically based on their reaction with LDH. The latter was immobilized on controlled pore glass (CPG) by cross-linking with glutaraldehyde and packed within the RVC. The concentrations of NAD(+) and NADH in the tissues, estimated using different electron mediators such as ferricyanide (FCN), meldola blue (MB) and TBO have also been compared. The effects of flow rate, pH, applied potential (versus Ag/AgCl reference) and adsorption of the mediators have also been investigated. Based on the measurements of NAD(+) and NADH in normal and cancer tissues it has been concluded that the NADH concentration is lower, while the NAD(+) concentration is higher in cancer tissues. Amongst the electron mediators TBO was found to be a more stable mediator for such measurements.

Flow-injection determination of trace hydrogen peroxide or glucose utilizing an amperometric biosensor based on glucose oxidase bound to a reticulated vitreous carbon electrode.

An electron transfer mediator, 8-dimethylamino-2,3-benzophenoxazine (Meldola Blue), dissolved in the carrier solution in a flow-injection system, was found to reduce the oxidation potential for hydrogen peroxide from 600-1200 mV without mediator to-100 mV vs. Ag/AgCl with the mediator present. The very low background current of reticulated vitreous carbon (RVC) at this potential makes it possible to detect very low levels of hydrogen peroxide or glucose. Glucose oxidase was covalently coupled with carbodiimide to RVC, and the RVC was formed into a column inserted in a flow-injection system. The calibration curve was linear from 30 nM to 10 microM glucose with 5 microM mediator. At higher mediator concentrations, the linear range was extended to 1000 microM, but with a much higher background current. The sample throughput was about 60 h(-1). The current response decreased to 50% of the original response after 20 days. The coulometric yield was high because the sample was pumped through the pores of the RVC. It was 16% and 55% at a flow rate of 1 ml min(-1) at mediator concentrations of 5 and 50 microM respectively.

DC-resistometric urea sensitive device utilizing a conducting polymer film for the gas-phase detection of ammonia

The construction of an urea sensitive device based on a conducting polymer is described. The conducting polymer (polypyrrole) was exposed only to a gas-phase; this led to a significant improvement in its stability. A DC-resistometric measuring principle was used, which required only simple measuring equipment. The device responded rapidly to between 0 and 500 mM urea, with resistance changes of between 0 and 400 Ohms. The measurement of the urea content of a sample and subsequent regeneration took 20 min.