This paper is a review of the authors' publications concerning the development of biosensors based on enzyme field-effect transistors (ENFETs) for direct substrates or inhibitors analysis. Such ...biosensors were designed by using immobilised enzymes and ion-selective field-effect transistors (ISFETs). Highly specific, sensitive, simple, fast and cheap determination of different substances renders them as promising tools in medicine, biotechnology, environmental control, agriculture and the food industry. The biosensors based on ENFETs and direct enzyme analysis for determination of concentrations of different substrates (glucose, urea, penicillin, formaldehyde, creatinine, etc.) have been developed and their laboratory prototypes were fabricated. Improvement of the analytical characteristics of such biosensors may be achieved by using a differential mode of measurement, working solutions with different buffer concentrations and specific agents, negatively or positively charged additional membranes, or genetically modified enzymes. These approaches allow one to decrease the effect of the buffer capacity influence on the sensor response in an aim to increase the sensitivity of the biosensors and to extend their dynamic ranges. Biosensors for the determination of concentrations of different toxic substances (organophosphorous pesticides, heavy metal ions, hypochlorite, glycoalkaloids, etc.) were designed on the basis of reversible and/or irreversible enzyme inhibition effect(s). The conception of an enzymatic multibiosensor for the determination of different toxic substances based on the enzyme inhibition effect is also described. We will discuss the respective advantages and disadvantages of biosensors based on the ENFETs developed and also demonstrate their practical application.
An assessment of the toxicity of methyl parathion and products of its photodegradation in water samples was performed using conductometric biosensors based on thin films planar electrodes and ...immobilized cholinesterases. All photodegradation experiments were carried out using a quartz reaction cell equipped with a 125
W high pressure mercury arc lamp as source of radiation. The photodegradation kinetics can be described as a first-order degradation curve
C=C
0
e
−kt
, where
C
0=(9.1±0.1)×10
−5
M,
k=0.026±0.001
min
−1 and
t
1/2=26
min. The high performance liquid chromatography (HPLC) was used to identify and quantify methyl paraoxon and 4-nitrophenol as main methyl parathion photoproducts. It was shown for the first time by using conductometric enzyme biosensor that the inhibition effect on the acetyl cholinesterase (AcChE) activity increases dramatically as soon as the photodegradation of methyl parathion begins. Methyl paraoxon is about 10 times more toxic than methyl parathion. The toxicity curve does not exactly follow the appearance of methyl paraoxon. The results obtained could be explained by a strong synergistic effect of methyl parathion and methyl paraoxon on the enzyme activity.
The paper is a review of the researches of Biomolecular Electronics Laboratory concerning the development of biosensors based on electrochemical transducers (amperometric and conductometric ...electrodes, potentiometric pH-sensitive field effect transistors) and different biorecognition molecules (enzymes, cells, antibodies), biomimics (molecularly imprinted polymers), as sensitive elements for direct analysis of substrates or inhibitory analysis of toxicants. Highly specific, sensitive, simple, fast and cheap detection of different substances renders them as promising tools for needs of health care, environmental control, biotechnology, agriculture and food industries. Diverse biosensor formats for direct determination of different analytes and inhibitory enzyme analysis of a number of toxins have been designed and developed. Improvement of their analytical characteristics may be achieved by using differential mode of measurement, negatively or positively charged additional semipermeable membranes, nanomaterials of different origin, genetically modified enzymes. These approaches have been aimed at increasing the sensitivity, selectivity and stability of the biosensors and extending their dynamic ranges. During the last 25 years more than 50 laboratory prototypes of biosensor systems based on mono- and multibiosensors for direct determination of a variety of metabolites and inhibitory analysis of different toxic substances were created. Some of them were tested in real samples analysis. The advantages and disadvantages of the biosensors developed are discussed. The possibility of their practical application is considered.
Aim. To identify a type of inhibition of immobilized acetylcholinesterase by aflatoxin B1. Methods. A bioselective element of the potentiometric biosensor was created using acetylcholinesterase, ...which was covalently immobilized on the surface of the pH-FET sensor by glutaraldehyde crosslinking with bovine serum albumin. Results. Optimal conditions for the potentiometric biosensor operation such as pH-optimum of the enzyme action and its inhibition were defined. An apparent Michaelis constant, as well as a maximum initial reaction rate of immobilized acetylcholinesterase as a part of the biosensor were determined. The type of reversible inhibition of immobilized acetylcholinesterase by aflatoxin B1 in potentiometric biosensor was identified by using a new graphical “degree of inhibition” method and the obtained result was confirmed with one of the tradi-tional methods, such as the Lineweaver-Burk plot. Conclusions. This study helps to understand the mechanisms of enzyme inhibition in biosensors and brings the biosensor implementation closer.
The differential method of conductometric measurements does not fully solve the problem of the influence of changes in the background electrical conductivity of the working buffer solutions on the ...results of conversion of the biosensor responses. The variation in the background electrical conductivity of the buffer solution upon addition of the highly conductive analyte acts as a common mode interference and causes the additive error. Here we present a new measurement method and structure of the device for quantification of the analytes that provide a significant decrease in the measurement error associated with a change in the background electrical conductivity caused by the introduction of the analyte to the working solution prior to the generation of the informative signal. A block diagram of the device and a vector model of the balancing process of its measuring circuit are presented. The advantages of the developed method and biosensor analyzer for application within the possible changes of the transducer parameters and measurement conditions are demonstrated. References 28, figures 4.
Highly sensitive biosensors based on pH-sensitive field effect transistors and cholinesterases for detection of solanaceous glycoalkaloids have been developed, characterised and optimised. The main ...analytical characteristics of the biosensors developed have been studied under different conditions and an optimal experimental protocol for glycoalkaloids determination in model solution has been proposed. Using such a biosensor and an enzyme reversible inhibition effect, the total potato glycoalkaloids content can be determined within the range of 0.2–100 μM depending on the type of alkaloid, with lowest detection limits of 0.2 μM for α-chaconine, 0.5 μM for α-solanine and 1 μM for solanidine. The dynamic ranges for the compounds examined show that such biosensors are suitable for a quantitative detection of glycoalkaloids in real potato samples. High reproducibility, operational and storage stability of the biosensor developed have been shown.
A novel electrochemical microbiosensor for protein determination based on an all-solid-state microelectrodes with a Ppy 3,3-Co(1,2-C
2B
9H
11)
2 solid internal contact and immobilized trypsin and ...carboxypeptidase B was developed. The specificity of the sensor signal was investigated using artificial ester and amide substrates of the proteinases:
Nα-benzoyl-
l-arginine ethyl ester hydrochloride (BAEE),
Nα-benzoyl-
dl-arginine-
p-nitroanilide hydrochloride (BAPNA), and Hippuryl-Arg. Study on a possibility of protein detection showed that quantitative proteins determination can be performed with the microbiosensor in the concentration range of 1–60
μg/ml.
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