A Novel Conductometric Urea Biosensor with Improved Analytical Characteristic Based on Recombinant Urease Adsorbed on Nanoparticle of Silicalite

Development of a conductometric biosensor for the urea detection has been reported. It was created using a non-typical method of the recombinant urease immobilization via adsorption on nanoporous particles of silicalite. It should be noted that this biosensor has a number of advantages, such as simple and fast performance, the absence of toxic compounds during biosensor preparation, and high reproducibility (RSD = 5.1 %). The linear range of urea determination by using the biosensor was 0.05–15 mM, and a lower limit of urea detection was 20 μM. The bioselective element was found to be stable for 19 days. The characteristics of recombinant urease-based biomembranes, such as dependence of responses on the protein and ion concentrations, were investigated. It is shown that the developed biosensor can be successfully used for the urea analysis during renal dialysis

Determination of total creatine kinase activity in blood serum using an amperometric biosensor based on glucose oxidase and hexokinase

International audienceCreatine kinase (CK: adenosine-5-triphosphate-creatine phosphotransferase) is an important enzyme of muscle cells; the presence of a large amount of the enzyme in blood serum is a biomarker of muscular injuries, such as acute myocardial infarction. This work describes a bi-enzyme (glucose oxidase and hexokinase based) biosensor for rapid and convenient determination of CK activity by measuring the rate of ATP production by this enzyme. Simultaneously the biosensor determines glucose concentration in the sample. Platinum disk electrodes were used as amperometric transducers. Glucose oxidase and hexokinase were co-immobilized via cross-linking with BSA by glutaraldehyde and served as a biorecognition element of the biosensor. The biosensor work at different concentrations of CK substrates (ADP and creatine phosphate) was investigated; optimal concentration of ADP was 1 mM, and creatine phosphate - 10 mM. The reproducibility of the biosensor responses to glucose, ATP and CK during a day was tested (relative standard deviation of 15 responses to glucose was 2%, to ATP - 6%, to CM - 7-18% depending on concentration of the CK). Total time of CM analysis was 10 min. The measurements of creatine kinase in blood serum samples were carried out (at 20-fold sample dilution). Twentyfold dilution of serum samples was chosen as optimal for CM determination. The biosensor could distinguish healthy and ill people and evaluate the level of CM increase. Thus, the biosensor can be used as a test-system for CM analysis in blood serum or serve as a component of multibiosensors for determination of important blood substances. Determination of activity of other kinases by the developed biosensor is also possible for research purpose

Influence of Composition of Zeolite/Enzyme Nanobiocomposites on Analytical Characteristics of Urea Biosensor Based on Ion-Selective Field-Effect Transistors

Zeolite/enzyme nanobiocomposites of different compositions were tested in this work for the improvement of biosensor analytical characteristics. The bioselective element based on urease immobilized by cross-linking with glutaraldehyde was used as a model. The working characteristics of biosensors based on various zeolite/enzyme nanocomposites were examined and compared with those of urease-based biosensors. An optimal concentration of zeolytes beta (BEA) in bioselective elements is determined to be 1.5%. It ensures as wide linear range of measurement without remarkable loss in biosensor sensitivity to urea. The BEA zeolite-based biosensors were shown to have better working parameters in comparison with those based on zeolites A (LTA). A decrease in biosensor sensitivity to heavy metal ions was demonstrated for all zeolites used, which testifies to probable increase in stability of urea measurement in real environmental samples

Elaboration of Urease Adsorption on Silicalite for Biosensor Creation

A possibility of efficient urease adsorption on silicalite for the purpose of biosensor creation was investigated. The procedure of urease adsorption on silicalite is notable for such advantages as simple and fast performance and non-use of toxic or auxiliary compounds. Optimal conditions for modifying transducer surfaces with silicalite and subsequent urease adsorption on these surfaces were selected. The working parameters of the created biosensor were optimized. The developed biosensor with adsorbed urease was characterized by good intra-reproducibility (RSD 4.5?%), improved inter-reproducibility (RSD of urea determination is 9?%) and operational stability (less than 10?% loss of activity after 10 days). Besides, the developed method for enzyme adsorption on silicalite was compared with the traditional methods of urease immobilization in biosensorics. Working conditions of the produced biosensor (pH and ionic strength) were shown to be close to those of the biosensor based on urease immobilized in GA vapor. For these reasons, it was concluded that the method of enzyme adsorption on silicalite is well-suited for biosensor standardization aimed at its further manufacture

Application of butyrylcholinesterase‐based biosensor for simultaneous determination of different toxicants using inhibition and reactivation steps

Fast and simple detection of toxic compounds in aqueous solutions is important for agriculture and pollution monitoring. Enzyme‐based electrochemical biosensors are a promising platform for this task, but they usually lack the ability to distinguish between toxicants if more than one toxicant is present in the sample. Herein, we propose a new method of detection of various toxic compounds in complex multi‐component water samples using an electrochemical biosensor and additional stages of enzyme reactivation. The biosensor is based on butyrylcholinesterase immobilized on the surface of conductometric transducers using glutaraldehyde cross‐linking. It was shown that the biosensor is sensitive to organophosphorus pesticides, heavy metals ions, glycoalkaloids, but has a limited sensitivity to mycotoxins and surfactants. We propose a procedure for the analysis of complex samples with several reactivation stages to be able to determine which category of toxicants is present. Glycoalkaloids are reversible inhibitors and biosensor's activity is restored by washing in working buffer; heavy metal ions and pesticides are irreversible inhibitors and biosensor's activity is restored by incubation in EDTA and PAM‐2 solutions, correspondingly. This biosensor can be used for the detection of separate toxicants or for analysis of their mixtures in aqueous samples. It can be also used for the evaluation of total toxicity of the samples and applied for water quality monitoring

A Novel Conductometric Urea Biosensor with Improved Analytical Characteristic Based on Recombinant Urease Adsorbed on Nanoparticle of Silicalite

Development of a conductometric biosensor for the urea detection has been reported. It was created using a non-typical method of the recombinant urease immobilization via adsorption on nanoporous particles of silicalite. It should be noted that this biosensor has a number of advantages, such as simple and fast performance, the absence of toxic compounds during biosensor preparation, and high reproducibility (RSD = 5.1 %). The linear range of urea determination by using the biosensor was 0.05-15 mM, and a lower limit of urea detection was 20 mu M. The bioselective element was found to be stable for 19 days. The characteristics of recombinant urease-based biomembranes, such as dependence of responses on the protein and ion concentrations, were investigated. It is shown that the developed biosensor can be successfully used for the urea analysis during renal dialysis

Improvement of urease based biosensor characteristics using additional layers of charged polymers

NRC publication: Ye

Application of enzyme/zeolite sensor for urea analysis in serum

Urea biosensor based on zeolite-adsorbed urease was applied for analysis of blood serum samples. It should be noted, that this biosensor has a number of advantages, such as simple and fast performance, the absence of toxic compounds during biosensor preparation, high reproducibility and repeatability (RSD = 9% and 4%, respectively). The linear range of urea determination by using the biosensor was 0.003-0.75 mM, and the limit of urea detection was 3 mu M. The method of standard addition was used for analysis of serum samples with 500-fold dilution. Total time of analysis was 10 min. Good reproducibility of urea determination in real samples was demonstrated (RSD = 10%). Biosensor results were verified by using a common method of urea determination (diacetyl monoxime reaction). It was shown that by using this biosensor distinguishing healthy people from people with renal dysfunction becomes easier