Application of zeolites and zeolitic imidazolate frameworks in the biosensor development.

Biosensors are advanced devices for analysis of composition of blood, urine, environmental samples, and many other media. Their current development is tightly linked with nanomaterials, such as zeolites and zeolitic imidazolate framework (ZIFs). The present review describes electrochemical (amperometric, conductometric, ISFET) and optical (fluorescent and colorimetric) biosensors that incorporate zeolites and ZIFs in their biorecognition elements. The biosensors are based on immobilized enzymes (such as glucose oxidase, urease, and acetylcholinesterase), antibodies, DNA, and aptamers. The review present reasons for application of these nanomaterials, and discusses advantages of zeolite- and ZIF-containing biosensors over other biosensors. In most cases, the biosensors have improved sensitivity, better limit of detection, wider linear range, and other improved characteristics. It is demonstrated that immobilization of biomolecules such as enzymes or antibodies on the surface of zeolites and ZIFs enables creation of unique advanced biosensors that have a potential for further development and practical applications.

Electrochemical biosensors based on multienzyme systems: Main groups, advantages and limitations - A review.

In the review, the principles and main purposes of using multienzyme systems in electrochemical biosensors are analyzed. Coupling several enzymes allows an extension of the spectrum of detectable substances, an increase in the biosensor sensitivity (in some cases, by several orders of magnitude), and an improvement of the biosensor selectivity, as showed on the examples of amperometric, potentiometric, and conductometric biosensors. The biosensors based on cascade, cyclic and competitive enzyme systems are described alongside principles of function, advantages, disadvantages and practical use for real sample analyses in various application areas (food production and quality control, clinical diagnostics, environmental monitoring). The complications and restrictions regarding the development of multienzyme biosensors are evaluated. The recommendations on the reasonability of elaboration of novel multienzyme biosensors are given.

A highly selective amperometric biosensor array for the simultaneous determination of glutamate, glucose, choline, acetylcholine, lactate and pyruvate.

The work was aimed at the development of a biosensor array for the simultaneous determination of six solutes (glutamate, glucose, choline, acetylcholine, lactate, and pyruvate) in aqueous solutions. Enzymes selective for these substrates were immobilized on the surface of amperometric platinum disc electrodes and served as bioselective elements of a biosensor array. Direct enzymatic analysis by the developed biosensors provided high sensitivity to the tested substrates (limits of detection were 1-5 µM). The linear ranges of the biosensors were from 0.001-0.01 mM to 0.2-2.5 mM. The influence of solution pH, ionic strength and buffer capacity on the biosensor responses was investigated; the conditions for simultaneous operation of all the bioselective elements were optimized. The absence of any cross-influence of the substrates of enzymatic systems used was shown as well as a high selectivity of the biosensors and the absence of any impact of interfering substances (ascorbic acid, dopamine, cysteine, paracetamol). The developed biosensor array had good response reproducibility and storage stability. The array is suitable for rapid (0.5-1 min) and simple simultaneous determination of glutamate, glucose, choline, acetylcholine, lactate, and pyruvate in aqueous (biological) samples; furthermore, the creation of a single chip with six sensitive elements is possible as well as the addition of other biosensors.

Advances in nanomaterial application in enzyme-based electrochemical biosensors: a review.

Electrochemical enzyme-based biosensors are one of the largest and commercially successful groups of biosensors. Integration of nanomaterials in the biosensors results in significant improvement of biosensor sensitivity, limit of detection, stability, response rate and other analytical characteristics. Thus, new functional nanomaterials are key components of numerous biosensors. However, due to the great variety of available nanomaterials, they should be carefully selected according to the desired effects. The present review covers the recent applications of various types of nanomaterials in electrochemical enzyme-based biosensors for the detection of small biomolecules, environmental pollutants, food contaminants, and clinical biomarkers. Benefits and limitations of using nanomaterials for analytical purposes are discussed. Furthermore, we highlight specific properties of different nanomaterials, which are relevant to electrochemical biosensors. The review is structured according to the types of nanomaterials. We describe the application of inorganic nanomaterials, such as gold nanoparticles (AuNPs), platinum nanoparticles (PtNPs), silver nanoparticles (AgNPs), and palladium nanoparticles (PdNPs), zeolites, inorganic quantum dots, and organic nanomaterials, such as single-walled carbon nanotubes (SWCNTs), multi-walled carbon nanotubes (MWCNTs), carbon and graphene quantum dots, graphene, fullerenes, and calixarenes. Usage of composite nanomaterials is also presented.

Conductometric biosensor for arginine determination in pharmaceutics.

A new conductometric biosensor based on coimmobilized urease and arginase has been developed for arginine determination in pharmaceutics. First, the main parameters of the selected method of immobilization (concentrations of arginase, urease, and glutaraldehyde, time of incubation) were optimized. An influence of the solution parameters (buffer ionic strength, capacity, pH, Mn2+ concentration) on the biosensor operation was studied, working conditions were optimized. After biosensor optimization, the main analytical characteristics were as follows. The limit of detection - 2.5 µM, the linear range - 2.5-500 µM, the sensitivity to arginine 13.4 ±â€¯2.4 µS/mM, the response time - 20 s. The signals repeatability and operational stability in continuous exploitation were studied over one working day and during one week. Additionally, the selectivity of the developed biosensor towards arginine was essayed relative to other amino acids. The developed biosensor has been used to measure arginine concentrations in some drugs. The results obtained were in high correlation with the characteristics declared by producers.

An amperometric glutamate biosensor for monitoring glutamate release from brain nerve terminals and in blood plasma.

An excess of the excitatory neurotransmitter, glutamate, in the synaptic cleft during hypoxia/ischemia provokes development of neurotoxicity and originates from the reversal of Na+-dependent glutamate transporters located in the plasma membrane of presynaptic brain nerve terminals. Here, we have optimized an electrochemical glutamate biosensor using glutamate oxidase and developed a biosensor-based methodological approach for analysis of rates of tonic, exocytotic and transporter-mediated glutamate release from isolated rat brain nerve terminals (synaptosomes). Changes in the extracellular glutamate concentrations from 11.5 ±â€¯0.9 to 11.7 ±â€¯0.9 µΜ for 6 min reflected a low tonic release of endogenous glutamate from nerve terminals. Depolarization-induced exocytotic release of endogenous glutamate was equal to 7.5 ±â€¯1.0 µΜ and transporter reversal was 8.0 ±â€¯1.0 µΜ for 6 min. The biosensor data correlated well with the results obtained using radiolabelled L-[14C]glutamate, spectrofluorimetric glutamate dehydrogenase and amino acid analyzer assays. The blood plasma glutamate concentration was also tested, and reliability of the biosensor measurements was confirmed by glutamate dehydrogenase assay. Therefore, the biosensor-based approach for accurate monitoring rates of tonic, exocytotic and transporter-mediated release of glutamate in nerve terminals was developed and its adequacy was confirmed by independent analytical methods. The biosensor measurements provided precise data on changes in the concentrations of endogenous glutamate in nerve terminals in response to stimulation. We consider that the glutamate biosensor-based approach can be applied in clinics for neuromonitoring glutamate-related parameters in brain samples, liquids and blood plasma in stroke, brain trauma, therapeutic hypothermia treatment, etc., and also in laboratory work to record glutamate release and uptake kinetics in nerve terminals.

Improvement of amperometric transducer selectivity using nanosized phenylenediamine films.

In this work, we studied the conditions of deposition of a semipermeable polyphenylenediamine (PPD)-based membrane on amperometric disk platinum electrodes. Restricting an access of interfering substances to the electrode surface, the membrane prevents their impact on the sensor operation. Two methods of membrane deposition by electropolymerization were compared-at varying potential (cyclic voltammetry) and at constant potential. The cyclic voltammetry was shown to be easier in performing and providing better properties of the membrane. The dependence of PPD membrane effectiveness on the number of cyclic voltammograms and phenylenediamine concentration was analyzed. It was shown that the impact of interfering substances (ascorbic acid, dopamine, cysteine, uric acid) on sensor operation could be completely avoided using three cyclic voltammograms in 30 mM phenylenediamine. On the other hand, when working with diluted samples, i.e., at lower concentrations of electroactive substances, it is reasonable to decrease the phenylenediamine concentration to 5 mM, which would result in a higher sensitivity of transducers to hydrogen peroxide due to a thinner PPD layer. The PPD membrane was tested during continuous operation and at 8-day storage and turned out to be efficient in sensor and biosensors.

A Novel Amperometric Glutamate Biosensor Based on Glutamate Oxidase Adsorbed on Silicalite.

In this work, we developed a new amperometric biosensor for glutamate detection using a typical method of glutamate oxidase (GlOx) immobilization via adsorption on silicalite particles. The disc platinum electrode (d = 0.4 mm) was used as the amperometric sensor. The procedure of biosensor preparation was optimized. The main parameters of modifying amperometric transducers with a silicalite layer were determined along with the procedure of GlOx adsorption on this layer. The biosensors based on GlOx adsorbed on silicalite demonstrated high sensitivity to glutamate. The linear range of detection was from 2.5 to 450 µM, and the limit of glutamate detection was 1 µM. It was shown that the proposed biosensors were characterized by good response reproducibility during hours of continuous work and operational stability for several days. The developed biosensors could be applied for determination of glutamate in real samples.

A novel conductometric biosensor based on hexokinase for determination of adenosine triphosphate.

The paper presents a simple and inexpensive reusable biosensor for determination of the concentration of adenosine-5'-triphosphate (ATP) in aqueous samples. The biosensor is based on a conductometric transducer which contains two pairs of gold interdigitated electrodes. An enzyme hexokinase was immobilized onto one pair of electrodes, and bovine serum albumin-onto another pair (thus, a differential mode of measurement was used). Conditions of hexokinase immobilization on the transducer by cross-linking via glutaraldehyde were optimized. Influence of experimental conditions (concentration of magnesium ions, ionic strength and concentration of the working buffer) on the biosensor work was studied. The reproducibility of biosensor responses and operational stability of the biosensor were checked during one week. Dry storage at -18 °C was shown to be the best conditions to store the biosensor. The biosensor was successfully applied for measurements of ATP concentration in pharmaceutical samples. The proposed biosensor may be used in future for determination of ATP and/or glucose in water samples.

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

Creatine 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 1mM, 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 CK - 7-18% depending on concentration of the CK). Total time of CK 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 CK determination. The biosensor could distinguish healthy and ill people and evaluate the level of CK increase. Thus, the biosensor can be used as a test-system for CK 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 purposes.

Development of novel enzyme potentiometric biosensor based on pH-sensitive field-effect transistors for aflatoxin B1 analysis in real samples.

This study aimed at the development and optimization of a potentiometric biosensor based on pH-sensitive field-effect transistors and acetylcholinesterase for aflatoxin B1 determination in real samples. Optimal conditions for bioselective elements operation were defined and analytical characteristics of the proposed biosensor were studied. The proposed biosensor characterized high operational stability and reproducibility of signal. Selectivity of acetylcholinesterase-biosensor to aflatoxins in relation to other groups of toxic substances was analyzed. The developed biosensor was applied to the determination of aflatoxin B1 in real samples (sesame, walnut and pea).

Monitoring of the velocity of high-affinity glutamate uptake by isolated brain nerve terminals using amperometric glutamate biosensor.

Glutamate is the major excitatory neurotransmitter in the central nervous system, which is involved in the main aspects of normal brain functioning. High-affinity Na(+)-dependent glutamate transporters is key proteins, which transport extracellular glutamate to the cytoplasm of nerve cells, thereby preventing continuous activation of glutamate receptors, and thus the development of neurotoxicity. Disturbance in glutamate uptake is involved in the pathogenesis of major neurological disorders. Amperometric biosensors are the most promising and successful among electrochemical biosensors. In this study, we developed (1) amperometric glutamate biosensor, (2) methodological approach for the analysis of glutamate uptake in liquid samples of isolated rat brain nerve terminals (synaptosomes). The basal level of glutamate, the initial velocity of glutamate uptake and time-dependent accumulation of glutamate by synaptosomes were determined using developed glutamate biosensor. Comparative analysis of the data with those obtained by radioactive analysis, spectrofluorimetry and ion exchange chromatography was performed. Therefore, the methodological approach for monitoring of the velocity of glutamate uptake, which takes into consideration the definite level of endogenous glutamate in nerve terminals, was developed using glutamate biosensor.

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 µM. 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.

Urease-based ISFET biosensor for arginine determination.

In this work a novel biosensor for arginine determination based on the urease inhibition effect has been proposed. Ion-selective field effect transistors were used as transducers. Urease immobilized in glutaraldehyde vapor served as a biorecognition element of the biosensor. Significant part of the work was aimed at proving the urease inhibition by arginine. Optimal concentration of urea for arginine determination was chosen. Detection limit for arginine was 0.05 mM. The biosensor selectivity towards different amino acids was studied. The results of quantitative determination of l-arginine in the real sample (a drinkable solution "Arginine Veyron") were in good agreement with the producer's data (a relative error was 5.2%). The biosensor showed a good reproducibility of arginine determination.

Development of conductometric biosensor array for simultaneous determination of maltose, lactose, sucrose and glucose.

The aim of this work was to develop an array of biosensors for simultaneous determination of four carbohydrates in solution. Several enzyme systems selective to lactose, maltose, sucrose and glucose were immobilised on the surface of four conductometric transducers and served as bio-recognition elements of the biosensor array. Direct enzyme analysis carried out by the developed biosensors was highly sensitive to the corresponding substrates. The analysis lasted 2 min. The dynamic range of substrate determination extended from 0.001 mM to 1.0-3.0mM, and strongly depended on the enzyme system used. An effect of the solution pH, ionic strength and buffer capacity on the biosensors responses was investigated; the conditions of simultaneous operation of all biosensors were optimised. The data on cross-impact of the substrates of all biosensors were obtained; the biosensor selectivity towards possible interfering carbohydrates was tested. The developed biosensor array showed good signal reproducibility and storage stability. The biosensor array is suited for simultaneous, quick, simple, and selective determination of maltose, lactose, sucrose and glucose.

Development and optimization of a novel conductometric bi-enzyme biosensor for L-arginine determination.

A highly sensitive conductometric biosensor for l-arginine determination was developed by exploiting the unique biorecognition capacities of two enzymes of urea cycle - arginase (E.C. 3.5.3.1) and urease (E.C. 3.5.1.5). The enzymes were co-immobilized in a single bioselective membrane on the working sensor, while a lysine rich bovine serum albumin (BSA) membrane was immobilized on the reference sensor, allowing differential measurements. The optimum percentage ratio of arginase and urease within the bioselective membrane was determined when the biosensor sensitivity to l-arginine and urea was optimum. Analytical characteristics of the conductometric biosensor for l-arginine determination were compared for two types of enzyme immobilization (cross-linking with glutaraldehyde (GA) and entrapment in the polymeric membrane). The optimum features in terms of the sensitivity, the linear range, and the detection limit (4.2 µS/mM, 0.01-4mM, and 5.0 × 10(-7)M, respectively) were found for l-arginine biosensor based on enzyme cross-linking with GA. A quantitative determination of l-arginine in the real sample (a drinkable solution "Arginine Veyron") gave a satisfactory result compared to the data provided by the producer (a relative error was 4.6%). The developed biosensor showed high operational and storage stability.

Novel conductometric biosensor based on three-enzyme system for selective determination of heavy metal ions.

A differential pair of planar thin-film interdigitated electrodes, deposited on a ceramic pad, was used as a conductometric transducer. The three-enzyme system (invertase, mutarotase, glucose oxidase), immobilized on the transducer surface, was used as a bioselective element. The ratio between enzymes in the membrane was found experimentally considering the highest biosensor sensitivity to substrate (sucrose) and heavy metal ions. Optimal concentration of sucrose for inhibitory analysis was 1.25 mM and incubation time in the investigated solution amounted to 10-20 min. The developed biosensor demonstrated the best sensitivity toward ions Hg(2+) and Ag(+). A principal possibility of the biosensor reactivation either by EDTA solution after inhibition with silver ions or by cysteine solution after inhibition with mercury ions was shown.

Microbiosensor based on glucose oxidase and hexokinase co-immobilised on platinum microelectrode for selective ATP detection.

ATP determination is of great importance since this compound is involved in a number of vital biological processes. To monitor ATP concentration levels, we have developed a microbiosensor based on cylindrical platinum microelectrode, covered with a layer of poly-m-phenylendiamine (PPD), and layer of co-immobilised glucose oxidase and hexokinase. Conditions for biosensor measurement of ATP (pH, Mg(2+) and substrates concentration) in vitro and microbiosensor characteristics such as sensitivity, selectivity, reproducibility, storage stability were studied and optimized. Under optimal conditions the microbiosensor can measure ATP concentrations down to a 2.5 microM detection limit with response time about 15 s. Interferences by electroactive compounds like biogenic amines and their metabolites, ascorbic acid, uric acid and L-cystein are rejected in general by the PPD layer. The microbiosensor developed is insensitive to ATP analogues (or substances with similar structure), such as ADP, AMP, GTP and UTP, too. It can be used for ATP analysis in vitro in the reactions consuming or producing macroergic triphosphate molecules to study kinetics of the process and in drug design concerning development of inhibitors specific to target kinases and others target enzymes.

[Kinetic properties of butyrylcholinesterases immobilised on pH-sensitive field-effect transistor surface and inhibitory action of steroidal glycoalkaloids on these enzymes].

The inhibitory action of steroid glycoalkaloids alpha-solanine, alpha-chaconine and tomatine on horse and human serum butyryl cholinesterases immobilized on the pH-sensitive field-effect transistors has been studied. Using acetyl- and butyryl choline as substrates, the optimal pH and the apparent kinetic parameters (< K(m) >, < V(max) >) of immobilized butyryl cholinesterases have been calculated in the absence of inhibitors. The affinity of each enzyme to glycoalkaloids has been estimated from calculation of apparent inhibition constants < K(i) > and inhibition coefficients i(0.5). Application of the studied cholinesterases for biosensoric determination of glycoalkaloids in the wide range of concentrations (10(-7)-10(-4) M) in different media has been discussed.

Highly selective microbiosensors for in vivo measurement of glucose, lactate and glutamate.

An alternative approach to production of amperometric microbiosensors, which combines electrochemical electrometallization and electropolymerisation of phenylene diamine film with covalent binding enzymes, is presented. In this respect, for a sensitive detection of hydrogen peroxide (HP) at +0.4V versus Ag/AgCl (detection limit of 0.5 microM, s/n=3), carbon fiber microelectrodes (30 microm in diameter and 500 microm long) were covered with ruthenium. To obtain a highly selective detection of HP, in the presence of different interfering compounds (ascorbic acid, uric acid, etc.), an additive semi-permeable polymer film was formed on the top of the ruthenium layer by electropolymerisation of m-phenylene diamine (m-PD). The enzymatic selective layers were formed by covalent cross-linking the enzymes (glucose oxidase, lactate oxidase or glutamate oxidase) with BSA by glutaraldehyde in the presence of ascorbate oxidase. An additional polymeric layer based on polyurethane and Nafion was deposited on the top of the enzymatic membrane (glucose oxidase, lactate oxidase, or glutamate oxidase) in order to extend the dynamic range of biosensors up to 4mM for glucose (R=0.997; Y[nA]=-0.22+9.68x[glucose, mM]), 1.75mM for lactate (R=0.991; Y[nA]=0.43+15.36x[lactate, mM]) and 0.25 mM for glutamate (R=0.999; Y[nA]=0.02+29.14x[glutamate, mM]). The developed microbiosensors exhibited also negligible influences from interfering compounds at their physiological concentrations. Microbiosensors remained stable during 10h in a flow injection system at 36 degrees C and pH 7.4. The microbiosensors developed are now used in vivo and, as an example, we report here the data obtained with the glucose biosensor.