Multi-walled carbon nanotubes functionalized with a new Schiff base containing phenylboronic acid residues: application to the development of a bienzymatic glucose biosensor using a response surface methodology approach.

An innovative supramolecular architecture is reported for bienzymatic glucose biosensing based on the use of a nanohybrid made of multi-walled carbon nanotubes (MWCNTs) non-covalently functionalized with a Schiff base modified with two phenylboronic acid residues (SB-dBA) as platform for the site-specific immobilization of the glycoproteins glucose oxidase (GOx) and horseradish peroxidase (HRP). The analytical signal was obtained from amperometric experiments at - 0.050 V in the presence of 5.0 × 10-4 M hydroquinone as redox mediator. The concentration of GOx and HRP and the interaction time between the enzymes and the nanohybrid MWCNT-SB-dBA deposited at glassy carbon electrodes (GCEs) were optimized through a central composite design (CCD)/response surface methodology (RSM). The optimal concentrations of GOx and HRP were 3.0 mg mL-1 and 1.50 mg mL-1, respectively, while the optimum interaction time was 3.0 min. The bienzymatic biosensor presented a sensitivity of (24 ± 2) × 102 µA dL mg-1 ((44 ± 4) × 102 µA M-1), a linear range between 0.06 mg dL-1 and 21.6 mg dL-1 (3.1 µM-1.2 mM) (R2 = 0.9991), and detection and quantification limits of 0.02 mg dL-1 (1.0 µM) and 0.06 mg dL-1 (3.1 µM), respectively. The reproducibility for five sensors prepared with the same MWCNT-SB-dBA nanohybrid was 6.3%, while the reproducibility for sensors prepared with five different nanohybrids and five electrodes each was 7.9%. The GCE/MWCNT-SB-dBA/GOx-HRP was successfully used for the quantification of glucose in artificial human urine and commercial human serum samples.

Evaluation of the operational conditions of the glucose oxidase and catalase multienzymatic system through enzyme co-immobilization on amino hierarchical porous silica.

Hexaric acids have attracted attention lately because they are platform chemicals for synthesizing pharmaceuticals. In particular, gluconic acid is one of the most studied because it is readily available in nature. In this work, operational conditions like temperature and pH were evaluated for the enzymatic production of gluconic acid. For this purpose, glucose oxidase (GOx) and catalase (CAT) were individually immobilized and co-immobilized using amino-silica as support. The catalytic performance of the enzymes both as separate biocatalysts (GOx or CAT) and as an enzymatic complex (GOx-CAT) was assessed in terms of enzymatic activity and stability at temperatures 45 °C and 50 °C and pH 6 to 8. The results show that CAT is a key enzyme for gluconic acid production as it prevents GOx from being inhibited by H2O2. However, CAT was found to be less stable than GOx. Therefore, different GOx to CAT enzymatic ratios were studied, and a ratio of 1-3 was determined to be the best. The highest glucose conversion conditions were 45 °C and pH 7.0 for 24 h. Regarding the biocatalyst reuse, GOx-CAT retained more than 70% of its activity after 6 reaction cycles. These results contribute to further knowledge and application of oxidases for hexaric acid production and shed greater light on the role of the glucose oxidase/catalase pair in better catalytic performance. Both enzymes were immobilized in one pot, which is relevant for their potential use in industry; an enzyme system was obtained in a single step.

Development of a Redox-Polymer-Based Electrochemical Glucose Biosensor Suitable for Integration in Microfluidic 3D Cell Culture Systems.

The inclusion of online, in situ biosensors in microfluidic cell cultures is important to monitor and characterize a physiologically mimicking environment. This work presents the performance of second-generation electrochemical enzymatic biosensors to detect glucose in cell culture media. Glutaraldehyde and ethylene glycol diglycidyl ether (EGDGE) were tested as cross-linkers to immobilize glucose oxidase and an osmium-modified redox polymer on the surface of carbon electrodes. Tests employing screen printed electrodes showed adequate performance in a Roswell Park Memorial Institute (RPMI-1640) media spiked with fetal bovine serum (FBS). Comparable first-generation sensors were shown to be heavily affected by complex biological media. This difference is explained in terms of the respective charge transfer mechanisms. Under the tested conditions, electron hopping between Os redox centers was less vulnerable than H2O2 diffusion to biofouling by the substances present in the cell culture matrix. By employing pencil leads as electrodes, the incorporation of these electrodes in a polydimethylsiloxane (PDMS) microfluidic channel was achieved simply and at a low cost. Under flow conditions, electrodes fabricated using EGDGE presented the best performance with a limit of detection of 0.5 mM, a linear range up to 10 mM, and a sensitivity of 4.69 µA mM-1 cm-2.

Integrating machine learning and electrochemistry to develop a glucose biosensor assembled with Ganoderma applanatum lectin.

Fungal lectins have enormous biotechnological potential, but limited knowledge about their biochemical and biophysical features prevents their proper use. Herein, we report an innovative alternative to use Ganoderma applanatum lectin (GAL) as a glucose biorecognition element, after identifying the ideal electroanalytical conditions by machine learning studies performed with a homologous agglutinin from the same macrofungus. The research revealed that GAL has moderate resistance to pH (4-8) and temperature (20-60 °C) variations, but its hemagglutinating activity (376.5 HU mg-1 GAL at 20 °C) was better conserved under physiological conditions. Integrating electrochemical data and semi-empirical molecular modeling, biocompatible and electrostatically favorable conditions were found to immobilize the lectin on Prussian blue-modified glassy carbon electrode, after thermal activation of the metal-complex film. The glucose dose-response relationship obtained with the developed biosensor, defined as GAL/ta-PB/GCE, showed a typical Hill equation correlation, suggesting electrodic interactions represented by a sigmoidal mathematical function. GAL/ta-PB/GCE achieved remarkable electroanalytical performance, with emphasis on the detection limit (10.2 pM) and sensitivity (0.012 µA µM-1cm-2). The biosensor was successfully used to quantify glucose in pharmaceutical formulations, reiterating that the association of theoretical and experimental information drives important advances in bioelectrochemical studies.

PEDOT-Polyamine-Based Organic Electrochemical Transistors for Monitoring Protein Binding.

The fabrication of efficient organic electrochemical transistors (OECTs)-based biosensors requires the design of biocompatible interfaces for the immobilization of biorecognition elements, as well as the development of robust channel materials to enable the transduction of the biochemical event into a reliable electrical signal. In this work, PEDOT-polyamine blends are shown as versatile organic films that can act as both highly conducting channels of the transistors and non-denaturing platforms for the construction of the biomolecular architectures that operate as sensing surfaces. To achieve this goal, we synthesized and characterized films of PEDOT and polyallylamine hydrochloride (PAH) and employed them as conducting channels in the construction of OECTs. Next, we studied the response of the obtained devices to protein adsorption, using glucose oxidase (GOx) as a model system, through two different strategies: The direct electrostatic adsorption of GOx on the PEDOT-PAH film and the specific recognition of the protein by a lectin attached to the surface. Firstly, we used surface plasmon resonance to monitor the adsorption of the proteins and the stability of the assemblies on PEDOT-PAH films. Then, we monitored the same processes with the OECT showing the capability of the device to perform the detection of the protein binding process in real time. In addition, the sensing mechanisms enabling the monitoring of the adsorption process with the OECTs for the two strategies are discussed.

Correlation of fasting blood, salivary glucose and malondialdehyde in subjects with & without type 2 diabetes

Oxidative stress is identified as the common pathogenic factor that leads to insulin resistance in diabetics. Malondialdehyde is a product of lipid peroxidation. Aim: The aim of this study was to determine the variation in the Salivary malondialdehyde (MDA) among subjects with and without T2DM in comparison to the fasting blood and Salivary glucose. Methods: This study involved 29 healthy participants as Controls (group I) and 29 participants with Type 2 Diabetes Mellitus as Cases (group II). Salivary Glucose was analysed by glucose oxidase end-point assay. Thiobarbituric acid (TBA) assay method was considered for estimation of MDA in fasting saliva. Data was Statistically analysed using SPSS20. Parametric test was performed to analyse the data. Results: The correlation calculated between FBG with FSG level was found to be highly significant. A positive correlation between MDA levels with FBG was found. The relationship between FBG and FSG (r = 0.7815, p < 0.05), FBG and MDA (r =0.3678, p < 0.05) and FSG and MDA (r = 0.2869, p < 0.05) were found to be positively significant. Conclusion: Saliva as a unique body fluid can serve as a medium for biochemical analysis only in standard settings and with multiple measures to be used as a diagnostic tool in par with the gold standard serum. Salivary MDA levels can be considered as one of the oxidative stress markers in Type 2 Diabetic condition

Mechanisms of Enzymatic Transduction in Nanochannel Biosensors.

The immobilization of enzymes in solid-state nanochannels is a new avenue for the design of biosensors with outstanding selectivity and sensitivity. This work reports the first theoretical model of an enzymatic nanochannel biosensor. The model is applied to the system previously experimentally studied by Lin, et al. (Anal. Chem. 2014, 86, 10546): a hourglass nanochannel modified by glucose oxidase for the detection of glucose. Our predictions are in good agreement with experimental observations as a function of the applied potential, pH and glucose concentration. The sensing mechanism results from the combination of three processes: i) the establishment of a steady-state proton concentration gradient due to a reaction-diffusion mechanism, ii) the effect of that gradient on the charge of the adsorbed enzymes and native surface groups, and iii) the effect of the resulting surface charge on the ionic current. Strategies to improve the sensor performance based on this mechanism are identified and discussed.

Poly-L-lysine-modified with ferrocene to obtain a redox polymer for mediated glucose biosensor application.

Ferrocene-based polymers as redox mediators are considered versatile and important in the study of glucose biosensors. Poly-L-lysine (PLL), as a cationic polymer, possesses good properties including biocompatibility, biodegradation and water solubility. In this work, PLL was modified with ferrocene carboxylate in a very simple way by activating the carboxyl group of Fc, which reacted with the amino groups of the polymer. The resulting product was analysed by FTIR. Performance as a redox mediator (Fc-PLL) with the enzyme glucose oxidase was tested by cyclic voltammetry and showed an increase in the oxidation current in the presence of glucose in PBS pH 7.4. Additionally, performance as a biosensor was evaluated by amperometry and gave a linear range of 0-10 mM, a limit of detection of 23 µM, a sensitivity of 6.55 µA/cm2 mM and high selectivity. To evaluate the charged regions of Fc-PLL/GOx on the electrode surface, analysis by scanning electrochemical microscopy showed remarkable activity. The Fc-PLL redox polymer as a glucose biosensor has been well accepted as this kind of material, and the results showed remarkable activity as an electron transfer mediator between the redox polymer and the GOx enzyme.

Highly sensitive and selective nanostructured microbiosensors for glucose and lactate simultaneous measurements in blood serum and in vivo in brain tissue.

Highly sensitive and selective nanostructured lactate and glucose microbiosensors for their in vivo simultaneous determination in rat brain were developed based on carbon fiber microelectrodes (CFM) modified with nanoporous gold (NPG) using the Dynamic Hydrogen Bubble Template (DHBT) method. Electrodeposition of platinum nanoparticles (PtNP) onto the NPG film enhances the sensitivity and the electrocatalytic properties towards H2O2 detection. The nanostructured microelectrode platform was modified by glucose oxidase (GOx) and lactate oxidase (LOx) enzyme immobilization. High selective measurements were achieved by covering with a perm-selective layer of electropolymerized m-phenylenediamine, deposition of a Nafion® film and by using a null sensor. The morphological characteristics and electroanalytical performance of the microbiosensors were assessed, by scanning electron microscopy and electrochemical techniques, respectively. The PtNP/NPG/CFM shows a high sensitivity to H2O2 (5.96 A M-1 cm-2) at 0.36 V vs. Ag/AgCl, with a linear range from 0.2 to 200 µM, and an LOD of 10 nM. The microbiosensors were applied to the simultaneous determination of lactate and glucose in blood serum samples. Moreover, the basal extracellular concentrations of lactate and glucose were measured in vivo in four different rat brain structures. These results support the potential of the microbiosensor to be used as a valuable tool to investigate brain neurochemicals in vivo.

Biosensing Membrane Base on Ferulic Acid and Glucose Oxidase for an Amperometric Glucose Biosensor.

A biosensing membrane base on ferulic acid and glucose oxidase is synthesized onto a carbon paste electrode by electropolymerization via cyclic voltammetry in aqueous media at neutral pH at a single step. The developed biosensors exhibit a linear response from 0.082 to 34 mM glucose concentration, with a coefficient of determination R2 equal to 0.997. The biosensors display a sensitivity of 1.1 µAmM-1 cm-2, a detection limit of 0.025 mM, and 0.082 mM as glucose quantification limit. The studies reveal stable, repeatable, and reproducible biosensors response. The results indicate that the novel poly-ferulic acid membrane synthesized by electropolymerization is a promising method for glucose oxidase immobilization towards the development of glucose biosensors. The developed glucose biosensors exhibit a broader linear glucose response than other polymer-based glucose biosensors.

Catalytic effects of magnetic and conductive nanoparticles on immobilized glucose oxidase in skin sensors.

Wearable skin sensors is a promising technology for real-time health care monitoring. They are of particular interest for monitoring glucose in diabetic patients. The concentration of glucose in sweat can be more than two orders of magnitude lower than in blood. In consequence, the scientific and technological efforts are focused in developing new concepts to enhance the sensitivity, decrease the limit of detection (LOD) and reduce the response time (RT) of glucose skin sensors. This work explores the effect of adsorbed superparamagnetic magnetite nanoparticles (MNPs) and conductive nanoparticles (CNPs) on carbon nanotube substrates (CNTs) used to immobilize glucose oxidase enzyme in the working electrode of skin sensors. MNPs and CNPs are made of magnetite and gold, respectively. The performance of the sensors was tested in standard buffer solution, artificial sweat, fresh sweat and on the skin of a healthy volunteer during an exercise session. In the case of artificial sweat, the presence of MNPs accelerated the RT from 7 to 5 s at the expense of increasing the LOD from 0.017 to 0.022 mM with slight increase of the sensitivity from 4.90 to 5.09µAm M-1cm-2. The presence of CNPs greatly accelerated the RT from 7 to 2 s and lowered the LOD from 0.017 to 0.014 mM at the expense of a great diminution of the sensitivity from 4.90 to 4.09µAm M-1cm-2. These effects were explained mechanistically by analyzing the changes in the concentration of free oxygen and electrons promoted by MNPs and CNPs in the CNTs and its consequences on the the glucose oxidation process.

Enhanced in vitro anticancer activity of yeast expressed recombinant glucose oxidase versus commercial enzyme.

Cancer treatments continue to have many disadvantages. Reactive oxygen species, such as H2O2, in high concentrations, can cause cytotoxicity to cells, being even greater in cancer cells. One of the H2O2-producing enzymes is glucose oxidase; its application in cancer treatment should be explored. In this work, the extracellular expression of the mutated recombinant enzyme glucose oxidase was carried out in the eukaryotic expression system Pichia pastoris SMD1168, through the modification and optimization of the gox gene of Aspergillus niger to improve its expression in yeast and its purification. Also, the secretion signal of the alpha-mating factor from Saccharomyces cerevisiae was added to the gene for extracellular expression, and it was inserted into the expression vector pPIC3.5k. The extracellular expression of the enzyme facilitated purification by anion exchange chromatography; the purification was corroborated by SDS-PAGE, with a molecular weight of its subunit between 63 kDa and 100 kDa. The mutated recombinant enzyme glucose oxidase showed greater anticancer activity compared to the commercial glucose oxidase and could have potential for cancer treatment. KEY POINTS: • Pichia pastoris is an excellent eukaryotic expression system for proteins that need post-translational modifications. • Extracellular expression facilitates protein purification. • Glucose oxidase has potential application in cancer treatment.

PEDOT-polyamine composite films for bioelectrochemical platforms - flexible and easy to derivatize.

We report a straightforward route for the preparation of flexible, electrochemically stable and easily functionalizable poly(3,4-ethylenedioxythiophene) (PEDOT) composite films deposited on PET foils as biosensing platforms. For this purpose, poly(allylamine) hydrochloride (PAH) was blended with PEDOT to provide amine-bearing sites for further biofunctionalization as well as to improve the mechanical properties of the films. The conducting PEDOT-PAH composite films were characterized by cyclic voltammetry, UV-vis and Raman spectroscopies. An exhaustive stability study was carried out from the mechanical, morphological and electrochemical viewpoint. Subsequent sugar functionalization of the available amine groups from PAH allowed for the specific recognition of lectins and the subsequent self-assembly of glycoenzymes (glucose oxidase and horseradish peroxidase) concomitant with the prevention of non-specific protein fouling. The platforms presented good bioelectrochemical performance (glucose oxidation and hydrogen peroxide reduction) in the presence of redox mediators. The developed composite films constitute a promising option for the construction of all-polymer biosensing platforms with great potential owing to their flexibility, high transmittance, electrochemical stability and the possibility of glycosylation, which provides a simple route for specific biofunctionalization as well as an effective antifouling strategy.

Enzymatically functionalized polyaniline thin films produced with one-step electrochemical immobilization and its application in glucose and urea potentiometric biosensors.

Glucose and urea enzymatic biosensors were fabricated. One-step electrochemical immobilization process was used to produce thin polyaniline films with entrapped enzymes. Chronopotentiometric analysis, scanning electron microscopy, electrochemical impedance spectroscopy and optical reflectance spectroscopy were used to determine the structure-property relationship of the functionalized polymeric thin films. The device has a recognition stage connected to a potentiometric field-effect-transistor stage and is based on the measurement of microenvironment pH variation or locally produced ions. Optimization of biosensor fabrication and effective measurement conditions were performed. The optimized films presented sensitivity, linearity and detection range to glucose of 14.6 ± 0.4 mV/decade, 99.8% and from 10-4 M to 10-1 mol/L. Two different biosensors were produced based on the enzymatic reaction of urea with selectivity to ammonium or hydroxyl ions. For ammonium ion selective film, the sensor's parameters were 14.7 ± 0.9 mV/decade, 98.2% and from 10-5 to 10-1 mol/L. For the hydroxyl ion selective film, the same parameters were 7.4 ± 0.5 mV/decade, 98.1% and from 10-5 to 10-1 mol/L. The change in the oxidation state of the polymeric matrix explains: i) the large loss of functionality of glucose biosensor in time, ii) the conservation of functionality to the hydroxyl ions for urea biosensor and iii) the selectivity variation of the ammonium ion selective urea biosensor. The results indicate that the polymeric matrix has indeed changeable selectivity, what can be applied in different situations for biosensors production.

Designing electrochemical interfaces based on nanohybrids of avidin functionalized-carbon nanotubes and ruthenium nanoparticles as peroxidase-like nanozyme with supramolecular recognition properties for site-specific anchoring of biotinylated residues.

We are reporting an original supramolecular architecture based on a rationally designed new nanohybrid with enhanced peroxidase-like activity and site-specific biorecognition properties using avidin-functionalized multi-walled carbon nanotubes (MWCNTs-Av) and Ru nanoparticles (RuNPs). The nanohybrid-electrochemical interface was obtained by drop-coating of MWCNTs-Av dispersion at glassy carbon electrodes (GCE) followed by solvent evaporation and further electrodeposition of RuNPs (50 ppm RuCl2 for 15 s at -0.600 V). The simultaneous presence of MWCNTs and RuNPs produces a synergic effect on the non-enzymatic catatalytic reduction of H2O2 and allows the quantification of H2O2 in a wide linear range (from 5.0 × 10-7 M to 1.75 × 10-3 M) with a low limit of detection (65 nM). The avidin residues present in MWCNTs-Av/RuNPs hybrid nanomaterial allowed the anchoring by bioaffinity of biotinylated glucose oxidase (biot-GOx) as proof-of-concept of the analytical application of MWCNTs-Av platform for biosensors development. The resulting nanoarchitecture behaves as a bienzymatic-like glucose biosensor with a competitive analytical performance: linear range between 2.0 × 10-5 M and 1.23 × 10-3 M, sensitivity of (0.343 ±â€¯0.002) µA mM-1 or (2.60 ±â€¯0.02) µA mM-1 cm-2, detection limit of 3.3 µM, and reproducibility of 5.2% obtained with five different GCE/MWCNTs-Av/RuNPs/biot-GOx bioplatforms prepared the same day using the same MWCNTs-Av dispersion, and 9.1% obtained with nine biosensors prepared in different days with nine different MWCNTs-Av dispersions. The average concentrations of glucose in Gatorade®, Red bull® and Pepsi® with the biosensor demonstrated excellent agreement with those reported in the commercial beverages.

A novel disposable self-adhesive inked paper device for electrochemical sensing of dopamine and serotonin neurotransmitters and biosensing of glucose.

In this work, we detail the progress of a novel electrochemical disposable device, which has a relatively low cost and easy production, with a novel conductive ink, that consists of graphite and automotive varnish mixture, deposited over a self-adhesive paper, granting an easy production with relatively low cost. The electrode surface was characterized by scanning electron microscopy, X-ray powder diffraction and Fourier transforms infrared and Raman, cyclic voltammetry and electrochemical impedance spectroscopies. In addition, the proposed electrode was applied for individual electrochemical determination of dopamine and serotonin. The device achieved a linear response between 30 and 800 µmol L-1 and a limit of detection (LOD) of 0.13 µmol L-1, by square wave voltammetry for dopamine and a linear range from 6.0 to 100 µmol L-1, with a LOD of 0.39 µmol L-1, by differential pulse voltammetry for serotonin. Later, the working electrode was modified with glucose oxidase and dihexadecyl phosphate film in order to obtain a biosensor. At this stage, CV was applied to detect glucose in the range of 1.0-10 µmol L-1 and LOD of 0.21 µmol L-1. By three different techniques and analytes, the sensoring and biosensoring processes presented high reproducibility. The proposed adhesive electrode is easy to prepare, disposable, within non-restrictive nature, which allows an approach of a new device for electrochemical sensing and biosensing.

Design of enzymatic cascade processes for the production of low-priced chemicals.

While the application of enzymes to synthetic and industrial problems continues to grow, the major development today is focused on multi-enzymatic cascades. Such systems are particularly attractive, because many commercially available enzymes operate under relatively similar operating conditions. This opens the possibility of one-pot operation with multiple enzymes in a single reactor. In this paper the concept of modules is introduced whereby groups of enzymes are combined in modules, each operating in a single reactor, but with the option of various operating strategies to avoid any complications of nonproductive interactions between the enzymes, substrates or products in a given reactor. In this paper the selection of modules is illustrated using the synthesis of the bulk chemical, gluconic acid, from lignocellulosic waste.

Uncovering the Formation of Color Gradients for Glucose Colorimetric Assays on Microfluidic Paper-Based Analytical Devices by Mass Spectrometry Imaging.

This study describes the use of mass spectrometry imaging with matrix-assisted laser desorption/ionization (MALDI) and desorption electrospray ionization (DESI) to understand the color gradient generation commonly seen in microfluidic paper-based analytical devices (µPADs). The formation of color gradients significantly impacts assay sensitivity and reproducibility with µPADs but the mechanism for formation is poorly understood. The glucose enzymatic assay using potassium iodide (KI) as a chromogenic agent was selected to investigate the color gradient generated across a detection spot. Colorimetric measurements revealed that the relative standard deviation for the recorded pixel intensities ranged between 34 and 40%, compromising the analytical reliability. While a variety of hypotheses have been generated to explain this phenomenon, few studies have attempted to elucidate the mechanisms associated with its formation. Mass spectrometry imaging using MALDI and DESI was applied to understand the nonuniform color distribution on the detection zone. MALDI experiments were first explored to monitor the spatial distribution of the glucose oxidase and horseradish peroxidase mixture, before and after lateral flow assay with and without KI. MALDI(+)-TOF data revealed uniform enzyme distribution on the detection spots. On the other hand, after the complete assay DESI(-) measurements revealed a heterogeneous shape indicating the presence of iodide and triiodide ions at the zone edge. The reaction product (I3-) is transported by lateral flow toward the zone edge, generating the color gradient. Mass spectrometry imaging has been used for the first time to prove that color gradient forms as result of the mobility small molecules and not the enzyme distribution on µPAD surface.

Impact of ultrasound on galactooligosaccharides and gluconic acid production throughout a multienzymatic system.

Galactooligosaccharides (GOS), recognised prebiotic, can be industrially produced from lactose and commercial ß-galactosidase (ß-gal) from Kluyveromyces lactis. Residual lactose and glucose limit GOS applications. To handle this problem, a multienzymatic system, with ß-gal and glucose oxidase (Gox), was proposed to reduce glucose content in reaction media through its oxidation to gluconic acid (GA). Besides, ultrasound (US) probe effect over the multienzymatic system to produce GOS and GA has been evaluated. A production around 40% of GOS was found in all treatments after the first hour of reaction. However, glucose consumption and GA production was significantly higher (P < 0.05) for sequential reaction assisted by US, obtaining the best production of GOS (49%) and GA (28%) after 2 h of reaction. The conformational and residual activity changes of enzymes under US conditions were also evaluated, Gox being positively affected whereas in ß-gal hardly any change was found.

Establecimiento de condiciones más eficientes para la producción de biosensores de glucosa / Establishment of more efficient conditions for the production of glucose biosensors

Los biosensores son dispositivos móviles que permiten detectar de forma rápida y sencilla enfermedades del metabolismo e infecciones víricas de interés veterinario y clínico, como el rotavirus y la hepatitis B y C. Este trabajo tuvo como objetivo determinar las variables significativas del proceso de producción de los biosensores de glucosa fabricados en el Centro de Inmunoensayo (La Habana, Cuba). Se produjeron ocho corridas experimentales teniendo en cuenta los procedimientos normativos de operación implementados en la planta de producción de biosensores y se realizaron las evaluaciones de calidad correspondientes (pruebas de exactitud) para liberar analíticamente los lotes producidos. Los experimentos realizados proporcionaron información acerca de cuáles variables deben controlarse con más cuidado durante la producción a fin de evitar altos niveles de productos no conformes o el comportamiento errático del proceso. Las variables seleccionadas para el estudio fueron las relacionadas con la preparación de la solución enzimática. Con los resultados obtenidos se realizó un análisis de regresión múltiple para determinar los factores estadísticamente significativos del modelo, obteniéndose un coeficiente de determinación superior al 90 por ciento, logrando explicar el 98,637 por ciento de la variación entre los valores de porcentaje de exactitud y la media. Los factores que resultaron ser significativos fueron la concentración de la enzima glucosa oxidasa, la concentración del mediador eléctrico y la conductividad del agua ultrapura para un nivel de confianza del 95 por ciento. El análisis realizado arrojó resultados satisfactorios demostrando que variando parámetros del proceso productivo es posible disminuir los valores del porcentaje de exactitud(AU)

Biosensors are mobile devices that allow rapid and easy detection of metabolic diseases and viral infections of veterinary and clinical interest, such as rotavirus and hepatitis B and C. The objective of this work was to determine the significant variables of the production process of the glucose biosensors manufactured in the Immunoassay Center (Havana, Cuba). Eight experimental runs were carried out taking into account the normative operating procedures of the biosensor production plant. Accuracy tests were carried out to release produced batches. The experiments provided information about which factors should be carefully controlled during the manufacture procedure in order to avoid high levels of faulty products or the erratic behavior of the process. The factors selected for the study were those related with the preparation of the enzymatic solution. A multiple regression analysis was carried out to determine the statistically significant factors of the model. The coefficient of determination was higher than 90 percent, and the 98.637 percent of the variation between the values of percentage of accuracy and the mean value could be explained. The significant factors were the concentration of the glucose oxidase enzyme, the electric mediator concentration, and the ultrapure water conductivity (95 percent confidence level). The analysis carried out showed satisfactory results. In the present study, it was demonstrated that varying parameters of the production process it is possible to decrease the accuracy percentage values(AU)