Sustainable Electropolymerization of Zingerone and Its C2 Symmetric Dimer for Amperometric Biosensor Films.

Polymeric permselective films are frequently used for amperometric biosensors to prevent electroactive interference present in the target matrix. Phenylenediamines are the most commonly used for the deposition of shielding polymeric films against interfering species; however, even phenolic monomers have been utilized in the creation of these films for microsensors and biosensors. The purpose of this paper is to evaluate the performances of electrosynthesized polymers, layered by means of constant potential amperometry (CPA), of naturally occurring compound zingerone (ZING) and its dimer dehydrozingerone (ZING DIM), which was obtained by straight oxidative coupling reaction. The polymers showed interesting shielding characteristics against the main interfering species, such as ascorbic acid (AA): actually, polyZING exhibited an AA shielding aptitude comprised between 77.6 and 99.6%, comparable to that obtained with PPD. Moreover, a marked capability of increased monitoring of hydrogen peroxide (HP), when data were compared with bare metal results, was observed. In particular, polyZING showed increases ranging between 55.6 and 85.6%. In the present work, the molecular structures of the obtained polymers have been theorized and docking analyses were performed to understand their peculiar characteristics better. The structures were docked using the Lamarckian genetic algorithm (LGA). Glutamate biosensors based on those polymers were built, and their performances were compared with biosensors based on PPD, which is the most widespread polymer for the construction of amperometric biosensors.

New perspective for an old drug: Can naloxone be considered an antioxidant agent?

Background: Experimental evidence indicates that Naloxone (NLX) holds antioxidant properties. The present study aims at verifying the hypothesis that NLX could prevent oxidative stress induced by hydrogen peroxide (H2O2) in PC12 cells. Methods: To investigate the antioxidant effect of NLX, initially, we performed electrochemical experiments by means of platinum-based sensors in a cell-free system. Subsequently, NLX was tested in PC12 cells on H2O2-induced overproduction of intracellular levels of reactive-oxygen-species (ROS), apoptosis, modification of cells' cycle distribution and damage of cells' plasma membrane. Results: This study reveals that NLX counteracts intracellular ROS production, reduces H2O2-induced apoptosis levels, and prevents the oxidative damage-dependent increases of the percentage of cells in G2/M phase. Likewise, NLX protects PC12 cells from H2O2- induced oxidative damage, by preventing the lactate dehydrogenase (LDH) release. Moreover, electrochemical experiments confirmed the antioxidant properties of NLX. Conclusion: Overall, these findings provide a starting point for studying further the protective effects of NLX on oxidative stress.

Inhibitory Effect of Curcumin-Inspired Derivatives on Tyrosinase Activity and Melanogenesis.

Tyrosinase is a well-known copper-containing metalloenzyme typically involved in the synthesis of melanin. Recently, curcumin and several synthetic derivatives have been recognized as tyrosinase inhibitors with interesting anti-melanogenic therapeutic activity. In this study, three curcumin-inspired compounds 1, 6 and 7 were prepared in yields ranging from 60 to 88 % and spectrophotometric, electrochemical, in vitro and in silico analyses were carried out. The viability of PC12 cells, a rat pheochromocytoma derived-cell line, with compounds 1, 6 and 7, showed values around 80% at 5 µM concentration. In cell proliferation assays, compounds 1, 6 and 7 did not show significant toxicity on fibroblasts nor melanoma cells up to 10 µM with viability values over 90%. The inhibition of tyrosinase activity was evaluated both by a UV-Vis spectroscopic method at two different concentrations, 0.2 and 2.0 µM, and by amperometric assay with IC50 for compounds 1, 6 and 7 ranging from 11 to 24 nM. Melanin content assays on human melanoma cells were performed to test the capability of compounds to inhibit melanin biosynthesis. All compounds exerted a decrease in melanin content, with compound 7 being the most effective by showing a melanogenesis inhibition up to four times greater than arbutin at 100 µM. Moreover, the antioxidant activity of the selected inhibitors was evaluated against H2O2 in amperometric experiments, whereby compound 7 was about three times more effective compared to compounds 1 and 6. The tyrosinase X-ray structure of Bacterium megaterium crystal was used to carry out molecular docking studies in the presence of compounds 1, 6 and 7 in comparison with that of kojic acid and arbutin, two conventional tyrosinase inhibitors. Molecular docking of compounds 6 and 7 confirmed the high affinity of these compounds to tyrosinase protein.

Quality control methods in musculoskeletal tissue engineering: from imaging to biosensors.

Tissue engineering is rapidly progressing toward clinical application. In the musculoskeletal field, there has been an increasing necessity for bone and cartilage replacement. Despite the promising translational potential of tissue engineering approaches, careful attention should be given to the quality of developed constructs to increase the real applicability to patients. After a general introduction to musculoskeletal tissue engineering, this narrative review aims to offer an overview of methods, starting from classical techniques, such as gene expression analysis and histology, to less common methods, such as Raman spectroscopy, microcomputed tomography, and biosensors, that can be employed to assess the quality of constructs in terms of viability, morphology, or matrix deposition. A particular emphasis is given to standards and good practices (GXP), which can be applicable in different sectors. Moreover, a classification of the methods into destructive, noninvasive, or conservative based on the possible further development of a preimplant quality monitoring system is proposed. Biosensors in musculoskeletal tissue engineering have not yet been used but have been proposed as a novel technology that can be exploited with numerous advantages, including minimal invasiveness, making them suitable for the development of preimplant quality control systems.

Epidemiology, Clinical Aspects, Laboratory Diagnosis and Treatment of Rickettsial Diseases in the Mediterranean Area During COVID-19 Pandemic: A Review of the Literature.

The purpose of the present review is to give an update regarding the classification, epidemiology, clinical manifestation, diagnoses, and treatment of the Rickettsial diseases present in the Mediterranean area. We performed a comprehensive search, through electronic databases (Pubmed - MEDLINE) and search engines (Google Scholar), of peer-reviewed publications (articles, reviews, and books). The availability of new diagnostic tools, including Polymerase Chain Reaction and nucleotide sequencing has significantly modified the classification of intracellular bacteria, including the order Rickettsiales with more and more new Rickettsia species recognized as human pathogens. Furthermore, emerging Rickettsia species have been found in several countries and are often associated with unique clinical pictures that may challenge the physician in the early detection of the diseases. Rickettsial infections include a wide spectrum of clinical presentations ranging from a benign to a potentially life treating disease that requires prompt recognition and proper management. Recently, due to the spread of SARS-CoV-2 infection, the differential diagnosis with COVID-19 is of crucial importance. The correct understanding of the clinical features, diagnostic tools, and proper treatment can assist clinicians in the management of Rickettsioses in the Mediterranean area.

Synthesis and Studies of the Inhibitory Effect of Hydroxylated Phenylpropanoids and Biphenols Derivatives on Tyrosinase and Laccase Enzymes.

The impaired activity of tyrosinase and laccase can provoke serious concerns in the life cycles of mammals, insects and microorganisms. Investigation of inhibitors of these two enzymes may lead to the discovery of whitening agents, medicinal products, anti-browning substances and compounds for controlling harmful insects and bacteria. A small collection of novel reversible tyrosinase and laccase inhibitors with a phenylpropanoid and hydroxylated biphenyl core was prepared using naturally occurring compounds and their activity was measured by spectrophotometric and electrochemical assays. Biosensors based on tyrosinase and laccase enzymes were constructed and used to detect the type of protein-ligand interaction and half maximal inhibitory concentration (IC50). Most of the inhibitors showed an IC50 in a range of 20-423 nM for tyrosinase and 23-2619 nM for laccase. Due to the safety concerns of conventional tyrosinase and laccase inhibitors, the viability of the new compounds was assayed on PC12 cells, four of which showed a viability of roughly 80% at 40 µM. In silico studies on the crystal structure of laccase enzyme identified a hydroxylated biphenyl bearing a prenylated chain as the lead structure, which activated strong and effective interactions at the active site of the enzyme. These data were confirmed by in vivo experiments performed on the insect model Tenebrio molitur.

Real-time telemetry monitoring of oxygen in the central complex of freely-walking Gromphadorhina portentosa.

A new telemetric system for the electrochemical monitoring of dissolved oxygen is showed. The device, connected with two amperometric sensors, has been successfully applied to the wireless detection of the extracellular oxygen in the central complex of freely-walking Gromphadorhina portentosa. The unit was composed of a potentiostat, a two-channel sensor conditioning circuit, a microprocessor module, and a wireless serial transceiver. The amperometric signals were digitalized and sent to a notebook using a 2.4 GHz transceiver while a serial-to-USB converter was connected to a second transceiver for completing the communication bridge. The software, running on the laptop, allowed to save and graph the oxygen signals. The electronics showed excellent stability and the acquired data was linear in a range comprised between 0 and -165 nA, covering the entire range of oxygen concentrations. A series of experiments were performed to explore the dynamics of dissolved oxygen by exposing the animals to different gases (nitrogen, oxygen and carbon dioxide), to low temperature and anesthetic agents (chloroform and triethylamine). The resulting data are in agreement with previous O2 changes recorded in the brain of awake rats and mice. The proposed system, based on simple and inexpensive components, can constitute a new experimental model for the exploration of central complex neurochemistry and it can also work with oxidizing sensors and amperometric biosensors.

The Presence of Polysaccharides, Glycerol, and Polyethyleneimine in Hydrogel Enhances the Performance of the Glucose Biosensor.

The use of amperometric biosensors has attracted particular attention in recent years, both from researchers and from companies, as they have proven to be low-cost, reliable, and very sensitive devices, with a wide range of uses in different matrices. The continuous development of amperometric biosensors, since their use involves an enzyme, is specifically aimed at keeping and increasing the catalytic properties of the loaded protein, so as to be able to use the same device over time. The present study aimed to investigate the impact of glycerol and polysaccharides, in the presence of polycationic substances to constitute a hydrogel, in enhancing the enzymatic and analytic performance of a glucose biosensor. Initially, it was possible to verify how the deposition of the starch-based hydrogel, in addition to allowing the electropolymerization of the poly(p-phenylenediamine) polymer and the maintenance of its ability to shield the ascorbic acid, did not substantially limit the permeability towards hydrogen peroxide. Moreover, different biosensor designs, loading a mixture containing all the components (alone or in combination) and the enzyme, were tested in order to evaluate the changes of the apparent enzyme kinetic parameters, such as VMAX and KM, and analytical response in terms of Linear Region Slope, highlighting how the presence of all components (starch, glycerol, and polyethyleneimine) were able to substantially enhance the performance of the biosensors. The surface analysis of the biosensors was performed by scanning electron microscope (SEM). More, it was shown that the same performances were kept unchanged for seven days, proving the suitability of this biosensor design for short- and mid-term use.

Low-Temperature Storage Improves the Over-Time Stability of Implantable Glucose and Lactate Biosensors.

Molecular biomarkers are very important in biology, biotechnology and even in medicine, but it is quite hard to convert biology-related signals into measurable data. For this purpose, amperometric biosensors have proven to be particularly suitable because of their specificity and sensitivity. The operation and shelf stability of the biosensor are quite important features, and storage procedures therefore play an important role in preserving the performance of the biosensors. In the present study two different designs for both glucose and lactate biosensor, differing only in regards to the containment net, represented by polyurethane or glutharaldehyde, were studied under different storage conditions (+4, -20 and -80 °C) and monitored over a period of 120 days, in order to evaluate the variations of kinetic parameters, as VMAX and KM, and LRS as the analytical parameter. Surprisingly, the storage at -80 °C yielded the best results because of an unexpected and, most of all, long-lasting increase of VMAX and LRS, denoting an interesting improvement in enzyme performances and stability over time. The present study aimed to also evaluate the impact of a short-period storage in dry ice on biosensor performances, in order to simulate a hypothetical preparation-conservation-shipment condition.

Use of ß-cyclodextrin as enhancer of ascorbic acid rejection in permselective films for amperometric biosensor applications.

Interference rejection in amperometric biosensors can be more effective introducing some modifiers during electro-deposition of permselective film. Addition of ß-cyclodextrin (ßCD), a cyclic oligosaccharide composed of seven glucose units, to the ortho-phenylendiamine (oPD) monomer were already demonstrated to provide an enhancement in ascorbic acid (AA) rejection. Here we evaluated the improvement in permselectivity of poly-eugenol and poly-magnolol films electro-polymerized in presence of different amounts of ßCD or eugenol-ßCD inclusion complex for amperometric biosensor application. Starting from Pt-Ir wire as transducer several microsensors were covered with polymeric films doped with ßCD-based modifiers through constant potential amperometry. Characterization of modified polymers was achieved by scanning electron microscopy and permselectivity analysis. Poly-magnolol film in combination with ßCD showed a worsening in permselectivity compared to poly-magnolol alone. In contrast, the introduction of ßCD-based modifier enhanced the interference rejection toward the archetypal interferent AA, while slightly affecting permeability toward H2O2 compared to the poly-eugenol without modifier. The AA rejection seems to be influenced by the availability of ßCD cavity as well as film performance due to concentration of ßCD-Eugenol inclusion complex. A poly-eugenol film co-polymerized with 2 mM ßCD-eugenol inclusion complex showed a permselectivity equal to poly-orthophenylendiamine film (PPD), with a lower permeability to AA, likely to be related with a self-blocking mechanism. Based on these results, a biosensor for glutamate was constructed with a poly-eugenol doped with ßCD-eugenol as permselective layer and its permselectivity, stability and lifetime were determined.

The role of molecular breast imaging in predicting complete tumor response to treatment and residual tumor extent following neoadjuvant therapy.

The aim of the present study was to investigate the usefulness of molecular breast imaging (MBI) in predicting complete tumor response to treatment and residual tumor extent following neoadjuvant therapy. A consecutive series of 43 female patients with large or locally advanced primary breast cancer, scheduled for surgery following neoadjuvant therapy, was retrospectively reviewed. Prior to surgery, all patients underwent MBI using a high­resolution semiconductor­based device for image acquisition. MBI data were correlated with surgical histopathological findings. Spearman's correlation coefficient was calculated to assess differences in residual tumor size with MBI and histopathological examination. From the images obtained using MBI, 7 patients were negative for residual tumors with pathological complete response (specificity, 100%) and positive in 34/36 patients with residual disease (sensitivity, 94.4%), including 26/27 patients with unifocal and 8/9 patients with multicentric/multifocal tumors, 5 of which exhibited multiple microscopic foci scattered in a fibrotic area. Overall accuracy was 95.3% and the positive predictive value (PPV) and negative predictive value (NPV) were 100 and 77.8%, respectively. MBI was false­negative in one patient with a 2.5­cm invasive ductal carcinoma located close to the chest wall and in one patient with microscopic foci of epithelial carcinoma. In the patients with unifocal residual tumors, correlation of tumor size between MBI and histopathology was r=0.981 (P<0.0001); however, MBI overestimated the number of lesions in one of these cases. In the patients with multifocal/multicentric tumors, MBI adequately assessed residual tumor extent in 5/8 positive cases, overestimating the number of lesions in one case and underestimating tumor extent in 2 further cases with microscopic foci scattered in a fibrotic area. MBI proved to be a highly accurate diagnostic tool in predicting complete tumor response to neoadjuvant therapy and residual tumor extent, correlating with surgical histopathological findings in 86% of overall cases. A positive result was always associated with the presence of residual disease and MBI tumor size was strongly correlated with histopathological analysis mainly in unifocal residual tumors.

A MOF-based carrier for in situ dopamine delivery.

MIL-88A (Fe) MOF crystals were nucleated and grown around a polymer core containing superparamagnetic nanoparticles to assemble a new class of biocompatible particles for magnetophoretic drug delivery of dopamine. The carrier enabled efficient targeted release, dopamine protection from oxidative damage, long-term delivery and improved drug delivery cost-efficiency. After loading, dopamine was stable within the carrier and did not undergo oxidation. Drug release monitoring via spectrofluorimetry revealed a shorter burst effect and higher release efficiency than silica based carriers. The in vitro cytotoxicity at different MOF concentrations and sizes was assessed using PC12 cells as the neuronal cell model. The drug was directly uptaken into the PC12 cells avoiding possible side effects due to oxidation occurring in the extracellular environment.

Synthesis of Nitric Oxide Donors Derived from Piloty's Acid and Study of Their Effects on Dopamine Secretion from PC12 Cells.

This study investigated the mechanisms and kinetics of nitric oxide (NO) generation by derivatives of Piloty's acid (NO-donors) under physiological conditions. In order to qualitatively and quantitatively measure NO release, electron paramagnetic resonance (EPR) was carried out with NO spin trapping. In addition, voltammetric techniques, including cyclic voltammetry and constant potential amperometry, were used to confirm NO release from Piloty's acid and its derivatives. The resulting data showed that Piloty's acid derivatives are able to release NO under physiological conditions. In particular, electron-withdrawing substituents favoured NO generation, while electron-donor groups reduced NO generation. In vitro microdialysis, performed on PC12 cell cultures, was used to evaluate the dynamical secretion of dopamine induced by the Piloty's acid derivatives. Although all the studied molecules were able to induce DA secretion from PC12, only those with a slow release of NO have not determined an autoxidation of DA itself. These results confirm that the time-course of NO-donors decomposition and the amount of NO released play a key role in dopamine secretion and auto-oxidation. This information could drive the synthesis or the selection of compounds to use as potential drugs for the therapy of Parkinson's disease (PD).

Is catalase involved in the effects of systemic and pVTA administration of 4-methylpyrazole on ethanol self-administration?

The oxidative metabolism of ethanol into acetaldehyde involves several enzymes, including alcohol dehydrogenase (ADH) and catalase-hydrogen peroxide (H2O2). In this regard, while it is well known that 4-methylpyrazole (4-MP) acts by inhibiting ADH in the liver, little attention has been placed on its ability to interfere with fatty acid oxidation-mediated generation of H2O2, a mechanism that may indirectly affect catalase whose enzymatic activity requires H2O2. The aim of our investigation was twofold: 1) to evaluate the effect of systemic (i.p. [intraperitoneal]) and local (into the posterior ventral tegmental area, pVTA) administration of 4-MP on oral ethanol self-administration, and 2) to assess ex vivo whether or not systemic 4-MP affects liver and brain H2O2 availability. The results show that systemic 4-MP reduced ethanol but not acetaldehyde or saccharin self-administration, and decreased the ethanol deprivation effect. Moreover, local intra-pVTA administration of 4-MP reduced ethanol but not saccharin self-administration. In addition, although unable to affect basal catalase activity, systemic administration of 4-MP decreased H2O2 availability both in liver and in brain. Overall, these results indicate that 4-MP interferes with ethanol self-administration and suggest that its behavioral effects could be due to a decline in catalase-H2O2 system activity as a result of a reduction of H2O2 availability, thus highlighting the role of central catalase-mediated metabolism of ethanol and further supporting the key role of acetaldehyde in the reinforcing properties of ethanol.

123I-ioflupane brain SPECT and 123I-MIBG cardiac planar scintigraphy combined use in uncertain parkinsonian disorders.

We evaluated the clinical usefulness of the combined use of I-ioflupane brain single photon emission computed tomography (SPECT) and I-metaiodobenzylguanidine (MIBG) cardiac scintigraphy in discriminating uncertain parkinsonism with vascular lesions in striatal nuclei at magnetic resonance imaging (MRI). Forty-three consecutive patients with uncertain parkinsonism and vascular lesions at MRI in striatal nuclei were retrospectively evaluated; the uncertain differential diagnosis was between Parkinson's disease and vascular parkinsonism (PD/VP) in 22 patients, between PD and other neurodegenerative parkinsonism (PD/PS) in 11 patients and between Lewy body dementia and Alzheimer disease (LBD/AD) in the remaining 10 cases. All patients underwent I-ioflupane SPECT with striatal dopaminergic activity determination as binding potentials (BP; cut-off: 3.3). I-MIBG cardiac planar scintigraphy was performed 2 weeks later, in early (15 minutes) and delayed (240 minutes) phases also calculating heart to mediastinum (H/M) ratio (cut-off: 1.56). I-Ioflupane uptake was normal in 9 patients with BP values >3.3, while it was reduced in 34/43 cases with BP values <3.3 at least in one of the striatal nuclei. I-MIBG uptake was normal in 21/43 patients (5 of whom with normal and 16 with I-ioflupane striatal defects) showing the H/M ratio >1.56 in all cases; the uptake was reduced in 22/43 cases, (4 of whom were normal and 18 were with I-ioflupane striatal defects) with the H/M ratio <1.56 in all cases. No statistical differences were found when early and delayed H/M ratios were mutually compared. Combining the 2 radioisotopic procedures, a more reliable diagnosis was achieved in 39/43 cases properly classifying 13 PD, 10 VP, 7 PS, 5 LBD, and 4 AD. However, the diagnosis remained uncertain in four patients with normal I-ioflupane and reduced I-MIBG uptake. The results of the present study confirmed that in uncertain parkinsonian syndromes associated with vascular lesions in striatal nuclei, brain I-ioflupane SPECT alone did not prove able to discriminate between the different forms of disease. Only the association with I-MIBG cardiac scintigraphy, also with the early acquisition alone, allowed the most appropriate diagnosis in 90.7% of our cases. However, patients with normal I-ioflupane and reduced I-I-MIBG uptakes need a close clinical and instrumental follow-up as sympathetic damage could precede striatal disorders in the early stage of PD and LBD.

Hydroxylated biphenyls as tyrosinase inhibitor: A spectrophotometric and electrochemical study.

A small collection of C2-symmetry hydroxylated biphenyls was prepared by straightforward methods and the capability to act as inhibitors of tyrosinase has been evaluated by both spectrophotometric and electrochemical assays. Our attention was focused on the diphenolase activity of this enzyme characterized by the absence of the characteristic lag time of enzymatic reaction of its monophenolase activity. To this purpose, we evaluated the capability of tyrosinase to oxidize a natural o-diphenol substrate to o-quinone analyzing the changes in the UV-Vis spectrum of a solution of caffeic acid and the reduction of the cathodic current in a tyrosinase-biosensor, respectively. Results of both the methods were comparable. Most of the compounds possessed higher inhibitory activity compared to compound 1, a known hydroxylated biphenyl inhibitor of tyrosinase.

Development of a biosensor telemetry system for monitoring fermentation in craft breweries.

The development and applications of biosensors in the food industry has had a rapid grown due to their sensitivity, specificity and simplicity of use with respect to classical analytical methods. In this study, glucose and ethanol amperometric biosensors integrated with a wireless telemetry system were developed and used for the monitoring of top and bottom fermentations in beer wort samples. The collected data were in good agreement with those obtained by reference methods. The simplicity of construction, the low cost and the short time of analysis, combined with easy interpretation of the results, suggest that these devices could be a valuable alternative to conventional methods for monitoring fermentation processes in the food industry.

Real-time monitoring of glucose and phenols intestinal absorption through an integrated Caco-2TC7cells/biosensors telemetric device: Hypoglycemic effect of fruit phytochemicals.

An integrated device for real-time monitoring of glucose and phenols absorption, that consists of a sensors/biosensors system (SB) and a Caco-2TC7 human intestinal cell culture, is described in this study. The SB is composed of a glucose oxidase-based biosensor, a sentinel platinum sensor, a laccase/tyrosinase-based biosensor and a sentinel carbon sensor, all located in the basolateral compartment (BC) of a cell culture plate. Caco-2TC7 cells, differentiated on culture inserts, separated the apical compartment that simulates the intestinal lumen, from the BC which represented the bloodstream. The system recorded currents relative to glucose (1mM) absorption, obtaining bioavailability values (5.1%) comparable to HPLC analysis (4.8%). Phloridzin and phloretin, specific phenolic inhibitors of SGLT1 and GLUT2 glucose transporters, reduced the glucose transport of almost 10 times. They were minimally absorbed in the BC with a bioavailability of 0.13% and 0.49% respectively. The hypoglycemic potential of blueberry and pomegranate juices was also studied. In particular, the amount of glucose absorbed through the Caco-2TC7 monolayer was 8‰ for pomegranate and 1.7‰ for blueberry, demonstrating the potential hypoglycemic effect of the juices. Polyphenols absorption was also monitored by the SB and an increase was recorded during the first 50min in presence of both blueberry and pomegranate juices, then a constant decrease occurred. The proposed device has been developed as innovative tool for the dynamic monitoring of natural compounds effects on glucose absorption, in order to manage postprandial hyperglycemia.

Enzyme Biosensors for Biomedical Applications: Strategies for Safeguarding Analytical Performances in Biological Fluids.

Enzyme-based chemical biosensors are based on biological recognition. In order to operate, the enzymes must be available to catalyze a specific biochemical reaction and be stable under the normal operating conditions of the biosensor. Design of biosensors is based on knowledge about the target analyte, as well as the complexity of the matrix in which the analyte has to be quantified. This article reviews the problems resulting from the interaction of enzyme-based amperometric biosensors with complex biological matrices containing the target analyte(s). One of the most challenging disadvantages of amperometric enzyme-based biosensor detection is signal reduction from fouling agents and interference from chemicals present in the sample matrix. This article, therefore, investigates the principles of functioning of enzymatic biosensors, their analytical performance over time and the strategies used to optimize their performance. Moreover, the composition of biological fluids as a function of their interaction with biosensing will be presented.