Magnetic separation and concentration of Aß 1-42 molecules dispersed at the threshold concentration for Alzheimer's disease diagnosis in clinically-relevant volumes of sample.

BACKGROUND: Alzheimer's disease (AD) is the leading cause of dementia and loss of autonomy in the elderly, implying a progressive cognitive decline and limitation of social activities. The progressive aging of the population is expected to exacerbate this problem in the next decades. Therefore, there is an urgent need to develop quantitative diagnostic methodologies to assess the onset the disease and its progression especially in the initial phases. RESULTS: Here we describe a novel technology to extract one of the most important molecular biomarkers of AD (Aß1-42) from a clinically-relevant volume - 100 µl - therein dispersed in a range of concentrations critical for AD early diagnosis. We demonstrate that it is possible to immunocapture Aß1-42 on 20 nm wide magnetic nanoparticles functionalized with hyperbranced KVLFF aptamers. Then, it is possible to transport them through microfluidic environments to a detection system where virtually all (~ 90%) the Aß1-42 molecules are concentrated in a dense plug of ca.50 nl. The technology is based on magnetic actuation by permanent magnets, specifically designed to generate high gradient magnetic fields. These fields, applied through submillimeter-wide channels, can concentrate, and confine magnetic nanoparticles (MNPs) into a droplet with an optimized shape that maximizes the probability of capturing highly diluted molecular biomarkers. These advancements are expected to provide efficient protocols for the concentration and manipulation of molecular biomarkers from clinical samples, enhancing the accuracy and the sensitivity of diagnostic technologies. CONCLUSIONS: This easy to automate technology allows an efficient separation of AD molecular biomarkers from volumes of biological solutions complying with the current clinical protocols and, ultimately, leads to accurate measurements of biomarkers. The technology paves a new way for a quantitative AD diagnosis at the earliest stage and it is also adaptable for the biomarker analysis of other pathologies.

Organotypic Hippocampal Slice Cultures from Adult Tauopathy Mice and Theragnostic Evaluation of Nanomaterial Phospho-TAU Antibody-Conjugates.

Organotypic slice culture models surpass conventional in vitro methods in many aspects. They retain all tissue-resident cell types and tissue hierarchy. For studying multifactorial neurodegenerative diseases such as tauopathies, it is crucial to maintain cellular crosstalk in an accessible model system. Organotypic slice cultures from postnatal tissue are an established research tool, but adult tissue-originating systems are missing, yet necessary, as young tissue-originating systems cannot fully model adult or senescent brains. To establish an adult-originating slice culture system for tauopathy studies, we made hippocampal slice cultures from transgenic 5-month-old hTau.P301S mice. In addition to the comprehensive characterization, we set out to test a novel antibody for hyperphosphorylated TAU (pTAU, B6), with and without a nanomaterial conjugate. Adult hippocampal slices retained intact hippocampal layers, astrocytes, and functional microglia during culturing. The P301S-slice neurons expressed pTAU throughout the granular cell layer and secreted pTAU to the culture medium, whereas the wildtype slices did not. Additionally, cytotoxicity and inflammation-related determinants were increased in the P301S slices. Using fluorescence microscopy, we showed target engagement of the B6 antibody to pTAU-expressing neurons and a subtle but consistent decrease in intracellular pTAU with the B6 treatment. Collectively, this tauopathy slice culture model enables measuring the extracellular and intracellular effects of different mechanistic or therapeutic manipulations on TAU pathology in adult tissue without the hindrance of the blood-brain barrier.

Mortalidad en adultos con COVID-19: experiencia de la Unidad de Cuidados Intensivos de un Hospital de Tercer Nivel de Paraguay

Introducción: diversos factores pueden influir en la mortalidad de pacientes hospitalizados por COVID-19. Objetivo: determinar la mortalidad y los factores asociados a esta en adultos con COVID-19 hospitalizados en la unidad de cuidados intensivos de un Hospital de Tercer Nivel de Paraguay. Metodología: estudio observacional, descriptivo, de asociación cruzada, de corte transversal y temporalmente retrospectivo. Se incluyó historias clínicas de pacientes adultos, de ambos sexos, que tenían diagnóstico confirmado (por prueba de antígeno y/o PCR) de infección por SARS-CoV-2 y que estaban hospitalizados en la unidad de cuidados intensivos de un Hospital General de Tercer Nivel de Paraguay. Resultados: Se incluyeron 116 pacientes, de los cuales el 54 % correspondió al sexo masculino. La edad media fue de 57±12,9 años. El 51 % tenía hipertensión arterial y el 29 % diabetes mellitus. El requerimiento de ventilación mecánica se dio en un 85% de pacientes. El 75 % de los pacientes ventilados tuvo un desenlace fatal. Se encontró una asociación estadísticamente significativa entre la presencia de infecciones bacterianas y requerimiento de hemodiálisis y el desenlace fatal (p=0,0074 y p=0,00011, respectivamente). La media de las edades de los pacientes fallecidos fue de 59,5 años, mientras que el grupo de pacientes que recibieron el alta desde la unidad de cuidados intensivos arrojó una media de 54,2 años. La diferencia entre estas edades en relación con el óbito fue significativa, con una p<0,05. Discusión: La mortalidad general debido a COVID-19 fue de más de 6 por cada 10 pacientes, siendo más alta en aquellos pacientes con ventilación. Aquellos pacientes que presentaron sobreinfección bacteriana o requirieron de hemodiálisis durante el curso de la hospitalización presentaron un peor desenlace en comparación con los pacientes que no presentaron este tipo de complicaciones.

Introduction: Several factors may influence mortality in patients hospitalized with COVID-19. Objective: This research aimed to determine mortality and associated factors in adults with COVID-19 hospitalized in the intensive care unit of a Third Level Hospital in Paraguay. Methodology: Observational, descriptive of cross-association, cross-sectional, and retrospective study. We included medical records of adult patients, of both sexes, who had a confirmed diagnosis (by antigen and/or PCR test) of SARS-CoV-2 infection and who were hospitalized in the intensive care unit of a Third Level General Hospital in Paraguay. Results: We included 116 patients, 54% of whom were male. The mean age was 57 ± 12.9 years. Of participants, 51% had hypertension and 29% diabetes mellitus. Mechanical ventilation was required in 85% of the patients. Of ventilated patients, 75% had a fatal outcome. A statistically significant association was found between the presence of bacterial infections and hemodialysis requirement and fatal outcome (p=0.0074 and p=0.00011, respectively). The mean age of the deceased patients was 59.5 years, while the group of patients discharged from the intensive care unit had a mean age of 54.2 years. The difference between these ages in relation to death was significant, with a p<0.05. Discussion: Overall mortality due to COVID-19 was more than 6 per 10 patients, being higher in those patients with ventilation. Those patients who presented bacterial superinfection or required hemodialysis during hospitalization had a worse outcome compared to patients who did not present this type of complications.

Reporte preliminar de las características sociodemográficas y clínicas de los pacientes con hemopatías malignas: experiencia de un hospital universitario

Introducción: debido a su amplia diversidad clínica y biológica, es de gran importancia para el médico conocer cómo se presentan los pacientes diagnosticados con una hemopatía maligna. Objetivo: describir las características sociodemográficas y clínicas de los pacientes con diagnóstico de una hemopatía maligna que consultaron en un Hospital Universitario de Paraguay. Metodología: estudio observacional, retrospectivo, descriptivo, de corte transversal, con muestreo no probabilístico, de casos consecutivos. Se incluyó a pacientes adultos, de ambos sexos, registrados en el archivo de Hematología del Hospital de Clínicas de la Facultad de Ciencias Médicas de la Universidad Nacional de Asunción (Paraguay), con diagnóstico confirmado por anatomía patológica de una neoplasia hematológica, según la clasificación de la Organización Mundial de la Salud, durante el periodo comprendido entre enero 2019 y agosto de 2021. Se estudiaron variables sociodemográficas y clínicas. Resultados: se estudió a 129 pacientes. El 58 % correspondía al sexo masculino. La edad promedio de los pacientes fue de 50,3±17,38. La mayoría de los pacientes procedía del Departamento Central de Paraguay (56,58 %). La mayoría de los pacientes (n=37) fue diagnosticado con un linfoma de tipo no Hodgkin o con una neoplasia con origen en células plasmáticas (n=32). El tiempo promedio de aparición de los síntomas hasta la consulta fue de 3,5 meses. El síntoma más reportado fue la astenia (35 %). La palidez fue el hallazgo positivo más frecuente al examen físico. Las comorbilidades más frecuentes fueron la hipertensión arterial (39,44 %) y la diabetes mellitus (15,59 %). Discusión: el paciente típico con una hemopatía que consulta en el Hospital de Clínicas es hombre, mayor de 50 años, proveniente del Departamento Central, consulta por astenia, presenta palidez al examen físico y ha tardado más de 3 meses en consultar. Se le ha diagnosticado un linfoma no Hodgkin y tiene comorbilidades cardiovasculares o metabólicas.

Introduction: due to their wide clinical and biological diversity, it is of great importance for the physician to know how patients diagnosed with a hematological malignancy present. Objective: to describe the sociodemographic and clinical characteristics of patients diagnosed with a hematological malignancy who consulted at a University Hospital in Paraguay. Methods: observational, retrospective, descriptive, cross-sectional, retrospective, descriptive, cross-sectional study, with non-probabilistic sampling of consecutive cases. We included adult patients, of both sexes, registered in the Hematology file of the Hospital de Clínicas of the Faculty of Medical Sciences of the National University of Asunción (Paraguay), with diagnosis confirmed by pathological anatomy of a hematological neoplasm, according to the classification of the World Health Organization, during the period from January 2019 to August 2021. Sociodemographic and clinical variables were studied. Results: 129 patients were studied. Fifty-eight percent were male. The mean age of the patients was 50.3±17.38. Most of the patients were from the Central Department of Paraguay (56.58 %). Most of the patients (n=37) were diagnosed with a non-Hodgkin's lymphoma or a neoplasm with plasma cell origin (n=32). The average time from symptom onset to consultation was 3.5 months. The most reported symptom was asthenia (35%). Pallor was the most frequent positive finding on physical examination. The most frequent comorbidities were arterial hypertension (39.44%) and diabetes mellitus (15.59%). Discussion: the typical patient with a hemopathy who consults at the Hospital de Clínicas is male, over 50 years of age, from the Central Department, consults for asthenia, presents pallor on physical examination and has taken more than 3 months to consult. He has been diagnosed with non-Hodgkin's lymphoma and has cardiovascular or metabolic comorbidities.

Exploring the Effect of Halogenation in a Series of Potent and Selective A2B Adenosine Receptor Antagonists.

The modulation of the A2B adenosine receptor is a promising strategy in cancer (immuno) therapy, with A2BAR antagonists emerging as immune checkpoint inhibitors. Herein, we report a systematic assessment of the impact of (di- and mono-)halogenation at positions 7 and/or 8 on both A2BAR affinity and pharmacokinetic properties of a collection of A2BAR antagonists and its study with structure-based free energy perturbation simulations. Monohalogenation at position 8 produced potent A2BAR ligands irrespective of the nature of the halogen. In contrast, halogenation at position 7 and dihalogenation produced a halogen-size-dependent decay in affinity. Eight novel A2BAR ligands exhibited remarkable affinity (Ki < 10 nM), exquisite subtype selectivity, and enantioselective recognition, with some eutomers eliciting sub-nanomolar affinity. The pharmacokinetic profile of representative derivatives showed enhanced solubility and microsomal stability. Finally, two compounds showed the capacity of reversing the antiproliferative effect of adenosine in activated primary human peripheral blood mononuclear cells.

Role of Dipolar Interactions on the Determination of the Effective Magnetic Anisotropy in Iron Oxide Nanoparticles.

Challenging magnetic hyperthermia (MH) applications of immobilized magnetic nanoparticles require detailed knowledge of the effective anisotropy constant (Keff ) to maximize heat release. Designing optimal MH experiments entails the precise determination of magnetic properties, which are, however, affected by the unavoidable concurrence of magnetic interactions in common experimental conditions. In this work, a mean-field energy barrier model (ΔE), accounting for anisotropy (EA ) and magnetic dipolar (ED ) energy, is proposed and used in combination with AC measurements to a specifically developed model system of spherical magnetic nanoparticles with well-controlled silica shells, acting as a spacer between the magnetic cores. This approach makes it possible to experimentally demonstrate the mean field dipolar interaction energy prediction with the interparticle distance, dij , ED ≈ 1/dij 3 and obtain the EA as the asymptotic limit for very large dij . In doing so, Keff uncoupled from interaction contributions is obtained for the model system (iron oxide cores with average sizes of 8.1, 10.2, and 15.3 nm) revealing to be 48, 23, and 11 kJ m-3 , respectively, close to bulk magnetite/maghemite values and independent from the specific spacing shell thicknesses selected for the study.

Synthesis and Characterization of a Biocompatible Nanoplatform Based on Silica-Embedded SPIONs Functionalized with Polydopamine.

Superparamagnetic iron oxide nanoparticles (SPIONs) have gained increasing interest in nanomedicine, but most of those that have entered the clinical trials have been withdrawn due to toxicity concerns. Therefore, there is an urgent need to design low-risk and biocompatible SPION formulations. In this work, we present an original safe-by-design nanoplatform made of silica nanoparticles loaded with SPIONs and decorated with polydopamine (SPIONs@SiO2-PDA) and the study of its biocompatibility performance by an ad hoc thorough in vitro to in vivo nanotoxicological methodology. The results indicate that the SPIONs@SiO2-PDA have excellent colloidal stability in serum-supplemented culture media, even after long-term (24 h) exposure, showing no cytotoxic or genotoxic effects in vitro and ex vivo. Physiological responses, evaluated in vivo using Caenorhabditis elegans as the animal model, showed no impact on fertility and embryonic viability, induction of an oxidative stress response, and a mild impact on animal locomotion. These tests indicate that the synergistic combination of the silica matrix and PDA coating we developed effectively protects the SPIONs, providing enhanced colloidal stability and excellent biocompatibility.

Significant Surface Spin Effects and Exchange Bias in Iron Oxide-Based Hollow Magnetic Nanoparticles.

Exchange bias (EB) properties have become especially important in hollow magnetic nanoparticles (MNPs) due to the versatility and reduced size of these materials. In this work, we present the synthesis and study of the EB properties of iron-oxide-based hollow MNPs and their precursors Fe/iron oxide MNPs with core/void/shell structure. The two mechanisms involved in EB generation were investigated: the frozen spins present in the nanograins that form the nanoparticles and the surface spins. The effect of external parameters on the coercivity (HC), remanence (MR), exchange bias field (HEB) and frozen spins, such as cooling field (HFC) and temperature, was investigated. Both HC and HEB present a maximum threshold above which their values begin to decrease with HFC, showing a new trend of HEB with HFC and allowing modulation on demand. The existence of surface spins, present on the outer and inner surfaces, was demonstrated, and an intrinsic EB phenomenon (HEB = 444 Oe for hollow iron oxide-based MNPs of 13.1 nm) with significant magnetization (MS~50 emu/g) was obtained. Finally, core/void/shell MNPs of 11.9 nm prior to the formation of the hollow MNPs showed a similar behavior, with non-negligible HEB, highlighting the importance of surface spins in EB generation.

Versatile Mesoporous Nanoparticles for Cell Applications.

Mesoporous silica nanostructures are emerging as a promising platform able to deal with challenges of many different applications in fields such as biomedicine and nanotechnology. The versatile physical and functional properties of these materials like high specific surface area, ordered porosity, chemical stability under temperature and pH variations, and biocompatible performance, offers new approaches to many biomedical applications ranging from drug delivery systems to biosensing, cell applications and tissue engineering. Their morphology, size and textural properties can be easily tailored by means of chemical control, giving rise to a variety of nanostructures with hexagonal (SBA15, MCM41) or cubic (SBA16) arrangement of channels and pore size ranging from 1.3 to 10 nm. Based on the versatility of their silane surface, a plethora of hybrid mesoporous matrices can be prepared incorporating new functionalities like contrast enhancement for magnetic resonance imaging, magnetic/plasmonic hyperthermia, drug delivery or cell applications by the simple grafting of superparamagnetic metal oxides (Fe3O4, transition metal ferrites) nanoparticles, noble metal (Au, Ag) nanoparticles, fluorescent moieties (fluorescein, rhodamine) or biological agents (mAb, mRNA, etc). The goal of this work is to present the development, by a facile soft template method, of size tailored mesoporous silica nanospheres from 20 to 350 nm (by means of chemical control), and highlight its versatility for surface grafting (with rhodamine and polydopamine) and their biological compatibility and efficient uptake by cultured HeLa cells. The combined, physicochemical and biological, properties indicate that MSNs are good candidates for cell tagging, gene transfer or targeted therapies.

Carbon-Coated Superparamagnetic Nanoflowers for Biosensors Based on Lateral Flow Immunoassays.

Superparamagnetic iron oxide nanoflowers coated by a black carbon layer (Fe3O4@C) were studied as labels in lateral flow immunoassays. They were synthesized by a one-pot solvothermal route, and they were characterized (size, morphology, chemical composition, and magnetic properties). They consist of several superparamagnetic cores embedded in a carbon coating holding carboxylic groups adequate for bioconjugation. Their multi-core structure is especially efficient for magnetic separation while keeping suitable magnetic properties and appropriate size for immunoassay reporters. Their functionality was tested with a model system based on the biotin-neutravidin interaction. For this, the nanoparticles were conjugated to neutravidin using the carbodiimide chemistry, and the lateral flow immunoassay was carried out with a biotin test line. Quantification was achieved with both an inductive magnetic sensor and a reflectance reader. In order to further investigate the quantifying capacity of the Fe3O4@C nanoflowers, the magnetic lateral flow immunoassay was tested as a detection system for extracellular vesicles (EVs), a novel source of biomarkers with interest for liquid biopsy. A clear correlation between the extracellular vesicle concentration and the signal proved the potential of the nanoflowers as quantifying labels. The limit of detection in a rapid test for EVs was lower than the values reported before for other magnetic nanoparticle labels in the working range 0-3 × 107 EVs/µL. The method showed a reproducibility (RSD) of 3% (n = 3). The lateral flow immunoassay (LFIA) rapid test developed in this work yielded to satisfactory results for EVs quantification by using a precipitation kit and also directly in plasma samples. Besides, these Fe3O4@C nanoparticles are easy to concentrate by means of a magnet, and this feature makes them promising candidates to further reduce the limit of detection.

Simultaneous in vivo PET/MRI using fluorine-18 labeled Fe3O4@Al(OH)3 nanoparticles: comparison of nanoparticle and nanoparticle-labeled stem cell distribution.

BACKGROUND: Mesenchymal stem cells (MSCs) have shown potential for treatment of different diseases. However, their working mechanism is still unknown. To elucidate this, the non-invasive and longitudinal tracking of MSCs would be beneficial. Both iron oxide-based nanoparticles (Fe3O4 NPs) for magnetic resonance imaging (MRI) and radiotracers for positron emission tomography (PET) have shown potential as in vivo cell imaging agents. However, they are limited by their negative contrast and lack of spatial information as well as short half-life, respectively. In this proof-of-principle study, we evaluated the potential of Fe3O4@Al(OH)3 NPs as dual PET/MRI contrast agents, as they allow stable binding of [18F]F- ions to the NPs and thus, NP visualization and quantification with both imaging modalities. RESULTS: 18F-labeled Fe3O4@Al(OH)3 NPs (radiolabeled NPs) or mouse MSCs (mMSCs) labeled with these radiolabeled NPs were intravenously injected in healthy C57Bl/6 mice, and their biodistribution was studied using simultaneous PET/MRI acquisition. While liver uptake of radiolabeled NPs was seen with both PET and MRI, mMSCs uptake in the lungs could only be observed with PET. Even some initial loss of fluoride label did not impair NPs/mMSCs visualization. Furthermore, no negative effects on blood cell populations were seen after injection of either the NPs or mMSCs, indicating good biocompatibility. CONCLUSION: We present the application of novel 18F-labeled Fe3O4@Al(OH)3 NPs as safe cell tracking agents for simultaneous PET/MRI. Combining both modalities allows fast and easy NP and mMSC localization and quantification using PET at early time points, while MRI provides high-resolution, anatomic background information and long-term NP follow-up, hereby overcoming limitations of the individual imaging modalities.

Tribological Behavior of Nanolubricants Based on Coated Magnetic Nanoparticles and Trimethylolpropane Trioleate Base Oil.

The main task of this work is to study the tribological performance of nanolubricants formed by trimethylolpropane trioleate (TMPTO) base oil with magnetic nanoparticles coated with oleic acid: Fe3O4 of two sizes 6.3 nm and 10 nm, and Nd alloy compound of 19 nm. Coated nanoparticles (NPs) were synthesized via chemical co-precipitation or thermal decomposition by adsorption with oleic acid in the same step. Three nanodispersions of TMPTO of 0.015 wt% of each NP were prepared, which were stable for at least 11 months. Two different types of tribological tests were carried out: pure sliding conditions and rolling conditions (5% slide to roll ratio). With the aim of analyzing the wear by means of the wear scar diameter (WSD), the wear track depth and the volume of the wear track produced after the first type of the tribological tests, a 3D optical profiler was used. The best tribological performance was found for the Nd alloy compound nanodispersion, with reductions of 29% and 67% in friction and WSD, respectively, in comparison with TMPTO. On the other hand, rolling conditions tests were utilized to study friction and film thickness of nanolubricants, determining that Fe3O4 (6.3 nm) nanolubricant reduces friction in comparison to TMPTO.

Development of Superparamagnetic Nanoparticles Coated with Polyacrylic Acid and Aluminum Hydroxide as an Efficient Contrast Agent for Multimodal Imaging.

Early diagnosis of disease and follow-up of therapy is of vital importance for appropriate patient management since it allows rapid treatment, thereby reducing mortality and improving health and quality of life with lower expenditure for health care systems. New approaches include nanomedicine-based diagnosis combined with therapy. Nanoparticles (NPs), as contrast agents for in vivo diagnosis, have the advantage of combining several imaging agents that are visible using different modalities, thereby achieving high spatial resolution, high sensitivity, high specificity, morphological, and functional information. In this work, we present the development of aluminum hydroxide nanostructures embedded with polyacrylic acid (PAA) coated iron oxide superparamagnetic nanoparticles, Fe3O4@Al(OH)3, synthesized by a two-step co-precipitation and forced hydrolysis method, their physicochemical characterization and first biomedical studies as dual magnetic resonance imaging (MRI)/positron emission tomography (PET) contrast agents for cell imaging. The so-prepared NPs are size-controlled, with diameters below 250 nm, completely and homogeneously coated with an Al(OH)3 phase over the magnetite cores, superparamagnetic with high saturation magnetization value (Ms = 63 emu/g-Fe3O4), and porous at the surface with a chemical affinity for fluoride ion adsorption. The suitability as MRI and PET contrast agents was tested showing high transversal relaxivity (r2) (83.6 mM-1 s-1) and rapid uptake of 18F-labeled fluoride ions as a PET tracer. The loading stability with 18F-fluoride was tested in longitudinal experiments using water, buffer, and cell culture media. Even though the stability of the 18F-label varied, it remained stable under all conditions. A first in vivo experiment indicates the suitability of Fe3O4@Al(OH)3 nanoparticles as a dual contrast agent for sensitive short-term (PET) and high-resolution long-term imaging (MRI).

Discovery of Potent and Highly Selective A2B Adenosine Receptor Antagonist Chemotypes.

Three novel families of A2B adenosine receptor antagonists were identified in the context of the structural exploration of the 3,4-dihydropyrimidin-2(1H)-one chemotype. The most appealing series contain imidazole, 1,2,4-triazole, or benzimidazole rings fused to the 2,3-positions of the parent diazinone core. The optimization process enabled identification of a highly potent (3.49 nM) A2B ligand that exhibits complete selectivity toward A1, A2A, and A3 receptors. The results of functional cAMP experiments confirmed the antagonistic behavior of representative ligands. The main SAR trends identified within the series were substantiated by a molecular modeling study based on a receptor-driven docking model constructed on the basis of the crystal structure of the human A2A receptor.

Pyrazin-2(1H)-ones as a novel class of selective A3 adenosine receptor antagonists.

BACKGROUND: A3AR antagonists are promising drug candidates as neuroprotective agents as well as for the treatment of inflammation or glaucoma. The most widely known A3AR antagonists are derived from polyheteroaromatic scaffolds, which usually show poor pharmacokinetic properties. Accordingly, the identification of structurally simple A3AR antagonists by the exploration of novel diversity spaces is a challenging goal. RESULTS: A convergent and efficient Ugi-based multicomponent approach enabled the discovery of pyrazin-2(1H)-ones as a novel class of A3AR antagonists. A combined experimental/computational strategy accelerated the establishment of the most salient features of the structure-activity and structure-selectivity relationships in this series. CONCLUSION: The optimization process provided pyrazin-2(1H)-ones with improved affinity and a plausible hypothesis regarding their binding modes was proposed.

Different enzymatic activities in carp (cyprinus carpio L.) as potential biomarkers of exposure to the pesticide methomyl.

This study investigated the influence of the pesticide methomyl on different enzymatic activities in carp. The fish were exposed to a sub-lethal concentration (0.34 mg L-1) of methomyl for 15 days. On days 4 and 15, catalase (CAT) and glutathione-S-transferase (GST) activities were measured in the liver and gills. Acetylcholinesterase (AChE) activity in brain and muscle was also determined. Liver catalase activity slightly increased in exposed fish when compared to controls, but it was statistically significant only at the beginning of the experiment. No changes in CAT activity in the gills of exposed and control animals were observed (mean values were in the range 10.7-11.7 nmol min-1 per mg of protein). Liver GST activity was slightly inhibited in the exposed animals at the beginning of the study; however, it was significantly inhibited in the gills. Brain AChE activity was diminished throughout the experiment and significantly decreased after 96 h of exposure compared to controls (0.041 vs. 0.075 nmol min1 per mg of protein; p<0.001). Our findings suggest that CAT, GST, and AChE are reliable biomarkers of effect after exposure to methomyl.