Exposición aguda a fentión en juveniles de cachama blanca (Piaractus brachypomus): efectos tóxicos, cambios en actividad colinesterasa y uso potencial en monitoreo ambiental

El fentión es un insecticida organofosforado usado en todo el mundo cuya aplicación puede derivar en contaminación de aguas y efectos potenciales en salud humana y animal. Objetivos: para evaluar los efectos tóxicos, la actividad colinesterasa plasmática tipo butiril (BChE) y el uso potencial en monitoreo ambiental se distribuyeron al azar treinta y cuatro juveniles de cachama blanca (Piaractus brachypomus) en tres tratamientos (control, 0,13 ppm y 2 ppm fentión) por un período de 96 horas. Materiales y métodos: los ejemplares fueron expuestos al fentión en tanques de diez galones, evaluando sintomatología, cambios post-mortem, índice viscero-somático (IVS) y actividad BChE. Se compararon las medias de cada variable para determinar significancia estadística (ANAVA, α=0,05). Resultados: se presentaron signos clínicos severos (nado frenético, tremores musculares y pérdida de eje de nado) en tres individuos (2 ppm) hacia las 11 y 34 horas. En 0,13 ppm hubo signos leves a moderados (tremores y movimiento de aleta caudal). Los IVS (control = 5,3 ± 0,5; 0,13 ppm = 6,9 ± 0,3 y 2 ppm = 7,3 ± 0,6) fueron significativamente diferentes entre los expuestos a fentión y los controles. La actividad BChE (control = 185,0 ± 20,4; 0,13 ppm = 12,5 ± 2,3 y 2 ppm = 9,8 ± 1,8 nmoles / ml plasma / min) mostró inhibición significativa en los expuestos a fentión con respecto a controles. Conclusiones: se demostraron los efectos tóxicos agudos del fentión en juveniles de cachama blanca. Los resultados en BChE mostraron su utilidad en monitoreo ambiental por exposición a insecticidas organofosforados.

Polimorfismos del gen Butirilcolinesterasa responsables de reacciones adversas en pacientes consumidores de “cocaína”

La butirilcolinesterasa humana (BChE; EC 3.1.1.8) es una enzima polimórfica sintetizada en el hígado y en el tejido adiposo, ampliamente distribuida en el organismo y encargada de hidrolizar algunos ésteres de colina como la procaína, ésteres alifáticos como el ácido acetilsalicílico, fármacos como la metilprednisolona, el mivacurium y la succinilcolina y drogas de uso y/o abuso como la heroína y la cocaína. Es codificada por el gen BCHE (OMIM 177400), habiéndose identificado más de 100 variantes, algunas no estudiadas plenamente, además de la forma más frecuente, llamada usual o silvestre. Diferentes polimorfismos del gen BCHE se han relacionado con la síntesis de enzimas con niveles variados de actividad catalítica. Las bases moleculares de algunas de esas variantes genéticas han sido reportadas, entre las que se encuentra las variantes Atípica (A), fluoruro-resistente del tipo 1 y 2 (F-1 y F-2), silente (S), Kalow (K), James (J) y Hammersmith (H). En este estudio, en un grupo de pacientes se aplicó el instrumento validado Lifetime Severity Index for Cocaine Use Disorder (LSI-C) para evaluar la gravedad del consumo de “cocaína” a lo largo de la vida. Además, se determinaron Polimorfismos de Nucleótido Simple (SNPs) en el gen BCHE conocidos como responsables de reacciones adversas en pacientes consumidores de “cocaína” mediante secuenciación del gen y se predijo el efecto delos SNPs sobre la función y la estructura de la proteína, mediante el uso de herramientas bio-informáticas. El instrumento LSI-C ofreció resultados en cuatro dimensiones: consumo a lo largo de la vida, consumo reciente, dependencia psicológica e intento de abandono del consumo. Los estudios de análisis molecular permitieron observar dos SNPs codificantes (cSNPs) no sinónimos en el 27.3% de la muestra, c.293A>G (p.Asp98Gly) y c.1699G>A (p.Ala567Thr), localizados en los exones 2 y 4, que corresponden, desde el punto de vista funcional, a la variante Atípica (A) [dbSNP: rs1799807] y a la variante Kalow (K) [dbSNP: rs1803274] de la enzima BChE, respectivamente. Los estudios de predicción In silico establecieron para el SNP p.Asp98Gly un carácter patogénico, mientras que para el SNP p.Ala567Thr, mostraron un comportamiento neutro. El análisis de los resultados permite proponer la existencia de una relación entre polimorfismos o variantes genéticas responsables de una baja actividad catalítica y/o baja concentración plasmática de la enzima BChE y algunas de las reacciones adversas ocurridas en pacientes consumidores de cocaína.

Esterase inhibition in tadpoles of Scinax fuscovarius (Anura, Hylidae) as a biomarker for exposure to organophosphate pesticides

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

New selective acetylcholinesterase inhibitors designed from natural piperidine alkaloids

Five new piperidine alkaloids were designed from natural (-)-3-O-acetyl-spectaline and (-)-spectaline that were obtained from the flowers of Senna spectabilis (sin. Cassia spectabilis, Leguminosae). Two semi-synthetic analogues (7 and 9) inhibited rat brain acetylcholinesterase, showing IC50 of 7.32 and 15.1 mu M, and were 21 and 9.5 times less potent against rat brain butyrylcholinesterase, respectively. Compound 9 (1 mg/kg, ip) was fully efficacious in reverting scopolamine-induced amnesia in mice. The two active compounds (7 and 9) did not show overt toxic effects at the doses tested in vivo. (c) 2005 Elsevier Ltd. All rights reserved.

Determination of carbamate residues in crop samples by cholinesterase-based biosensors and chromatographic techniques

An amperometric biosensor based on cholinesterase (ChE) has been used for the determination of selected carbamate insecticides in vegetable samples. The linear range of the biosensor for the N-methylcarbamates (aldicarb, carbaryl, carbofuran, methomyl and propoxur) varied from 5 x 10(-5) to 50 mg kg(-1). Limits of detection were calculated on the basis that the ChE enzymes were 10% inhibited and varied, depending of the combination ChE (as acetyl- or butyrylcholinesterase) vs. inhibitor (pesticide), from 1 x 10(-4) to 3.5 mg kg(-1). The biosensor-based carbamate determination was compared to liquid chromatography/UV methods. Three vegetable samples were spiked with carbofuran and propoxur at 125 mu g kg(-1) followed by conventional procedures. Good correlations were observed for carbofuran in the vegetable extracts (79, 96 and 91% recoveries for potato, carrot and sweet pepper, respectively), whereas for propoxur unsatisfactory results were obtained. Potato and carrot samples were spiked with 10, 50 and 125 mu g kg(-1) carbofuran, followed by direct determination by the amperometric biosensor. The fortified sampler; resulted in very high inhibition values, and recoveries were: 28, 34 and 99% for potato, and 140, 90 and 101% for carrot, respectively, at these three fortification levels. (C) 1998 Elsevier B.V. B.V.

Evaluation of biochemical and redox parameters in rats fed with corn grown in soil amended with urban sewage sludge

The increased production of urban sewage sludge requires alternative methods for final disposal. A very promising choice is the use of sewage sludge as a fertilizer in agriculture, since it is rich in organic matter, macro and micronutrients. However, urban sewage sludge may contain toxic substances that may cause deleterious effects on the biota, water and soil, and consequently on humans. There is a lack of studies evaluating how safe the consumption of food cultivated in soils containing urban sewage sludge is. Thus, the aim of this paper was to evaluate biochemical and redox parameters in rats fed with corn produced in a soil treated with urban sewage sludge for a long term. For these experiments, maize plants were grown in soil amended with sewage sludge (rates of 5, 10 and 20. t/ha) or not (control). Four different diets were prepared with the corn grains produced in the field experiment, and rats were fed with these diets for 1, 2, 4, 8 and 12 weeks. Biochemical parameters (glucose, total cholesterol and fractions, triglycerides, aspartate aminotransferase and alanine aminotransferase) as well the redox state biomarkers such as reduced glutathione (GSH), malondialdehyde (MDA), catalase, glutathione peroxidase and butyrylcholinesterase (BuChE) were assessed. Our results show no differences in the biomarkers over 1 or 2 weeks. However, at 4 weeks BuChE activity was inhibited in rats fed with corn grown in soil amended with sewage sludge (5, 10 and 20. t/ha), while MDA levels increased. Furthermore, prolonged exposure to corn cultivated in the highest amount per hectare of sewage sludge (8 and 12 weeks) was associated with an increase in MDA levels and a decrease in GSH levels, respectively. Our findings add new evidence of the risks of consuming food grown with urban sewage sludge. However, considering that the amount and type of toxic substances present in urban sewage sludge varies considerably among different sampling areas, further studies are needed to evaluate sludge samples collected from different sources and/or undergoing different types of treatment. © 2013 Elsevier Inc.

Avaliação de neurotoxicidade de formas enantioméricas de praguicidas organifosforados: estudos in vitro, in vivo e de neuroproteção

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The enigmatic role of cholinergic reflex in the pathogenesis of Chagas disease

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Oxidative stress and inflammatory response biomarkers in dogs with mammary carcinoma

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Novel markers of inflammatory response and hepatic dysfunction in canine leishmaniasis

Dogs are the main host of Leishmania infantum, and the clinical presentation may range from asymptomatic to systemic manifestations. The immune mechanisms in infected, but clinically healthy dogs, prevails Th1 response mediated by cytokines. In this sense, adenosine deaminase (ADA) and butyrylcholinesterase (BChE) are considered as key enzymes in several physiological processes, including the modulation of inflammatory process. Considering the variable immune response against Leishmania and the known participation of ADA and BChE, the aim of this study was to assess the relation between these two enzymes with the inflammatory response as well as hepatic function in dogs naturally infected with L. infantum. For this purpose, the activity of ADA and BChE was assessed in sera of 24 dogs naturally infected with L. infantum, plus 17 healthy dogs. The naturally infected dogs had clinical signs compatible with leishmaniasis and sera activities of ADA (P<0.01) and BChE (P<0.05) decreased, when compared to the healthy group. The reduction of ADA activity probably represented an effect on inflammatory response, especially due to the decreased hydrolysis of extracellular adenosine, might in order to protect against tissue damage and, also, setting a down-regulation on pro-inflammatory cytokines. BChE enzyme had no effect on modulating the immune response in leishmaniasis, but it decreased, a fact may related to deficiency of synthesis in the liver. Therefore, ADA and BChE activities reduced probably in order to protect against extra tissue damage and due failure in synthesis, respectively.

Indicadores diagnósticos da síndrome metabólica, butirilcolinesterase e estado de condicionamento físico em adolescentes obesos e eutróficos: uma breve explanação do atual estado da arte

Damage to health caused by excess body fat also generating a negative economic impact, with significant increase in public spending. The metabolic syndrome, which also gets several other names, such as plurimetabolic syndrome or syndrome X, is nothing more than the combined incidence of some diseases or metabolic disorders in which obesity, besides being one of them, seems to promote parallel effects that contribute to the development of other chronic diseases such as diabetes and cardiovascular disease. Researchers agree that hyper caloric diets associated with a sedentary lifestyle are the main triggers of disease, including the increasing on genetic predisposition to this disease in children and adolescents. In the case of children and adolescents the diagnosis is complicated by the lack of a consensus accepted by the scientific community. In addition to behavioral and environmental factors unfavorable to health, in a more detailed analysis also found hereditary aspects or simply genetic, such as hepatic enzyme Butyrylcholinesterase. When compared to eutrophic, obese adolescents, like adults obese, have higher serum concentration values as well as major activity for this enzyme. Increasing evidence suggests that excess body weight assumes an important role in the variation of metabolic functions in adolescents, favoring the emergence of early diagnostic indicators of metabolic syndrome.

Síntese de potencial inibidor de acetilcolinesterase para tratamento da Doença de Alzheimer

A doença de Alzheimer (DA) é a forma mais comum de demência, representando cerca de 80% dos casos. A DA é caracterizada por um processo de declínio progressivo e irreversível das funções cognitivas e da memória, que se estende para a desorganização do comportamento. Atualmente, 46,8 milhões de pessoas em todo o mundo foram diagnosticadas com demência. Embora vários fatores tenham sido implicados na DA, sua etiologia ainda não é completamente conhecida. Do ponto de vista neuropatológico, é observado no cérebro de indivíduos com DA atrofia cortical difusa, presença de grande número de placas senis, emaranhados neurofibrilares, processo inflamatório e perda neuronal. A progressão dos sintomas está associada a mudanças estruturais nas sinapses colinérgicas em certas regiões do cérebro, que consequentemente, apresentam neurotransmissão colinérgica reduzida. Os vários eventos patológicos interligados contribuem para o avanço da doença e direcionam diversas pesquisas na busca por tratamentos multialvos com base no processo multifatorial de DA. Assim o presente trabalho descreve a síntese de derivados híbridos dual binding site de donepezila-tacrina (fármacos inibidores de acetilcolinesterase), com potencial para agir em dois alvos terapêuticos pela (i) inibição da acetilcolinesterase em ambos os sítios ativo e periférico, como demonstrado pelos estudos de modelagem molecular, e (ii) na agregação do peptídeo A? neurotóxico induzido pela acetilcolinesterase, na tentativa de interromper a progressão da doença. A estratégia sintética envolveu a condensação da 5,6-dimetóxiindanona com a unidade 4-piperidinil carbaldeído, a qual forneceu o intermediário 5,6- dimetóxindan-1-ona-4-piperidinil-metileno-1-[(4-cloroquinolin-2-il)metil], seguido de redução da dupla ligação, gerada na reação de condensação anterior, e substituição do átomo de cloro-quinolina por amino para obtenção do produto final, ou manutenção da função olefina, seguido de substituição do átomo de cloro-quinolina por azido ou amino, gerando cinco híbridos estruturalmente correlacionados. Os híbridos foram testados em ensaio de inibição de acetilcolinesterase e butirilcolinesterase pelo método de Ellman, e o híbrido insaturado, contendo a função amino-quinolina foi o mais ativo da série com IC50 na faixa de nanomolar (0,014 ?M). Futuramente, os intermediários da reação e produto final serão submetidos ao ensaio de inibição da agregação do peptídeo A? neurotóxico pelo método da tioflavina T. Neste trabalho, também são descritos os testes de predição in vitro para permeação pela barreira hematoencefálica, bem como sua absorção intestinal, pelo método PAMPA.

Size, blood organochlorine pesticide concentrations and biochemical indicators measured in hawksbill sea turtles (Eretmochelys imbricata) nesting at Punta Xen (Campeche, Mexico) in August 2010

The impact of environmental pollution on the homeostasis of sea turtles remains scarce, particularly in the southern Gulf of Mexico. As many municipalities do not rely on a waste treatment plant along the coastline of the Yucatan Peninsula, the vulnerability of these specimens could results enhanced. We searched for relationships between presence of organochlorine pesticides (OCP) and the level of several oxidative and pollutant stress indicators of the hawksbill sea turtle (Eretmochelys imbricata) during the egg-laying period 2010 at Punta Xen (Campeche, Mexico). Endosulfans, aldrin related (aldrin, endrin, dieldrin, endrin ketone, endrin aldehyde) and dichlorodiphenyldichloroethylene (DDT) families were detected in 17, 21 and 26 of the 30 sampled sea turtles, respectively. Significant correlation existed between the size of sea turtles with the concentration of methoxychlor, cholinesterase activity in plasma and heptachlors family, and catalase activity and hexachlorohexane family. Cholinesterase activity in washed erythrocytes and lipid peroxidation were positively correlated with glutathione reductase activity. Antioxidant enzyme actions seem adequate as no lipids damages were correlated with any OCPs. Future studies are necessary to evaluate the effect of OCPs on males of the area because of the significant detection of methoxychlor that target endocrine functioning and increase its concentration with size of the sea turtles.

S-Alkylated/aralkylated 2-(1H-indol-3-yl-methyl)-1,3,4- oxadiazole-5-thiol derivatives. 2. Anti-bacterial, enzymeinhibitory and hemolytic activities

Purpose: To evaluate the antibacterial, enzyme-inhibitory and hemolytic activities of Salkylated/ aralkylated 2-(1H-indol-3-ylmethyl)-1,3,4-oxadiazole-5-thiol derivatives. Methods: Antibacterial activities of the compounds were evaluated using broth dilution method in 96 well plates. Enzyme inhibitory activities assays were investigated against α-glucosidase, butyrylcholinesterase (BchE) and lipoxygenase (LOX) using acarbose, eserine and baicalien as reference standards, respectively. A mixture of enzyme, test compound and the substrate was incubated and variation in absorbance noted before and after incubation. In tests for hemolytic activities, the compounds were incubated with red blood cells and variations in absorbance were used as indices their hemolytic activities. Results: The compounds were potent antibacterial agents. Five of them exhibited very good antibacterial potential similar to ciprofloxacin, and had minimum inhibitory concentrations (MIC) of at least 9.00 ± 4.12 μM against S. aureus, E.coli, and B. subtilis. One of the compounds had strong enzyme inhibitory potential against α-glucosidase, with IC50 of 17.11 ± 0.02 μg/mL which was better than that of standard acarbose (IC50 38.25 ± 0.12 μg/mL). Another compound had 1.5 % hemolytic activity. Conclusion: S-Alkylated/aralkylated 2-(1H-indol-3-ylmethyl)-1,3,4-oxadiazole-5-thiol deviratives with valuable antibacterial, anti-enzymatic and hemolytic activities have been successfully synthesized. These compounds may be useful in the development of pharmaceutical products.

Phytochemical profile and some biological activities of three Centaurea species from Turkey

Purpose: To characterise the phytochemical profile of whole plants of Centaurea balsamita, C. depressa and C. lycopifolia with LC-ESI-MS/MS, and as well as their antioxidant, anticholinesterase and antimicrobial activities. Methods: Organic and aqueous extracts of the three Centaurea species were evaluated for DPPH free radical, ABTS cation radical scavenging and cupric reducing antioxidant capacity (CUPRAC). Acetyland butyryl-cholinesterase enzyme inhibition abilities of the extracts using petroleum ether, acetone, methanol and water were studied to determine anticholinesterase activity, while antimicrobial activity was determined by disc diffusion method using appropriate antimicrobial standards and organisms. The phytochemical components of the methanol extracts were assessed by LC-MS/MS. Results: The methanol extract of C. balsamita exhibited much higher DPPH free and ABTS cation radicals scavenging activities (with IC50 of 62.65 ± 0.97 and 24.21 ± 0.70 mg/ml, respectively) than the other extracts. The petroleum ether extracts of the plant species exhibited moderate inhibitory activity against butyrylcholinesterase enzymes while the acetone extract of C. balsamita showed good antifungal activity against Candida albicans. Quinic acid (17513 ± 813 μg/g, 63874 ± 3066 μg/g and 108234 ± 5195 μg/g) was the major compound found in the methanol extracts of C. balsamita, C. depressa and C. Lycopifolia, respectively. Conclusion: These results indicate quinic acid is the major compound in the three plant species and that Centaurea balsamita has significant antioxidant, anticholinesterase and antimicrobial properties. Further studies to identify the compounds in the extracts responsible for the activities are required.