Araújo Marlyete Chagas de, Assis Caio Rodrigo Dias, Silva Luciano Clemente, Machado Dijanah Cota, Silva Kaline Catiely Campos, Lima Ana Vitória Araújo, Carvalho Luiz Bezerra, Bezerra Ranilson de Souza, Oliveira Maria Betânia Melo de
Departamento de Bioquímica, Universidade Federal de Pernambuco, Recife, PE, Brazil.
Departamento de Bioquímica, Universidade Federal de Pernambuco, Recife, PE, Brazil.
Aquat Toxicol. 2016 Aug;177:182-9. doi: 10.1016/j.aquatox.2016.05.019. Epub 2016 May 24.
This contribution aimed to characterize physicochemical and kinetic parameters of the brain cholinesterases (ChEs) from Parachromis managuensis and investigate the in vitro effects of pesticides and metal ions on its activity intending to propose as biomarker. This species is suitable for this investigation because (1) it was recently introduced in Brazil becoming invasive (no restrictions on capture) and (2) occupies the top of the food chain (being subject to bioaccumulation). The enzyme extract was exposed to 10 metal ions (Al(3+), Ba(2+), Cd(2+), Cu(2+), Hg(2+), Mg(2+), Mn(2+), Pb(2+), Fe(2+) and Zn(2+)) and ChEs selective inhibitors (BW284c51, Iso-OMPA, neostigmine and serine). The extract was also incubated with organophosphate (dichlorvos) and carbamate pesticides (carbaryl and carbofuran). Inhibition parameters (IC20, IC50 and ki) were determined. Selective inhibitors and kinetic parameters confirmed acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) -like as responsible for the ChE activities, most AChE. The IC50 values for pesticides were: 1.68μM (dichlorvos); 4.35μM (carbaryl) and 0.28μM (carbofuran). Most of the analyzed ions did not show significant effect at 1mM (p=0.05), whereas the following ions inhibited the enzyme activity in the order: Hg(2+)>Cu(2+)>Cd(2+)>Zn(2+). Mercury ion strongly inhibited the enzyme activity (IC20=0.7μM). The results about allow to conclude that P. managuensis brain AChE is a potential biomarker for heavy metals and pesticides under study, mainly for the carbamate carbofuran once it was capable to detect 6-fold lower levels than the limit concentration internationally recommended.
本研究旨在表征马那瓜副丽鱼脑胆碱酯酶(ChEs)的物理化学和动力学参数,并研究农药和金属离子对其活性的体外影响,以期将其作为生物标志物。该物种适合此项研究,原因如下:(1)它最近被引入巴西并成为入侵物种(捕获无限制);(2)处于食物链顶端(易发生生物累积)。酶提取物分别与10种金属离子(Al(3+)、Ba(2+)、Cd(2+)、Cu(2+)、Hg(2+)、Mg(2+)、Mn(2+)、Pb(2+)、Fe(2+)和Zn(2+))以及ChEs选择性抑制剂(BW284c51、异稻瘟净、新斯的明和丝氨酸)接触。提取物还与有机磷农药(敌敌畏)和氨基甲酸酯类农药(西维因和克百威)一起孵育。测定了抑制参数(IC20、IC50和ki)。选择性抑制剂和动力学参数证实,乙酰胆碱酯酶(AChE)和丁酰胆碱酯酶(BChE)样酶是ChE活性的主要来源,其中大部分为AChE。农药的IC50值分别为:1.68μM(敌敌畏);4.35μM(西维因)和0.28μM(克百威)。大多数分析的离子在1mM时未显示出显著影响(p = 0.05),而以下离子对酶活性的抑制顺序为:Hg(2+)>Cu(2+)>Cd(2+)>Zn(2+)。汞离子强烈抑制酶活性(IC20 = 0.7μM)。研究结果表明,马那瓜副丽鱼脑AChE是所研究的重金属和农药的潜在生物标志物,主要针对氨基甲酸酯类克百威,因为它能够检测到比国际推荐限量浓度低6倍的水平。