The present study investigates the toxicity of the herbicide tribenuron methyl (TBM) as an anthropogenic agent and ammonia as an abiotic factor on Daphnia magna at environmentally relevant concentrations. These stressors may coexist in surface waters in agricultural regions. To achieve this objective, D. magna were exposed to TBM at a nominal concentration of 0.81 μg/L in association with a low ammonia (LA) concentration of 0.65 mg/L and a high ammonia (HA) concentration of 1.61 mg/L in acute toxicity tests of 96-h duration and chronic toxicity tests of 21-day duration. The D. magna also were exposed to TBM, HA, and LA singly. The D. magna were analysed for various biomarkers of sublethal toxicity. Glutathione peroxidase (GPx), glutathione S-transferase (GST), cholinesterase (ChE) enzyme activities, and levels of thiobarbituric acid reactive substances (TBARS) and total protein were determined spectrophotometrically. Mitochondrial membrane potential (MMP) was analysed by microscopy with fluorescence staining. Cytochrome c and 5′ AMP-activated protein kinase (AMPK) were analysed by Western blotting. Morphometric properties were examined microscopically. This is the first study in which AMPK, an indicator of intracellular energy, was measured in D. magna. GST and ChE enzyme activities and TBARS and total protein levels did not change during acute exposures (i.e., 96 h) in all treatments. GPx activity increased in D. magna from the HA + TBM treatment compared with single-exposure groups. The level of cytochrome c protein was elevated in D. magna from the LA and LA + TBM treatments. AMPK protein levels increased in all treatments with daphnids, except in the LA group. MMP was depolarised in D. magna from all treatments, whereas the most notable change was observed in HA + TBM mixture group in chronic exposures. The results show that GST and ChE may not be sensitive biomarkers for evaluating the sublethal toxic effects to D. magna exposed to environmentally relevant concentrations of ammonia and TBM. Acute and chronic exposure to ammonia and TBM probably caused an energetic crisis in D. magna. Therefore, AMPK and MMP are promising biomarkers for these toxicants.

本研究调查了在环境相关浓度下，除草剂丁苯磺隆（TBM）作为人为源毒物和氨作为非生物因子对大型蚤（Daphnia magna）的毒性。这些压力源可能共存于农业地区的地表水中。为了实现这一目标，将D. magna暴露于名义浓度为0.81μg/ L的TBM中，同时将低浓度（0.65 mg / L）和高浓度（1.61 mg / L）的氨暴露于TBM中。持续96小时的急性毒性试验和持续21天的慢性毒性试验。该大型溞也暴露于TBM，HA和LA单。该大型溞分析了亚致死毒性的各种生物标志物。用分光光度法测定了谷胱甘肽过氧化物酶（GPx），谷胱甘肽S-转移酶（GST），胆碱酯酶（ChE）酶活性以及硫代巴比妥酸反应性物质（TBARS）和总蛋白的含量。线粒体膜电位（MMP）通过显微镜与荧光染色进行了分析。通过Western印迹分析细胞色素c和5'AMP激活的蛋白激酶（AMPK）。用显微镜检查形态学性质。这是在其中AMPK，细胞内能量的指标，在测定第一项研究D.蚤。在所有治疗中，急性暴露（即96小时）期间，GST和ChE酶活性以及TBARS和总蛋白水平均未改变。GPx活动在与单次接触组相比，来自HA + TBM处理的D. magna。通过LA和LA + TBM处理，D。magna中的细胞色素c蛋白水平升高。除LA组外，在所有使用水蚤的治疗中，AMPK蛋白水平均升高。在所有处理中，D。magna中的MMP去极化，而在慢性暴露中，HA + TBM混合物组中观察到最显着的变化。结果表明，GST和ChE可能不是评估暴露于环境相关浓度的氨和TBM的对D. magna的亚致死毒性作用的敏感生物标志物。急性和慢性暴露于氨水和TBM可能会导致D. magna发生精力充沛的危机。因此，AMPK和MMP是这些有毒物质的有前途的生物标志物。