Evaluation of the toxicity of glyphosate and its different formulations using the cell line human HepG2.

CHAUFAN G,; COALOVA I,; RÍOS DE MOLINA M.C.

Milan, Italia

Congreso; SETAC Europe 21st Annual Meeting; 2011

SETAC Europe

The herbicide N-phosphonomethyl glycine, commercially known as glyphosate, is one of the most widely used agrochemicals in the fields of Argentina and the world. The use of different glyphosate formulations for weed control in agriculture produces herbicide excess in the environment that may cause undesirable toxic effects in the biota. Although humans are not a direct target, they are in risk of glyphosate exposure. Because the molecular mechanism of gliphosate toxicity is poorly understood, this study will assess the possible adverse effects and the risk of human exposure to glyphosate. We studied the glyphosate toxicity in an in vitro model of human cells (hepatocytes cultures, HepG2). The human hepatoma cell line Hep G2 was cultured in minimal essential medium (MEM) supplemented with 10% heat-inactivated fetal bovine serum (FBS), 100 units/ml penicillin, 100 mg/ml streptomycin and 2.5 mg/ml amphotericin B. Cells were cultured at 37 ºC in a humidified atmosphere of 5% CO2–95% air and the medium was renewed twice a week. For all experiments, confluent attached cells were removed from cell culture dishes with 0.25% sterile trypsin, diluted with MEM/10% FBS and replated into 96-well plates (0.2 ml; 2 x104 cells/well) for 24, 48 and 72h for the MTT assay and into 6 well plates (3ml; 3 x 105 cells/well) for 24h for the caspase 3/7 activity assay, and lipid peroxidations as thioborbituric acid reactive substance (TBARs) level. We measured the cytotoxicity (LC50) of Roundup® and of Atanor® glyphosate formulations with or not the addition of Impact® adjuvant, and the technical grade glyphosate in its acid form on cell line HepG2. All parameters were measured at sub-agricultural doses with all formulations.The LC50 of different glyphosate presentations at 24, 48 and 72h was determinate: 41.2 (37.6 - 45.1), 35.2 (34.9 - 35.4), 34.7 (34.1 - 35.3) mg/L for Roundup; 93.9 (75.0 -117.4), 40.4 (31.2 - 52.2), 29.9 (17.8 - 50.4) mg/L for Atanor® with Impacto®; 445.7 (416.4 - 477.1), and 418.7 (405.1 - 432.8), 383.5 (372.8 - 394.4) mg/L for Atanor®. Not significant effect was observed at sub-agricultural doses with technical grade glyphosate in its acid form. The caspases 3/7 are significantly activated with doses of Roundup® (35 mg/L) up to 240% in 24h. Not significant effect was observed at this dose for TBARs levels.We conclude that cytoxicity and apoptotic effects were more dependent on the formulations than on the glyphosate concentration.