¹HNMR-based study on characteristic metabolites in urine of mice following 90d of flurochloridone exposure

Background  Flurochloridone (FLC), a kind of selective preemergence herbicides, is mainly used to control a variety of broad-leaved and gramineous weeds, commonly applied in sunflower, potato, carrot, wheat, and corn.

Objective  This study aims to describe the urinary metabolites in rats after 90 d of FLC exposure, identify characteristic metabolites, and explore the toxicological mechanism of FLC.

Methods  Forty-eight Wistar rats were randomly divided into four groups by weight: 31.25, 125.0, and 500.0 mg·kg^-1 FLC exposure groups and a solvent control group, with 12 rats in each group, half male and half female. Urine samples were collected after 90 d of intragastric administration. After 600 MHz ¹H nuclear magnetic resonance (¹HNMR) spectroscopy, onedimensional hydrogen spectrum was obtained, and the spectral region was integrated into regions with chemical shifts δ over an interval of 0.04. Principal component analysis and partial least squares discriminant analysis (PLS-DA) were applied to determine the characteristic metabolites of FLC.

Results  The PLS-DA cross validation analysis results showed that the male rat model was reasonable (P < 0.05), with obvious classification of scattered points among the three dose groups and the solvent control group, while the female rat model was unreasonable (P > 0.05), with obscure classification results. The PLS-DA intragroup analysis results showed that there was a significant difference between male and female rats in the solvent control group (P < 0.05), but not in the three exposure groups (P > 0.05). The PLS-DA model between single FLC exposure group and the solvent control group was reasonable (P < 0.001), with accuracy > 0.6 and area under curve of Receiver Operating Characteristic > 0.75, providing a clear classification between groups. Thirty-five metabolites were identified by urinary nuclear magnetic resonance, and the significantly changed metabolites after FLC exposure were proline, creatine/creatinine, inositol, trimethylamine oxide, taurine, mannitol, and phenylalanine; 15 characteristic metabolites were obtained between the male rats in the high-dose FLC group and the solvent control group, including increased acetate and uric, and decreased phenylalanine, lactic acid, taurine, 3-hydroxybutyric acid, proline, trimethylamine oxide, dimethylamine, methylamine, creatine, creatinine, inositol, glucose, mannitol, and citrate. However, only citrate, creatine, creatinine, and phenylalanine were significantly decreased in the female rats after FLC exposure

Conclusion  Male rats are more sensitive to FLC as the changes of metabolites are more obvious, showing a dose-response relationship. With the increase of FLC exposure dose, the differences of metabolites are reduced between male and female rats in the same group. The 15 characteristic metabolites in male rats exposed to high dose of FLC are mainly involved in carbohydrate metabolism and amino acid metabolism, indicating hepatotoxicity and nephrotoxicity.