concentrations (Fig. 1C, E). The bo

concentrations (Fig. 1C, E). The body lengths at 96 hpf wereapparently inhibited by prothioconazole at concentrations of0.0750 and 0.1500 mg/L. In addition, the deformity symptoms wereproduced at 48hpf in the groups treated with prothioconazole,including pericardial oedema (Pe) and yolk sac oedema (Yse)(Fig. 2B). As the concentration increases, the symptom of Pe and Ysebecame serious increasingly (Fig. 2A), the rate of Pe had a signifi-cant increase at 0.0750 and 0.1500 mg/L treatment groups, and therate of Yse increased apparently in the 0.1500 mg/L treatmentgroup (Fig. 2B).3.2. Effects of prothioconazole on oxidative stress in zebrafish larvaeThe alterations of oxidative stress in zebrafish larvae wereshowed inFig. 3. Compared with the control group, the activities ofSOD and CAT showed no statistically significant differences (Fig. 3Band C). There was a significant decrease of the levels of GSH in the0.1500 mg/L treatment group (Fig. 3A). Additionally, the levels ofMDA of zebrafish larvaewere significantly increased with exposurethe 0.0375, 0.0750 and 0.1500 mg/L of prothioconazole (Fig. 3D).3.3. Effects of prothioconazole on metabolism profiles in zebrafishlarvaeThe representation of 600 MHz1H NMR spectra of zebrafishlarvae was shown inFig. S1. A trend of good separation was shownin the 0.1500 mg/L treatment group by the PCA score plot as awhole (Fig. 4), which indicated that the metabolism profiles ofzebrafish larvae were disturbed with prothioconazole exposure.But the significant separation was not shown in other two lowerconcentrations treatment groups. A total of 23 metabolites wereidentified, which were shown inFig. S1(Jia et al., 2019;Qiao et al.,2019;Teng et al., 2018b;Yan et al., 2018). Then, the heatmap withcluster analysis was generated by 19 significantly altered metabo-lites based on the VIP values obtained from the PLS-DA model andFig. 1.Developmental effects of prothioconazole on embryos. A: The number of spontaneous movements at 24 hpf. B: Heart rates of zebrafish embryos in the control and pro-thioconazole -treated groups at 48hpf. C: The hatching rate at each observation time point. D: Heart rates of zebrafish embryos in the control and prothioconazole treated groups at72hpf. E: Body lengths of hatched larvae at 96 hpf in the control and prothioconazole-treated groups. F: The hatching rate at each observation time point. Asterisks denote sig-nificant difference between the treatments and control (*p