Effects of diclofenac on the gametes and embryonic development of Arbacia lixula

Figures & data

Figure 1. Scheme of experimental design.

Table I. HPLC-UV/DAD instrument conditions

Parameter	Value
Loop	50 μl
Flux	1.2 ml/min
Column temperature	40°C
Autosampler temperature	15°C
Stop time	20 min
Detector	UV 240–254-278-290 nm

Table II. The gradients of the eluents after the HPLC column was conditioned in the mobile phase for 1 h

Time (min)	Acetonitrile (%)	Phosphoric acid (%)
0	35	65
2	50	50
4	75	25
15	75	25
16	50	50
20	35	65

Figure 2. A, B and C: Box-plots obtained using the Kruskal-Wallis test on the developed embryos and the unfertilised and degenerated eggs (from left to right). The circles represent the outlier; a, b, c: microscopic photographs of the embryos and the unfertilised and degenerated eggs. Scale bar 50 µm. The asterisks represent statistical differences (** p < 0.01, ***p < 0.001).

Figure 3. DCF accumulation in embryos. Values are expressed as the mean ± SD. The asterisks represent statistical differences (** p < 0.01).

Figure 4. DCF accumulation in A. lixula eggs (A) and sperm (B) measure by HPLC analysis. The circles represent the outlier. Significant accumulation were observed at concentrations over 50 mg/L. The asterisks represent statistical differences (***p < 0.001).

Table III. Concentrations of DCF absorbed in the sperm and egg samples, expressed as the mean ± standard deviation

DCF treatments	0 mg/L	0.05 mg/L	0.5 mg/L	5 mg/L	50 mg/L
Egg	0.06 ± 0.04	0.05 ± 0.05	0.06 ± 0.06	0.21 ± 0.20	0.91 ± 0.80
Sperm	0.1 ± 0.09	0.07 ± 0.05	0.06 ± 0.04	0.10 ± 0.03	0.40 ± 0.24