Perfection v550 photo

The multi-species microbial assay (MARA) was used to toxicity of bisphenol derivatives, respectively. In the MARA assay, lyophilized microorganisms placed in row H of the microplates were rehydrated and pre-incubated for 4 h at 30 °C. Series of six dilutions of the initial solution of each bisphenol derivatives (1, 0.5, 0.125, 0.0625, 0.0312, and 0.0156 mg L⁻¹) were placed in rows G-B of the microplates. Pure medium was introduced into row A as a strain control. Subsequently, microorganisms from row H were added to each sample dilution. Microplates were incubated at 30 °C (18 h) and subsequently scanned in a flatbed scanner (Epson Perfection V550 Photo). The results were processed using an image analysis program that facilitates calculation of the average MTC and minimum MTC (microbial toxic concentration) values (mg L⁻¹). Moreover, based on the growth inhibition results for each strain, the EC₅₀ value (half-maximal effective concentration) was calculated for each bisphenol and classified as: EC₅₀ < 1 mg L⁻¹ — very toxic to aquatic organisms, 1–10 mg L⁻¹ — toxic to aquatic organisms, 10–100 mg L⁻¹ harmful to aquatic organisms, and > 100 mg L⁻¹ — not classified as harmful to aquatic organisms (European Commission (EC) 1993 ).

For RSA analysis, roots from 10- or 14-day-old plants were drawn on transparent plastic sheets and root images were scanned at a resolution of 200 dpi using an Epson scan (Epson perfection V550 photo). The images were first opened with the ImageJ software to invert the color. Finally, root images were analyzed using EZ-Rhizo software (Figure 1(a)) [34 (link)]. The root traits analyzed were the main root length (MRL), number of lateral root (NLR), total root size (TRS), lateral root size (LRS), lateral root density of the main root (LRD-MR), basal zone length (Basal); branched zone length (Branched), and apical zone length (Apical) (Figure 1(b)).