(A) For scanning electron microscopy (SEM), NPMAs were diluted 1:5000 and mounted on plastic coverslips (Thermanox), metalized with gold in an evaporator (Bal-Tec, model MD20) and analyzed by SEM with secondary electrons, backscattered electron imaging and (B) Atomic force microscopy (AFM) image using non-contact mode, sample diluted 1:5000 and deposited in silicon plates.

Colloidal parameters as size, PDI and zeta potential were evaluated as a function of time using the DLS technique (n = 3). All parameters were considered stable, indicating the suitability of the nanoformulations.

The systems were maintained under magnetic stirring. Samples were collected during 2 hours and BENZ concentration was determined by HPLC. The experiments were conducted in triplicate at room temperature. The percentage of BENZ released was less than 20% for both nanocapsules, showing sustained drug release. No statistical difference was observed for the BENZ release curves (*p>0.05).

(A) Anesthetic induction time (B) Recovery time of zebrafish adults.

In vivo fluorescence images of zebrafish exposes to NPMAs.

Fish gills, liver and intestine images showed fluorescence only under NPMA treatment.

(A), (B) and (C) show fish sections without relevant pathological alterations. (L) liver; (P) pancreas and (I) intestine (H&E staining).

Survival rate curves of embryos exposed to BENZ, NP and NPMA at concentrations of 3.9, 15.6 and 62.5 μg/L^-1. The Kaplan Meier curve was constructed and performed the log-rank (Mantel-Cox) statistical test to compare the groups.

(A-C) control embryos presenting wildtype morphology. (D-F) Embryos exposed to BENZ revealed tail malformation (D and E) and pericardial edema (F). (G-I) Embryo contact with NP developed curved tail. (G and H) Nanoparticles formed aggregates deposition on the chorion surface. (J-L) NPMA exposure led to general malformation of embryos at 3.9 μg/L^-1.