Immersion of nanostructured lipid carriers loaded with 17-alpha methyltestosterone for masculinization of red tilapia (Oreochromis sp.)

The present study aimed to develop novel masculinization agents for enhancing sex-reversal in red tilapia (Oreochromis sp.) by immersion administration using 17α-methyltestosterone-loaded nanostructured lipid carriers (MT-NLC). Moreover, the surface charge of the MT-NLC was modified with chitosan (CS-MT-NLC) to improve their mucoadhesive property. Dynamic light scattering (DLS) and transmission electron microscopy (TEM) were used to characterize the morphology of the nanoparticles. Furthermore, we assessed the toxicity associated with immersion-induced masculinization in red tilapia. Subsequently, we examined alterations in gill morphology, determined the percentage of masculinization, analyzed their length-weight relationship, and investigated the expression of genes related to sex differentiation. MT-NLC had an average size, polydispersity index, and negatively charged surface of 140.8 ± 2.6 nm, <0.2, and −26.9 ± 0.3 mV, respectively, with higher stability at 4 °C and 25 °C compared to CS-MT-NLC. The release mechanisms of NLC and CS-NLC were regulated by diffusion processes, which were characterized using Avrami's equation. In contrast, the release mechanism of non-encapsulated MT (MT-ET) was governed by burst release kinetic. Furthermore, tests assessing mucin binding confirmed that CS-MT-NLC, with a positive charge, exhibited superior mucoadhesive properties compared to MT-NLC, with a negative charge. Importantly, CS-MT-NLC immersion was found to be toxic to red tilapia fry; therefore, it has been discontinued for further experiments. The immersions of both MT-NLC and MT-ET were non-toxic to red tilapia fry, with an LC₅₀ exceeding 800 μg/L, and the morphological appearance of the gills after treatment was normal. The MT-NLC-400 and MT-ET-800 groups exhibited elevated expression levels of dmrt1 and sox9 compared to cyp19a1a and soxl2. These higher expression levels corresponded to significantly higher percentages of males (98.3% and 95.7%) in both the MT-NLC and MT-ET groups compared to the control group (54.7%). The MT-NLC and MT-ET groups displayed a negative allometric growth behavior in red tilapia fry, with a relative condition factor > 1. Adopting MT-NLC-400 immersion treatment for masculinizing red tilapia could serve as a beneficial strategy for commercial tilapia farms to reduce the required dosage and the time needed to treat fish.