Evaluation of oral supplementation of free and nanoencapsulated Minthostachys verticillata essential oil on immunological, biochemical and antioxidants parameters and gut microbiota in weaned piglets

Abstract

Early weaning is an important stressor that impairs the piglet´s health, and essential oils appear as promising candidates to improve it. The aim of this study was to evaluate the effect of oral supplementation of free and nanoencapsulated Minthostachys verticillata essential oil (EO and NEO, respectively) on immunological, biochemical and antioxidants parameters as well as on gut microbiota in weaned piglets. EO was extracted by hydrodistillation and nanoencapsulation was performed by highenergy method using Tween 80 and Span 60 as surfactants. EO and NEO were chemically analyzed by gas chromatography-mass spectrometry (GC-MS). The cytotoxic effects of both EO and NEO was evaluated on Caco-2 cell line. For in vivo assay, piglets were randomly distributed in six groups of six animals each (n = 6) and received orally EO (10 mg/kg/day) or NEO (2.5, 5 and 10 mg/kg/day) for 30 consecutive days. Animals not treated or treated with surfactants mixture were evaluated as control and vehicle control. Subsequently, histological, hematological and biochemical parameters, cytokines production, oxidative markers, CD4 + /CD8 + T cells and gut microbiota were evaluated. GC-MS analysis was similar in both EO and NEO. The NEO was more toxic on Caco-2 cells than EO. Oral supplementation of EO or NEO (10 mg/kg/day) increased growth performance compared to control group or NEO (2.5 or 5 mg/kg/day) (p < 0.05) groups. NEO (2.5, 5 and 10 mg/kg/day) did not alter the morpho-physiology of digestive organs and decreased MDA levels in liver (p < 0.05), resulting safer than EO. In addition, NEO (10 mg/kg/day) showed an increase in CD4 + /CD8 + T cells ratio (p < 0.001), and induced the highest serum levels of IL-10 (p < 0.01). Serum triglycerides levels were signi cantly lower in animals treated with EO or NEO (2.5, 5 and 10 mg/kg/day) compared to control group (p < 0.001). Gut microbiota analysis showed that NEO (10 mg/kg/day) favor the development of bene cial intestinal microorganisms to maintain an anti-in ammatory microenvironment. In conclusion, EO and NEO improved parameters altered by early weaning in piglets however, NEO was safer and powerful. Therefore, NEO should be further studied to be applied in swine health. nanoencapsulated M. verticillata EO showed adjuvant properties of the immune response (Campra et al. 2022) or antimicrobial activity (Cecchini et al. 2021), higher than free EO.Therefore, the aim of this study was to evaluate the effect of oral supplementation of free and nanoencapsulated Minthostachys verticillata essential oil (EO and NEO, respectively) on immunological, biochemical and antioxidants parameters as well as on gut microbiota in weaned piglets. Materials and methods Plant materialThe aerial parts (leaves, thin stems and owers) of Minthostachys verticillata (Griseb.) Epling (Lamiaceae) (WFO 2023) were collected from cultivated plants grown in a eld located in