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Herbal medicines with anthelmintic effects are alternatives for the sustainable control and prevention of disease caused by gastrointestinal parasites. The nanoencapsulation of essential oils has been proposed to enhance the absorption of their constituents and improve their efficacy. The present study aimed to evaluate the efficacy of free and nanoencapsulated Eucalyptus citriodora essential oil (EcEO) on the control of gastrointestinal nematodes of small ruminants in vitro and in vivo. Chitosan was used as a matrix for the formulation of a nanoemulsion. Chromatographic and physico-chemical analyses of EcEO were performed. Egg hatch (EHT) and larval development (LDT) tests were conducted to evaluate the effectiveness of nanoencapsulated and free EcEO on the eggs and larvae of Haemonchus contortus. Acute toxicity of free and nanoencapsulated EcEO was evaluated using mice. Finally, nanoencapsulated EcEO efficacy on the control of gastrointestinal nematodes was calculated by fecal egg count reduction test (FECRT) treating 30 sheep naturally infected with 250 mg/kg of free and nanoencapsulated EcEO. In vitro tests were analyzed by an analysis of variance (ANOVA) followed by comparison with the Tukey test. The efficacy of FECRT was calculated by the BootStreet program through arithmetic average, using the formula 100 (1-XT/XC). To compare the differences between epg, the data were transformed to log(x+1) and subjected to an ANOVA to compare the significant differences between groups by Tukey's. The level of significance was P<0.05. The free (4 mg/ml concentration) and nanoencapsulated (2mg/ml concentration) EcEO inhibited larvae hatching by 97.2% and 92.8%, respectively. Free and nanoencapsulated EcEO at 8 mg/ml inhibited larval development by 99.8% and 98.1%, respectively. In the acute toxicity test, the LD10 and LD50 of free EcEO was 1999 and 2653 mg/kg, respectively, while the LD10 and LD50 of nanoencapsulated EcEO was 1121 and 1681 mg/kg, respectively. Nanoencapsulated and free EcEO reduced FEC similarly by 40.5% and 55.9%, respectively at 10 days post-treatment. Nanoencapsulated EcEO did not obtain the expected efficacy in vivo.

Phytotherapy has been described as an alternative method for the control of gastrointestinal nematodes in small ruminants. Goal of the encapsulation of essential oils in biopolymer matrices is to optimize the biological effects of these oils. The aim of the present study was to evaluate the in vitro and in vivo anthelmintic activity of encapsulated Eucalyptus staigeriana essential oil (EncEs) on the eggs and larvae of Haemonchus contortus. Therefore, the egg hatching test (EHT), larval development test (LDT) and worm load evaluation were performed to evaluate Meriones unguiculatus experimentally infected with H. contortus. The chemical constituents of E. staigeriana essential oil (EsEO) and the in vitro oil release profile from the chitosan matrix at a pH of 1.2 and 7.0 were also characterized. EncEs and EsEO inhibited larval hatching by 97.19% and 99.96% at doses of 1.5 and 1.0 mg ml(-1), respectively. In the LDT, EncEs and EsEO induced a larvicidal effect greater than 95% at concentrations of 5.8 and 8 mg ml(-1), respectively. EncEs and EsEO decreased H. contortus load in M. unguiculatus by 40.51% and 46.44%, respectively. The major chemical constituents of EsEO were (+)-Limonene (72.9%), 1,8-Cineole (9.5%) and o-Cimene (4.6%). The release profile of EsEO was 30% in acid and 25% at neutral pH. The similar efficacy of EncEs and EsEO demonstrates that there was no optimization of anthelmintic action following the encapsulation process. Therefore, the use of new encapsulation matrices with controlled release in the pH of the abomasum should be investigated.

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