In vitro maturation (IVM) process aims to provide oocytes with the necessary conditions to complete a set of processes in the nucleus and cytoplasm, which enable meiotic progression and the production of mature and potentially fertilizable oocytes [1]. During IVM, the cumulus-oocyte complexes (COCs) are exposed to several adverse conditions such as light, temperature, oxygen tension, pH variation, and composition of the media used for manipulation and in vitro culture [2,3]. These conditions can cause metabolic imbalances leading to reactive oxygen species (ROS) over-production and oxidative stress, damaging DNA, RNA, and proteins structures, resulting in oocyte viability reduction [4,5]. An effective strategy that might mitigate those oxidative stress deleterious effects during IVM is the media supplementation using compounds with antioxidant activity. Antioxidant agents can neutralize ROS through simple electron transference [6], or by converting highly oxidizing molecules into products with lower oxidative potential, thus reestablishing the balance between ROS, and their removal by antioxidant enzymes and small-molecular-weight antioxidants [7].

Several substances are known for their antioxidant potential during IVM, such as polyphenols - resveratrol [8,9], rosmarinic acid [10], and eugenol [11]; flavonoids - quercetin and taxifolin [12,13]; vitamins and minerals - selenium, tocopherol [14] and ascorbic acid [15]; carotenoids - β-carotene [16]; and aromatic compounds, such as anethole [17,18]. Anethole, or trans-anethole (1-methoxy-4 [1-propenyl]-benzene), is a natural phenylpropanoid obtained mostly from essential oil of some plant species such as star anise (Illicium verum), anise (Pimpinella anisum), and fennel (Foeniculum vulgare), comprising around 90 % of their composition [19,20].

In vivo studies, anethole oral administration increased endogenous glutathione and superoxide dismutase levels as well as total antioxidant capacity in mice [21] and broiler chickens [22], respectively. In vitro, anethole enable lipid peroxidation inhibition by ROS levels modulation and mTOR/PPARγ regulation in human mesenchymal stem cells [23]. Anethole antioxidant properties were also reported after supplementation for goat ovarian tissue fragments in vitro culture media [24]; isolated secondary follicles [25]; and early antral follicles [26], bovine COCs [17,18], and bovine [17] and porcine embryos [27].

During IVM in bovine species, different anethole concentrations (30, 300, or 2000 μg/mL) were tested [17,18]. In a study with bovine COCs, adding 300 μg/mL of anethole into maturation medium promoted antioxidant activity increasing FRAP levels and oocytes mitochondrial activity after IVM, as well as increased rates of morulae and blastocysts after in vitro embryo culture (IVEC) [18]. On the other hand, another study also in bovine species used different base media for IVM and IVEC, adding during IVM or IVM and IVEC 300 μg/mL of anethole, producing no effects on ROS levels or blastocyst production [17]. In this study, beneficial effect of anethole on blastocyst production was observed only when 30 μg/mL anethole was added to IVEC medium, accompanied by ROS reduction [17]. The effect of anethole supplementation has also been investigated in swine where different anethole concentrations (0, 300, 500, or 1000 μg/mL) were added to IVEC medium of porcine and influenced embryo production rates, ROS, and glutathione levels, with the best results obtained when the 500 μg/mL was used [27]. In general, bovine oocytes, bovine and porcine embryos, as well as goat ovarian follicles were tolerant to different anethole concentrations added to the culture media. However, other studies have shown that anethole can exert a concentration-dependent cytotoxic effect on both cancer cells and normal cells cultured in vitro [28].

Based on literature, it is possible to infer that the effect of anethole varies according to concentration, moment of addition (IVM versus IVEC), structure studied (ovarian follicles, COCs and embryos), and species. However, up to now, there are no studies investigating the effect of anethole addition during caprine IVM and neither its possible impacts on in vitro embryo production. Thus, the aim of this study was to evaluate the effects of adding different anethole concentrations to a standard IVM medium, on goat oocyte viability and maturation, oxidative stress, and embryo development in vitro after oocytes parthenogenetic activation.