Desenvolvimento e caracterização de microemulsão a base do óleo de alfavaca (Ocimum gratissimum) para avaliação da atividade antioxidante, citotóxica e antimicrobiana

Abstract

Ocimum gratissimum, popularly known in Brazil as “alfavaca,” “manjericão,” “alfavacão,” or “alfavaca-cravo,” belongs to the Lamiaceae family, is native to Africa and Asia, and is widely distributed throughout Brazil. Studies on its therapeutic applications have revealed significant pharmacological diversity, with antibacterial, antifungal, hypoglycemic, and antioxidant properties documented in different experimental models. Several biologically active monoterpenes and sesquiterpenes have been described for this genus, especially regarding its antimicrobial potential. In this context, the aim of this study was to develop a microemulsion containing Ocimum gratissimum essential oil (OGEO), obtained by hydrodistillation of the leaves. The essential oils were analyzed by gas chromatography coupled with mass spectrometry (GC-MS) and subsequently evaluated regarding seasonal and circadian influences on chemical composition, as well as antioxidant, cytotoxic, antibacterial, and antifungal activities. Significant variations were observed in both yield and concentration of the essential oil constituents throughout the evaluated periods. On average, 23 compounds were identified, with carvacrol (38.4%–63.5%), p-cymene (9.3%– 19.3%), and γ-terpinene (8.7%–27.9%) being the major constituents in all samples. For microemulsion development, a phase diagram was constructed, allowing the identification of the most stable system proportion, composed of 18.2% oil, 72.7% surfactant/cosurfactant, and 9.1% water. The selected formulation (ME6) was characterized in terms of particle size (43.82 ± 0.319 nm), polydispersity index (0.416 ± 0.021), zeta potential (−26.2 mV), thermal stability, conductivity (76.8 μS/cm), and pH (6.42–5.37). Headspace GC-MS analysis confirmed the incorporation of the essential oil into the microemulsion matrix, highlighting carvacrol (40.39%), γ-terpinene (22.51%), and p-cymene (12.57%) as the major constituents present in ME6. The microemulsion exhibited superior biological performance compared to the Negative Control (ME-Negative Control) and the isolated essential oil (EO) against all tested microorganisms, both in terms of bacteriostatic activity (MIC) and bactericidal activity (MBC). Notably, it demonstrated strong inhibitory effects against Staphylococcus aureus and Klebsiella pneumoniae, maintaining high microbial growth inhibition even at a concentration of 2000 mg/mL. Microbiological analyses demonstrated that the microemulsion (ME6) presented MIC values ranging from 15.62 to 250 mg/mL against the tested bacteria, whereas the isolated essential oil exhibited MIC values ranging from 250 to 2000 mg/mL, indicating up to a 16- fold increase in antimicrobial potency after microemulsification. Antifungal activity was also enhanced, with a reduction in the minimum fungicidal concentration (MFC) from 500 mg/mL (EO) to 15.62 mg/mL (ME6) against Candida albicans and Candida parapsilosis strains. In the antioxidant assay, the microemulsion exhibited an IC50 value of 39.38 mg/mL, whereas the isolated essential oil showed an IC50 value of 70.51 mg/mL, confirming a 44.14% increase in antioxidant capacity after microemulsification. In hemolysis assays, OGEO showed values below 10% only at concentrations lower than 500 μg/mL, ranging from 9.8% (500 μg/mL) to 9.6% (15.62 μg/mL). In contrast, the microemulsion maintained hemolysis values below 10% even at higher concentrations (2000 μg/mL), reaching 0.3% at the lowest tested concentration (15.62 μg/mL), demonstrating a considerably milder cytotoxic profile compared to the free essential oil. Taken together, these results demonstrate that the microemulsion containing Ocimum gratissimum essential oil exhibited clear superiority over the non-formulated essential oil, presenting enhanced antimicrobial and antioxidant activities associated with a significant reduction in cytotoxicity. This combination of high physicochemical stability, improved biological efficiency, and enhanced safety profile reinforces the potential of this formulation as a promising candidate for the development of topical pharmaceutical products with antibacterial and antifungal properties. The findings of this study contribute to the advancement of pharmaceutical technologies applied to natural products and highlight relevant perspectives for future clinical and biotechnological applications.