Desenvolvimento de emulgel à base de R-(+)-limoneno para tratamento de úlceras traumáticas da cavidade oral

Abstract

Traumatic ulcers (UTs) of the oral mucosa consist of a group of common changes in the oral cavity that cause painful symptoms and affect the well-being of the patient. The therapies available for the treatment of these injuries are isolated in terms of application and the occurrence of adverse effects. In this context, the search for new therapeutic alternatives from medicinal plants, such as limonene, a terpene with properties of pharmacological interest, stands out. This study evaluated in silico and in vitro, the antifungal activity of R-(+)-limonene isolated on Candida spp, and incorporated into an emulgel its physicochemical properties, antifungal, toxicological, anti-inflammatory and healing action of this emulgel. Through the in silico assay, the probability of the biological potential of the monoterpene, its theoretical oral bioavailability and acute oral toxicity were evaluated. The following strains were used: C. albicans ATCC 10231, C. glabrata ATCC 90031, C. krusei ATCC 6258, C. tropicalis ATCC 750, C. parapsilosis (ATCC 22019) and clinical isolates of C. albicans A1, and C. albicans A2. The Minimum Inhibitory Concentration (MIC), Minimum Fungicide Concentration (MFC) was determined by microdilution, its effect on the growth kinetics of C. albicans and its antifungal potential by logarithmic reduction. The physical-chemical properties of the emulgel based on R-(+)-limonene (ELI) were determined by means of pH, resistance to centrifugation, accelerated centrifugation and spreadability. Acute oral toxicity and clastogenic and aneugenic potential were evaluated in Mus musculus mice (2000mg/kg). Wistar rats were submitted to the application of 5% acetic acid in the buccal mucosa to evaluate the anti-inflammatory and healing potential of the formulation, through clinical and histopathological analysis. Data were analyzed using descriptive and inferential statistics, analyzed using GraphPad Prism 5.0® software, San Diego, CA, USA. The in silico assays estimated that R-(+)-limonene has a broad spectrum of action on microorganisms of great clinical importance. The MIC of R-(+)-limonene ranged from 656.25 µg/mL to 2625 µg/mL and was classified as fungistatic on the C. albicans strain (ATCC 10231), while for the other species its action was fungicide. R-(+)-limonene had a potentially significant effect on the growth kinetics of C. albicans (p<0.05). The ELI showed an acidic pH of 5.71 ± 0.11; 5.71 ± 0.09 and 5.97 ± 0.05 for formulation of the emulgel with R-(+)-limonene (ELI) at 1.2%, 2.4% and 5%, respectively, and 5.74 ± 0.21 for the control formulation, good stability and spreadability, as well as good viscosity. Through analysis of growth kinetics and logarithmic reduction, it can be seen that the product significantly inhibited the growth of C. albicans in all the different concentrations tested when compared to the growth control. ELI did not induce signs of acute toxicity in mice submitted to gavage treatment at a single dose of 2000 mg/kg and produced a statistically significant lower number of micronuclei compared to the positive control (cyclophosphamide 50 mg/kg) in mouse erythrocytes. R-(+)-limonene has been shown to be a potential antifungal agent against Candida species, both as an isolated compound and incorporated into the emulgel. The ELI showed low toxicity and mutagenicity, in addition to good anti-inflammatory and healing action in the treatment of UTs in the oral cavity of Wistar rats. Therefore, the developed formulation was considered promising for the treatment of UTs in the oral cavity.