AVALIAÇÃO DO POTENCIAL ANTIMICROBIANO E ANTIBIOFILME DE UM INIBIDOR DE TRIPSINA EXTRAÍDO DAS SEMENTES DE POINCIANELLA PYRAMIDALIS (TUL) L.P. QUEIROZ

Abstract

Microorganisms have evolved various resistance mechanisms to survive, which has led to the rise of antimicrobial resistance. This global issue poses a significant threat to public health, resulting in economic damage and high mortality rates. Given this, there's a clear need to search for new compounds with antimicrobial properties. Peptidase inhibitors (PIs) are polypeptides or proteins that play a role in plant defense and have potential biotechnological applications, such as the trypsin inhibitor derived from Poincianella pyramidalis (PpyTI). Despite its proven insecticidal activity, there are no reports in the literature regarding its potential antimicrobial and antibiofilm properties. Therefore, this study aimed to determine the antimicrobial and antibiofilm activity of PpyTI. PpyTI was isolated using chromatographic techniques followed by SDS-PAGE for protein electrophoresis. The minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and minimum fungicidal concentration (MFC) were determined for Gram-positive and Gram-negative bacteria as well as yeasts. Sorbitol, ergosterol, and the fluorescent dye Sytox™ Green were used to investigate potential mechanisms of action of the inhibitor. PpyTI showed no activity against Gram-positive and Gram-negative bacteria but exhibited significant fungicidal activity, with MIC and MFC values of 3.28 μM for C. tropicalis and MIC and MFC values of 6.56 μM for C. albicans. When used in combination with fluconazole, the inhibitor displayed a synergistic effect, reducing the MIC of both compounds against C. albicans. The addition of ergosterol led to an increase in the MIC for both strains, suggesting that the mechanism of action of PpyTI involves altering the integrity of the fungal plasma membrane. These findings were supported by results from Sytox™ Green, a nucleic acid stain that cannot penetrate intact membranes. Additionally, PpyTI was able to inhibit biofilm formation by damaging the cell membrane of the fungal biofilm cells, reducing its mass. These results were superior to those observed with the antifungals amphotericin B and fluconazole. The data presented in this study provide evidence that PpyTI could be used in developing therapeutic strategies for treating fungal infections.