Chitosan Nanogel Design on Gymnema sylvestre Essential Oils to Inhibit Growth of Candida albicans Biofilm and Investigation of Gene Expression ALS1, ALS3

Abstract

Chitosan (CS) is a polycation with a pka of about 6.3 with a charge density dependent on pH and the %DA-value which can interact with polyanions to form complex and gels. Nanoparticles (CNPs) in addition can increase the antifungal potential of bioactive compounds like essential oils by increasing cellular interactions between them and the fungal as a result of the very small size that enhances cellular uptake. In this study, was set to investigate the encapsulation of the Gymnema sylvestre essential oils (G.EOs) using Chitosan and Myrestic acid made Nanogel in order to enhance its antifungal activity and stability to the oil against C. albicans strain (ATCC 10231). To procedure this, the self-assembled process of Chitosan and Myrestic acid Nanogel (CS-MA) through the 1- ethyl - 3- (3 dimethyl aminoprophyl) carbodiimide (EDC) was designed. Its physicochemical properties were determined by Fourier Transforms Infrared spectroscopy (FT-IR), X-ray diffraction (XRD), and microscopic methods by Atomic force microscopy (AFM), Transmission Electron Microscopy (TEM), and Scanning Electron Microscopy (SEM). Minimum inhibitory concentration (MIC) at 18.7 to 37.5 µg/ml and 2.3 to 4.6 µg/ml and minimal fungicidal concentration (MFC) at 75 µg/ml and 5.38 µg/ml using by broth micro dilution (BMD) method for G. sylvestre oils (G.EOs) and oil-loaded Nanogels (G.OLNPs) were measurement. The susceptibility of C. albicans biofilm to fractions was examined by 2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-[8phenylamino) carbonyl] - 2H - tetrazolium hydroxide (XTT) quantitatively.The concentration of G.OLNPs required to inhibit 50 % biofilm formation was 4.68 µg/ml, while that to remove 90 % biofilm growth was 18.07 µg/ml. In addition, it was observed that cell uptake of G.OLNPs was much higher compared with free G.EOs. Reverse transcription polymerase chain reaction (RT-PCR) analysis was performed to determine the effect of sub-MIC concentrations of G.EOs and G.OLNPs on expression of the biofilm–related gene ALS1 / ALS3, and indicated the G.OLNPs down-regulated the expression of hypha-specific gene ALS3. Furthermore, the data strongly suggested that G.OLNPs more effective suppressed C. albicans planktonic cells and reduction biofilm biomass.