Candida albicans is a commensal organism that can cause disease in individuals with weakened and compromised immune systems. These fungi can cause superficial or invasive infections associated with high mortality. The aim of this study was to compare the activity of the new antifungal compound Venetin-1, obtained from the coelomic fluid of the earthworm Dendrobaena veneta, with the effects of the antibiotic amphotericin B. Flow cytometry, fluorescence microscopy, scanning electron microscopy, atomic force microscopy, and FTIR spectroscopic methods were used for comparative analysis. After using selected preparations at specified concentrations, it was found that AmB reduced fungal cell survival to a greater extent than Venetin-1. The present study shows that both Venetin-1 and AmB induced fungal cell necrosis and proliferation. SEM analysis showed that both compounds caused morphological changes in cells, including the presence of multiple bundles at the cell poles and the formation of unseparated cell chains. Congo red fluorochrome analysis showed that both compounds exposed the B-glucan layer of the cell wall, which was revealed by red fluorescence. Flow cytometric analysis of the cell wall showed that V-1 caused exposure of the β-glucan and chitin layers in the fungal cells. FTIR analysis confirmed molecular remodelling of the cell wall structure after exposure to both compounds. AFM analysis indicated an increase in the elasticity of the fungal cell wall after treatment with both compounds. The comparison of Venetin-1 and amphotericin B revealed pronounced differences in their electrokinetic behaviour, originating from their distinct chemical structures and modes of interaction with electrolyte ions. Venetin-1 exhibited significantly more negative zeta potential values, particularly in alkaline media and in the presence of Cl⁻ ions, indicating stronger electrostatic stabilisation and higher electrokinetic stability than Amphotericin B. Considering its effect on Candida cells and the fact that the Venetin-1 complex has no toxic effect, this compound is a promising candidate for further biomedical research on new antifungal preparations.