Abstract Palindromic peptide LfcinB (21–25) Pal : RWQWRWQWR was synthesized by Solid Phase Peptide Synthesis (SPPS‐Fmoc/ t Bu), purified by Reverse Phase Solid Phase Extraction (RP‐SPE) and characterized by Reverse Phase High Performance Liquid Chromatography (RP‐HPLC) and Matrix‐Assisted Laser Desorption/Ionization‐Time Of Flight Mass Spectrometry (MALDI‐TOF MS). The antifungal activity of LfcinB (21–25) Pal against both ATCC strains and clinical isolates of C. albicans, C. glabrata, C. krusei, C. auris and C. tropicalis was evaluated. The palindromic peptide exhibited fungistatic and fungicidal activity against all yeast evaluated. The antifungal activity was dependent on peptide concentration in all cases. Additionally, LfcinB (21–25) Pal (25–50 μg/mL) combined with fluconazole exhibited a synergistic antifungal effect against C. tropicalis 883 and C. krusei 6258 (resistant to fluconazole). This study showed that the palindromic peptide derived from Lactoferricin B (LfcinB) exhibited significant antifungal activity against Candida spp, suggesting that this peptide could have a therapeutic application solely or in combination with fluconazole.