This study evaluated the effectiveness of three disinfectants used in Dentistry for decontamination of gutta-percha cones. Sixty gutta-percha cones were contaminated with standardized pure cultures of five species of microorganisms (Enterococcus faecalis ATCC 29212, Staphylococcus aureus ATCC 25923, Candida albicans ATCC CBS-ICB/USP 562, Bacillus subtilis spores ATCC 6633 and Streptococcus mutans ATCC 25175). The cones were treated with 10% polyvinylpyrrolidone-iodine aqueous solution (PVP-I; Groups 1 and 2), 5.25% aqueous sodium hypochlorite (Groups 3 and 4) and paraformaldehyde tablets (Group 5). All chemical agents were efficient for the cold sterilization of gutta-percha cones in short time periods.