Objective: To evaluate the efficiency of humic acid substances on removing micro-organisms from denture base materials.Background: Old denture wearer needs effective, easy-use and safe denture-cleaning material.Material and Methods: Square-shaped, heat-polymerised acrylic resin specimens (n = 550) were prepared and divided into five groups (n = 110 for each) corresponding to the microbial contamination (Candida albicans, Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, Pseudomonas aeruginosa). Contaminated specimens were randomly assigned to the application of five different denture-cleaning agents as follows (n = 20 for each): Kloroben, Corsodyl, Steradent, Corega, experimental solution with humic acid. Ten specimens were assessed as an experimental control carried out simultaneously for the treatment groups for each micro-organism. It was divided into two groups: negative control and positive control (n = 5 for each). All acrylic specimens were incubated 37°C for 24 h (for bacterial strains) and 37°C for 48 h (for yeast strains). After incubation period, all brain-heart infusion broths (BHI) which contain disinfectant acrylic specimens were cultured on 5% sheep blood agar (for bacteria) and Sabouraud dextrose agar (SDA) for yeast using loop. The numbers of colony-forming units per millilitre (CFU/ml) were calculated. The results were analysed by Mann-Whitney U-test and Kruskal-Wallis tests (p = 0.05).Results: Corsodyl and Kloroben completely eliminated the adherence of all investigated micro-organisms (100%) and showed the highest removal activity compared with other cleaning agents (p < 0.05). There was no statistically significant difference between Corsodyl and Kloroben (p ≥ 0.05), and there was no statistically significant difference between Corega, Steradent and experimental solution (p ≥ 0.05).Conclusion: Humic acid could be used as an alternative 'natural' solution for denture-cleaning agent.