AIM: The aim of this study was to analyze the cytotoxicity of ciprofloxacin (CIP) loaded on chitosan bioactive glass scaffold on human periodontal ligament stem cells (PLSCs) in vitro.MATERIALS AND METHODS: PLSCs obtained from human third molars, cultures treated with medium containing 15 x 15 mm chitosan/bioactive glass scaffolds without/with different concentration 0, 5, 10, and 20 % of CIP. A total of 15 x 10^3 cells were plated in 6 well plates. The attached cells of each group were harvested from the plates after 1, 4 and 8 days of culture to detect the viability of cells. The cell number was determined using a hemocytometer and the trypan blue dye-exclusion assay. Data was analyzed using normality using Shapiro-Wilk test. Comparisons between groups were made using One-way ANOVA complemented by Tukey's test.RESULTS: When comparing the proliferation rate of cells in the four groups, no statistically significant difference was found (P = 0.633). With regards to cell viability, no statistical difference was found between the 0, 5, and 10 % CIP concentrations, while the 20 % CIP concentration demonstrated the least viability with a high statistically significant difference (P = 0.003).CONCLUSION: Twenty percentages CIP demonstrated the least proliferation rate and viability.