The objective of this study is to determine the in vitro cytotoxicity effect of chitosan hydrogel-based metronidazole on fibroblast. Chitosan hydrogel-based metronidazole(0.12%, 0.15%, 0.25%) and chitosan without metronidazole are made by dissolving lactic acid to reach a gel consistency, and then adding powdered metronidazole according to the concentration. 3T3 fibroblast cells were cultured in well mikroplate containing RPMI-1640 media inside an incubator. The cell viability tests were carried out using theMTT assay method and repeated five times. The colour degradation was read with a microplate reader. The mean viability of fibroblasts mixed with chitosan hydrogel-based metronidazole at concentration of 0.12%, 0.15%, and 0.25%, and chitosan without metronidazole were 96%, 93%, 91% and 98.8% respectively. ANOVA statistical test results for the 0.12% concentration group, where p < 0.05 represents the cell's viability, significantly differs from the results for the 0.15% and 0.25% concentration groups. Chitosan hydrogel-based metronidazole shows non toxic effects on fibroblast cells.