Objective: To investigate the contraceptive effect of intratesticular injection of chlorhexidine gluconate in dogs and compare it with that of zinc gluconate neutralized with arginine.Methods: Twenty-four sexually mature male mongrel dogs were randomly divided by replicate into four groups (n=6 per group). Group I received intra-testicular injection of 2 mL zinc gluconate (10 mg/mL) neutralized with arginine. Group II received intratesticular injection of 2 mL chlorhexidine gluconate (5% w/v). Group Ⅲ received intratesticular injection of 2 mL chlorhexidine gluconate (4% w/v). Group Ⅳ did not receive any treatment and served as the control group. Testicular morphometry was conducted on day 0, 7, 15 and 30 after treatment. Semen was collected and evaluated on day 0 and 30. Data were analyzed using repeated measures analysis of variance.Results: There was no difference in the mean values of various parameters between dogs treated with zinc gluconate and those treated with chlorhexidine gluconate at any of the time points. In dogs treated with zinc gluconate or chlorhexidine gluconate, there was a significant increase in the testicular morphometric parameters on day 7 followed by a significant reduction thereafter (day 15 and 30). In contrast, there was no change in any of the parameters in the control untreated dogs during the course of the study. Compared to the pre-treatment values, the mean scrotal circumference and the mean paired testicular volume and testicular weight on day 30 were significantly lower in the treated dogs. Semen samples collected on day 30 from treated dogs were found to be azoospermic, whereas no change in semen quality was observed in the control untreated dogs. Conclusions: Intratesticular injection of chlorhexidine gluconate (5% w/v and 4% w/v) is equally as effective as zinc gluconate neutralized with arginine for chemical contraception in dogs.