While traditional periodontal treatments like scaling and root planing, antibiotics, and surgical intervention remain the primary approaches, herbal medicine is continuously evolving as an alternative for the management of periodontal diseases. This study focused on the evaluation of the cytotoxicity of Ocimum sanctum (OS)-based hydrogel for its use in local drug deliveryin periodontitis. OS-based hydrogel was prepared using 50 g of OS powder and 200 ml of ethanol, along with carboxymethyl cellulose gel and sorbitol. The prepared formulation was evaluated for its cytotoxicity by using the MTT assay, cell viability, cell morphology, and brine shrimp egg assessment. Cell viability was assessed, and it was above 95% for the control and 85% for the OS hydrogel by the 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide assay. Brine shrimp egg assessment also showed a survival rate of 80% at low concentrations. The cell morphology test showed round and uniform cells growing in a monolayer shape. The findings of this study confirmed that OS-based hydrogel is cytocompatible and, hence, can be used as a local drug delivery agent for periodontitis management, especially in resource-constrained settings where affordable and natural treatment options are highly valued.