Untapped potential lies in the solid waste of freshwater mussel shells. Indeed, the shells possess a substantial amount of calcium. Valorization is essential for enhancing added value by utilizing calcium as an abrasive ingredient in toothpaste. This study aimed to investigate the characteristics of toothpaste containing abrasive components derived from freshwater mussel shells. The research procedure consisted of the production of CaO flour and the synthesis of hydroxyapatite through the calcination method. The test parameters consisted of evaluation of toothpaste preparations, analysis of calcium content, and antibacterial activity of S. aureus in 4 toothpaste formulations, namely T1 (CaO flour), T2 (commercial CaCO 3), T3 (hydroxyapatite), and T4 (commercial toothpaste). Except for treatment T1, the results demonstrated that toothpaste’s pH, spreadability, and foam formation fulfilled toothpaste quality standards. Every treatment’s toothpaste had a homogenous paste with good cleaning abilities. Treatment T2 had the greatest calcium level in toothpaste, measuring 2132.5 ppm. Subsequently, toothpaste T3 and T4 had antibacterial properties, however T1 and T2 did not possess any antibacterial action. The toothpaste formulation including hydroxyapatite as an abrasive, known as treatment T3, can be determined as the most effective.