## 4 Discussion
gingivalis_, a 99.98% inhibition rate was observed, and, at 10 mg/mL, a 99.99% inhibition rate indicated significant antibacterial effects; however, complete bacterial eradication occurred at a concentration of 40 mg/mL. This suggests that the anaerobic nature of _P. gingivalis_, along with its ability to emit strong secretions of toxins such as ammonia, hydrogen sulfide, amines, and lipopolysaccharidesaccharides that degrade collagen [37], require higher concentrations for complete bacterial extinction compared to other oral-disease-causing bacteria. HGF plays a crucial role in the onset of periodontal disease through its relationship with various infiltrating inflammatory cells and exhibits regenerative capabilities during wound healing, thus playing an important role in the maintenance and repair of periodontal tissues [27]. Therefore, a toxicity assessment was conducted to ensure safe application of _Lespedeza cuneata_ extract while minimizing its impact on HGF. The WST-1 analysis method used in this study was developed to quantitatively assess cell proliferation and survival across a wide range of pharmacological and functional applications [38]. When quantifying cell viability using the WST-1 assay, it was found that concentrations of _Lespedeza cuneata_ extract above 20 mg/mL caused cell death in HGF cells, indicating a cytotoxic effect, and suggesting that concentrations below 20 mg/mL may be appropriate. This study confirmed that applying 10 mg/mL of _Lespedeza cuneata_ extract did not affect cell viability. The SRB analysis method is based on measuring cell protein content and is advantageous for confirming cell proliferation inhibition and cytotoxicity as it provides excellent linearity, sensitivity, and stable results [39, 40]. In this study, the extract affected changes in HGF cell morphology in a concentration-dependent manner. Regarding treatments up to 10 mg/mL, cell density and number decreased, but no changes in cell morphology were detected, and detached or dead cells were also not observed. However, morphological changes began to appear in cells treated with 20 mg/mL of _Lespedeza cuneata_ extract, confirming that 10 mg/mL is a safe concentration. Based on the results above, _Lespedeza cuneata_ extract has shown antibacterial efficacy against _P. gingivalis_, the causative bacterium of periodontal disease. For stable application, using a concentration of 10 mg/mL, which corresponds to the half-maximal inhibitory concentration (IC50), will not affect normal oral periodontal cells and may be safely and effectively applied as a natural antibiotic for the prevention and improvement of periodontal disease without chemical side effects. A limitation of this study is that only _P. gingivalis_, a representative bacterium causing periodontal disease, was examined, indicating a need for further research on the antibacterial effects against various other bacteria that contribute to the occurrence of periodontal disease. Future studies will be needed to fractionate _Lespedeza cuneata_ extract and evaluate the most active components to determine their specific contribution. Additionally, clinical studies applying _Lespedeza cuneata_ extract within a safe concentration range in products related to periodontal disease improvement, such as mouthwashes and toothpastes, will be necessary to evaluate its practicality and clinical applicability. This study not only demonstrates the antibacterial effect of _Lespedeza cuneata_ extract against _P. gingivalis_, a major contributor to periodontal disease, but also verifies its safe functioning at clinically relevant concentrations, thus providing foundational data for the development of periodontal treatment agents containing _Lespedeza cuneata_ extract and establishing a basis for practical application.