Purpose: To evaluate the potential cytotoxic effects of four one-step self-etching dental adhesives [Adper Easy One (AEO), iBond (IB), Clearfil S³ Bond (CSB), and G-Bond (GB)] on cultured human periodontal ligament fibroblasts. Materials and Methods: Cured adhesives were immersed in complete DMEM or deionized water and maintained at 37°C for 24 h, followed by sterilization. The deionized water-based extract was used for Fourier transform infrared spectroscopy analysis. The DMEM-based extract was diluted into various concentrations for cytotoxicity tests. The viability, integrity, and apoptosis of cultured human periodontal ligament fibroblasts upon treatment with the extracts were determined using the CCK-8 assay, microscopy, and flow cytometry. Results: All of the four adhesives induced cell viability loss, cell morphology alteration, and cell death. GB showed the greatest cytotoxicity by inducing cell apoptosis and necrosis, while IB had the weakest cytotoxic effect on the cultured cells. Conclusion: All tested dental adhesives have significant adverse effects on cell viability. Therefore, precautions should be taken to protect the periodontal tissues when dental adhesives are applied in the clinic. Keywords: one-step self-etching adhesive, cytotoxicity, human periodontal ligament fibroblasts