Background <p>The aim of the study is to evaluate saliva levels of Soluble urokinase plasminogen activator receptor (suPAR), Hypoxia-inducible factor-1 alpha (HIF-1α) and tumor necrosis factor-alpha (TNF-α) in stage III grade A, grade B, grade C periodontitis and periodontal health and to understand the roles of these molecules in periodontal inflammation process and also to compare the three biomarkers' discriminative efficacy in periodontal disease.</p> Methods <p>A total of 80 individuals, 20 with stage III grade A periodontitis (group A), 20 with stage III grade B periodontitis (group B), 20 with stage III grade C periodontitis (group C) and 20 with healthy periodontium (group H) were recruited for this study. Full-mouth clinical periodontal measurements were recorded in periodontal charts. Whole saliva samples were collected to determine the levels of suPAR, HIF-1α and TNF-α in study groups using enzyme-linked immunosorbent assay (ELISA) method.</p> Results <p>The saliva concentration of suPAR, HIF-1α, and TNF α was significantly higher in group A, group B, and group C compared with group H (<i>p</i> &lt; .05). Additionally, salivary suPAR concentration was significantly higher in group C than in groups A and B (<i>p</i> &lt; .05). Positive statistically significant correlations were observed between three biomarkers and all clinical parameters (<i>p</i> &lt; .05).</p> Conclusions <p>Increased levels of saliva suPAR, HIF-1α, and TNF α suggest that these molecules may play a role in periodontitis. In addition, the higher salivary suPAR levels in grade C periodontitis compared to other grades suggest that suPAR may be one of the potential molecules that can be used to predict disease progression and periodontal disease classification.</p> Trial registration <p>Before starting the study, the study plan was uploaded to clinicaltrials.gov.tr ​​and an identification number was obtained (Date: 21.05.2024, İdentification number: NCT06430450).</p>