The study included 268 patients with benign paroxysmal positional vertigo (BPPV) and 268 age- and sex-matched healthy controls. Patients with BPPV had significantly higher serum levels of the inner ear biomarkers Otolin-1 (median 12.64 ng/ml vs. 7.58 ng/ml) and otoconin-90, which were independently associated with the likelihood of BPPV (adjusted OR for Otolin-1: 2.08, 95% CI: 1.49–2.91; for otoconin-90: 1.71). Poorer sleep quality according to the Pittsburgh Sleep Quality Index (PSQI) was associated with greater severity of dizziness as measured by the Dizziness Handicap Inventory (DHI) (β = 2.14 per PSQI point, 95% CI: 1.56–2.72). Higher PSQI scores were also associated with increased levels of both biomarkers, which were independently associated with vertigo severity. Including biomarkers in the models weakened the PSQI–DHI association (β from 2.14 to 1.28) and improved the explanatory power of the model (R² from 0.26 to 0.38). Exploratory mediation analyzes showed that Otolin-1 and Otokonin-90 statistically explained approximately 40% and 29% of the association between sleep quality and symptoms, respectively.