Enrichment of Platelet-Rich Fibrin Using Cobalamin and Alpha-Tocopherol to Modulate Growth Factor Release

Platelet-rich fibrin (PRF) is an autologous biomaterial widely used in regenerative dentistry due to its biocompatibility and sustained release of growth factors. Recent research has shifted toward biologically modulating PRF to enhance its regenerative performance without altering its fundamental preparation principles. This study presents a protocol for biologically fine-tuning PRF through enrichment with cobalamin and alpha-tocopherol prior to centrifugation and evaluates their effects on growth factor and proinflammatory cytokine release under in vitro conditions. Peripheral blood samples obtained from healthy volunteers were processed using a fixed centrifugation protocol. Cobalamin or alpha-tocopherol was added to whole blood before centrifugation to obtain bioactively modified PRF matrices. The release profiles of platelet-derived growth factor (PDGF), transforming growth factor-β1 (TGF-β1), vascular endothelial growth factor (VEGF), insulin-like growth factor-1 (IGF-1), fibroblast growth factor-2 (FGF-2), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α) were quantified at 24 h and 72 h using an enzyme-linked immunosorbent assay. Cobalamin-enriched PRF was associated with significant intergroup differences in TGF-β1 levels and reduced release of proinflammatory cytokines, indicating a shift toward a more favorable regenerative microenvironment. In contrast, increases observed in FGF-2 and IGF-1 were time-dependent and detected across all groups, without demonstrating a cobalamin-specific effect. Alpha-tocopherol-enriched PRF exhibited a selective increase in VEGF release, suggesting a potential proangiogenic influence. These findings demonstrate that targeted vitamin enrichment enables controlled modulation of PRF bioactivity while preserving its autologous and anticoagulant-free nature. This protocol provides a practical framework for biologically fine-tuning PRF and supports its use as a tunable regenerative platform for future translational and clinical applications.