Supplementary Material for: Irradiation with 265-nm ultraviolet light-emitting diodes to plasma: Alterations of hemostatic parameters and von Willebrand factor

Introduction: Among pathogen inactivation methods for blood products, an ultraviolet C (UVC) irradiation has been attracting attention. We previously reported that 265 nm UV-LED (UVC-LED) effectively prevents bacterial infection. However, the effect of UVC-LED on plasma was not considered. This study aimed to explore the effects of high-dose UVC-LED irradiation, required for viral inactivation, on plasma hemostatic potential. Methods: After irradiation of plasma with high-dose UVC-LED for 60 min, the activities of coagulation factors and coagulation inhibitors and von Willebrand factor (VWF) antigen levels were determined. VWF multimer assay and global hemostatic analysis using reconstituted blood under flow were also conducted. Results: UVC-LED irradiation prolonged PT and APTT in a fluence-dependent manner. Fibrinogen levels and factors V and XI activities decreased to 40–47% of pre-irradiation levels. VWF multimer analysis revealed a striking reduction in high-molecular-weight VWF multimers, irrespective to ADAMTS13 activity, while VWF antigen levels showed a slight decrease. Electrophoresis under reducing conditions indicated minimal changes in the 250 kDa-VWF subunit band, suggesting that UVC-LED irradiation disrupts inter-subunit disulfide bonds in VWF multimers. Global hemostatic analysis using reconstituted blood with irradiated plasma showed a marked prolongation in primary hemostasis, indicating impaired platelet adhesion to collagen, reflecting reduced high-molecular-weight VWF multimers. Conclusions: We demonstrated for the first time that a high-dose UVC irradiation to plasma cleaves the inter-subunit disulfide bonds of VWF multimers and reduces the molecular size. In plasma products, the balance between maintaining hemostatic activity and inactivating various pathogens by UVC irradiation is a challenge for the future.