Lyophilization for long-term storage of perfluorocarbon-based artificial oxygen carriers

Allogenic blood transfusions are indispensable in modern medicine, yet limited. Perfluorocarbon (PFC)-based artificial oxygen carriers present a promising alternative due to high oxygen capacity. Emulsification renders PFC compatible for intravenous use. For application, long-term storage stability is crucial. Despite colloidal instability in PFC/water emulsions, lyophilization offers a gentle way to dry sensitive products, ensuring a stable formulation for clinical use.

The nanoemulsion consisted of a perfluorodecalin core and a mixed shell of albumin and lecithin (LENOX). Emulsification was carried out using a high-pressure homogenizer. After the synthesis of LENOX, different concentrations of trehalose were added as a cryoprotectant and the product was lyophilized, by using a protocol developed for a precursor formulation of LENOX. After a storage period of 24 h or 14 days the particles were reconstituted and subjected to physico-chemical characterization. Particle size was determined using dynamic light scattering, while viscosity was measured using a rheometer and oxygen capacity was measured using the Oroboros O2k respirometer.

Particle size measurements 24 h post-lyophilization revealed no significant increase compared to non-lyophilized LENOX stored at 4 °C for the same duration (control emulsion). The particles of the control emulsion exhibited significant growth during the 14 days of storage at 4 °C, unlike the lyophilized particles. After 14 days, the particles of the control emulsion were significantly larger than the lyophilized particles. The rheological analysis indicated that the lyophilization did not affect emulsion viscosity. Furthermore, the determination of oxygen capacity showed no loss of functionality in the lyophilized particles after 14 days of storage compared to freshly synthesized LENOX and to the control emulsion.

The data indicate that lyophilization improved the stability of LENOX compared to the control emulsion, as evidenced by the absence of particle growth and viscosity changes. Additionally, LENOX maintain their functionality post-lyophilization. Thus, lyophilization represents a promising storage option for the artificial oxygen carrier enabling long-term storage required for a clinical application.

Some of the authors are named as inventors in the patent of the lecithin modified nanoscale oxygen carrier (LENOX) with the application number EP 22193221.3, filed on Aug. 31st, 2022, that was used for this work.