Scalable Production\nand Biophysical Characterization\nof High-Molecular-Weight Relaxed and Tense Quaternary State Polymerized\nHuman Hemoglobin as Potential Red Blood Cell Substitutes

High-molecular-weight (HMW) (>500 kDa) glutaraldehyde-polymerized\nhuman hemoglobin (PolyhHb) is a promising hemoglobin-based oxygen\ncarrier (HBOC) due to its decreased risk of vasoconstriction and oxidative\ntissue injury. Previously, HMW tense (T) quaternary state PolyhHb\nwas synthesized at the pilot scale with tangential flow filtration\n(TFF) for the removal of low-molecular-weight species. However, T-state\nPolyhHb is limited to specific biomedical applications due to its\nlow oxygen affinity, thus motivating the need to produce high oxygen\naffinity relaxed (R) quaternary state PolyhHb at the pilot scale.\nThis study explored the pilot-scale synthesis and extensive biophysical\ncharacterization of both HMW T- and R-state PolyhHb. The resultant\ncharacterization demonstrated the successful synthesis of low and\nhigh oxygen affinity PolyhHb with increased molecular weight (∼1000–1500\nkDa). Overall, T- and R-state PolyhHb provides a platform for manufacturing\noxygen therapeutics with a diverse range of oxygen affinities and\npotential biomedical applications.