Polymerized human hemoglobin (PolyhHb) is being studied as a possible red blood cell (RBC) substitute for use in scenarios where blood is not available. While the O2 carrying capacity of PolyhHb makes it appealing as an O2 therapeutic, the commercial PolyhHb PolyHeme? (Northfield Laboratories Inc., Evanston, IL) was never approved for clinical use due to the presence of large quantities of low molecular weight polymeric (LMW) hemoglobin (Hb) species (< 500 kDa), which have been shown to elicit vasoconstriction, systemic hypertension, and oxidative tissue injury in vivo. Previous bench?top scale studies in our lab demonstrated the ability to synthesize and purify PolyhHb using a two?stage tangential flow filtration (TFF) purification process to remove almost all undesirable Hb species (>0.2 ?m and < 500 kDa) in the material, in order to create a product that should be safer for transfusion. Therefore, in order to enable future large animal studies and eventual human clinical trials, PolyhHb synthesis and purification processes need to be scaled up to the pilot scale. Hence in this study, we describe the pilot scale synthesis and purification of PolyhHb. Characterization of pilot scale PolyhHb showed that PolyhHb could be successfully produced to yield biophysical properties conducive for its use as an RBC substitute. Size exclusion chromatography showed that pilot scale PolyhHb yielded a high MW Hb polymer containing a small percentage of LMW Hb species (< 500 kDa). Additionally, the auto?oxidation rate of pilot scale PolyhHb was even lower than that of previous generations of PolyhHb. Taken together, these results demonstrate that PolyhHb has the ability to be seamlessly manufactured at the pilot scale to enable future large animal studies and clinical trials.This article is protected by copyright. All rights reserved.

» Publication Date: 19/09/2022

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