Poly(ethylene glycol)-Conjugation and Deoxygenation Enable Long-Term Preservation of Hemoglobin-Vesicles as Oxygen Carriers in a Liquid State

Abstract

The stability of hemoglobin vesicles (HbV) as an oxygen infusion was tested during the storage for 1 year at 4, 23, and 40 degrees C. The surface of the HbV was modified with poly(ethylene glycol) (PEG), and the suspension was deoxygenated with nitrogen bubbling. The samples stored at 4 and 23 degrees C showed a stable dispersion state for 1 year, though the sample stored at 40 degrees C showed the precipitation and decomposition of vesicular components, a decrease in pH, and 4% leakage of total Hb after 1 year. The PEG chains on the vesicular surface stabilize the dispersion state and prevent the aggregation and fusion due to their steric hindrance. The original metHb content (ca. 3%) before the preservation gradually decreased to less than 1% in all the samples after 1 month due to the presence of homocysteine inside the vesicles which consumed the residual oxygen and gradually reduced the trace amount of metHb. The rate of metHb formation was strongly dependent on the partial pressure of oxygen, and no increase in metHb formation was observed due to the intrinsic stability of the deoxygenated Hb. Preservation at 4 and 23 degrees C slightly reduced P(50) (increased the oxygen affinity) from 38 Torr to 32 and 31 Torr, respectively. These results indicate the possibility that HbV suspension can be stored at room temperature for at least 1 year.

Systemic and microvascular responses to hemorrhagic shock and resuscitation with Hb vesicles

“…Carbonylhemoglobin was converted to oxyhemoglobin by exposure to visible light in an O 2 atmosphere. HbVs were suspended in a physiological salt solution and filtered through sterilizable filters (pore size: 0.45 m, Dismic, Toyo Roshi; Tokyo, Japan) and deoxygenated with N 2 bubbling for storage (29). Physicochemical parameters of the HbVs are listed in Table 1.…”

Use of Hemoglobin Vesicles During Cardiopulmonary Bypass Priming Prevents Neurocognitive Decline in Rats

“…HbVs were suspended in a physiological salt solution at [Hb]ϭ10 g/dL, sterilized with filters (Dismic, ToyoRoshi, Tokyo, Japan, pore size, 0.45 m), and deoxygenated with N 2 bubbling for storage. 5 Before use, the HbV suspension ([Hb]ϭ10 g/dL, 8.6 mL) was mixed with a solution of human serum albumin (1.4 mL; Nipro, Osaka, Japan) to adjust the albumin concentration in the vesicle suspending medium to 5 g/dL. Under these conditions, the colloid osmotic pressure of the suspension is Ϸ20 mm Hg (Wescor 4420 Colloid Osmometer; Wescor, Logan, Utah).…”

“…Third, HbV is very stable and can be stored as a powder for a long time. 5 Fourth, HbVs are captured by phagocytes in the reticuloendothelial system and are metabolized within Ϸ7 days, without iron or lipid deposition. 6 The only concern posed by HbV for clinical use is its short half-life of only 35 hours in the circulating blood.…”

Fluid Resuscitation with Hemoglobin Vesicles PreventsEscherichia coliGrowth via Complement Activation in a Hemorrhagic Shock Rat Model

“…HbV has been shown to possess a number of positive characteristics: the absence of viral contamination (Sakai et al, 1993), a long-term storage period of more than 2 years at room temperature (Sakai et al, 2000b;Sou et al, 2000), low toxicity (blood compatibility, no nephrotoxicity) (Sakai et al, 2000a), and good metabolic performance (Taguchi et al, 2009b(Taguchi et al, , 2010. Moreover, the pharmacological effects of HbVs have been reported to be equivalent to that of RBCs, when evaluated in a model of hemorrhagic shock in rats (Sakai et al, 2004b.…”