Cross-linked hemoglobin as a potential membrane for an artificial red blood cell

A cross-linked hemoglobin membrane has been created with discerning permeability between dissolved hemoglobin and small molecules. Such a membrane could be used to enclose a sphere of hemoglobin solution thereby allowing the entire "cell" to transport oxygen. The hemoglobin membrane was cross-linked on a polycarbonate support; the mechanical support was necessary for diffusion experiments in this study and would not be used during any sphere preparation. A 30% methemoglobin solution in phosphate buffer was used to fill the pores of the 10 microm polycarbonate support, then cross-linked with a homobifunctional cross-linking agent. The cross-linked hemoglobin within the support was evaluated for hemoglobin and benzoic acid permeability in a side-by-side diffusion cell. Disuccinimidyl glutarate, disuccinimidyl suberate and disuccinimidyl tartrate were used as cross-linking agents. Disuccinimidyl glutarate, 4.65 mM, created a hemoglobin-impermeable membrane after cross-linking for 20 minutes, reducing the concentration to 0.46 mM required a cross-linking time to 60 minutes. Benzoic acid, representing a typical small molecule, was capable of diffusing through the disuccinimidyl glutarate cross-linked hemoglobin membrane at 87.2% of its diffusion through buffer.