Oxygen storage materials: a new type of hemoglobin based oxygen carrier (HBOC)

Hemoglobin-based oxygen carriers (HBOCs) have been developed in last years as an alternative to red blood cells (RBCs) mostly composed of Hemoglobin (Hb) that is the natural carrier for oxygen transport in mammals. To overcome the problem of tissues necrosis in the inner regions of 3D scaffolds with low porosity and static conditions due to the major consumption of nutrients respect to the diffusion, we decided to realize a new type of HBOC through chemical conjugation between bovine gelatine non-porous microbeads (75-150 ?m diameter, 5% w/w cross-linked with glyceraldehyde-6-phosphate) and hemoglobin A0 ferrous stabilized. The goal is to create a carrier of molecular oxygen as natural as possible without loss of Hb to insert inside 3D scaffolds or to utilize it as oxygen releasing biomaterial with which to build the scaffolds themselves. Tissue engineering has always characterized by the death of the cells in the inner parts of the scaffolds due to the major consumption of nutrients respect to the diffusion. In the other cases, diffusion causes a wash out of extracellular matrix as an undesirable effect. As known, oxygen is the fundamental nutrient for the cells and we realized a new type of HBOC trough EDC/NHS and N,N?-Disuccinimidyl Carbonate (DSC) chemistries. We made several PQM test to evaluate if the HMsB (conjugated microbeads through DSC reaction) have better performance of HMsA (conjugated microbeads through EDC/NHS reaction) and we evaluated molecular oxygen absorption and release. Until now, no studies are published about the surface characterization and Hb-functionalization of non-porous-gelatin microbeads. In fact, we first characterized the surface of gelatin microbeads and then, after calculating the average amino Molar Degree of superficial functionalization (?), we conjugated hemoglobin to the surface of the microbeads and performed experiments to observe Hb presence and how it works. We observed that our HBOC release oxygen in hypoxic conditions and adsorb oxygen in O2 saturation conditions. Besides, DSC chemistry resulted more efficient than EDC/NHS one and HMsB have better performances than HMsA.