Abstract: The present invention provides a hemoglobin molecule (Hb) having six±one PEG chains, wherein two of said PEG chains are bound to Cys-93 (?) of Hb, and the remaining PEG chains are bound to thiol groups introduced on ?-NH2 of Hb. The present invention also provides a process for preparing a modified hemoglobin molecule (Hb), comprising the steps of: (a) reacting Hb with 8-15 fold excess of iminothiolane to form thiolated Hb; and (b) reacting the thiolated Hb with 16-30 fold excess of PEG functionalized with a maleimide moiety, to form the modified Hb.

Abstract: Novel modified hemoglobins comprising polyalkylene glycols and novel methods for making those hemoglobins are provided. One group of modified hemoglobins comprise polyalkylene glycols bonded to the hemoglobin with an amide linkage at Glu-43(?). Additional polyalkylene glycols can also be bonded to the Glu-22(?) and/or the Asp-47(?). These hemoglobins are made by a novel amidation procedure. A second group of modified hemoglobins comprise a polyalkylene glycol covalently bonded to the hemoglobin at the ?-amino of a Val-1(?) or a Val-1(?). Additional polyalkylene glycols can optionally be covalently bonded to a limited number of ?-amino groups. This second group of hemoglobins is made using a novel reductive alkylation procedure. A third group of modified hemoglobins comprise a polyalkylene glycol bonded to a thiol group of the hemoglobin through a phenylsuccinimido linkage, wherein no polyalkylene glycol is bonded to a Cys-93(?).

Abstract: The present invention is directed to pegylated red blood cells comprising a polyethylene glycol (PEG) attached to a thiolated amino group on a membrane protein, and to compositions comprising the pegylated red blood cells. The invention also provides methods of preparing pegylated red blood cells comprising reacting red blood cells with a compound that produces a thiolated amino group on a red blood cell membrane protein, and reacting the thiolated red blood cell with a PEG. The invention further provides methods of treatment using pegylated red blood cells.

Abstract: The present invention provides pegylated hemoglobins comprising a thiocarbamoyl-phenyl-polyethylene glycol (PEG) attached to hemoglobin, and comprising a polyethylene glycol (PEG) attached to hemoglobin by an acyl group. The invention also provides methods of preparing pegylated hemoglobins using isothiocyanato phenyl carbamate of PEG and using isothiocyanato phenyl di-PEG carbamate. The invention further provides compositions and blood substitutes comprising pegylated hemoglobins and methods of treating a subject which comprise administering to the subject blood substitutes comprising non-hypertensive pegylated hemoglobins.