Biotest-serum-institut Gmbh

Klaus Bonhard

Norbert Kothe

Uwe Boysen

Bertram Eichentopf

Hans Schleubner

Crosslinked hemoglobin preparations of extended shelf life and high oxygen transport capacity can be made by reducing a hemoglobin solution with an oxygen consuming reducing agent down to an oxygen partial pressure of 0 mbar, then adding an effector, crosslinking with a dialdehyde, reducing with a carbonyl-group-specific reducing agent, circulating through an ultrafilter, and stabilizing the product by adding a reducing agent thereto.

Crosslinked hemoglobin of extended shelf life and high oxygen transport

A process for obtaining hepatitis-safe, sterile, pyrogen- and stroma-free hemoglobin solutions of low potassium content comprising stirring an erythrocyte concentrate with about 1 to 6-times its volume of an about 5 to 15% aqueous solution of a sugar, sugar alcohol or high-molecular-weight colloid, adjusting the pH of the resulting suspension to about 5 to 6.5 with an acid which is physiologically tolerable as salt, allowing the erythrocytes to settle, removing the supernatant from the sediments, adjusting the pH of the sediment to about 7 to 8, homogenizing the sediment, adjusting its hematocrit to about 55 to 60%, treating the mass with a dilute beta-propiolactone solution, hemolyzing the residual solids and forming a suspension thereof, contacting the suspension with a cation exchanger in H.sup.+ form until the pH has decreased to about 5 to 5.5 thereby to precipitate the stroma, separating from the liquid the cation exchanger and precipitated stroma, and adjusting the pH to about 7.2 to 7.6.

Process for obtaining hepatitis-safe, sterile hemoglobin solutions free

Hemoglobin molecules are coupled so as to increase their intravascular residence times without significantly diminishing their oxygen transport ability. This is achieved by coupling hemoglobin molecules to one another and/or to serum proteins and gelatin derivatives using dialdehydes such as aliphatic dialdehydes of 3 to 8 carbon atoms, optionally followed by addition of pyridoxal phosphate. The desired material isolated by ammonium sulfate precipitation, the sulfate being added before, simultaneously with, or after the dialdehyde. Solution of the precipitate followed by dialysis and/or ion exchange can also be undertaken.

Preparation of coupled hemoglobin molecules

Hemoglobin preparation suited for intravenous injection with increased oxygen release in relation to erythrocytes or hemoglobin solution, comprising a substantially pyrogen-free condensation product of hemoglobin and pyridoxal phosphate having a retention time in the blood vessel from about 2 to 9 hours. It is produced by washing human erythrocytes at least four times with a weakly alkaline solution, then hemolyzing and treating the material with a cation exchange resin in the H+ form until the pH value has dropped to about 5 to 5.5, separating the material from resin and any precipitated stroma, diluting the material to a hemoglobin concentration of about 5 to 9%, adjusting the pH to about 7 to 9, displacing any oxygen therein, and adding about 0.25 to 1.25 g of pyridoxal-5-phosphate per liter of hemoglobin solution of 5 to 9% concentration. To increase the retention time still further there can be included the further steps of treating the solution with a borohydride and then with a dialdehyde thereby to cross-link the hemoglobin molecules in said erythrocytes, and separating uncross-linked hemoglobin from the residual solution of the desired cross-linked hemoglobin.

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Hanau, Germany

Klaus Bonhard