Search

The reducing effect of ascorbic acid and of borohydride upon ferrihaemoglobin present in native and chemically modified human and bovine stroma-free hemoglobins was investigated. Ferrihaemoglobin which had been freshly prepared from oxyhaemoglobin by treatment with potassium ferricyanate was fully reduced to ferrohaemoglobin. Full reduction of ferrihaemoglobin, however, could not be achieved with those haemoglobin samples which had a partially deteriorated conformation due to long time storage or chemical modification.

Stroma-free solutions of human haemoglobin modified with pyridoxal-5-phosphate, glutaraldehyde, borohydride and serum albumin were injected into the artery of an isolated rat heart perfused with Krebs-Henseleit solution under hypoxic conditions. About 70% of the oxygen transported by the modified haemoglobin was found to be utilized for a marked increase in the force of heart contraction. The results were in general correlation to the analysis of oxygenation curves of haemoglobin samples under study and confirmed the oxygen offloading ability.

Sucrose is one of the most effective substances which protect haemoglobin from spontaneous methaemoglobin formation during lyophilization and subsequent storage. The dry haemoglobin-sucrose system was treated under different conditions of temperature (up to 85 degrees C), time of storage (up to 6 years) and residual moisture (less than 0.1-7.5% H2O), in order to reveal the main features of the hitherto unclear mechanisms of both spontaneous methaemoglobin formation and of the stabilizing effect of sucrose. In conclusion, a dry sucrose network was recognized as a significant support to the native ferrous structure of oxyhaemoglobin, while the presence of water molecules, of assumed peroxidic radicals and the action of thermal vibrations favour the oxidation and denaturation of haemoglobin.

The stability of experimental haemoglobin solution for infusion (HSI) was investigated during one year storage at temperatures 20 degrees to 25 degrees C, 4 degrees C, -8 degrees to -12 degrees C, and -195.8 degrees C, respectively. Main attention was devoted to the formation of precipitate, oxidation to haemiglobin and general changes in the absorption spectrum between 550 and 650 nm. Changes were almost negligible in HSI samples stored in the frozen state. This procedure now seems the most feasible to keep the HSI injectable for at least one year. Storage at 4 degrees C led to a marked increase of haemiglobin and to changes of the absorption spectrum. Storage at 20 degrees to 25 degrees C caused profound changes of all parameters tested. Addition of EDTA, ascorbic acid or glutathione, or storage in N2 atmosphere under the given conditions failed to increase the stability of HSI. Storage of HSI samples at pH values between 7.0 and 7.8 led to similar results.