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Irradiation of human insulin in aqueous solution, first step towards radiosterilization.

The degradation of irradiated human insulin in aqueous solutions was investigated in order to protect the protein against ionizing radiation. The influence of the drug concentration, excipients and irradiation temperature were studied. Aqueous solutions at pH 2 were irradiated by gamma rays or by accelerated electrons. Two different high-performance liquid chromatography (HPLC) methods were used: reverse-phase high-performance liquid chromatography (RP-HPLC)/UV and size exclusion liquid chromatography (SEC/UV) to investigate both the fragmentation and the formation of higher molecular weight proteins. In solution without excipients irradiated at ambient temperature at 10 kGy, the loss of human insulin is almost complete. Addition of radio-protecting excipients (free radicals scavengers) and cryo-irradiation allowed to decrease insulin degradation. The best radio-protector used was ascorbic acid in aqueous solution and oxidized glutathione in the frozen solutions. Only the combination of these two approaches (addition of scavenger and freezing) enables the irradiated human insulin in aqueous solution to meet the European Pharmacopoeia requirements for chemical potency (>or=90%).

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