Preparation of Bleomycinic Acid from Bleomycin A2
Twenty-two grams of bleomycin A2 (copper-containing hydrochloride powder) was heated at 100°C under diminished pressure (15 mm Hg) for 16 hr and then dissolved in 200 ml of 0.05 M ammonium chloride. The resulting solution was applied to an 8OQ-ml CM-Sephadex C-25 column pretreated with the same solvent. Then 690 ml of 0.1 M ammonium chloride was passed through the column, and the blue eluate was collected. The eluate was passed through a 500-ml column of activated charcoal, and the column was washed with water to remove inorganic salts. A 1:1 0.02 M HCI-acetone mixture was then passed through the column, and the blue eluate containing bleomycin demethyl-A2 (copper-containing hydrochloride) was collected. A basic anion-exchange resin, Dowex 44 (OH- form) was added to the collected fraction and the pH was adjusted to 6.0. After removing the solvent by distillation and drying, 8.91 g of a blue powder of bleomycin demethyl-A2 (copper-containing hydrochloride), mp 201-212°C (decomposion upon melting), was obtained.