Preparation of oxytetracycline
A mutant strain, CBS M-108-OX, of Streptomyces alboflavus, which gave increased yields of oxytetracycline (I), was obtained by uv irradiation. It differed in colonial morphology and in pigmentation from the parent strain and from S. rimosus. When grown in submerged culture on a production medium contg. starch 6 kg., corn-steep liquor 2.7 kg., CaCO3 0.72 kg., (NH4)2SO4 0.8 kg., NH4Cl 0.2 kg., CoCl2 0.18 g., CuSO4.5H2O 0.6 g., ZnSO4.7H2O 6.0 g., FeSO4.7H2O 0.18 g., sunflower oil 0.2 kg., and H2O 120 1., at 26.degree., starting pH 7.2, for 150 hrs., the yield of I was 6.6 g./l. Addn. of 3 aliquots of N,N'-dibenzylethylenediamine (II) increased the yield to 8.1 g./l. Recovery was by acidification of the broth to pH 1.5 with H2SO4, filtration, washing of the mycelium with H2O, addn. of 2 g./l. of EDTA, raising the pH to 6 with NH4OH, addn. of 150 g. (in 240 l.) of II, and raising the pH to 9.5. After 3 hrs. stirring, the ppt. was filtered, washed, and dried. It was resuspended in twice its wt. of MeOH, stirred 30 min., adjusted to pH 2(HCl gas), then to pH 1.5 with 36% HCl, and stirred. The ppt. was filtered, washed with MeOH, and dried. The yield of I, as hydrochloride, was 81%. It could be further purified and converted into the free base.