12β-Hydroxylation of Digitoxin by Suspension-culturedDigitalis lanataCells. Production of Deacetyllanatoside C Using a Two-Stage Culture Method1

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Abstract
Suspension-cultured Digitalis lanata cells, known to form β-methyldigoxin from β-methyldigitoxin without any by-products, were not able to 12β-hydroxylate digitoxin efficiently when cultivated in the cell culture medium devised by Murashige and Skoog. Most of the substrate added was merely glucosylated at its 16′-O-position leading to purpureaglycoside A as the main biotransformation product after 9 days of incubation. An 8% glucose solution (pH 5.5) turned out to be a suitable production medium for an efficient 12β-hydroxylation of digitoxin. A two-stage procedure was developed in which Digitalis cells were grown in a modified Murashige and Skoog medium for 10 days and then transferred into 8% glucose medium. With regard to an effective 12β-hydroxylation of digitoxin, maximum productivity was achieved when the cell line K 3 OHD was used with an initial cell density of about 20%. The substrate was added in one batch (190 mg digitoxin per flask, i.e. 0.5 gl-1) 3 days after transfer of cells to production medium. Under these conditions all of the digitoxin added was biotransformed within 12 days of incubation yielding the main product deacetyllanatoside C (88%) together with purpureaglycoside A (12%) both of which accumulated in the cells.

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