PROOF FOR THE BIOSYNTHETIC CONVERSION OF L-[INDOLE-15N]TRYPTOPHAN TO [10-15N]ANTHRAMYCIN USING (13C, 15N) LABELLING IN CONJUNCTION WITH 13C-NMR AND MASS SPECTRAL ANALYSIS

Using 13C-NMR and mass spectral analysis we have demonstrated that the N-10 nitrogen of anthramycin is biosynthetically derived from the indole-nitrogen of tryptophan. Our experimental approach was to bring a 15N atom, which is derived from L-[indole-15N]tryptophan, and a 13C atom which is derived from DL-[1-13C]tyrosine, into adjacent positions of anthramycin. From resonance intensities and 13C-15N spin-spin coupling in the 13C-NMR spectrum of didehydroanhydroanthramycin, a derivative of anthramycin, we could then determine the 13C enrichment at C-11 and the proportion of 13C bonded to 15N at N-10. These results when combined with mass spectral analysis and isotopic dilution measurements proved that the indole nitrogen of tryptophan was completely retained at N-10 of anthramycin.