Butirosins are broad-spectrum antibiotics with activity against Pseudomonas strains and are less toxic than other aminoglycoside antibiotics. Studies on the inactivating mechanism of aminoglycoside antibiotics prompted us to prepare new butirosin analogs effective against resistant Gram-negative bacteria. We now communicate the preparation of 6'-N-methyl and 6'-C-methyl derivatives of butirosins via the technique of "mutational biosynthesis".