A new erythromycin, designated erythromycin F, was isolated from the mother liquors of an early commercial strain of Streptomyces erythreus. Erythromycin F was isolated by HPLC, and its high resolution mass spectrum gave a weak molecular ion at m/z 749.4570, indicating a molecular formula of C₃₇H₆₇NO₁₄. Structural analysis using PMR and CMR spectra revealed that erythromycin F has a C-2 methyl group oxidized to a CH₂O group and a C-16 methylene group. The antibacterial activity of erythromycin F was about equal to that of erythromycin E or 2-8 fold less active than erythromycin A. Although not established experimentally, erythromycin F may be a biosynthetic precursor of erythromycin E.