Abstract: AIM To study the antibacterial activity against erythromycin-resistant organisms of 3-hydroxy-6-O-methylerythromycin-9-O-substituted oxime derivatives, a new route of synthesis with 6 steps was designed. METHODS The starting material, erythromycin A (1), was reacted with NH₂OH·HCI to give 2, which reacted with BzBr to give 3. Selective methylation of C-6 hydroxy group using iodomethane afforded 4, which was hydrolyzed with loss of the 3-cladinosyl to give 5. Compound 5 was reduced by H₂ to provide 6, which was treated with substituted benzyl chlorides to provide 7 and 8. RESULTS Four unreported compounds (5-8) were synthesized. The antibacterial activity of the new compounds were tested in vitro against both erythromycin-susceptible and erythromycin-resistant organisms. The compounds 5 (MIC=1 μg·mL^-1) and 6 (MIC=1 μg·mL^-1) showed significant activity against Staphylococcus epidermidis 26069 compared with erythromycin (MIC=4 μg·mL^-1). Compounds 5 (MIC=16, 4 μg·mL^-1), 7 (MIC=32, 64 μg·mL^-1) and 8 (MIC=64, 32 μg·mL^-1) showed better activity against Streptococcus pneumoniae 64 and Staphylococcus aureus 9525 than erythromycin (MIC>128, 128 μg·mL^-1). CONCLUSION 3-hydroxy-6-O-methylerythromycin-9-O-substituted oxime derivatives have stronger antibacterial activity against some erythromycin resistant organisms than erythromycin A.