A series of novel positional oximes (2a-2d), O-methyl oxime ethers (3a-3d), and O-benzyl oxime ethers (4a-4d) were synthesized in high yields starting from their corresponding methyl 3-, 4-, 6-, and 13-keto tetradecanoates. The synthesized compounds were characterized by H-1 NMR, C-13 NMR, FT-IR, mass, and elemental analyses for their structures and (E) /(Z) isomerizations. Their DNA binding abilities were investigated in vitro by agarose gel electrophoresis. The antibacterial and antifungal activities were tested also in vitro against eleven bacterial strains and three fungal strains. The relationship between the structure and the mentioned biological activities was discussed. Compound 2a showed good antibacterial activity against five types of bacteria. Compounds 2b, 2c, 2d, and 4d were effective against several microorganisms. Among these, 2a showed the best DNA binding, antibacterial, and antifungal activities. Therefore, 2a can be a pro-drug as an anticancer, antibacterial, and antifungal agent.