New unsymmetrical Schiff bases (L3a-L3c) were synthesized by using a two-stage method. The starting Schiff bases were prepared by the reaction of 2-aminophenol (or 2amino-4-methylphenol or 2-amino-4-chlorophenol) with 1-nitro-2-naphthaldehyde. In these compounds, the nitro groups were reduced to amino groups using selective reducing agent and then 2-hydroxybenzaldehyde were added. Thus, three new unsymmetric Schiff base ligands were obtained. The antimicrobial activity of the synthesized unsymmetric Schiff bases were evaluated against some pathogenic strains (Shigella dysenteria type 7, Listeria monocytogenes 4b, Escherichia coli, Salmonella typhi H, Staphylococcus epidermis, Brucella abortus, Micrococcus luteus, Bacillus cereus sp., Pseudomonas putida sp.) and yeast (Candida albicans).