Juglans regia L. (Juglandeaceae) is used medicinally for centuries, but it also is used in pharmaceutical and cosmetic industries. The extract of walnut green husks and walnut leaves Juglans regiaL. (Juglandeaceae) were screened in vitro for their antimicrobial activity against Gram-positive, Gram-negative bacteria and two yeasts. The chemical composition of walnut leaves and green husks were analyzed. Disk diffusion method was used for antimicrobial activity. GC/MS analysis was tested for chemical composition of walnut green husks and walnut leaves extract. The aqueous extract of green husks showed antimicrobial activity against five of the seven Gram-positive bacteria and six of the nine Gram-negative bacteria. Contents of the aqueous extract of walnut green husks were ethylene oxide (83.67%), cyclotrisiloxane hexamethyl (5.04%), and walnut leaves were ethylene oxide (14.74%), cyclotrisiloxane hexamethyl (17.89%). The aqueous extract of the walnut leaves was effective against four Gram-positive and one Gram-negative organism. Both walnut green husks and leaves extracts exhibited antifungal activity. The results of the present in vitro work indicate that aqueous extract of walnut leaves could be used as natural antimicrobial agents in the food preservation and human health for pathogenic and/or antibiotic resistant bacteria because of contain minimum percentage of ethylene oxide (14.74%) compared to walnut green husks.