The objectives of this study was to improve poly-gamma-glutamic acid (gamma-PGA) production by Bacillus subtilis D7 isolated from a Korean traditional fermented food and to assess its antioxidant activity for applications in the cosmetics and pharmaceutical industries. Strain D7 produced gamma-PGA in the absence of l-glutamic acid, indicating l-glutamic acid-independent production. However, the addition of l-glutamic acid increased gamma-PGA production. Several tricarboxylic acid cycle intermediates and amino acids could serve as the metabolic precursors for gamma-PGA production, and the addition of pyruvic acid and d-glutamic acid to culture medium improved the yield of gamma-PGA markedly. The maximum yield of gamma-PGA obtained was 24.93 +/- 0.64 g/l in improved medium, which was about 5.4-fold higher than the yield obtained in basal medium. gamma-PGA was found to have 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity (46.8 +/- 1.5 %), hydroxyl radical scavenging activity (52.0 +/- 1.8 %), 2,2'-azinobis-3-ethylbenzothiazoline-6-sulfonate (ABTS) radical scavenging activity (42.1 +/- 1.8 %), nitric oxide scavenging activity (35.1 +/- 1.3 %), reducing power (0.304 +/- 0.008), and metal chelating activity (91.3 +/- 3.5 %). These results indicate that gamma-PGA has a potential use in the food, cosmetics, and biomedical industries for the development of novel products with radical scavenging activity. As far as we are aware, this is the first report to describe the antioxidant activityof gamma-PGA produced by bacteria.