Batch fermentation of Penicillium chrysogenum, MTCC 5108 was carried out using potato dextrose broth medium prepared in seawater: distilled water (1:1). Biomass as dry weight was determined by gravimetric analysis. Citrinin, the main secondary metabolite, is produced in large quantities during the stationary phase of growth. The yield amounted to approximately 530 mg 1 sup(-1). After optimization of culture conditions, P. chrysogenum was mass cultured and citrinin was isolated and purified from the medium using a combination of chromatographic techniques (Thin layer and column chromatography). Citrinin, thus obtained was characterized on the basis of its spectral data (UV, Proton Nuclear Magnetic Resonance and Electrospray Ionization Mass spectra). The present study consists report on the effect of the concentration of citrinin on the growth of the culture and antibiotic activities assayed by disc diffusion method using clinical pathogens