Fermentative production of spiramycins by Streptomyces ambofaciens has been performed using fermentation media of different chemical compositions. Medium I was selected from nine media as the best for production of high titres of spiramycins. Biochemical changes which occurred during fermentative production of spiramycins revealed that adjustment of the initial pH value of the medium was very important. The initial pH value of the fermentation medium which allowed the organism to produce a good yield of antibiotic was 6.5. The fermentation period affected the formation of spiramycins, and the maximum incubation period required for the fermentation process was 120 h. The role of inoculum on spiramycin yield showed that it was better to inoculate the fermentation medium with vegetative cells of Streptomyces ambofaciens rather with spores. The carbon source influenced spiramycin biosynthesis; dextrin was the best carbon source and stimulated the organism to form high titres of antibiotics. The best concentrations of dextrin and glucose for increased antibiotic yields were 25 and 15 g l - 1, respectively. Organic sources in the fermentation medium were more efficient than inorganic nitrogen sources for spiramycin formation. Fodder yeast was the best organic nitrogen source in fermentative production of spiramycins. The maximal concentrations or fodder yeast, soybean meal, peptone, Ca(NOa) 2 and NH4NO 3 for increased antibiotic yield were 6.5, 6.0, 4.0, 10.0 and 4.0g l- , respectively.