The production of toxic shock syndrome toxin-1 (TSST-1) was studied in batch and continuous culture of Staphylococcus aureus strain 1169 in a carbohydrate-free chemically defined medium (CDM). In continuous culture oxygen- and arginine-limitation were required for steady-state TSST-1 synthesis. Aeration suppressed toxin synthesis. The amount of TSST-1 per mg dry weight (specific toxin) at dilution rates from 0.05 to 0.15 h-1 was inversely proportional to the dilution rate. Protease activity increased with increasing dilution rates. In batch culture, TSST-1 began to accumulate in the medium towards the end of the exponential phase of growth, after a critical cell mass was attained. Maximum specific toxin production was observed in medium with an initial pH between 6.5 and 7.0. Growth and toxin synthesis took place in anaerobic conditions when CDM was supplemented with pyruvate and uracil. The Mg++ concentration had no effect on the specific toxin in anaerobic conditions. In aerobic conditions, specific toxin increased c. 23-fold as the Mg++ concentrations increased to 0.4 mM. Further increases in the Mg++ concentration resulted in a reduction in specific toxin.