The effect of ion-supplemented medium on peroxidase excretion from horseradish (Armoracia rusticana) hairy roots was studied. Supplementation of mannitol instead of ions revealed that the excretion was stimulated not by osmotic pressure in the medium but by ionic properties. Extracellular peroxidase activity per dry cell was proportionally correlated with the ionic strength of the cations. CaCl2 or MgCl2 was found to be the most effective agent for excretion among other combinations. CaCl2 supplementation at the beginning of the culture caused higher peroxidase production in the medium without a significant loss of final cell mass compared with CaCl2 addition during the culture. Repeated batch culture with 50 m M CaCl2 supplementation allowed a continuous retention of cell viability over 149 days and produced a great amount of extracellular peroxidase, 12-fold higher than that achieved in a 40-day-old batch culture with 50 m M CaCl2 supplementation.