Three strains of Lactococcus lactis ssp. lactis, a dairy strain 65.1, a type strain ATCC 19435, and a mutant AS 211, were grown on glucose and on maltose under chemostat conditions. When the culture was shifted from glucose-limiting to non-limiting conditions, the product shifted from mixed acids to lactate. Mixed acids were obtained in all maltose cultures; however, an enhanced lactate formation was observed in 19435 and AS 211. An inorganic-phosphate (P-i)-dependent maltose phosphorylase activity was found to be responsible for the initial conversion of maltose. The activation of maltose phosphorylase by P-i was strain-specific. When growth was on maltose under non-limiting conditions, a correlation was found between high initial maltose phosphorylase and beta-phosghoglucomutase activities and lactate production. No such correlation was observed in maltose-limited cells. In glucose-grown cells under non-limiting conditions, home-fermentative lactate formation coincided with high concentrations of fructose 1,6-bisphosphate (Fru1,6P(2)) and pyruvate (Pyr) and low concentrations of phosphoenolpyruvate (PPyr). Under limiting conditions, mixed acid formation coincided with low concentrations of Fru1,6P(2) and Pyr and high concentrations of PPyr. In maltose-grown cells there was no correlation between intracellular intermediary metabolite concentrations and product formation. Therefore, in addition to intracellular intermediary metabolite concentrations, the product formation on maltose is suggested to be regulated by the transport and initial phosphorylating steps.