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Two aminopeptidases, designated as aminopeptidase A and B, were purified 1720- and 1950-fold, respectively, from the culture filtrate of Brevibacterium linens F by ammonium sulphate fractionation, a series of column chromatography steps on DEAE-Sephadex, DEAE-Trisacryl M and Mono-Q, and gel filtration on Superose-6. Aminopeptidase A accounted for 85% of the aminopeptidase activity remaining by the end of purification. The purified enzymes were homogeneous as judged by disc gel electrophoresis. Aminopeptidases A and B showed the same pH optimum of 9·3 and apparent temperature optimum of 40 °C, although the former showed slightly higher pH stability than the latter. NaCl or KC1 concentrations to 2 m did not affect the activities or stabilities of aminopeptidases A and B. Both enzymes were completely inhibited by incubation with EDTA, indicating that they are metalloenzymes, and were reactivated by incubation with Ca2+, Co2+, Mg2+, Zn2+ or Mn2+. The M r values of aminopeptidases A and B were estimated as 150000 and 110000, respectively, by gel filtration on Superose-6, and as 36000 and 26000, respectively, by SDS-PAGE. It was thus assumed that each of the native enzymes exists as a tetramer. The K m values of aminopeptidase A and B for l-leucine-p-nitroanilide were calculated to be 16·1 and 15·2 mm, respectively. The substrate specificities of aminopeptidases A and B were similar to each other.