High amounts of β-glucosidase (MW 74, 000) and β-galactosidase (MW 122, 000) were isolated from the periplasmic space of Rhizobium trifolii 4S (infectious strain), compared with the cell homogenate. The characterization of β-glucosidase and β-galactosidase was determined. Both enzymes were inhibited by the addition of Cu²⁺, Fe²⁺, Zn²⁺, Hg²⁺ and p-chloromercuribenzoic acid. The β-glucosidase exhibited a strong hydrolytic activity on cellobiose, sophorose and laminaribiose to glucose, and the β-galactosidase degraded lactose and O-β-D-galactosyl-1, 3-D-arabinoside to their respective components. Both enzymes hydrolyzed polysaccharide only slightly.