Hydrolysis of α-d-Galactosyl Oligosaccharides in Soymilk by α-d-Galactosidase ofBifidobacterium breve203

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Abstract
Bifidobacterium breve 203 grew well on soymilk, reaching a cell number of 2 × 109 cells/ml on 24 hr cultivation, at which time the medium had solidified and its pH had decreased to 4.0. Stachyose and raffinose in the medium were assimilated in preference to sucrose. A crude extract of B. breve 203 hydrolyzed the oligosaccharides in soymilk almost completely into their constituent monosaccharides. The α-d-galactosidase responsible for the first reaction in the degradation of the α-d-galactosyl oligosaccharides was isolated from a crude extract of the organism (500-fold purification, 2% yield) by means of ammonium sulfate-fractionation and column chromatographies on DEAE-cellulose, DEAE-cellulofine, Sepharose 6B, hydroxylapatite, Sephacryl S-300 and Phenyl-Sepharose 6B. The enzyme was a homo-octamer with a molecular weight of about 330,000, Λ and its isoelectric point was 3.7. It reacted optimally at pH 5.5 and hydrolyzed stachyose (Km 8.6 mm), raffinose (Km 4.4 mm) and melibiose (Km 3.0 mm), with 50 ~ 60% of its reactivity toward p- nitrophenyl α-d-galactoside (Km 0.16mm). The reaction of the purified α-d-galactosidase with soymilk and guar gum was also investigated.