Transgalactosylation. Galacto-oligosaccharides are the products of transgalactosylation reactions catalyzed by â-galactosidases when using lactose or other structurally related galactosides as the substrate. â-Galactosidases are generally classified as hydrolases. In fact, hydrolysis of the glycosidic bond is a special case of transgalactosylation in which the galactosyl acceptor is water (6). Scheme 1 illustrates the possible lactose conversion reactions catalyzed by â-galactosidases. Transgalactosylation is thought to involve intermolecular as well as intramolecular reactions. Intramolecular or direct galactosyl transfer to D-glucose yields regioisomers of lactose. The glycosidic bond of lactose [â-D-Galp-(1f4)-D-Glc] is cleaved and immediately formed again at a different position of the glucose molecule before it diffuses out of the active site. This is how allolactose [â-D-Galp-(1f6)-D-Glc], the presumed natural inducer of â-galactosidases in certain microorganisms, can be formed even in the absence of significant amounts of free D-glucose (5, 7, 8). By intermolecular transgalactosylation, di-, tri-, and tetrasaccharides and eventually higher oligosaccharides are produced. Any sugar molecule in the reaction mixture can be the nucleophile to accept the galactosyl moiety from the galactosyl-enzyme complex. The GOS produced can be regarded as kinetic intermediates as they are also substrates