The 2'-,4'-, and 6'-deoxy derivatives of methyl beta-lactoside have been synthesised by deoxygenation at positions 2', 4', and 6', and the 3'-deoxy derivative was obtained by a glycosylation reaction. The 2'-O-methyl, 2'-O-benzyl, 2'-amino-2'-deoxy, and 1'-deuterio derivatives have been synthesized also. Only the 6'-deoxy and 1'-deuterio derivatives were substrates for the beta-D-galactosidase from E. coli, and the 2'-deoxy- and 2'-amino-2'-deoxy derivatives were potent inhibitors for the hydrolysis of methyl beta-lactoside by the enzyme.