Demethylthio-, de-S-methyl-, and ethylthio-derivatives of the alpha-mannosidase inhibitor, mannostatin A, have been synthesized and evaluated for their inhibition of Jack bean alpha-mannosidase in order to elucidate the roles of the methylthio group. All derivatives had lowered inhibitory potentials. However, a mannostatin A analogue with a methoxyl instead of the methylthio, exhibited about 2-fold enhancement of the activity, indicating an importance for the methyl group rather than the sulfur atom.