A new type of bacterial enzyme hydrolyzed alternan (Leuconostoc mesenteroides NRRL B-1355 fraction S dextran, an alternating α-1,3–α-1,6-d-glucan) to give rise to a series of oligosaccharides. The oligosaccharide formed in the greatest proportion was a cyclic tetrasaccharide of d-glucosyl residues linked in an alternating α-1,3–α-1,6 fashion. Other saccharide products included isomaltose and α-d-glucopyranosyl-1,3–α-d-glucopyranosyl-1,6-d-glucose. Oligosaccharides of higher degrees of polymerization were also formed, and included α-d-glucosylated derivatives of the cyclic tetrasaccharide. This is the first report of a naturally produced cyclic tetrasaccharide.