Two β-glucans were prepared from the aqueous extract of the endosperm of naked barley. The extraction was carried out under conditions which obviated enzyme action and eliminated the contamination of starch fraction. The precipitate obtained with 20% concentration of ammonium sulfate of the extract was adsorbed on DEAE-cellulose (borate form) column and fractionated into two fractions by the elutions with H2O and 0.5 N NaOH, respectively. From 5 kg. of the endosperm the yield of the former was 13.3 g. (0.27% of the endosperm), the latter was 2.0 g. (0.04%). Both showed only one peak ultracentrifugally, were slightly levorotatory, not degraded with α-amylase, and consisted of only d-glucose. These data suggest that they are both β-glucans. β-Glucan consumed in periodate oxidation 0.74 mole periodate per 1 mole of anhydro- glucose. Hydrolysate of the methylated β-glucan gave 2, 3,6-tri-O-methyl-d-glucose and 2, 4, 6-tri-O-methyl-d-glucose, and methanolysate of the methylated β-glucan gave methyl 2, 3, 6-tri-O-methyl-β-d-glucopyranoside and methyl 2, 4, 6-tri-O-methyl-β-d-glucopyranoside, although α-anomers were not separated. From sedimentation and diffusion coefficients the molecular weight of the β-glucan was calculated as 220,000. These results indicated that the β-glucan was consisted of β-1, 4 and β-1, 3 linked glucopyranose residues with proportions of 2.5 : 1 to 3:1 (β-1, 4 linkage to be predominant) and that the β-glucan had a linear structure.