To clarify a low level of cellulose biosynthesis of the in vitro cultured cells, the effects of several biochemical factors such as carbon sources (sucrose, maltose, and UDPG), antioxidants (ascorbic acid and glutathione) and physical factors such as artificial pressure, high gravity, on the cellulose production in barley callus and suspension cultures were investigated. In the suspension culture of two barley strains, the supplement of different concentrations (0, 1.5, 3.0, and 4.5%) of sucrose or maltose into the medium for 30 days did not promote the cellulose production and 4.5% of sugar supplement was rather inhibitory in one strain. However, in the presence of sucrose at 3%, UDPG (3 or 10 mM) supplement, as a precursor for cellulose, promoted 1.2∼1.3 fold of the production in two strains. A low concentration (3 mM) of ascorbic acid and glutathione promoted 1.5 and 1.2 fold of the production in two strains, respectively. These results suggest that low cellulose biosynthesis of the in vitro cultured cells is due to a decreased level of the UDPG in the cytosol, and that the oxidative condition of external medium impedes cellulose synthesis in some manners. Artificial pressure applied to the callus promoted 1.4 fold of the cellulose production. High gravity (5,000 or 10,000 g) applied to the suspension-cultured cells by centrifugation did not cause a substantial change.