Hairy roots of Hyoscyamus albus were induced from the shoot cultures by transformation with A. rhizogenes strain MAFF 03-01724. The hairy root cultures were tested for growth and alkaloid production (5 tropane alkaloids simultaneously quantified by HPLC) in various media. Half-strength MS and Gamborg BS media containing 3% sucrose gave the highest alkaloid content (ca 2% dry wt), but only poor growth. On the other hand, fast growth was observed in Woody Plant medium containing 8% sucrose, while the production of tropane alkaloids was relatively low. The addition of KNO3 (1-50 mM) to Woody Plant medium (3% sucrose) enhanced the growth, and consequently the alkaloid yield in the hairy roots increased remarkably. The hairy roots cultured in the improved medium (Woody Plant medium containing 3% sucrose and 15 mM KNO3) for 28 days produced over 30 mg tropane alkaloids in one 100 ml flask (ca 0.8 mg 7β-hydroxyhyoscyamine, 1.7 mg littorine, 1.7 mg scopolamine, 7 mg 6β-hydroxyhyoscyamine and 19 mg hyoscyamine in 50 ml liquid medium). © 1991.