The effect of different concentrations of NaCl (6, 8 and 10 dS/m) on growth and alkaloids production in 21 days-old callus of Egyptian henbane (Hyoscyamu muticus L.) was studied. Results showed that 10 dS/m of NaCl increased fresh and dry weight by 4.1 and 2.8 fold more than control, respectively. Two fold Total alkaloids increased by 2 fold at 10 dS/m compared to control or wild leaves. In contrary, wild leaves had the highest amount of hyosyamine (2.8 mg/g DW) followed by 6 dS/m stressed-callus (1.14 mg/g DW) as detected by HPTLC. Peroxidase and superoxide dismutase activity were increased by 2.6 and 2.3 fold, respectively in highly salt stressed callus than control while catalase activity (CAT) was decreased. Anthocyanin concentration was increased 3 fold in 10 dS/m of NaCl stressed callus compared to control. Glutathione content was increased by 33.8% under low level of NaCl more than control. Reduction of photosynthetic pigments under all NaCl levels led to decrease the total carbohydrate and protein content. Protein band with molecular weight 109 KDa was a unique band in 10dS/m of NaCl treated callus with high expression of protein bands with MW 44, 38 and 32. Histologically, callus cells under high level of NaCl had amoeboid shape and start in senescence with formation of lysiogenous intercellular spaces.