Background: Different species of Hyoscymaus are rich sources of tropane alkaloids, especially hyoscyamine and scopolamine. These compounds medicinally are important tropane alkaloids that exclusively are produced by plants.

Objective: Evaluation of hyoscyamine and scopolamine tropane alkalolids content in shoots and roots of Hyoscyamus species including H. kotschyanus, H. squarrosus, H. pusillus, H. kurdicus, H. scoparia, H. othocarpus.

Methods: This research was done as a factorial experiment in a completely randomized design with three replications. Seeds of Hyoscyamus species were grown in growth room under 25 C° temperature and 16/8 h light/dark period conditions. Samples of dried shoot and underground organs of 6 to 7 month plants were extracted and then content of hyoscyamine and scopolamine was evaluated by HPLC.

Results: Hyoscyamine and scopolamine content in H. orthocarpus were more than the other species and total of these two compounds in H. orthocarpus (15.81 mg/gr dry weight) was approximately 2 times more than H. kotschyanus (7.45 mg/gr dry weight) and manifold of the other studied species. Hyoscyamine was the main compound in H. orthocarpus. The least amount of hyoscyamine and scopolamine was in H. pusillus.

Conclusion: H. orthocarpus and H. kotschyanus because of higher content of hyoscymine and scopolamine with more studies can be used as rich sources of these tropane alkaloids in pharmacy industries.