Effect of Biotic Elicitors on Tropan Alkaloids Production in Hairy Roots Culture of Datura (Datura innoxia)

Document Type : Full Paper

Authors

• ahmad sharifi ¹
• mahboobeh yazdi ²
• sara khosravinia ²
• asma zahedi ²
• Mahdiyeh Kharrazi ¹
• Azadeh Khadem ¹

¹ Horticulture Biotechnology Research Group, Department of Industrial Biotechnology, Khorasan Razavi branch of Academic Center for Education, Culture and Research, Mashhad, Iran.

² Department of Biotechnology and Plant Breeding, Faculty of Agriculture, Ferdowsi University of Mashhad, Iran.

Abstract

Keywords

• Datura
• Elicitor
• Hairy Root
• Hyoscyamine
• Scopolamine

Main Subjects

• Medicinal plant science

Extended Abstract

Introduction

Hairy root culture, a promising technique in plant biotechnology, has garnered significant interest in recent years for producing herbal medicinal compounds. Datura, known for its therapeutic potential in treating various diseases, particularly cancer and digestive system issues, was the focus of this research. This study aimed to optimize the cultivation conditions of Datura innoxia hairy roots to enhance the production of the alkaloids hyoscyamine and scopolamine.

Material and Methods

To conduct this experiment, Datura innoxia seeds were first surface-sterilized and cultivated in MS medium. Leaf explants from sterile seedlings were then co-cultivated with Agrobacterium rhizogenes strains A4, 15834, and MSU in LB liquid medium. Induced roots, approximately 2 to 3 cm in length, were excised and transferred to B5 liquid medium on a shaker at 90 rpm in the dark. After two months, the rate and stability of root growth were assessed. The bacterial species were confirmed by amplifying the rolC gene, located in the T-DNA region of the hairy root-inducing plasmid, using PCR with specific primers. The effects of different elicitors—methyl jasmonate, salicylic acid, and yeast extract—on the growth and alkaloid production were investigated. Preliminary identification of tropane alkaloids in the samples was done using Dragendorff reagent. The alkaloid content in each sample was carefully measured using spectrophotometry. To enhance accuracy in quantifying alkaloids across different treatments, HPLC analysis was also performed on the samples with the highest alkaloid content.

Result and Discussion

The study on the effect of methyl jasmonate on the growth rate and alkaloid production revealed that methyl jasmonate effectively enhanced alkaloid production, increasing it by 30% compared to the control. Salicylic acid and yeast extract did not significantly affect the growth rate of hairy roots, as there was no substantial difference in the dry weight of hairy roots across treatments. However, alkaloid production varied significantly between treatments. Yeast extract did not change alkaloid content after 48 hours compared to the control. In contrast, salicylic acid treatment resulted in a significant increase in alkaloid content after 48 hours. These findings indicate that elicitors can influence tropane alkaloid production in Datura hairy roots, but their effectiveness depends on optimization alongside other environmental factors. Additionally, HPLC analysis showed a significant increase in hyoscyamine content with methyl jasmonate treatment, whereas the increase in scopolamine was not significant.

Conclusion

The results of this experiment demonstrated that adding an elicitor as a stimulating factor enhances tropane alkaloid production in Datura hairy roots. Specifically, methyl jasmonate increased alkaloid production by 30% compared to the control treatment.