Influence of nano‐zinc oxide on tropane alkaloid production, <em>h6h</em> gene transcription and antioxidant enzyme activity in <em>Hyoscyamus reticulatus</em> L. hairy roots

Kamal Asl

Bahman Hosseini

Ali Sharafi

Javier Palazon

^{Department of Horticulture, Faculty of Agriculture, Urmia University, Iran,}

^{Zanjan Applied Pharmacology Research Center, Zanjan University of Medical Sciences, Zanjan, Iran,}

^{Department of Plant Physiology, Faculty of Pharmacy, University of Barcelona, Barcelona, Spain,}

Bahman Hosseini, Email: ri.ca.aimru@iniessoh.b.

^{Corresponding author.}

^{Correspondence}: Dr. Bahman Hosseini (ri.ca.aimru@iniessoh.b), Department of Horticulture, Faculty of Agriculture, Urmia University, 165, Urmia, Iran

Received 2018 May 15; Revised 2018 Oct 8; Accepted 2018 Oct 24.

Abstract

The use of nanotechnology and biotechnology to improve the production of plant bioactive compounds is growing. Hyoscyamus reticulatus L. is a major source of tropane alkaloids with a wide therapeutic use, including treatment of Parkinson's disease and to calm schizoid patients. In the present study, hairy roots were obtained from two‐week‐old cotyledon explants of H. reticulatus L. using the A7 strain of Agrobacterium rhizogenes. The effects of different concentrations of the signaling molecule nano‐zinc oxide (ZnO) (0, 50, 100 and 200 mg/L), with three exposure times (24, 48 and 72 h), on the growth rate, antioxidant enzyme activity, total phenol contents (TPC), tropane alkaloid contents and hyoscyamine‐6‐beta‐hydroxylase (h6h) gene expression levels were investigated. Growth curve analysis revealed a decrease in fresh and dry weight of ZnO‐treated hairy roots compared to the control. ANOVA results showed that the antioxidant activity of the enzymes catalase, guaiacol peroxidase and ascorbate peroxidase was significantly higher in the ZnO‐treated hairy roots than in the control, as was the TPC. The highest levels of hyoscyamine (37%) and scopolamine (37.63%) were obtained in hairy roots treated with 100 mg/L of ZnO after 48 and 72 h, respectively. Semi‐quantitative RT‐PCR analysis revealed the highest h6h gene expression was in hairy roots treated with 100 mg/L of ZnO after 24 h. It can be concluded that ZnO is as an effective elicitor of tropane alkaloids such as hyoscyamine and scopolamine due to its enhancing effect on expression levels of the biosynthetic h6h gene.

Keywords: Agrobacterium rhizogenes, Antioxidant enzymes, Hyoscyamine, Scopolamine, Nano‐zinc oxide

Abbreviations

APX: ascorbate peroxidase
CAT: catalase
GPX: guaiacol peroxidase
h6h: hyoscyamine‐6‐beta‐hydroxylase
PCR: polymerase chain reaction
TPC: total phenol content
ZnO: nano‐zinc oxide

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