Carboxylic acid-functionalized multiwalled carbon nanotubes (COOH-MWCNTs) improved production of atropine in callus of Datura inoxia by influencing metabolism, gene regulation, and DNA cytosine methylation; an in vitro biological assessment.

Atropine is a well-known tropane alkaloid commonly employed in medicine class called anticholinergics. This study intends to address biochemical and molecular responses of Datura inoxia calluses to fortifying culture medium with carboxylic acid-functionalized multi-walled carbon nanotubes (COOH-MWCNTs). The application of MWCNTs influenced callogenesis performance and biomass in a dose-dependent manner. The MWCNT at 5 mgL resulted in the highest biomass of calluses by 57%. While, MWCNTs at high concentrations were accompanied by cytotoxicity. On the other hand, MWCNTs at concentrations above 100 mgL exhibited cytotoxicity, decreased callogenesis performance, and reduced Atropine biosynthesis. The MWCNTs increased the activity of phenylalanine ammonia-lyase (PAL) and catalase enzymes. The concentrations of proline and soluble phenols displayed upward trends in response to using MWCNTs. According to the HPLC assessment, enriching culture medium with MWCNTs at 5 mgL elicited Atropine production in calluses by 64%. The quantitative PCR assessment referred to the upregulation in the transcription of the PAL gene. The expression of ornithine decarboxylase (ODC) and putrescine N-methyltransferase 1 (PMT) genes were also upregulated in calluses cultured in a medium supplemented with MWCNTs. Methylation Sensitive Amplification Polymorphism (MSAP) technique indicated that employing MWCNTs altered the DNA methylation profile, reflecting epigenetic modification. Overall, engineering plant cells with MWCNTs as a nano-elicitor can be suggested for large-scale synthesis of industrially-valuable secondary metabolites.