Isolation, identification, and characterization of α- asarone, from hydromethanolic leaf extract of Acorus calamus L. and its apoptosis-inducing mechanism in A549 cells

Abstract

Due to the presence of several active secondary metabolites, the traditional Indian and Chinese medicinal herb Acorus calamus L. has been utilized for both medical and culinary purposes since ancient times. A recent report has underscored the promising cytotoxic effect of A. calamus leaves extract against non-small cell lung cancer A549 cells. Thus, we want to separate the bioactive substance from the hydromethanolic extract of A. calamus leaves in the current investigation. Thin-layer chromatography was used to separate the compounds and different spectroscopic methods (UV, FTIR, NMR, and LCMS/MS) were used for the structure prediction. α-asarone was found to be the main bioactive compound present and it was isolated from A. calamus leaves extract. It exerted a good cytotoxic effect with an IC₅₀ value of 21.43 ± 1.27 μM against A549 cells and IC₅₀ value of 324.12 ± 1.32 μM against WI-38 cells. The induction of apoptosis in A549 cells by α-asarone was reaffirmed by the diverse differential staining methods including DAPI, Acridine Orange/Ethidium Bromide, and Giemsa staining. Additionally, α-asarone induced mitochondrial membrane potential (ΔΨ_m) dissipation with a concomitant increase in the production of ROS. Furthermore, it also increased expressions of caspase-3, caspase-9, caspase-8, DR4, and DR5 genes in A549 cells. In conclusion, α-asarone-induced apoptotic cell death in non-small lung cancer cells (A549) as a result of loss of mitochondrial function, increased ROS production, subsequent activation of an internal and extrinsic caspase pathway, and altered expression of genes controlling apoptosis. As a whole, α-asarone is a plausible therapeutic agent for managing lung cancer.

HIGHLIGHTS

• Isolation of bioactive compound from hydromethanolic leaves extract of Acorus calamus L. by thin layer chromatography.

• Structural elucidation of the bioactive compound was carried out using different methods like UV analysis, FTIR, NMR, and LC-MS/MS analysis.

• A plausible mode of action revealed that α-asarone can induce apoptosis in lung cancer cells (A549).

Communicated by Ramaswamy H. Sarma

Keywords:

• α- asarone
• apoptosis
• A549
• cytotoxicity
• Acorus calamus L

Acknowledgment

The Authors are thankful to the P. G. Department of Biosciences, P. G. Department of Applied and Interdisciplinary Sciences (IICISST), and DST-PURSE Programme, Sardar Patel University for the infrastructure and Instruments facilities and also thankful to SICART, CVM University for the NMR and FTIR analysis.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

The author(s) reported there is no funding associated with the work featured in this article.