Assessment of DNA-Damaging Potential of Acorus calamus Rhizome Oil

Author : Karishma Desai

Abstract : Background: Acorus calamus rhizome oil contains phenylpropanoid isomers (α-, β-, γ-asarone) that have been associated with genotoxic and carcinogenic effects in some experimental systems and have prompted regulatory restrictions. We performed a combined chemical and in vitro genotoxicity assessment of a hydro-distilled A. calamus rhizome oil (Kashmir origin) to inform a weight-of-evidence safety evaluation. Methods: The test oil was characterized by HPLC and LC–MS/MS for α-, β- and γ-asarone content. An in vitro genotoxicity battery was applied following OECD guidance: the bacterial reverse mutation (Ames) test (TA98, TA100, TA102, TA1535, TA1537; ±S9), the cytokinesis-block in vitro micronucleus test (IVMNT) in human peripheral blood lymphocytes (short ±S9 and extended -S9) and, the L5178Y TK+/- mouse lymphoma assay (MLA; short- and long-term exposures ±S9). Cytotoxicity, precipitation, pH and osmolality were recorded and acceptance criteria/historical control ranges were met. Results: Chemical analysis showed the sample to be β-asarone dominant (40.75%) with lower α-asarone (4.16%); γ-asarone was below the detection limit. In the Ames assay, no biologically or statistically reproducible increase in revertant colonies was observed in any strain up to the highest non-cytotoxic concentrations (no mutagenicity; cytotoxicity apparent ≥1,500 µg/plate). The IVMNT revealed no statistically significant or biologically relevant increases in micronucleated binucleated cells at concentrations evaluated (up to 175 µg/mL). In the MLA, mutant frequencies at all evaluable doses remained at or below concurrent control means and did not exceed the OECD Global Evaluation Factor (GEF); RTG and colony-size distributions were consistent with valid assay performance. Conclusions: Under the applied test conditions, this A. calamus rhizome oil sample showed no evidence of direct mutagenic, clastogenic or aneugenic activity in a validated, complementary in vitro test battery. However, because (i) asarone isomers can be bioactivated via hepatic pathways not fully recapitulated in vitro, (ii) the study represents a single batch/chemotype with high β-asarone content, and (iii) in vitro cytotoxicity limited the top dose in some assays, we recommend targeted follow-up: chemotype variability screening, human-relevant metabolic studies, and in vivo genotoxicity with TK measurements to define systemic exposures and refine risk assessment.

Keywords : Acorus calamus, genotoxicity, β-asarone, micronucleus test, Ames test, herbal safety, in vitro toxicology.

Conference Name : World Congress on Medicinal Plants & Natural Products Research (WCMPNPR - 25)

Conference Place : Vadodara, India

Conference Date : 20th Dec 2025