Evaluating the toxicity of the roots of Asarum heterotropoides var. mandshuricum extracted using the decoction method: Genotoxicity, single-dose toxicity, and 13-week repeated-dose toxicity studies

The roots of Asarum heterotropoides F. Maekawa var. mandshuricum F. Maekawa (AR) is a traditional herbal medicine used across Asia, including Korea, China, and Japan. AR exhibits a range of biological activities, such as anti-inflammatory, anti-cancer, cold treatment, and anti-nociceptive effects. Various extraction methods, including decoction, which utilizes traditional knowledge and techniques. The AR decoction extract expected to contain fewer toxicants and have reduced toxicity due to the use of hot water in the extraction process. However, scientific evidence on the toxicity of AR decoction extracts is lacking, necessitating further studies for safe usage. This study aimed to evaluate the genotoxicity and toxicity of single and repeated administration of AR decoction extracts. The genotoxicity was assessed using a bacterial reverse mutation (Ames test), an in vitro mammalian chromosome aberration test (CA test), and an in vivo micronucleus test (MN test) in Sprague-Dawley (SD) rats. The general toxicity was evaluated through single-dose and 13-week repeated-dose toxicity studies. In the single-dose toxicity study, 40 SD rats were orally administered AR decoction extract at doses of 1,000, 2,000, and 5000 mg/kg. In the 13-week repeated-dose toxicity study, 140 SD rats received daily oral doses of 0, 250, 500, 1,000, 2,000, and 5000 mg/kg of AR decoction extract. The genotoxicity tests revealed that AR decoction extract was not genotoxic. The single-dose toxicity study showed no changes in body weight, clinical pathology, or macroscopic findings, with the approximate lethal dose (ALD) exceeding 5000 mg/kg. The 13-week repeated-dose toxicity study demonstrated no treatment-related changes in body weight, general symptoms, hematology, clinical chemistry, or urinalysis. Histopathological findings revealed hyperplasia of squamous cells in the forestomach after AR decoction extract administration, a treatment-related effect that resolved during the recovery period. The no observed adverse effect level (NOAEL) for both male and female rats was estimated to be 2000 mg/kg. This study establishes the non-toxic dose of AR decoction extract, providing a foundation for further non-clinical and clinical evaluations AR safety.