Systematic Multi-Model Toxicological Evaluation and Phytochemical Signatures of Carthamus tinctorius L., Scabiosa comosa, and Their Mixture: Acute and 28-Day Oral Toxicity Studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Carthamus tinctorius L. and Scabiosa comosa Fisch. ex Roem. & Schult. are traditional medicinal plants used within the Traditional Chinese Medicine (TCM) and Traditional Mongolian Medicine (TMM) systems in China. Historically, the dried florets of C. tinctorius are administered as an aqueous decoction, typically at 3-10 g/day, and are prescribed to invigorate blood circulation and resolve blood stasis, whereas the capitula of S. comosa are prepared as a decoction or powder, typically at 6-9 g/day, and are utilized to clear heat and detoxify, with traditional indications for liver and pulmonary febrile diseases. However, comprehensive evaluations of their antioxidant properties and toxicological characteristics, particularly regarding their individual and combined applications, remain limited.

AIM OF THE STUDY: This study aimed to systematically characterize the phytochemical composition, antioxidant capacity, and safety profiles of C. tinctorius, S. comosa, and their combination using multiple extraction strategies and complementary biological models.

MATERIALS AND METHODS: Water extracts, ethanol extracts, and sequential water-ethanol extracts were prepared and analyzed by LC-MS/MS and total flavonoid content determination. Their antioxidant capacities were evaluated using complementary in vitro assays. The toxicological profiles were investigated using BRL-3A rat hepatocytes, zebrafish embryo toxicity assays, acute oral toxicity studies in mice, and a 28-day repeated-dose oral toxicity study in Wistar rats.

RESULTS: LC-MS/MS profiling revealed diverse chemical constituents, including phenolic acids, flavonoids, coumarins, and iridoids. Notably, C. tinctorius exhibited the highest total flavonoid content (TFC), and sequential extraction significantly increased TFC compared to single-solvent extractions. In the antioxidant evaluations, S. comosa demonstrated the most potent antioxidant capacity with the lowest EC50 values. Toxicity varied by concentration and extract type, with water extracts displaying the lowest toxicity. While high concentrations induced developmental abnormalities (e.g., mortality, oedema) in zebrafish, rodent models demonstrated high safety. Acute (5000 mg/kg) and 28-day repeated administrations caused no mortality or severe histopathological lesions, yielding only minor body weight and biochemical fluctuations.

CONCLUSIONS: The present study provides a systematic chemical and toxicological evaluation of C. tinctorius, S. comosa, and their combination. These extracts exhibited antioxidant potential and favorable safety profiles in multiple experimental models, providing important evidence for their further development as medicinal or functional food-related products.