Toxicological safety profiling of Berberis vulgaris hydroethanolic extract using rodent, zebrafish, and in silico models

The hydroethanolic extract of Berberis vulgaris root bark (BV) is extensively used in traditional medicine, particularly Homoeopathy, for treating renal and hepatic disorders, yet a systematic safety evaluation remains limited. This study aimed to establish a comprehensive preclinical safety profile of BV using in vivo rodent and zebrafish models, alongside in silico toxicity predictions. Phytochemical profiling was conducted using Liquid Chromatography–Mass Spectrometry (LC–MS). Acute and 28-day repeated-dose oral toxicity studies were performed in Wistar rats following OECD guidelines 423 and 407, respectively. Developmental toxicity was assessed in zebrafish embryos (OECD 236), and in silico toxicity predictions for identified phytoconstituents were generated using ProTox 3.0. LC-MS analysis identified 22 bioactive chemical components. In the acute oral toxicity study, BV administered at 2000 µL/kg caused no mortality or toxicity, indicating an LD₅₀ > 2000 µL/kg. The 28-day repeated-dose study showed no significant alterations in haematological, biochemical, or histological parameters at doses up to 1000 µL/kg/day, establishing a No Observed Adverse Effect Level (NOAEL) of ≥ 1000 µL/kg/day. While lower concentrations were safe in zebrafish, high concentrations (4 µL/2 mL) induced developmental abnormalities such as scoliosis and pericardial edema. Computational analysis predicted low-to-moderate toxicity for the majority of phytoconstituents. BV exhibits a wide safety margin in rodent models and is non-toxic at therapeutically relevant doses. However, observed developmental effects in zebrafish suggest caution at high concentrations, supporting the need for adherence to recommended dosages in traditional therapeutic contexts. • Berberis vulgaris hydralcoholic extract showed no acute or sub-chronic toxicity in rats. • No treatment-related changes in haematology, biochemistry, or histology. • Zebrafish embryos showed minimal developmental toxicity at high dose. • LC–MS identified 22 major alkaloid and flavonoid phytoconstituents. • In vivo and in silico findings confirm BV’s wide safety margin.