Reproductive toxicity of Fructus Psoraleae in zebrafish: material basis and implications for clinical safety dosing

Background: Fructus psoraleae (FP), a classical TCM with a medicinal history spanning several millennia, is esteemed for its therapeutic properties in regulating Qi, alleviating asthma, tonifying kidneys and reinforcing Yang. Previous studies have revealed that prolonged use of FP may induce reproductive toxicity, particularly in females. However, its safety evaluation has not yet been systematically conducted.

Purpose: To develop an integrated framework for elucidating the key toxic compounds, underlying toxicological mechanisms, and safety clinical dosage of FP.

Methods: Zebrafish were exposed to varying concentrations of FP (0.025, 0.05, 0.1 mg/mL) to evaluate its reproductive toxicity. The in-vitro and in-vivo chemical compounds of FP were comprehensively identified and quantified using liquid chromatography-mass spectrometry. Potential toxic compounds were determined through network pharmacology, literature mining, and cytotoxicity evaluation. The perturbations induced by FP and the potential toxic compounds on gene expression in zebrafish oocytes were elucidated through transcriptomic analysis. The metabolic profile of key toxic compounds in human females was visualized using GastroPlus™.

Results: In zebrafish, 21-day FP extract exposure induced vitellogenin reduction, hypothalamic-pituitary-gonadal axis gene dysregulation, and marked oocyte atresia. Subsequently, nine FP-derived potential toxic compounds were identified in the ovary, with five exhibiting BMDL (benchmark dose limit) values near or below their measured concentrations, implicating them as principal toxic contributors. Adverse Outcome Pathway (AOP) networks and non-negative matrix factorization (NMF) analysis revealed isopsoralen, isopsoralenoside, and bakuchiol as dominant effectors. First-in-human physiologically based pharmacokinetic (PBPK) modeling determined maximum permissible daily intakes for isopsoralen, isopsoralenoside, and bakuchiol over 21 days as 0.5 μg/kg, 0.021 μg/kg, and 2.5 μg/kg, respectively.

Conclusion: FP exhibited pronounced reproductive toxicity in zebrafish, with its underlying material basis, mechanistic pathways, and oral dosage preliminarily elucidated. Furthermore, a systematically toxicological assessment strategy was developed, providing a precise approach for identifying the potential toxicity or efficacy of TCM and its key material basis.