A novel bilirubin-lowering mechanism of Ginkgo Biloba terpene lactones discovered in safety assessment.

Against the backdrop of assessing the safety of nanocrystalline solid dispersion of component-based Chinese medicine of Ginkgo biloba leaves (GBCCM NC-SD), this study serendipitously observed a significant dose-dependent reduction in serum bilirubin levels in rats during a 28-day sub-chronic toxicity study (doses: 16.50-148.50 mg/kg). No obvious toxic symptoms, target organ damage, or treatment-related histopathological lesions were found, with key hematological, biochemical, and urinary parameters showing no biologically relevant abnormalities. The no observed adverse effect level (NOAEL) was determined to be >148.50 mg/kg for both genders. Subsequent validation in mice confirmed that terpene lactones (TLs) in GBCCM were the key active substances, reducing direct and indirect bilirubin proportionally. Integrative network pharmacology and proteomics revealed that TLs regulate key bilirubin metabolism-related pathways, including porphyrin-chlorophyll metabolism and metabolic pathways, and modulated key proteins: downregulating HMOX1 (bilirubin production), upregulating Gstm4/Gstm6/Fabp4 (bilirubin transport), and inducing UGT1A1/CYP450 isoforms (bilirubin transformation). This study completes the first comprehensive sub-chronic toxicity assessment of GBCCM NC-SD to support its clinical translation and elucidates a novel bilirubin-lowering mechanism of Ginkgo biloba TLs, providing potential therapeutic insights for hyperbilirubinemic disorders.