Processing-dependent hepatic risk and benefit of Polygonum Multiflorum Radix: Integrated chemical profiling, in vivo validation and candidate quality control markers.

To clarify how steaming and sun-drying cycles affect the hepatic risk-benefit profile of Polygoni Multiflori Radix (PMR), the herb was processed with black bean decoction for 1-9 cycles following the traditional "nine steaming and nine sun-drying" protocol. UPLC-MS analysis showed that anthraquinones and trans-THSG progressively decreased with increasing cycles and stabilized after five cycles, whereas cassialactone-8-O-β-D-glucoside and hyperoside emerged as new components during processing. In a low-dose LPS-sensitized mouse model mimicking immune-idiosyncratic liver injury, PMR processed for three or more cycles significantly alleviated liver damage. In an acetaminophen (APAP)-induced acute liver injury model, PMR processed for 5-7 cycles exhibited the strongest hepatoprotective effects. Spectrum-effect correlation analysis identified twelve constituents (e.g., emodin-8-O-β-D-glucoside) associated with liver injury, involving targets related to drug metabolism (CYP3A4), membrane transporters (ABCB1, SLC2A1), and oxidative stress (SOD1). Five components (e.g., cassialactone-8-O-β-D-glucoside) were linked to hepatoprotection, potentially through modulation of synthetic function (ALB) and inflammatory response (TNF, EGFR, HSP90AA1). The Cellular Thermal Shift Assay (CETSA) was employed to validate the molecular docking results, demonstrating that emodin-8-O-β-D-glucopyranoside and physcion-8-O-β-D-glucoside significantly enhanced the thermal stability of TP53. In parallel, emodin and hyperoside were confirmed to increase the thermal stability of albumin (ALB). Overall, processing PMR with black bean decoction for 5-7 cycles is optimal for reducing hepatic risk while preserving or enhancing hepatoprotective activity. These findings support a risk-benefit-oriented processing standard and propose candidate quality markers for standardized manufacturing, including trans-THSG, emodin, aloe-emodin, cassialactone-8-O-β-D-glucoside, and hyperoside.