HPLC–FLD guided multimodal chromatographic isolation of pancreatic lipase modulators from Saxifraga tangutica

HPLC-FLD enabled recognition and tracking of PL-affinity peaks during separation. • PL-affinity peaks remained traceable across reversed-phase and HILIC separations. • Three compounds from S. tangutica , including 1 activator and 2 inhibitors. Background Translating biological recognition into chromatographic information that can continuously guide targeted isolation remains challenging in natural products. Methods HPLC coupled with fluorescence detection (HPLC–FLD) was combined with multimodal preparative chromatography to recognize, track, and isolate candidate pancreatic lipase (PL)-affinity ligands from Saxifraga tangutica . Their effects on PL were evaluated by activity and kinetic assays, with DFT calculations and molecular docking as complementary analyses. Results Three flavonoid glycosides with chromatographic purities above 95% were isolated: Q3XG (160.8 mg), Q3AG (167.9 mg), and Q3GG (623.1 mg). Q3XG activated PL with an EC₅₀ of 4.78 ± 0.95 μg/mL, whereas Q3AG and Q3GG inhibited PL with IC₅₀ values of 38.33 ± 1.97 and 43.86 ± 2.87 μg/mL, respectively. Kinetic analysis showed that Q3XG decreased the apparent K m and slightly increased the apparent V max, whereas Q3AG and Q3GG showed the opposite changes, consistent with mixed-type inhibition. DFT and docking analyses indicated differences in electronic properties and predicted PL-ligand interaction patterns. Conclusion Integration of HPLC–FLD with multimodal chromatography preserved the link between affinity signals and target peaks throughout chromatographic tracking, selectivity optimization, and preparative isolation, providing a practical route to functionally validated ligands from complex natural products. Download: Download high-res image (162KB) Download: Download full-size image