Adsorptive Purification Behavior of Acylated Anthocyanins from Red Radish and Its Anti-Lipase Activity in vitro

Anthocyanins of red radish (ARR) are natural red pigments characterized by strong processing stability and diverse physiological activities. This study aimed to optimize the purification process of ARR and systematically elucidate its inhibitory mechanism against pancreatic lipase, thereby providing a theoretical foundation for the development of functional foods targeting hyperlipidemia.

The adsorption and desorption properties of six macroporous resins with different polarities on ARR were evaluated to select the optimal resin. The kinetic and thermodynamic characteristics of resin adsorbing ARR, as well as dynamic desorption characteristics at different ethanol concentrations, were investigated. Individual anthocyanins were identified by ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry, and the purification performance of dextran gel chromatography was also assessed. Furthermore, the inhibitory activity and inhibition type of ARR against pancreatic lipase were evaluated in vitro, while the molecular interaction mechanism was explored using Fourier transform infrared spectroscopy and molecular dynamics simulations.

The non-polar macroporous resin X-5 exhibited the highest recovery rate of ARR (81.17%) following sequential adsorption and desorption. The adsorption process followed pseudo-first-order kinetics and the Temkin isotherm model, indicating a spontaneous endothermic reaction with a maximum equilibrium adsorption capacity of 10.55 mg·g^-1 at 35 ℃. Ethanol (40%) was identified as the optimal desorption agent. Purification using X-5 resin alone yielded higher purity than the combined application of X-5 and dextran gel. A total of 22 anthocyanins were identified, including 15 polyacetylated, 4 monooacylated, and 3 common anthocyanins. The main components (58.14%) were pelargonidin-3-(feruloyl) diglucoside-5-(malonyl) glucoside, pelargonidin-3-(caffeoyl) diglucoside-5-(malonyl) glucoside, pelargonidin-3-(feruloyl) diglucoside-5-glucoside, and pelargonidin-3-(caffeoyl) diglucoside-5-glucoside. ARR functioned as a reversible and mixed-type inhibitor of pancreatic lipase with an IC₅₀ of 3.01 mg·mL^-1. ARR interacts with active site residues of pancreatic lipase (Phe77, His263, and Ser152) via hydrophobic, hydrogen bonding, and electrostatic interactions, inducing conformational changes in the secondary structure and inhibiting its activity.

High-purity ARR, efficiently prepared using X-5 macroporous resin, exerted a reversible mixed-type inhibitory effect on pancreatic lipase, mediated predominantly by hydrophobic and hydrogen-bonding interactions.