Synthesis of rhein derivatives and evaluation of anti-inflammatory activity

To conduct structural modification of rhein by using hydroxydecanoic acid and decanol as modifiers, increase lipid solubility of rhein, reduce toxic effects, and improve anti-inflammatory and antioxidant effects for potential clinical application.

Firstly, based on the structure-activity relationship of rhein, the carboxyl group in rhein was selected for esterification with ten-carbon hydroxydecanoic acid and decanol to synthesize four rhein derivatives including rhubarb-5-hydroxydecanol (R-5HD), rhubarb-hydroxydecanol (R-DA), rhubarbic-10-hydroxydecanoic acid (R-10HA), rhubarb-10-hydroxy-2-denoic acid (R-10HDA). The chemical structural characteristic of the rhein derivatives were investigated by ¹H nuclear magnetic resonance (¹H NMR) and Fourier transform infrared spectroscopy (FTIR). Secondly, the CCK-8 kit was used to determine the effects of rhein derivatives on viability of mouse mononuclear macrophage cells RAW 264.7. The most suitable concentration range of rhein and its derivatives was determined according to the cell viability. Finally, to further verify the anti-inflammatory activity of rhein derivatives, a lipopolysaccharide (LPS)-induced RAW 264.7 cell model was constructed, and the reactive oxygen species (ROS), nitric oxide (NO), tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) expression were determined. The antioxidant activity of the rhein derivatives were analyzed by the ROS fluorescent probe2,7-dichlorofluorescin diacetate (DCFH-DA).

Four rhein derivatives were prepared successfully and proved by ¹H NMR and FTIR. The in vitro anti-inflammatory model validation shows that the cytotoxicity of rhein after modification is significantly reduced; Four rhein derivatives significantly increased the cell viability of RAW 264.7 compared with rhein. The rhein derivatives had better anti-inflammatory and antioxidant activity in the LPS-treated RAW 264.7 cell model. All four rhein derivatives significantly reduced NO release, as well as expression of TNF-α, IL-6 and IL-β. All four rhein derivatives significantly inhibited ROS production compared to rhein.

This study focuses on the structural modification of rhein by hydroxydecanoic acid and decanol. In vitro experiments confirmes that R-DA has stronger anti-inflammatory and antioxidant activities, which provides a theoretical basis for the subsequent research and development of rhein derivatives.