Anti-oxidant and Anti-inflammatory Effects of Ethanol Extract from Polygala sibirica L. var megalopha Fr. on Lipopolysaccharide-Stimulated RAW264.7 Cells / 中国结合医学杂志

OBJECTIVE@#To investigate the anti-oxidant and anti-inflammatory effects of ethanol extract of Polygala sibirica L. var megalopha Fr. (EEP) on RAW264.7 mouse macrophages.@*METHODS@#RAW264.7 cells were pretreated with 0-200 µg/mL EEP or vehicle for 2 h prior to exposure to 1 µg/mL lipopolysaccharide (LPS) for 24 h. Nitric oxide (NO) and prostaglandin (PGE2) production were determined by Griess reagent and enzyme-linked immunosorbent assay (ELISA), respectively. The mRNA levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor α (TNF-α), interleukin-1beta (IL-1β), and IL-6 were determined using reverse transcription polymerase chain reaction (RT-PCR). Western blot assay was used to determine the protein expressions of iNOS, COX-2, phosphorylation of extracellular regulated protein kinases (ERK1/2), c-Jun N-terminal kinase (JNK), inhibitory subunit of nuclear factor Kappa B alpha (Iκ B-α) and p38. Immunofluorescence was used to observe the nuclear expression of nuclear factor-κ B p65 (NF-κ B p65). Additionally, the anti-oxidant potential of EEP was evaluated by reactive oxygen species (ROS) production and the activities of catalase (CAT) and superoxide dismutase (SOD). The 2,2-diphenyl-1-picrylhydrazyl (DPPH), hydroxyl (OH), superoxide anion (O2-) radical and nitrite scavenging activity were also measured.@*RESULTS@#The total polyphenol and flavonoid contents of EEP were 23.50±2.16 mg gallic acid equivalent/100 g and 43.78±3.81 mg rutin equivalent/100 g. With EEP treatment (100 and 150 µg/mL), there was a notable decrease in NO and PGE2 production induced by LPS in RAW264.7 cells by downregulation of iNOS and COX-2 mRNA and protein expressions (P<0.01 or P<0.05). Furthermore, with EEP treatment (150 µg/mL), there was a decrease in the mRNA expression levels of TNF-α, IL-1β and IL-6, as well as in the phosphorylation of ERK, JNK and p38 mitogen-activated protein kinase (MAPK, P<0.01 or P<0.05), by blocking the nuclear translocation of NF-κ B p65 in LPS-stimulated cells. In addition, EEP (100 and 150 µg/mL) led to an increase in the anti-oxidant enzymes activity of SOD and CAT, with a concomitant decrease in ROS production (P<0.01 or P<0.05). EEP also indicated the DPPH, OH, O2- radical and nitrite scavenging activity.@*CONCLUSION@#EEP inhibited inflammatory responses in activated macrophages through blocking MAPK/NF-κ B pathway and protected against oxidative stress.

Glycolipids from Polygala sibirica var. megalopha and their in vitro inhibitory activity on xanthine oxidase / 中草药

Objective: To investigate the glycolipids from Polygala sibirica var. megalopha and their structure-activity relationship of xanthine oxidase (XO) inhibitory activity. Methods: The 75% EtOH extract of P. sibirica var. megalopha was chromatographied with D101 macroporous resins column by water, 35% EtOH, 65% EtOH, and 95% EtOH, successively. Compared to other elution parts, the 65% EtOH elution part exhibited stronger XO inhibition. Further bioassay-guided separation led to the isolation of four known glycolipids, which demonstrated potent XO inhibition. Results: Four compounds were identified to be tenuifoliside A (1), 3,6'-disinapoyl sucrose (2), 3'-E-3,4,5-trimethoxycinnamoyl-6-benzoyl sucrose (3), and 3'-E-3,4,5-trimethoxycinnamoyl-4-benzoyl sucrose (4) through spectroscopic data analysis. All compounds were obtained from this plant for the first time. This enzyme inhibition was dose dependent and the IC50 values of compounds 1, 3, and 4 were 9.5, 10.2, and 7.7 μmol/L, respectively. Among them, compound 2 had significantly higher XO inhibitory activity than that of the control allopurinol (IC50=11.2 μmol/L). It was speculated that the styryl side chain linking with sugar base played an important role on the XO inhibitory activity. Conclusion: Glycolipides as a novel series of XO inhibitors, may be developed into a promising remedy for human gout.