Nitric oxide-scavenging compounds in Agrimonia pilosa Ledeb on LPS-induced RAW264.7 macrophages

The extract of Agrimonia pilosa Ledeb, with a high polyphenol content, inhibited nitrite accumulation as an indicator of nitric oxide (NO) in LPS-induced RAW264.7 macrophages. The NO inhibitory compounds in the extract were isolated using open column chromatography and HPLC, and five phenolic compounds, namely aromadendrin (AD), dihydrokaempferol 3-O-β-D-glucoside (DK3-O-glc), quercitrin (QC), aglimonolide-6-O-β-D-glucoside (AG6-O-glc) and loliolide (LL), were determined by 1H, 13C NMR and LC/MS analyses. This is the first time that these compounds have been isolated from this plant. 4-Ethyl-2-hydroxyamino-5-nitro-3-hexenamide (NOR3), as a NO donor, was used in the presence of these compounds and then the nitrite-level, as an index of NO, decreased, indicating that these compounds would potentially have a NO-scavenging activity. An ESR study of the system containing NOR3 and 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy-PTIO), as a NO detection reagent, with or without the compound, provided the evidence that these compounds directly scavenged NO and the scavenging activities of three flavonoids (AD, DK3-O-glc and QC) were remarkably high in comparison to those of the other phenolic compounds. These results indicated that one of the suppression mechanisms in cells would be the NO-scavenge of phenolic compounds.