One new flavonol diglycoside, 4',5-dihydroxy-3,3',7-trimethoxyflavone 4'-O-α-L-rhamnopyranosyl-(1 --> 6)-β-D-glucopyranoside (1), and two known compounds (2-3) were isolated from the methanolic extract of Anoectochilus annamensis Aver. aerial parts. The effects were evaluated of all isolated compounds (1-3) on LPS-induced production of the inflammatory mediator nitric oxide (NO) by RAW264.7 cells. 4',5-Dihydroxy-3,3',7-trimethoxyflavone (2) was the most active while the addition of a rutinoside at C4' (compound 1) decreased the inhibitory activity. This is the first report on the chemical composition and biological activity of A. annamensis.