A new flavonoid glycoside, herbacetin-8-O-methyl glucuronide, together with nine known flavonoids herbacetin-8-O-β-d-xylopyranoside, kaempferol, herbacetin-8-methylether, naringenin, tricin, aromadendrin, quercetin, myricetin, and 5,7,3',4',5'-pentahydroxy-flavanone, was isolated from Rhodiola fastigiata (Hook.f. & Thomson) Fu, Crassulaceae. Their structures were identified by comprehensive analyses of physicochemical properties and spectrometric data (NMR and HR-ESI–MS). According to the bioactive study, herbacetin-8-O-β-d-xylopyranoside showed significant in vitro antimalarial activity against Plasmodium falciparum 3D7 at 50 µM. Further studies indicated that herbacetin-8-O-β-d-xylopyranoside was able to reduce the mitochondrial membrane potential of P. falciparum 3D7 considerably.