One major flavone isolated from the rhizomes of Kaempferia parviflora, is 3, 5, 7, 3′, 4′-pentamethoxyflavone (PMF). Extracts from this rhizome have been used in Thai traditional medicine for many purposes including improvement of blood circulation. However there has been no scientific evidence to support this therapeutic claim. The present study aimed to establish if PMF had any effect on blood vessels and if it did what was its mode of action. Studies were performed in vitro on isolated rat thoracic aorta obtained from female rats (220-260g) after being killed by decapitation with a guillotine, and placed in oxygenated Krebs Heinseleit solution at 37oC (PSU-Ethic No. 22/52). Cumulative concentration-response (C-R) curves were recorded by a Grass Polygraph. Data are expressed as a mean ± S.E.M. (6 rats/group/each experiments) with significant differences (p ≤0.05) between tests and control identified using the Student’s paired t-test. PMF (0.5-50 µM) caused a concentration dependent dilatation of thoracic aortic rings preconstricted with 3 µM phenylephrine. NG-nitro-L-arginine (LNA, 300 µM), a nitric oxide synthase inhibitor, or removal of the endothelium caused a significant parallel shift of the PMF induced C-R curve to the right. In the endothelium-intact thoracic aortic ring, ODQ (soluble guanylate cyclase inhibitor, 10 µM) significantly caused a rightward shift of the PMF C-R curve. Glybenclamide (10 µM) potentiated the PMF C-R curve on the thoracic aorta that was endothelium independent. Tetraethylammonium (1 mM) had no effect on the PMF C-R curves. In normal Krebs solution with LNA and nifedipine, or in Ca2+-free Krebs solution with LNA, PMF produced a further inhibition of the Phe C-R curve on the thoracic aortic ring. With thoracic aortic rings treated with LNA and thapsigargin, PMF suppressed the C-R curve produced by Phe and a further suppression was found when nifedipine (3 µM) was also added. No further suppression was achieved with a sequential addition of SKF-96365 (100 µM). However, when Y-27632 (Rho-kinase inhibitor, 30 µM) was also added into the above incubation medium cocktail, a complete suppression of the Phe C-R curve on the thoracic aortic ring was obtained. These results indicated that PMF produced an endothelium independent dilatation of the thoracic aortic ring. The mechanism responsible for this effect would be that PMF inhibited intracellular Ca2+ mobilization, stimulated the soluble guanylate cyclase and also the release of the nitric oxide from the vascular endothelium. It is unlikely that PMF caused an opening of the Ca2+- or ATP sensitive K+ channels, neither as an L-type Ca2+ channel or store-operative Ca2+ channel blockers. These results partly support the claims that extracts from rhizomes of the K. parviflora would improve blood circulation in men.