The genus Anoectochilus (Orchidaceae) is a perennial herb, which comprise more than 35 species that are widespread in the tropical regions, from India through the Himalayas and southeast Asia to Hawaii. 1) Several species have been used in Chinese folk medicines.2,3) Among them, A. formosanus Hayata which is only found in Taiwan and Okinawa, has been used for hypertension, lung and liver diseases, and underdeveloped children as a folk remedy. 4) Since the natural sources of A. formosanus are becoming exhausted, other Anoectochilus species such as A. koshunensis Hayata and a different genus, Goodyera, are commercialized as substitutes used for the same purpose in the present market.5,6) Previously, Ito et al. isolated small amounts of kinsenoside (1) and the methyl ester of the carboxylic acid form of 1 (4) from A. koshunensis by Diaion and silica gel chromatography and high-performance liquid chromatography (HPLC).7) Interestingly, we isolated a great amount of 1 without 4 by silica gel column chromatography eluting with chloroform-ethanol solvent system. From these results it is easily suggested that some artificial conversion occurred during the separation procedure. This result inspired us to investigate the existence and concentrations of 1 and its analogues in Anoectochilus.This paper deals with the examination of these compounds in wild and cultured plants of A. formosanus as well as in wild A. koshunensis. Quantitative analysis of 1 was also achieved using an HPLC method as an index component of Anoectochilus. Moreover, compound 4 was confirmed to be an artifact induced during the separation procedure, and a simple and economical method was established for purification of 1. Finally the anti-hyperliposis effects of the major component 1 in Anoectochilus, and 2 (goodyeroside A) in Goodyera species, 8) have been determined by using high-fat diet rats and the anti-hyperliposis effect of 1 also was determined by using aurothioglucose-induced obese mouse.
MATERIALS AND METHODS
General Experimental Procedures1 H-and 13 C-NMR spectra were recorded on a JEOL EX 270 spectrometer. FAB-MS was recorded on a JEOL AX-500 spectrometer. HPLC was carried out on a Gasukuro Kogyo Model 576, with a Shodex RI SE-61 detector and a YMC-Pack NH 2 A-603 column. Silica gel 60 (70-230 and 230-400 mesh, Merck; 40-100 mm, Kanto Chemical Co., Inc.) were used for column chromatography. Kiesel-gel 60F 254 (Merck) was used for analytical TLC.Plant Material and Extraction A. formosanus was cultured in vitro in Seiwa Pharmaceuticals Ltd. 8,9) The wild A. formosanus and A. koshunensis were collected in Ishigaki and Taiwan, and G. schlechtendaliana was collected in Ishigaki, respectively. The voucher specimens have been deposited in the Herbarium of Medicinal Plant Garden of the Graduate School of Pharmaceutical Sciences, Kyushu University. The dried powders of whole plants of cultured A. formosanus (20 g) were extracted with MeOH at room temperature. The MeOH extract (M-1, 6.54 g) was partitioned between CHCl 3 and H 2 O to afford an H 2 O-...