Flavonoids from Tanacetum vulgare flowers

Abstract

Flowers of tansy (Tanacetum vulgare L.) are used in various medicinal forms as antihelminthic and cholegogic agents [1–5]. The pharmacological properties of tansy flower preparations are due mainly to the essential oil (thujone and other terpenoids) and flavonoids. According to various literature sources [3–5], apigenin, acacetin, luteolin, cinaroside, eupatilin, jaceidin, and jaceoside are the main flavonoids in tansy flowers. However, reports in the domestic and foreign literature are somewhat contradictory. The goal of the present work was to study the flavonoid composition of tansy flowers growing in Samara Oblast (Russia). We studied tansy flowers collected near Nizhnee Sancheleevo village, Samara Oblast (July, 2008). Flowers (150 g) were extracted exhaustively with EtOH (70%) by combining maceration (24 h) with subsequent thermal extraction at 85–90°C. The aqueous alcohol extracts were evaporated in vacuo to a thick residue (~50 mL). The condensed extract was dried over L 40/100 silica gel. The resulting powder (extract + silica gel) was placed on a layer of silica gel that was formed in CHCl3 and eluted by CHCl3 and CHCl3:EtOH in various ratios (97:3, 95:5, 93:7, 90:10, 88:12, 85:15, 80:20, 70:30). The separation of the compounds was monitored by TLC on Silufol UV 254 and Sorbfil PTSKh-AF-A-UV plates using CHCl3:EtOH (4:1) and CHCl3:MeOH:H2O (26:14:3). Fractions containing the dominant compound 1 were combined. The precipitate that formed in these was separated and recrystallized from EtOH to afford 1 in 0.2% yield of the air-dried raw material mass. Fractions containing compounds 2–4 (each separately) were transferred to a chromatographic column (sorbent height 4.0 cm; diameter, 5 cm). The respective chromatographic column was eluted by H2O and aqueous EtOH (20%, 40, 70, and 96). Purification over polyamide columns afforded 2 (96% EtOH eluent), 3 (40% EtOH eluent), and 4 (70% EtOH eluent). The compounds were purified additionally by recrystallization from aqueous EtOH. The chemical structures of flavonoids 1–4 were elucidated using PMR and UV spectroscopy and mass spectrometry in addition to chemical transformations. Flavonoids 1 and 3 were cleaved by acid hydrolysis (10% HCl, 100°C, 2 h) and -glucosidase (Fluka, Hungary) into glucose and the aglycons, which were identified by TLC as acacetin (5,7-dihydroxy-4 methoxyflavone) (2) and apigenin (5,7,4 -trihydroxyflavone) (4), respectively. Flavonoids 2 and 4 were also isolated from tansy raw material in the free state. The PMR spectrum of 1 showed two 2H doublets at 8.07 and 7.14 ppm with SSCC (J) 9 Hz that were assigned to H-2 ,6 and H-3 ,5 , respectively; two 1H doublets at 6.84 and 6.45 ppm with J = 2.5 Hz that were characteristic of ring A protons (H-8 and H-6), and a singlet for H-3 at 6.95 (flavone compound). A 3H singlet at 3.87 for the C-4 OCH3 group and a singlet for the 5-OH of the flavonoid were observed in the PMR spectrum. The carbohydrate was placed on the 7-OH group on the basis of these results and those from enzymatic hydrolysis, and UV spectroscopy (lack of a bathochromic shift of the short-wavelength absorption band in the presence of NaOAc) [6]. The glucose was bonded as -D-glucopyranosyl (characteristic doublet for the anomeric proton at 5.35 ppm with J = 7.2 Hz). The chemical studies and spectra data suggested that 1 had the structure 5,7-dihydroxy-4 -methoxyflavone 7-O-Dglucopyranoside (tilianin) [7]. The chemical structure of 3 was studied analogously. Based on UV, NMR, and mass spectral data in addition to results from chemical transformations it was identified as 5,7,4 -trihydroxyflavone 7-O-D-glucopyranoside (cosmosiin).