New steroidal alkaloids with anti-inflammatory and analgesic effects from Veratrum grandiflorum

Ethnopharmacological relevance: "Li-Lu", the roots and rhizomes of Veratrum grandiflorum (Melianthiaceae), has been historically used as a traditional folk medicine for the treatment of wrist pain, fractures, sores, and inflammation in Yunnan Province, China. However, the anti-inflammatory and analgesic studies of this plant have seldom reported. Aim of the study: To evaluate the anti-inflammatory and analgesic properties related to the traditional usage of V. grandiflorum both in vitro and in vivo, and further explore the accurate bioactive compounds from the medicinal plant. Materials and methods: Phytochemical investigation was carried out by chromatographic methods and their structures were established based on extensive spectra and comparison with corresponding data in the reported literatures. Anti-inflammatory activities were assessed by the suppression of lipopolysaccharide-activated inflammatory mediators in RAW 264.7 macrophage cells in vitro. Furthermore, anti-inflammatory and analgesic effects were evaluated based on carrageenan-induced paw edema and acetic acid-stimulated writhing in mice. Results: The methanol extract (ME) of V. grandiflorum significantly alleviated the paw edema caused by carrageenan and the writhing numbers induced by acetic acid. Subsequent phytochemical investigation led to isolated of 21 steroidal alkaloids, including seven new compounds, veragranines C-I (1-7). Anti-inflammatory test indicated that steroidal alkaloids could decrease the expression of cyclooxygenase-2 (COX-2), interleukin-1 beta (IL1 beta), and tumor necrosis factor alpha (TNF-alpha) in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophage cells at a concentration of 5.0 mu g/ml in vitro, comparable to DXM. Moreover, five new steroidal alkaloids (2, 4, 5, 6, and 7) and two major steroidal alkaloids (9 and 13) significantly decreased the numbers of writhing in mice at the doses of 0.5 and/or 1.0 mg/kg (p < 0.01/0.05), roughly comparable to DolantinTM at 10.0 mg/kg. Conclusions: The investigation supported the traditional use of V. grandiflorum and provided new steroidal alkaloids as potent analgesic agents.