Dammarane saponins from Gynostemma pentaphyllum Yongmei Hu a , Fanny C.F. Ip a , Guangmiao Fu a , Haihong Pang a , Wencai Ye b , Nancy Y. Ip a, * a Department of Biochemistry, Molecular Neuroscience Center, Biotechnology Research Institute, Department of Biochemistry, The Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong b Institute of Traditional Chinese Medicine and Natural Products, Jinan University, Guangzhou 510632, China article info Article history: Received 29 July 2009 Received in revised form 19 January 2010 Available online 17 May 2010 Keywords: Gynostemma pentaphyllum Cucurbitaceae Saponins Cortical neurons ERK phosphorylation abstract Dammarane-type saponins (1–7), together with five known compounds, were isolated from the aerial parts of Gynostemma pentaphyllum. Compounds 1–4, 6 and 7 induced the phosphorylation of ERK protein in primary rat cortical neurons, which indicates their potential neuroactivity. On the other hand, no induction of ERK phosphorylation was observed for HEK293 cells following treatment with saponins 1, 3, 4 and 7. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction Gynostemma pentaphyllum (Thunb.) Makino (Cucurbitaceae) has been used in China as a folk medicine with various therapeu- tic properties such as lowering cholesterol levels, regulating blood pressure and strengthening the immune system (Razmovski-Nau- movski et al., 2005). Studies on the chemical and pharmacological characterization of G. pentaphyllum have led to the designation of ‘‘Southern Ginseng” for the herb. Previous reports on this herb identified a series of dammarane-type glycosides (e.g., gypeno- sides I to LXXIX) that are structurally related to ginsenosides (Kuwahara et al., 1989; Liu et al., 2004, 2005; Yin et al., 2004, 2006a,b). Since many ginsenosides, such as the well-known active principles in Korean ginseng and Panax ginseng (Araliaceae), are known for their significant biological activities (Xue et al., 2006; Findeis, 2007; Fujita et al., 2007; Zhang et al., 2008), the gypeno- sides derived from G. pentaphyllum have also received much attention because of their similarities. While gypenosides have been reported to exhibit a potential apoptotic effect on various cancer cell lines (Wang et al., 2007; Chen et al., 2009), they also show an anti-oxidative effect on macrophages, endothelial and li- ver cells (Li et al., 1993; Tanner et al., 1999; Aktan et al., 2003). Studies on the active entity of gypenosides have established the effect of phanoside, TR1 and gypenoside XLIX on specific receptor activation or factor release in macrophages, endothelial and pancreatic cells (Norberg et al., 2004; Huang et al., 2005, 2007). However, few studies have reported the effect of gypenosides on neurons although G. pentaphyllum has been used for treatment of neurological diseases (e.g., headache and stroke) in China (Raz- movski-Naumovski et al., 2005). A recent study has also shown that the ethanol extract of G. pentaphyllum prevents rat brain slices from hypoxia/hypoglycemia damage and this effect was suggested to be beneficial for stroke and heart attack (Schild et al., 2009). On the other hand, two groups reported the protec- tive effect of gypenosides against glutamate neurotoxicity and reperfusion injury (Qi et al., 2000; Shang et al., 2006). In Qi’s study, it was demonstrated that gypenosides preserved DNA and RNA of rat neurons against ischemic injury. Furthermore, Shang showed that the neuroprotective effects of gypenosides on cortical neurons are mediated by multiple anti-oxidative ac- tions, e.g., via enhancing intracellular glutathione, suppressing glutamate-induced cytosolic Ca 2+ elevation, and blocking gluta- mate-induced apoptosis, indicating the potential role of gypeno- sides in the treatment of neurological diseases involving glutamate and oxidative stress (Shang et al., 2006). In addition, gypenoside has been shown to inhibit Na+/K+-ATPase activity in microsomal preparations of rat brain and heart (Han et al., 2007). All these data indicate that gypenosides may comprise a mixture of compounds with potential applications for neuroindi- cations. In our study, a systematic chemical investigation was car- ried out on G. pentaphyllum and which resulted in isolation of 12 dammarane-type saponins including seven new ones (1–7). The structure elucidation of the new compounds was achieved based on 2D-NMR spectroscopic analysis as well as chemical degrada- tion methods. The bioactivities of saponins were evaluated in the present work by investigating the induction of extracellular 0031-9422/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.phytochem.2010.04.003 * Corresponding author. Tel.: +852 2358 7304; fax: +852 2358 1552. E-mail address: boip@ust.hk (N.Y. Ip). Phytochemistry 71 (2010) 1149–1157 Contents lists available at ScienceDirect Phytochemistry journal homepage: www.elsevier.com/locate/phytochem