Chain conformation and anti-tumor activities of phosphorylated (1→3)-β-d-glucan from Poria cocos

Abstract

(1→3)-β-d-Glucan isolated from Poria cocos was phosphorylated to obtain a series of phosphorylated derivatives. Their structures, weight-average molecular weights (M_w), and chain conformation were studied by ¹³C NMR, ³¹P NMR, static laser light scattering and viscometry. The experimental results revealed that the phosphorylated glucan existed as relatively extended flexible chain in 0.15 M NaCl aqueous solution, and exhibited relatively strong inhibition against S-180 tumor cell in vitro and in vivo. In vivo, the fractions with relatively high molecular weight at low dosage exhibited stronger anti-tumor activities. The results revealed that the molecular weights and molecular conformation could influence the anti-tumor activities. The molecular weight ranging from 2.6 × 10⁴ to 26.8 × 10⁴ and the extended chain conformation were beneficial to enhance the anti-tumor activity, as a result of the increasing of the interaction between polysaccharide and immune system.

Introduction

The studies on the fungal polysaccharides have attracted attention in the fields of pharmacology because of their non-toxicity and anti-tumor effects (Jeong et al., 2008, Lavia et al., 2006, Ye et al., 2008, Zhang et al., 2007). Polysaccharides are regarded as biological response modifier (BRM) because they are harmless and help the body to adopt to environmental and biological stress (Moradali et al., 2007, Yan et al., 1999). Many anti-tumor drugs such as 5-fluorouracil can kill tumor cells as well as normal cells and are therefore toxic. Some polysaccharides can only destroy the tumor cells without affecting the normal ones, and their anti-tumor activities mostly resulted from their immunomodulating effects (Hsieh et al., 2008, Togola et al., 2008). Normally some polysaccharides can stimulate granulocytes, monocytes, macrophages and NK-cells and trigger the secretion of IFN-g, IL-6, IL-8 and IL-12 from macrophages, neutrophils and NK-cells (Ladanyi, Timar, & Lapis, 1993). It is noted that polysaccharide derivatives have shown stronger anti-tumor activities than the unmodified polysaccharide (Bao et al., 2007, Tao et al., 2006). Particularly, phosphorylated derivatives of polysaccharides have exhibited attenuates cardiac dysfunction, anti-inflammatory, antioxidant activity and antimicrobial immunity (Sherwood et al., 2001, Williams et al., 2004, Yuan et al., 2005). However, phosphorylated polysaccharides as anti-tumor agent have scarcely been published.

Polysaccharides with different structures have been found to exist as various chain conformations in solution (Huang and Zhang, 2005, Patel et al., 2008, Tao and Zhang, 2006, Yang and Zhang, 2008, Zhang et al., 2003). The molecular weight and chain conformation of the polysaccharides significantly affected their bioactivities (Falch et al., 2000, Kojima et al., 1986, Wolfgang et al., 1992). (1→3)-β-d-Glucan in the triple helical state can suppress growth of S-180 tumor, but its random coil state was found to be ineffective (Yanaki et al., 1983). The anti-tumor activities of triple helical lentinan decreases significantly with the transition to a single chain. (Maeda et al., 1988, Surenjav et al., 2006, Zhang, Li et al., 2005). Therefore, a basic understanding of the molecular weight, conformation of the phosphorylated polysaccharide in aqueous solution is essential for successful interpretation of the bioactivities mechanism of the polysaccharides.

Poria cocos is a kind of fungi and has been used as a traditional medicine (Chen et al., 2009). (1→3)-β-d-Glucan extracted from P. cocos sclerotium is its main component, but it is water-insoluble and shows no anti-tumor activity (Ding, Zhang, & Zeng, 1998). Usually, chemical modification of water-insoluble polysaccharide can improve their water solubility. The molecular weight, molecular conformation, and anti-tumor activity of the phosphorylated (1→3)-β-d-glucan of P. cocos have never been reported. In the present work, we attempted to synthesize phosphorylated (1→3)-β-d-glucan derivatives and to study the molecular weight, molecular chain conformation of the phosphorylated polysaccharide derivative as well as its anti-tumor activities in vitro and in vivo. It may contribute some information to anti-tumor mechanism of the polysaccharides.