Elsevier

Food Hydrocolloids

Volume 46, April 2015, Pages 1-9
Food Hydrocolloids

Preparation and structural characterization of a partially depolymerized beta-glucan obtained from Poria cocos sclerotium by ultrasonic treatment

https://doi.org/10.1016/j.foodhyd.2014.12.005Get rights and content

Highlights

  • Ultrasonication depolymerized insoluble β-glucan (PCS0) into water-soluble fractions.

  • PCS90 (ultrasonic-treated for 90-min) had lower MW and viscosity than PSC0.

  • PCS90 had stronger inhibition of SW620 than PCS0.

  • Compactly coiled PSC0 was opened up into a linear chain in PSC90 as shown by TEM.

  • PCS90 is a partially depolymerized bioactive beta-glucan.

Abstract

A water-insoluble polysaccharide (PCS) obtained by alkaline extraction from the sclerotium of a medicinal mushroom, Poria cocos (PC) was partially depolymerized by ultrasonication for various period of times up to 90 min. The untreated polysaccharide (PCS0) and two mushroom polysaccharides treated for 10 and 90 min (PCS10 and PCS90) were purified by membrane ultrafiltration. PCS90 as a hydrocolloid that had a lower intrinsic viscosity (about 30% of PCS0) and Mw (43 kDa) than PCS0. The 13C NMR spectra of PCS0 and PCS90 indicated that they both had a linear (1 → 3)-β-D-glucan which was further confirmed by methylation analysis. TEM results also showed that ultrasonic treatment could open up the compact structure of PCS into linear chain. PCS90 showed stronger inhibitory effects towards the proliferation of colorectal cancer cell SW620 than PCS0 (IC50 of PCS90 and PCS0 was 32.2 and 169.4 μg/mL, respectively). These results suggested that ultrasonic treatment might be an effective physical method for improving the solution properties of water-insoluble homoglycans such as β-glucan without changing its primary chemical structure and such modified polysaccharides had enhanced their antitumor activity.

Introduction

Polysaccharides from edible and medicinal fungi have attracted attention in the field of biopharmacological field due to their antitumor and immunomodulatory effects (Zeković et al., 2005, Zhang et al., 2007) including their direct tumor inhibition (Wasser, 2002). It has been reported that a polysaccharide extracted from Poria cocos significantly suppressed the proliferation of U937 and HL-60 cells (Chen & Chang, 2004). Polysaccharides isolated from Pleurotus tuber-regium, Pleurotus pulmonarius have been demonstrated to have potent cytotoxicity towards cancer cells (Xu et al., 2012, Zhang et al., 2004c). In vitro studies had indicated that incubation of mushroom polysaccharides with tumor cells could affect the intracellular signaling pathways that would lead to their inhibition of proliferation (Chen and Chang, 2004, Li et al., 2004, Lin et al., 2003).

Polysaccharides contain repetitive structural features which are polymers of monosaccharide residues joined to each other by glycosidic linkages and can be interconnected at several points to form a wide variety of branched or linear structures (Cheung, 2008). Numerous studies have been attempted to correlate biological activity and molecular weight of polysaccharides. Mizuno indicated that high molecular weight glucans appeared to be more effective than those of low molecular weight, especially (1-3)-β-glucans that ranged from 500 to 2000 kDa (Mizuno, 1996). Water solubility of polysaccharides is another factor that influences its bioactivity. Overall, the solubility of polysaccharides depends on their degree of polymerization and their physico-chemical properties (Zeković et al., 2005). It has been established that the immunomodulation of soluble and particulate beta-glucans is mediated by activation of CR3 and Dectin-1 receptors, respectively (Goodridge et al., 2011, Qi et al., 2011). However the insoluble nature of polysaccharides substantially limits their antitumor activity, as there have been studies suggesting that soluble polysaccharides exhibit more inhibitory effect than those insoluble ones (Chen et al., 2009, Chen et al., 2010). Chemical modifications of native polysaccharides by forming their carboxymethylated, hydroxylated, formylmethylated, aminoethylated and sulfated derivatives have been reported to enhance their aqueous solubility as well as improved antitumor activity (Wang et al., 2004a, Zhang et al., 2004a, Zhang et al., 2004b). Other than chemical methods, physical means such as ultrasonic, γ-ray, ultraviolet light and microwave treatments have been applied to modify the structure and properties of polysaccharides (Vodeničarová, Dřímalová, Hromádková, Malovíková, & Ebringerová, 2006). The main advantage of physical means over chemical methods is that no chemical reagents are required and hence removal of any excess chemicals is not necessary. Ultrasonic treatment is an effective method to partially depolymerize polysaccharides by glycosidic bond cleavage, the extent of which could be controlled by different power input and treatment time. A recent study on ultrasonic depolymerization of β-glucan from Cs-HK1, an exopolysaccharide (EPS) isolated from fermentation broth of a medicinal fungus Cordyceps sinensis, showed a significant increase in water solubility and antioxidant ability in the modified EPS (Chen, Siu, Cheung, & Wu, 2014).

The sclerotium of Poria cocos has been widely used in traditional Chinese medicine (Ríos, 2011). The main component of P. cocos sclerotium is a water-insoluble linear (1-3)-β-D-glucan, which had very modest antitumor activity (Ding et al., 1998, Wang et al., 2004a). Chemically modified polysaccharide derivatives from P. cocos sclerotium exhibited notably increased solubility and significant inhibitory effect towards S-180 tumor cells after phosphorylation (Chen et al., 2009). Till now, polysaccharide derivatives obtained from P. cocos sclerotium by ultrasonication have not been reported before. Therefore, in this study, water-insoluble polysaccharide extracted from the sclerotium of P. cocos by alkaline extraction was partially degraded using ultrasonic treatment to give a mushroom hydrocolloid. The chemical and physical properties of this mushroom hydrocolloid including water solubility, molecular weight, intrinsic viscosity and structural characteristics as well as its antitumor activity were determined. To our knowledge, this is the first report on modified β-glucan as a hydrocolloid prepared by ultrasonication from P. cocos sclerotium.

Section snippets

Alkaline extraction of polysaccharide

Sclerotium of P. cocos was originated from the Yunnan province, China. It was dried and pulverized into powders to pass through a screen with an aperture of 0.5 mm by using a cyclotech mill (Tecator, Hӧganäs, Sweden). Powders were defatted by ethyl acetate and then acetone, before subjected to a cold alkali extraction (1 M NaOH, room temperature, 16 h). The supernatant of the cold alkali-soluble fraction was first neutralized to pH 7.0 with 0.5 M HCl to yield precipitates which were washed for

Purification and chemical analysis of PCS0

In the present study, the untreated PCS sample (PCS0) obtained by membrane ultrafiltration (Mw cut-off 10 kDa) had significantly higher carbohydrate content (more than 20%) than the crude alkali mushroom extract (Fig. 1). Moreover, no protein and uronic acids were detected in PCS0 (data not shown), indicating it was only composed of neutral polysaccharide. The results indicated that the crude alkali mushroom extract could be purified by membrane ultrafiltration to give a reasonably pure

Discussion

P. cocos consists of three morphological forms, including mycelium, sclerotium and fruiting body, among which sclerotium has been studied extensively (Wang et al., 2004). In this project, an alkaline extraction was used since this PCS fraction would include the majority of polysaccharides (>84%) (Wang, Zhang, Ruan, et al., 2004) obtained from the sclerotium of P. cocos and this fraction has been demonstrated to exhibit anti-tumor or immunomodulatory activities after modification (Wang, Zhang,

Conclusion

A (1 → 3)-β-D-glucan (PCS) was purified by membrane ultrafiltration from a crude alkaline extract obtained from the sclerotium of P. cocos. Ultrasonic treatment was shown to be an effective means to partially depolymerize PCS, resulted in the reduction of Mw and viscosity as well as an increase in water solubility of the ultrasonic-treated PCS samples. Ultrasonication can also unfold the compact structure of PCS into an open chain conformation in aqueous medium. All the changes in the

Acknowledgments

This project was financially supported by the Research Committee of CUHK Directed Grant for Research (No. 2030435). The technical support provided by technicians from the School of Life Sciences and the Department of Physics of CUHK for FT-IR was gratefully acknowledged.

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