Structural Elucidation and Immune-Enhancing Effects of Novel Polysaccharide from Grifola frondosa

Abstract

Beta-glucan (β-glucan) is a macromolecule structure where glucose unit has bonded through β-glycosidic bond at 1 and 3 positions. It is well known as a natural immunomodulator without exhibiting any side effects via enhancing immunity. Mushroom contains a large amount of β-glucan and it has anticancerous and antioxidant efficacy. Structure and physical properties of β-glucan are highly influenced by the types of mushroom. In particular, Grifola frondosa has β-1, 3 and β-1, 6 bonds in their structure. It has been noted that β-glucan content also depends upon the size of mushroom particles. The exact content of β-glucan and their immunological activity by a particle size of G. frondosa have yet to be fully elucidated. Herein, β-glucan contents were analyzed according to the particle size of leaf mushroom followed by cell activation and immunoactivity analysis. The highest β-glucan content was observed at a particle size of 20-30 μm (27.65 ± 0.30 w/w). All samples showed ~ 103% cell activation compared to the control and greater cell activity was observed at higher concentration. The significant increase in cytokines secretion was observed in the presence of 20-30 μm particle size of G. frondosa compared to the control. This study suggested that 20-30 μm size is the suitable size of G. frondosa that can be used as a health supplement and food additive to act as an immune booster, hypotensive agent, and hypoglycemic agent.

Figure 1

Particle size analysis of the G. frondosa powder by air classifying mill.

Figure 2

A ¹H-NMR spectrum of β-glucan extracted from G. frondosa.

Figure 3

MALDI-TOF mass spectrum of G. frondosa glucans using 2, 5-DHB as a matrix. The gap with 162 Da between peaks represents a hexose unit in the extract. Fragmented ions appeared at DP = 4-15 with 667.8, 830.0, 1166.0, 1814.5, 1976.6, and 2406.0 Da, respectively.

Figure 4

WST-1 assay of hMSCs (bone cell) cultured for different particle sizes of G. frondosa. Values are means ± SD for three concentrations per sample. ∗p < 0.05 compared with the control group.

Figure 5

Representative profile of the release of cytokines from hMSCs activated by microparticulated G. frondosa. (a) Template showing the location of antibodies for protein spotted onto the human cytokine antibody array kit. (b) Representative expression of various antibodies in the cell-free supernatants with or without G. frondosa treated. (c) Relative protein levels detected with the cytokine array. Values are means ± SD for three concentrations per sample. ∗p < 0.05 compared with the control group.