International Journal of Biological Macromolecules 101 (2017) 910–921

Contents lists available at ScienceDirect

International Journal of Biological Macromolecules

journal homepage: www.elsevier.com/locate/ijbiomac

Polysaccharides in Grifola frondosa mushroom and their health

promoting properties: A review
Xirui He a,b,1 , Xiaoxiao Wang b , Jiacheng Fang b , Yu Chang a,∗ , Ning Ning a , Hao Guo a ,
Linhong Huang a,∗ , Xiaoqiang Huang a , Zefeng Zhao b
a
Hong-Hui Hospital, Xi’an Jiaotong University College of Medicine, Xi’an 710054, PR China
b
Key Laboratory of Resource Biology and Biotechnology in Western China (Northwest University), Ministry of Education, Xi’an 710069, PR China

a r t i c l e i n f o a b s t r a c t

Article history: Grifola frondosa (Dicks.) Gray is a widely consumed edible and medicinal fungus, Ancient books record
Received 20 February 2017 that it can boost qi and fortify the spleen, moisten the lung and protect the liver. Modern people mainly
Received in revised form 15 March 2017 use it to assist in the treatment of diabetes mellitus and various cancers. Over the past three decades,
Accepted 29 March 2017
G. frondosa polysaccharides were shown to possess various promising bioactivities, mainly including
Available online 31 March 2017
antitumor and immunomodulation, anti-oxidation, anti-hyperglycemia, and meanwhile can effectively
act on the skin and hematopoietic stem cells. The purpose of the present review is to provide system-
Keywords:
atically reorganized information on structural characteristics, biological activities, and structure-activity
MD-fraction
SX-fraction
relationship of G. frondosa polysaccharides to support their further therapeutic potentials and sanitarian
Maitake functions.
© 2017 Elsevier B.V. All rights reserved.

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 910
2. Physiochemical, structural features and structure-activity relationship . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 911
3. Biological activities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 915
3.1. Antitumor and immunomodulatory activities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 915
3.1.1. G. frondosa polysaccharides other than D-fraction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 915
3.1.2. D-fraction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 916
3.2. Effect on hematopoietic stem cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 917
3.3. Anti-oxidative activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 917
3.4. Hypoglycemic activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 917
3.5. Effect on the skin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 917
3.6. Others . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 918
4. Conclusions and future perspectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 918
Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 918
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 918

America), eikhaas (Holland), korallkjuke (Norway), polypore en

1. Introduction touffe (France), grib-baran (Russia) [1], is a fleshy polypore recog-
nized by its smoky brown, wavy caps, which are organized in large
Grifola frondosa (Dicks.) Gray, also known as Maitake/kumotake clusters of rosettes arising from a single, branched stem structure
(Japan), gray tree flower (China), laubporling/ram’s head [2] (Fig. 1). The mature fruiting body is fleshy dark grayish brown,
(Germany), signorina mushroom (Italy), hen of the woods (North and the color gradually becomes lighter gray with age [3]. Index
Fungorum [4] presents the currently adopted taxonomy of G. fron-
dosa (Dicks.) Gray as follows: Meripilaceae, Polyporales, Incertae
∗ Corresponding authors. sedis, Agaricomycetes, Agaricomycotina, Basidiomycota, Fungi. In
E-mail addresses: hxrhist@163.com (X. He), huanglhlab@163.com (L. Huang). natural habitats, clusters of large fruiting bodies mostly occur on the
1
First author ground at or near the base of stumps of tree trunks of dead, dying,

http://dx.doi.org/10.1016/j.ijbiomac.2017.03.177
0141-8130/© 2017 Elsevier B.V. All rights reserved.
 X. He et al. / International Journal of Biological Macromolecules 101 (2017) 910–921
Fig. 1. Mature fruiting bodies of Grifola frondosa (Dicks.) Gray. Photo taken by Andrew Khitsun at Mirror Lake State Park, Wisconsin, United States (a); overmature fruiting
bodies of G. frondosa. Photo from http://mycoweb-stv.ru/(b); local fleshy porous structures of G. frondosa. Photos from www.velutipes.com (c).
or aging hardwoods, such as oaks, elms, maples, black gums, beech,
and chestnut [5]. G. frondosa is mainly distributed in Northern tem-
perate regions in Japan, European countries and the northeastern
states of America [6], and also can be found at high elevations with
temperate climate in subtropics [7]. The first large-scale commer-
cial production was developed in Japan in 1981 [8]. Ten years later,
United States and China also began large-scale artificial cultivation.
In 1999, Japanese cultivators produced nearly 40,000 t and in 2001
China produced 14,600 t of G. frondosa [9,10]. G. frondosa is prized
in traditional Chinese herbology as a medicinal mushroom to boost
qi and fortify the spleen [11]. Shennong’s Classic of Materia Medica
[12] states that it can be used for improving spleen and stomach ail-
ments, calming nerves and mind. G. frondosa has also occupied an
important position in traditional Japanese medicine, many herbal
books record it as a medicinal and edible mushroom. It is sweet in
flavour, neutral in nature, non-toxic, and it can moisten the lung
and protect the liver, strengthen healthy qi [13]. Researchers have
considered edible mushrooms as healthy food because they are rich
sources of vitamins, minerals, proteins, carbohydrates, and pheno-
lic compounds, apart from low level of lipids and low caloric content
[14–16]. Nonvolatile taste components in culinary-medicinal G.
frondosa make its fruiting bodies and mycelia possess highly intense
umami taste [17]. Remarkably, G. frondosa is also a rich sources of
trehalose [17], which has been extensively used as food preserva-
tive, sweetener and vaccine stabilizer.
Unlike other culinary-medicinal mushrooms such as Hericium
erinaceus [18], although various kinds of chemical constituents,
including coumarins, terpene lactones, flavonoids, organic acids,
alkaloids, steroids, and triterpenes have been reported from G. fron-
dosa [19], the vast majority of pharmacological studies have focused
on the bioactive polysaccharides. It was reported that G. frondosa
contains an average of 33.53% and 47.84% carbohydrates on a dry
weight basis in fruiting bodies and mycelia, respectively [17]. The
predominant monosaccharide composition of crude G. frondosa
polysaccharides is glucose (72.2–75.6%), galactose (10.5–10.9%),
mannose (7.5–7.8%), fucose (5.8%), and ribose (1.3–2.2%) [20,21].
In addition, G. frondosa contains a markedly high level of about
3.8% water-soluble polysaccharides on a dry weight basis, and high
amount of (1 → 3,1 → 6)-ˇ-d-glucans, which account for 13.2% of
the water-soluble polysaccharides [21]. G. frondosa is also a rich
sources of ␣-d-glucans, and the matted mycelium contains less
␣-glucans compared with the fruit body [22]. Remarkably, the
molecular weight of water-soluble polysaccharides of G. frondosa
shows diverse distribution, two major macromolecular populations
are 722.7 kDa and 19.6 kDa [21]. To date, besides a total of more than
forty-seven polysaccharides, there are also a great deal of bioactive
fractions obtained from G. frondosa, mainly including MD-fraction
[23], MZ-fraction [24], SX-fraction [25], and Grifolans [26]. In par-
ticular, MD-fraction is a protein-bound polysaccharide having a
glucan/protein ratio ranging from 80:20 to 99:1 [27] (Fig. 2), it is
obtained from further purification of D-fraction and has been devel-
911
oped into complementary and alternative medicines and health
care products against various cancers for more than twenty years.
More importantly, D-fraction is indeed safe with negligible or few
side-effects on healthy human subjects [28]. This was supported
further by the fact that the U.S Food and Drug Administration had
exempted D-fraction from a phase I study of toxicology and had
also approved it for an investigational new drug application to con-
duct a phase II pilot study on patients with advanced breast and
prostate cancers [28]. The SX-fraction, a glycoprotein having a pro-
tein to saccharide ratio ranging from about 75:25 to about 90:10,
helps maintain healthy blood sugar levels and insulin sensitivity
and has also been commercially exploited. In addition, the China
Food and Drug Administration (CFDA) has also approved some
patent drugs and health care products [29]. And several other new
drugs which contain G. frondosa polysaccharides as the only medici-
nal ingredient are under review [30]. Pharmacological results have
revealed that some crude G. frondosa polysaccharides and other
different Maitake fractions possess various promising bioactivities,
including antitumor and immunomodulation, anti-oxidation and
hepatoprotection, anti-hyperglycemia, etc. Essential steps involved
in the preparation of different fractions from submerged cultivated
G. frondosa are provided in (Fig. 3). A variety of purification methods
of G. frondosa polysaccharides include precipitation, ion-exchange
chromatography, gel filtration, and affinity chromatography [31].
Research advances on mycelial growth and bioactive polysaccha-
ride production of G. frondosa by submerged fermentation have
mainly focused on two following aspects: (a) improvement and
optimization of G. frondosa self-strain, culture medium and cultur-
ing methods; (b) effects of some stimulators added into submerged
culture of G. frondosa [32–36].
Reviewing the available literatures, no review concerning G.
frondosa polysaccharides is available. In this review, we intend to
systematically summarize the research findings in the past thirty
years and eventually provide a comprehensive insight into the
structure characteristics and pharmacological effects of G. frondosa
polysaccharides to provide knowledge to people for better utiliza-
tion of them.
2. Physiochemical, structural features and
structure-activity relationship
Since the 1980s, a large number of studies have focused on the
isolation and structural identification of G. frondosa polysaccha-
rides because of their promising biological activities. Herein, we
listed the reported forty-seven G. frondosa polysaccharides and pro-
vided a comprehensive information with regard to their molecular
weight, monosaccharide composition, structures, bioactivities, and
associated references in Table 1.
The structures of polysaccharides from different biological
sources are with obvious species specificity, and meanwhile the
synthesis of polysaccharides is affected by the external factors, such
 912
Table 1
Composition of monosaccharides, molecular weight, structures, and bioactivities of polysaccharides isolated from Grifola frondosa.

No. Name Monosaccharide composition M.W. (Da) Structures Bioactivities Ref.

Glucan
1 F4 Glc 1.4 × 105 (1 → 6)-branched (1 → 3)-ˇ-glucan ND [37]
2 NMF-5N Glc 7.5 × 105 (1 → 6)-branched (1 → 3)-ˇ-glucan 20 ␮g/mL against sarcoma 180 tumor cells, [22]

X. He et al. / International Journal of Biological Macromolecules 101 (2017) 910–921

94% (inhibition ratio)
3 Grifolan LE Glc 5.0 × 106 (1 → 6)-branched (1 → 3)-ˇ-glucan 100 ␮g/mL against sarcoma 180 tumor cells, [38]
99% (inhibition ratio)
4 FI0 -a-ˇ1 Glc 1.0 × 106 (1 → 6)-branched (1 → 3)-ˇ-glucan against sarcoma 180, 5.8 mg/kg (IC50 ) [39]
5 FA-1a-ˇ1 Glc 5.0 × 105 ND against sarcoma 180, 12.9 mg/kg (IC50 ) [39]
6 MT-2 (MD-fraction) Glc 1.0−1.2 × 106 (1 → 3)-branched (1 → 6)-ˇ-glucan antitumor and immunomodulatory activity [40]
7 MT-3 Glc ND (1 → 3)-branched (1 → 6)-ˇ-glucan ND [40]
8 GLP-A-2 Glc 6.2 × 104 ˇ-(1 → 6), ˇ-(1 → 2), ␣-(1 → 2), and ND [42]
␣-(1 → 3)-branched (1 → 3)-ˇ-glucan
9 GLP Glc 8.3 × 104 (1 → 3,1 → 6)-ˇ-d-glucan ND [43]
10 G.F.-1 Glc 1.2 × 106 ˇ-(1 → 6)-branched (1 → 3,1 → 6)-ˇ-d-glucan ND [44]
11 GFP2 Glc 2.6 × 103 (1 → 3,1 → 6,1 → 4)-␣-d-glucan 9.0 mg/kg against sarcoma 180, 26.56 [47]
(inhibition ratio)
12 GF4F1 Glc 1.9 × 105 (1 → 3)-branched (1 → 3,1 → 6)-ˇ-d-glucan 10.0 mg/kg against sarcoma 180, 54.3 [48]
(inhibition ratio)
13 GRN-LE Glc ND (1 → 6)-branched (1 → 3)-ˇ-glucan ND [51]
14 GFPBW1 Glc 3.0 × 105 (1 → 6)-branched (1 → 3)-ˇ-glucan antitumor activity [55]
15 GFPBW2 Glc 2.6 × 104 (1 → 6)-branched (1 → 3,1 → 4)-ˇ-d-glucan activate macrophage [56]
16 GFLP Glc 3.0 × 106 ND ND [13]

Heteropolymer
17 FII-3 Glc, Xyl in a ratio of 100.0:82.0 5.0 × 104 xyloglucan, (1 → 6,1 → 2)-branched against sarcoma 180, 23.8 mg/kg (IC50 ) [39]
(1 → 3)-ˇ-glucan
18 FIII-1a Glc, Xyl, Man, Fuc in a ratio of 100:58:34:14 1.0−2.5 × 105 ND against sarcoma 180, 16.1 mg/kg (IC50 ) [39]
19 FIII-2a Glc and trace amounts of Xyl, Man, Fuc, Gal 1.0 × 106 ND against sarcoma 180, 38.5 mg/kg (IC50 ) [39]
20 FIII-2b Glc and trace amounts of Xyl, Man, Fuc, Gal 7.0−10.0 × 104 ND against sarcoma 180, 13.9 mg/kg (IC50 ) [39]
21 FIII-2c Glc and trace amounts of Xyl, Man, Fuc, Gal 2.0−5.0 × 104 ND against sarcoma 180, 9.3 mg/kg (IC50 ) [39]
22 FI0 -a-␣ Man, Fuc, Glc, Gal in a ratio of 1.9 × 104 ND 5 mg/kg/day against sarcoma 180, 73.4% [41]
1.00:0.31:0.10:0.40 (inhibition ratio)
23 FI0 -a-ˇ Man, Fuc, Gal in a ratio of 1.00:1.21:1.06 1.2 × 104 ND 10 mg/kg/day against sarcoma 180, 66.3% [41]
(inhibition ratio)
24 FA-2-b-␣ Man, Glc, Gal in a ratio of 1.00:0.39:0.72 5.5 × 103
ND 5 mg/kg/day against sarcoma 180, 78.5% [41]
(inhibition ratio)
25 FA-2-b-ˇ Man, Fuc, Xyl, Glc, Gal in a ratio of 1.2 × 104 ND 10 mg/kg/day against sarcoma 180, 6.9% [41]
1.00:0.54:0.28:2.53:0.64 (inhibition ratio)
Table 1 (Continued)

No. Name Monosaccharide composition M.W. (Da) Structures Bioactivities Ref.

26 FII-2 Man, Fuc, Xyl, Glc in a ratio of ND ND 5 mg/kg/day against sarcoma 180, 6.7% [41]
1.00:4.25:5.81:24.57 (inhibition ratio)
27 FII-3 Man, Fuc, Xyl, Glc, Gal, Rib, Ara in a ratio of 1.5 × 104 ND 10 mg/kg/day against sarcoma 180, −12.0% [41]
1.00:0.80:1.61:2.81:0.25:0.14:0.10 (inhibition ratio)
28 FIII-1-a Man, Fuc, Xyl, Glc in a ratio of 1.1 × 105
ND 5 mg/kg/day against sarcoma 180, 84.6% [41]

X. He et al. / International Journal of Biological Macromolecules 101 (2017) 910–921

1.00:3.22:7.39:3.51 (inhibition ratio)
29 FIII-1-b Man, Fuc, Xyl, Glc, Ara in a ratio of 1.6 × 104 ND 10 mg/kg/day against sarcoma 180, 88.8% [41]
1.00:2.26:2.80:1.41:0.13 (inhibition ratio)
30 FIII-1-c Man, Fuc, Xyl, Glc, Rib, Ara in a ratio of 1.3 × 104 ND 10 mg/kg/day against sarcoma 180, 33.1% [41]
1.00:1.92:2.59:1.55:0.19:0.11 (inhibition ratio)
31 FIII-2-a Man, Fuc, Xyl, Glc in a ratio of 6.5 × 105
ND 10 mg/kg/day against sarcoma 180, 100% [41]
1.00:1.56:3.31:1.92 (inhibition ratio)
32 FIII-2-b Man, Fuc, Xyl, Glc in a ratio of 1.7 × 105 ND 10 mg/kg/day against sarcoma 180, 92.9% [41]
1.00:1.21:1.56:0.50 (inhibition ratio)
33 FIII-2-c Man, Fuc, Xyl, Glc, Gal in a ratio of 1.0 × 105 ND 10 mg/kg/day against sarcoma 180, 72.1% [41]
1.00:0.99:1.27:0.47:0.18 (inhibition ratio)
34 G.F.-2 Glc, Man, Xyl, Gal in a ratio of 4.3:1.0:0.7:1.5 7.2 × 105 ˇ-(1 → 6)-branched (1 → 3,1 → 4)-ˇ-d-glucan ND [45]
35 PGF-2 Glc, Man, Gal in a ratio of 1.00:2.35:1.22 1.2 × 105 ␣-glycosidic linkages ND [46]
36 GFPS1b Glc, Gal, Ara in a ratio of 4:2:1 2.1 × 104 backbone composed of (1 → 4)-␣-d-Galp and inhibit proliferation of MCF-7 cells, nearly [49]
(1 → 3)-␣-d-Glcp residues 100 ␮g/mL (IC50 )
37 GFM2A Glc, Man, Xyl in a ratio of 9:2:1 1.1 × 105 backbone composed of (1 → 3)-␣-d-Glcp and ND [50]
(1 → 3)-␣-d-Manp residues
38 MZF Gal, Man, Fuc, Glc in a ratio of 2.3 × 104 ND antitumor activity [52]
1.24:1.00:0.95:0.88
39 Not Given Glc, Man, Gal in a ratio of 6.5:1.0:2.6 1.3 × 104
ˇ-glycosidic linkages beneficial for the treatment of skin disease [53]
40 GFP75-2-2B Fuc, Rha, Gal, Glc, Man in a ratio of < 8.0 × 103 ND enhance the release of nitric oxide from [54]
1.0:1.3:3.6:4.0:4.5 macrophages
41 GFPW Man, Fuc, Gal in a ratio of 0.41:0.44:1.00 1.6 × 104 (1 → 2)-branched backbone composed of ND [57]
(1 → 6)-␣-d-Galp residues
42 GP11 Man, Glc, Gal in a ratio of 1.00:5.04:2.61 6.9 × 103 ˇ-glycosidic linkages antitumor and immunomodulatory activity [58]
43 Se-GP11 Man, Glc, Gal in a ratio of 1.00:4.91:2.41 3.3 × 104 ND antitumor and immunomodulatory activity [59]
44 GFP1 Glc, Fuc in a ratio of 2.3:0.5 4.1 × 104 (1 → 3)-branched (1 → 6)-ˇ-glucan anti-EV (enterovirus) 71 activity [60]
45 GFP-A Rha, Ara, Xyl, Gal, Man, Glc a ratio of 8.5 × 105 (1 → 3)-branched backbone composed of immunomodulatory activity [61]
1.38:0.53:0.11:1.07:28.75:1.76 (1 → 4,1 → 6)-␣-d-Glcp and
(1 → 3,1 → 6)-␣-d-Manp residues
46 EXGFP-A Rha, Ara, Xyl, Man, Glc, Gal a ratio of ND ND immunomodulatory activity [62]
0.28:0.31:0.30:0.06:7.98:0.61
47 Se-GFP-22 Man, Glc, Gal in a ratio of 3.30:23.30:1.00 4.1 × 106 backbone chain of (1 → 4)-␣-d-Glcp units with antioxidant activity [63]
a branched point at C-6 of both
1,3,6-ˇ-d-Manp and 1,4,6-␣-d-Galp unit

Abbreviations: Ara, araban; Xyl, xylose; Man, mannose; Glc, glucose; Gal, galactose; Fuc, fucose; Rha, rhamnose. ND, not detected. IC50 , 50% inhibiting concentration.

913
914 X. He et al. / International Journal of Biological Macromolecules 101 (2017) 910–921

Fig. 2. The major structural fragment of the polysaccharide moiety of MD-fraction with a molecular weight distributed around 1,000,000 [23,27].

Fig. 3. Fractional preparation of various Grifola frondosa fractions from submerged cultivation and fruiting bodies [40,85,95,135] and US Patent 7,214,778 B2.

as nutrients, temperature and season. Under different conditions,
the molecular weight (Mw), structures and biological activities
of polysaccharides obtained from different sources and differ-
ent extraction processes vary significantly. Previous studies have
shown that long-term heating can gradually degrade G. frondosa ˇ-
d-glucans into small fragments [64], while high molecular weight
ˇ-d-glucans (8.0 × 105 ) were shown to have stronger immunomod-
ulatory and anti-tumor activities than the lower molecular weight
ˇ-d-glucans [65]. Recent evidences showed that different extrac-
tion temperatures (70 ◦ C, 100 ◦ C and 121 ◦ C) can significantly affect
(1 → 3,1 → 6)-ˇ-d-glucan content, molecular weight, and bioactiv-
ity of water soluble polysaccharides from G. frondosa [66]. As for
the correlation between antioxidant activity and molecular weight,
combined enzyme extracts that contain lower molecular weight G.
frondosa polysaccharides than that of boiling water extracts were
shown to exhibit stronger antioxidant activities [67]. Besides the
extraction by boiling water and enzymolysis, the effect of alkali
extraction on Mw was also extensively evaluated. The molecu-
lar weights of hot water-extracted, cold alkali-extracted and hot
alkali-extracted (1 → 3)-ˇ-d-glucan were shown to be 5.6 × 106 ,
7.5 × 105 and 1.2 × 106 , respectively [68]. In the same year, Iino
and colleagues reported that a ˇ-1,3-glucan obtained from the
hot sodium hydroxide extract of G. frondosa also has a molecular
weight of 1.2 × 106 [69]. However, the molecular weight decreased
to 1.4 × 105 using aqueous zinc chloride [37]. In addition, it has also
been reported that the molecular weights of water-insoluble acidic
xyloglucan and acidic hetero-glycan extracted through ammonium
oxalate and hot alkali are 5.0 × 104 and 1.0–2.5 × 105 , respectively
[70].
G. frondosa polysaccharides obtained from boiling water con-
tain ␣-1,4-, ␣-1,6-, ˇ-1,3-, and ˇ-1,6-glucosidic linkages [71], the
hot water extract contains a large amount of (1 → 4)-␣-glucans,
whereas the cold and hot alkali extracts contain a large amount
of ˇ-glucans [71]. Among various structurally different bioactive
polysaccharides, (1 → 3,1 → 6)-ˇ-d-glucans constitute the active
principles in the immunomodulatory and antitumor activities
[69,71]. In addition, ␣-amylase digestion of hot water extract did
not affect the antitumor activity, indicating that (1 → 4)-␣-glucan is
 X. He et al. / International Journal of Biological Macromolecules 101 (2017) 910–921
not involved in the antitumor activity of G. frondosa [71]. Moreover,
evidence has suggested that the structures of hot water-extracted,
cold alkali-extracted and hot alkali-extracted (1 → 3)-ˇ-glucan are
quite similar [68]. Grifolans composed of a (1 → 3)-ˇ-glucan back-
bone with a single (1 → 6)-ˇ-d-Glcp side branching unit on every
third residue have been reported from the cultured fruit bod-
ies [55,71], matted mycelia [22] and liquid culture supernatant
[38,51]of G. frondosa. D-fraction was reported to be mainly com-
posed of (1 → 3)-branched (1 → 6)-ˇ-glucans [40]. A glucan termed
GFPBW2 structurally different from Grifolans and D-fraction was
elucidated to possess a backbone consisting of (1 → 3,1 → 4)-ˇ-
Glcp residues with branches that were composed of (1 → 3)-Glcp
residues attached to O-6 [56]. Remarkably, Ohno and colleagues
reported that an antitumor (1 → 3)-ˇ-glucan obtained via various
extraction procedures from G. frondosa possesses two kinds of con-
formations in the solid state, i.e., helix (curdlan type) and native
(laminaran type) [72]. Further evidences demonstrated that the
(1 → 3)-ˇ-glucan conformation in the fruit body of G. frondosa has
high motional freedom and is the native form [73,74]. Another
exopolysaccharide termed GLP-A-2 was shown to probably exist
as a triple helical conformation in water [42]. In addition, two G.
frondosa heteropolysaccharides were shown to exist as a spherical
conformation with random coil structure [59,63].
Except for glucans, there are also some heteropolysaccharides
isolated from G. frondosa. Cui and colleagues [49] reported an
acidic heteropolysaccharide which has a backbone consisting of
(1 → 4)-␣-d-Galp and (1 → 3)-␣-d-Glcp residues substituted at O-6
with glycosyl residues composed of ␣-l-arabinose–(1 → 4)-␣-d-
glucose (1 → linked residues. Yu and colleagues [50] reported
a heteropolysaccharide which has a backbone consisting of
three consecutive (1 → 3)-␣-d-Glcp spaced one (1 → 3)-␣-d-Manp
residues substituted at O-6 with two glycosyl residues, includ-
ing (1 → 4)-ˇ-d-Glcp and (1 → 4)-␣-d-Xylp. Wang and colleagues
[57] reported a heteropolysaccharide which possesses a backbone
consisting of (1 → 6)-␣-Galp residues with branches that are com-
posed of (1 → 3)-␣-linked fucose residues and ␣-terminal mannose
attached to O-2. Recently, Zhao and colleagues [60] reported a
heteropolysaccharide which possesses a (1 → 6)-ˇ-glucan back-
bone with a single (1 → 3)-␣-d-Fucp side branching unit. Another
heteropolysaccharide was shown to possess a backbone chain of
(1 → 4)-␣-d-Glcp units with a branched point at C-6 of both 1,3,6-
ˇ-d-Manp and 1,4,6-␣-d-Galp units, and side chains were shown to
be composed of (1 → 4)-␣-d-Glcp oligosaccharides and terminated
by ␣-d-Glcp units [63]. Moreover, Ma and colleagues [61] reported
a heteropolysaccharide which consisted of (1 → 4,1 → 6)-␣-d-Glcp
and (1 → 3,6)-␣-d-Manp residues substituted at C-3 with branches
that are composed of (1 → 6)-␣-d-Galp and t-L-Rhap residues.
G. frondosa polysaccharides namely FI0 -a-␣, FI0 -a-ˇ, FA-1, and
FA-2-b-␣ in water-soluble fractions showed higher antitumor
effect on Sarcoma 180/mice than FIII-1-a, FIII-1-b, FIII-2-a, FIII-
2-b, and FIII-2-c in water-insoluble fractions [41]. A series of
chemically-modified polyaldehyde-, polyalcohol-, and formylated-
polysaccharides together with some formolysis products were
shown to exhibit higher antitumor activity than the original G.
frondosa polysaccharides [75]. Phosphorylation can significantly
improve anticancer activities (in vivo adjuvant effect and in vitro
growth inhibitory effect) of G. frondosa polysaccharide-peptides,
followed by esterification and acetylation [76]. 2,2,6,6-tetramethyl-
1-piperidine oxoammonium ion (TEMPO)-mediated oxidation
where carboxyl groups are substituted for C6 primary hydroxyl
groups can cause a decrease in the molecular weight and viscosity
of G. frondosa polysaccharides and an increase in the water solu-
bility. The in vitro treatment of human fibrosarcoma HT1080 cells
with 100% oxidized polysaccharide was shown to effectively inhibit
cell growth. However, the oxidation reduced the activity of super-
oxide dismutase up to 82% at a concentration of 2.5 mg/mL [77].
915
In addition, carboxymethylation and selenylation were shown to
play a great influence on enhancing the antioxidant and antitumour
activities of exopolysaccharides from G. frondosa, respectively [78].
Mao and colleagues have confirmed that the antioxidant activity
of Se-G. frondosa polysaccharides for the DPPH, ABTS and hydroxyl
radicals was higher than that of G. frondosa polysaccharides and
was highest for the hydroxyl radical [79].
3. Biological activities
3.1. Antitumor and immunomodulatory activities
3.1.1. G. frondosa polysaccharides other than D-fraction
Previous studies showed that many antitumor polysaccharides
work through a host-mediated mechanism rather than directly act
on tumor cells. G. frondosa polysaccharides not only as biological
response modifiers assist the host to endure adverse biological
stresses and to increase immunity against the development of
tumor cells, but also directly induce tumor cell apoptosis. In 1957,
Byerrum and colleagues first reported that mushroom polysaccha-
rides possess antitumor property. Since then, numerous antitumor
polysaccharides were isolated from mushrooms, and the antitu-
mor activities of polysaccharides were extensively investigated.
Lentinan is one of the most studied among them. In the 1980s, a
polysaccharide fraction termed GF-1 has a similar antitumor effect
as lentinan which is also a 6-branched ˇ-1,3-glucan, whereas some
differences between GF-1 and lentinan were observed as follows
[80]: (a) GF-1 exhibited an antitumor effect when given not only
intraperitoneally and intravenously, but also intratumorally; (b)
the antitumor effect was not observed when GF-1 was admin-
istered before the tumor inoculation; (c) GF-1 was effective to
solid form of Meth A fibrosarcoma in BALA/c mice. However,
GF-1 showed no direct cytocidal effect on tumor cells, and it
was considered that GF-1 exhibited antitumor activity via host-
mediated mechanism as evidenced the fact that GF-1 affected as
an immunomodulatory on the antibody response and carbon clear-
ance activity [81]. Grifolan NMF-5N, another ˇ-1,3-glucan from G.
frondosa, also did not show cytotoxic effect on cultures of tumor
cells. And it was concluded that the antitumor activity of gri-
folan NMF-5N was due to the activation of macrophages and T
cells [26,82]. It is well known that activated macrophages, killer
T cells, and natural killer cells play important roles in immunore-
sistance to tumors. A (1 → 3)-branched (1 → 6)-ˇ-glucan from G.
frondosa showed antitumor activity against several mouse syn-
geneic tumors. Evidences showed that it not only directly activated
various effector cells, including macrophages, killer T cells, and
natural killer cells, but also potentiated the activities of various
mediators, including lymphokines and IL-1 (the first mediator in
the activation of T cell lines) [83]. It is well known that IL-12 aug-
ments NK cell- and T cell-mediated cytotoxicity and stimulates
IFN- and IL-2 production by Th1 cells. Maitake Z fraction was
shown to not only increase the expression of IL-12, IFN-, and IL-2
in tumor-bearing mice, but also induce a Th1 response by increasing
the percentage of IFN--producing cells in both splenic CD4+ and
CD8+ T cells and cytotoxic activity in NK cells and CD8+ T cells [52].
In addition, Maitake Z fraction was also shown to promote the infil-
tration of CD4+ and CD8+ T cells in tumor sites [52]. More in-depth
in vitro study has further shown that Maitake Z fraction induced DC
maturation and antigen-specific Th1 response via enhancement of
IL-12 production by DCs [84].
The first step in triggering the immune-modulating effects by
ˇ-glucans requires the specific cellular receptors that recognize
and bind to them. In recent years, a number of research findings
has shown that several pattern-recognition receptors, such as den-
dritic cell associated C-type lectin (Dectin-1), Toll-like receptor
916 X. He et al. / International Journal of Biological Macromolecules 101 (2017) 910–921
4 (TLR-4), and complement receptor 3 (CR3) are likely involved
in the immunomodulatory and antitumor effects of G. frondosa
polysaccharides. For example, a homogeneous ˇ-glucan purified
from the fruit bodies of G. frondosa was shown to be a potential lig-
and of Dectin-1 and an effective inducer of macrophage activation
through triggering cytokine secretion, at least in part, via Dectin-
1 [56]. Another ˇ-glucan from G. frondosa was shown to induce
cytokine production in macrophages through the Dectin-1/Syk/NF-
B signaling pathway, thereby showing immunomodulatory and
antitumor activities [55]. Experimental work performed in vitro
with mouse macrophage cell line RAW264.7 has suggested that the
antitumor activity of a water-soluble G. frondosa polysaccharide
namely GP11 was likely to involve the TLR-4 mediated up-
regulation of NO and TNF-␣ [58]. Another neutral ␣-polysaccharide
from G. frondosa namely GFP-A was shown to stimulate the TLR-4-
and mitogen-activated protein kinases to NF-B/pathway [61]. Fur-
thermore, a hot water extract of G. frondosa mycelia, which mainly
composed of polysaccharides was capable of not only enhancing
phagocytosis of human polymorphonuclear neutrophils, but also
up-regulating the expression of CD11b, indicating that CR3 may
involve the augmentation of the innate immunity [85].
Experimental studies performed in vitro with SGC-7901 cells
have suggested that a polysaccharide-peptide and a glucan sulfate
from G. frondosa both can significantly induce cell apoptosis [86,87].
And the apoptotic mechanism was shown to be associated with
drop in mitochondrial transmembrane potential, Bax upregulation,
Bcl-2 downregulation, and caspase-3 activation [86]. Similarly,
other experimental findings showed that a chemically sulfated
polysaccharide from G. frondos can induce HepG2 cell apopto-
sis through a notch1/NF-B/p65-mediated caspase pathway [88].
Moreover, cell cycle analysis identified hepatocellular carcinoma
SMMC-7721 cell cycle arrest at the G2/M phase following a combi-
nation of D-fraction and vitamin C treatment [89]. Furthermore, it is
well known that the tumor growth and metastasis are angiogenesis
dependent, thus blockade of angiogenesis is a promising approach
against cancer. A study carried out to evaluate the antiangiogenic
activity of a sulfated G. frondosa hetero-polysaccharide has shown
that it can significantly inhibit endothelial cell proliferation, as well
as reduce endothelial cell migration and tube formation [57].
It is also noteworthy that addition of a glucan sulfate from G.
frondosa can accelerate the cytotoxic efficacy of cyclophosphamide
or 5-fluorouracil on human gastric carcinoma SGC-7901 cells and
improve the damaged immune-competence [90]. Ma and col-
leagues showed that a novel polysaccharide with molecular weight
of 889 kDa from G. frondosa namely GFP-A can significantly relieve
the inhibition of immune responses induced by cyclophosphamide
in vivo [91]. Selenium (Se) deficiencies play an important role in the
increasing number of cancer patients among various countries [92].
A homogenous Se-enriched G. frondosa polysaccharides termed Se-
GP11 was shown to significantly inhibit the growth of Heps tumor,
increase the relatively thymus and spleen weights as well as the
serum levels of TNF-␣ and IL-2, promote the phagocytosis and NO
production of RAW264.7 cells [59].
3.1.2. D-fraction
Among the various Maitake fractions prepared, the D-fraction
was found to be very promising for enhancing the immune sys-
tem whether oral administration or injection. Other than many
antitumor mushroom polysaccharides which are mainly composed
of ˇ-1,3-glucan as a main chain with ˇ-1,6 branches, the polysac-
charide moiety of D-fraction is (1 → 3)-branched (1 → 6)-ˇ-glucan.
Further purification of the D-fraction yielded the MD-fraction [27].
Essentially, although the MD-fraction provided superior results
over the D-fraction in antitumor tests described in the patent [27], it
should be noted that any references to the D-fraction equally apply
to the MD-fraction, since they are of the same ˇ-glucan configu-
rations derived from G. frondosa [93]. Likewise, since MD-fraction
has a core ˇ-glucan structure for its inherent bioactivities, all
immunomodulatory activities demonstrated with ˇ-glucans would
be substantially relevant to those exhibited by MD-fraction [94].
Early in 1987, evidences suggested that oral administration of
D-fraction can activate the stimulation of the immune response
system triggered by the tumor-bearing state, and potentiate the
delayed type hypersensitivity response [95]. The antitumor activity
of RAW cells in the presence of D-fraction was due to iNOS-
mediated NO production, but not direct cytotoxic activity [96].
Oral administration of D-fraction was also shown to elevate the
cytotoxic activity of NK cell for long term in cancer patients
[23]. Similar other study illuminated that D-Fraction represses
cancer progression and primarily exerts its effect through stim-
ulation of NK activity in patients suffering from lung, liver, and
breast tumors [97]. Since IL-12 is critical to the functions of NK
and T cells, the long-term NK cytotoxicity induced by D-fraction
could be at least partly attributed to the increased IL-12 release.
Intraperitoneal administration of normal C3H/Hej mice with D-
fraction was shown to indirectly activate NK cells through IL-12
production from macrophages and dendritic cells [98]. Antigen-
presenting cells (APCs) like dendritic cells (DCs) are known to
involve the activation and differentiation of T cells. D-fraction can
promote the differentiation into Th-1 or Th-2 cells of CD4+ T cells
through enhancement of IL-12p70 and IFN- production by APCs in
colon 26 carcinoma-bearing BALB/c mice [99,100]. Another study
showed that D-fraction can enhance a Th-1 dominant response,
including the cell-mediated immunity associated with cytotoxic
T cell activation [101]. In addition, D-fraction was also shown to
induce a Th-2 dominant response through macrophages activation,
thereby enhancing humoral immunity rather than cell-mediated
immunity [102]. Another in vivo study has shown that D-fraction
can enhance granulopoiesis and mobilization of granulocytes by
increasing granulocyte colony-stimulating factor production and
modulating CXCR4/SDF-1 expression [103].
Nanba showed that the cancer metastasis was prevented by
91.3% by the oral administration of D-fraction. Likewise, the other
test indicated its ability to prevent carcinogenesis induced by
N-nitrosodi-n-butylamine in normal cells [104]. Another study fur-
ther illuminated that Maitake D-Fraction inhibits tumor metastasis
not only by activating NK cells and APCs during systemic circu-
lation, but also by suppressing intercellular adhesion molecule
(ICAM)-1 expression leading to the inhibition of tumor cell adhe-
sion to vascular endothelial cells [105]. D-Fraction was shown to
exert pro-apoptotic effects and reduce breast cancer cell viability.
And the action was accompanied by an increase in pro-apoptotic
genes, which was further corroborated by an increase in BAK-1 gene
expression, cytochrome c release to the cytoplasm, and activation
of caspase 7 and caspase 1 [106]. Further in depth study showed
that certain genes, such as IGFBP-7, ITGA2, ICAM3, SOD2, CAV-1,
Cul-3, NRF2, Cycline E, ST7, and SPARC, which are responsible for
the suppression of the tumoral phenotype mechanism induced by
D-Fraction in breast cancer cells [107].
Furthermore, D-fraction can decrease the effective dosage of
the chemotherapeutic agent in tumor-bearing mice [108], and was
shown to not only significantly enhance cisplatin’s antitumor and
anti-metastatic activities, but also markedly reduce the myelosup-
pression and nephrotoxicity induced by cisplatin [109]. Addition
of D-fraction can also reduce the effective dosage of vancomycine
in the treatment of Listeria infection [110], indicating the possibil-
ity that immunotherapy using the D-fraction has a clinical benefit
for the treatment of opportunistic bacterial infection. In addition,
D-fraction also had a positive impact in HIV patients [111].
Intravenous administration of vitamin C has been often used to
increase physiological concentration of MD-fraction and the com-
bination of MD-fraction and vitamin C was shown to be a viable
 X. He et al. / International Journal of Biological Macromolecules 101 (2017) 910–921 917

Table 2
Effects of MD-fraction alone and its combination with vitamin C on various cancer cells [94].

Cancer cells Growth inhibition or cell death (%)

MD-fraction a MD-fraction/VC b

T24 (bladder) 50% CD with MD-fraction (500 g/ml) 95% CD

95% CD with MD-fraction (1000 g/ml)
MCF-7 (breast) 10% GI with MD-fraction (1000 g/ml) 50% GI
HepG2 (liver) 10% GI with MD-fraction (1000 g/ml) 50% GI
U-87 (brain) 90% GI with MD-fraction (250 g/ml) 95% CD
90% GI with MD-fraction (500 g/ml)
HL-60 (leukemia) 50% CD with MD-fraction (250 g/ml) 95% CD
90% CD with MD-fraction (500 g/ml)
CF33 (mammary gland) c 40% GI with MD-fraction (500 g/ml) 90% CD
65% GI with MD-fraction (1000 g/ml)
CF21 (connective tissue) c 30% GI with MD-fraction (500 g/ml) 90% CD
70% GI with MD-fraction (1000 g/ml)
c
CL−1 (lymphoma) 20% CD with MD-fraction (500 g/ml) 95% CD
90% CD with MD-fraction (1000 g/ml)

Abbreviations: CDcell death; GIgrowth inhibition; VCvitamin C.

a
Effective concentrations of MD-fraction tested (0–1000 g/ml) are shown.
b
Combination of MD-fraction (150 g/ml) and VC (200 M) was tested, but this VC alone had no effects.
c
Canine cancer cells.

therapeutic option [94]. Synergistic potentiation of MD-fraction
with vitamin C can improve the efficacy of currently ongoing cancer
therapies. For example, MD-fraction/vitamin C combination was
shown to be more effective than various chemotherapeutic drugs,
including vinblastine, 5-florouracil, methotrexate, etoposide, cis-
platin, cyclophosphamide, mitomycin C, and carmustine, which
currently being used in clinical cases [94]. Effects of MD-fraction
alone and its combination with vitamin C on various cancer cells
are summarized in Table 2. Another recent study demonstrated that
the combination of MD-fraction and vitamin C is highly cytotoxic,
thereby inducing severe cell death in ACHN cells. The cytotoxic
mechanism was shown to be primarily associated with oxidative
stress and G1 cell cycle arrest. Also, the modulation of apoptosis
regulators (Bcl2, Bax, and PARP) was likely linked to the apoptotic
cell death [112].
3.2. Effect on hematopoietic stem cells
Lin and colleagues demonstrated that D-fraction can directly
enhance bone marrow cells proliferation and differentiation
into granulocytes-macrophages, protect the colony formation
unit response of granulocytes-macrophages from doxorubicin-
induced hematopoietic suppression, and can increase the pro-
duction of granulocyte colony-stimulating factor by cord blood
CD33+ monocytes [113,114]. Thus, these results suggest that
D-fraction has a potential to induce hematopoietic cell differ-
entiation and protect those cells from the toxic effects exerted
by chemotherapy. In particular, D-fraction may have clini-
cal implications in treatment of myelosuppression and other
hematopoietic disorders or diseases. In facts, a phase II trial
has examined the effects of a Maitake extract on innate
immune function in myelodysplastic syndromes (MDS). And
the results showed that the Maitake extract can improve neu-
trophil and monocyte function in lower-risk MDS patients. The
enhanced ROS response to Escherichia coli ex vivo in response to
Maitake extract treatment suggested that Maitake may enhance
immune responses against bacterial infection in MDS patients
[115].
3.3. Anti-oxidative activity
A combination of in vitro assays that focused on evaluating the
scavenging effects on DPPH, hydroxyl, and superoxide radicals, as
well as the reducing power, the ferrous ions chelating effect, and
the ability to inhibit rat liver lipid oxidation have demonstrated
that purified fractions from G. frondosa polysaccharides possess
effective anti-oxidative activities [79,116–118].
3.4. Hypoglycemic activity
G. frondosa has been confirmed to contain substances with anti-
diabetic activity, and it was found to lower blood sugar due to the
inhibitory activity toward ␣-glucosidase [119,120]. Evidences sug-
gested that anti-diabetic activity of an ␣-glucan from G. frondosa
on KK-Ay mice was mainly due to its effects on insulin recep-
tors, thus enhancing insulin sensitivity and ameliorating insulin
resistance of peripheral target tissues [121]. More in-depth study
performed in vitro with HepG2 cells has suggested that G. fron-
dosa polysaccharides can enhance glucose uptake by HepG2 cells,
activate the insulin receptor protein in the cell membrane, and
increase phosphorylated-AktSer473 production, thereby relieving
insulin resistance [122]. The results indicated that the G. frondosa
polysaccharides increase the metabolism of glucose and stimulate
synthesis of intracellular glycogen through the Akt/GSK-3 path-
way. Another study demonstrated that G. frondosa polysaccharides
decreased fasting serum glucose levels through the improvement
of insulin sensitivity as a result of the increased protein levels
of phospho-IR (Try1361) and decreased levels of phospho-IRS-1
(Ser307) [123].
In addition, most of the research on this area has been per-
formed with G. frondosa SX-fraction, which has shown to exhibit
hypoglycemic activity in diabetic mice and type 2 patients in
clinic [124–128]. SX-fraction is a glycoprotein having a protein
to saccharide ratio ranging from about 75:25 to about 90:10, its
average molecular weight is 20,000. The saccharide portion of
SX-fraction was found to have Galactose, Mannose, Glucose, N-
acetylglucosamine, and Fucose (US Patent 7,214,778 B2). Evidences
suggested that SX-fraction can specifically target the insulin signal
pathway, and in particular the insulin receptor and insulin recep-
tor substrate 1 therein that trigger the subsequent signaling events.
Therefore, the possible hypoglycemic action of SX-fraction was
associated with the activation of impaired insulin signal transduc-
tion pathway, thereby ultimately facilitating glucose uptake [25].
Clinical study showed that all seven diabetic patients demonstrated
over 30% (30%–63%) decline in their fasting blood Glc levels under
a SX-fraction regimen of 2–4 weeks (Table 3).
918 X. He et al. / International Journal of Biological Macromolecules 101 (2017) 910–921

Table 3 and sanitarian functions. Over the past thirty years, phytochem-
Hypoglycemic effects of SX-fraction (SXF) on type 2 diabetic patients [25].
ical investigations have led to the isolation of a large number of
Patients Average FBG (mg/dL) % FBG decline with SXF a bioactive G. frondosa polysaccharides which were shown to pos-
sess various biological activities, mainly including antitumor and
Age Sex Before SXF After SXF
immunomodulatory activities, anti-oxidative activity, and hypo-
44 M 260 90–100 63
glycemic activity. Nowadays, the MD-fraction and SX-fraction have
41 M 210 100–110 50
37 M 180–200 120–140 32 been developed into several patent health care products and phar-
75 F 200 110–130 40 maceuticals for cancers and diabetes mellitus, respectively. In facts,
64 F 220 130–150 37 since many other fractions from G. frondosa polysaccharides also
53 F 170–190 100–110 42 have a core ˇ-glucan structure as MD-fraction for their inherent
25 F 150–180 110–120 30
bioactivities, they can be undoubtedly utilized by those patients
Abbreviations: F, female; M, male; FBG, fasting blood glucose. suffering from cancers as functional foods as well. Additionally,
a
As all patients were under oral medication therapy, their average FBG levels
considering the fact that plenty of carcinogenic foods exist in our
before SXF were with medications while those after SXF were with both medications
and SXF. Hence, the percent of FBG decline with SXF reflects the improved glycemic
today’s daily diet, it is encouraging that G. frondosa polysaccha-
control with SXF. rides are effective in reducing cancer occurrence risks. Likewise,
the immune-modulating action of G. frondosa polysaccharides is
especially valuable as a mild and non-invasive form of treat-
3.5. Effect on the skin
ment, prevention of metastatic tumors, and as a co-treatment with
chemotherapy and radiotherapy as well as commonly used anti-
Experimental work performed in vitro with human dermal
cancer drugs. However, the differences in the composition and
fibroblasts has suggested that a G. frondosa extracellular polysac-
bioactivity of various fractions obtained via different preparation
charide (GF-EPS) can protect cells from hydroxyl radical-induced
process are need further investigation, and meanwhile the fer-
DNA strand breaks, inhibit matrix metalloproteinase expression,
mentation process of G. frondosa is preferably done in submerged
stimulate fibroblasts proliferation, and prevent melanin formation.
cultures, especially fed-batch or step-up cultures.
In addition, GF-MPS can stimulate the biosynthesis of collagen,
Remarkably, there are some weaknesses in the current study of
and was shown to increase the mRNA level of type I collagen in
G. frondosa polysaccharides. For example: (a) an impaired insulin
human dermal fibroblasts [118,129,130]. The results indicated that
signal pathway was shown to be activated by SX-fraction, thereby
G. frondosa polysaccharides can be potent ingredients for cosmetic.
ultimately facilitating Glc uptake, while it is likely that the Glc
Moreover, G. frondosa polysaccharides were also suggested
uptake would presumably result from Akt-induced Glc transporter
to inhibit atopic dermatitis (AD)-like skin lesions via enhancing
4 (GLUT4) translocation to the plasma membrane. Therefore, the
immunity function. For example, a hetero-polysaccharide from
cellular localization of GLUT4 is worthy to be confirmed; (b) it
G. frondosa was shown to markedly enhance skin TNF-␣ lev-
is indicative that D-fraction may help reduce the risk of cardio-
els, reduce lgG content, T lymphocyte rate and caspase-3 mRNA
vascular disease and improve renal and liver function. However,
expression in aged rats [53]. Oral administration of NC/Nga mice
larger randomized studies are still required to further confirm
with a G. frondosa polysaccharide was shown to suppress 2,4-
these interesting findings and delineate the clinical effectiveness
dinitrochlorobenzene-induced AD-like skin lesions via modulating
and potential health benefits of D-fraction; (c) given the estab-
the Th-1/Th-2 response [131].
lished experimental findings regarding genomic expression [107],
investigating each of the pathways shown by this study and
3.6. Others demonstrating how D-fraction as a nutrigenomic agent can be
transformed into a therapeutic agent for breast cancer disease
Extracellular polysaccharides from G. frondosa mycelium can would be future research directions; (d) although a phase II trial
protect liver cells from CCl4 -induced injury. The hepatoprotective has confirmed the effects of D-fraction on innate immune function
effect was shown to be associated with the down-regulation of in myelodysplastic syndromes, larger studies of longer duration
expression of cytochrome P4502E1 and TNF-␣, cell cycle arrest, are still required to see if these observed effects will translate
decrease of the activities of aspartate aminotransferase and ala- into decreased rates of infection. Furthermore, (1 → 3,1 → 6)-ˇ-
nine aminotransferase, and inhibition of superoxide anion oxygen d-glucans which primarily constitute the active principles in G.
species and ROS [132]. frondosa polysaccharides show a high degree of branching, and
It was demonstrated that a hetero-polysaccharide from G. triple-helix structures were reported to exist in G. frondosa polysac-
frondosa mycelia can block enterovirus 71 viral replication and charides. Future research is required to determine the relationship
suppress viral VP1 protein expression and genomic RNA synthe- between the three-dimensional structures and functions of G. fron-
sis in infected cells. Moreover, it exhibited apoptotic activity via dosa polysaccharides. This knowledge will help scientists to design
suppressing enterovirus 71-induced caspase-3 cleavage and IB␣ more potential health promoting pharmaceuticals and functional
down-regulation [60]. Based on other findings, some researchers foods based on chemical modification.
showed that D-fraction can strengthen immune recognition and
response to hepatitis B virus [133].
Evidences have shown that a G. frondosa exopolymer possesses Acknowledgments
hypolipidemic effect as evidenced by the fact that it can signifi-
cantly reduce the levels of plasma triglyceride, total cholesterol, This work was supported by Shaanxi Foundation for Develop-
low-density lipoprotein cholesterol, phospholipid, as well as liver ment of Science and Technology, China (Grant No. 2012K16-02-02)
total cholesterol level [134]. and 2014 CAS “Light of West China” Program.

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