Structural characterization of a heteroglycan from an edible mushroom Termitomyces heimii

International Journal of Biological Macromolecules 151 (2020) 305–311

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Structural characterization of a heteroglycan from an edible mushroom Termitomyces heimii Prasenjit Maity a,b, Ashis K. Nandi a, Manabendra Pattanayak a, Dilip K. Manna a, Ipsita K. Sen c, Indranil Chakraborty d, Sunil K. Bhanja e, Atish K. Sahoo f, Nibha Gupta f, Syed S. Islam a,⁎ a

Department of Chemistry and Chemical Technology, Vidyasagar University, Midnapore 721102, West Bengal, India Department of Chemistry, Sabang Sajanikanta Mahavidyalaya, Lutunia, 721166 Paschim Medinipur, West Bengal, India Department of Chemistry, Sidhu Kanhu Birsa Polytechnic, Keshiary, 721133 Paschim Medinipur, West Bengal, India d Department of Chemistry, Kharagpur College, Kharagpur, 721305 Paschim Medinipur, West Bengal, India e Department of Chemistry, Government General Degree College, Kharagpur-II, Paschim Medinipur 721149, West Bengal, India f Division of Plant Pathology and Microbiology, Regional Plant Resource Centre, Bhubaneswar 751 015, Odisha, India b c

a r t i c l e

i n f o

Article history: Received 18 August 2019 Received in revised form 29 December 2019 Accepted 11 February 2020 Available online 18 February 2020 Keywords: Termitomyces heimii Heteroglycan NMR studies

a b s t r a c t A water soluble heteroglycan (THPS) of an average molecular weight ~1.98 × 105 Da was isolated from the aqueous extract of the fruit bodies of an edible mushroom Termitomyces heimii. Structural characterization of THPS was carried out using acid hydrolysis, methylation analysis, periodate oxidation, Smith degradation and 1D/2D NMR studies. Sugar analysis indicated the presence of glucose, mannose, galactose, and fucose in a molar ratio of nearly 6:2:2:1. The repeating unit of the THPS had a backbone consisting of four (1 → 3)-β-D-glucopyranosyl, one (1 → 6)-β-D-glucopyranosyl, two (1 → 3)-α-D-manopyranosyl, and two (1 → 6)-α-D-galactopyranosyl residues, out of which one (1 → 3)-β-D-glucopyranosyl residue was branched at O-6 position with terminal β-Dglucopyranosyl residue and one (1 → 6)-α-D-galactopyranosyl residue was branched at O-2 position with terminal α-L-fucopyranosyl residue. © 2020 Elsevier B.V. All rights reserved.

1. Introduction Mushrooms are nutritionally and medicinally renewable natural gift for humankind. Now a day, it's most important constituent like polysaccharides have drawn the attention of chemist and immunobiologist on account of their potent immunological application field [1,2]. Termitomyces heimii is a basidiomycete and agaric-type fungus belonging to the genus Termitomyces in the family of Lyophyllaceae [3,4]. It is known as “termite mushroom” due to its symbiotic relationship with termite infested soil and also it have been identified as an edible mushroom with a unique taste and flavor [4,5]. Naturally this mushroom was collected from India during monsoon season [6]. This mushroom is extensively used as a dietary calcium supplement in people suffering from hypocalcinaemia and osteoporosis. It is also used in regulation of blood pressure, blood lipid, immune response, inflammation, and apoptosis. Thus, the result supports the beneficial claims of this mushroom [7]. These mushrooms showed DPPH radical scavenging activity, hydroxyl radical scavenging activity, inhibition of lipid peroxidation, β-carotene bleaching assay, reducing power, and total antioxidant

⁎ Corresponding author. E-mail address: [email protected] (S.S. Islam).

https://doi.org/10.1016/j.ijbiomac.2020.02.120 0141-8130/© 2020 Elsevier B.V. All rights reserved.

activity [8–10]. Its protective role in human lymphocytes was reported [11]. This mushroom shows varying immune-stimulatory activities [12]. Decolorisation capacity of laccase from termitomyces heimii was reported [13]. T. heimii showed noticeable antibacterial activities [14]. This mushroom species may act as a potential source for the antihypertensive proteins [15]. So, T. heimii would be beneficial for health purposes and also used in medicine of selected diseases of humankind. Different mushrooms of the genus Termitomyces including Termitomyces striatus [16], T. eurhizus [17], T. microcarpus [18], T. robustus [19], T. robustus var [20], and T. clypeatus [21] have been identified and characterized by our group. Different extract of mushroom T. heimii, alkaline extract polysaccharide was identified as β-glucan [11] whereas aqueous extract polysaccharide was characterized as heteroglycan (THPS). THPS contain important sugar residue α-L-fucose which is used for human breast cancer [22] and infertility treatment [23]. Heteroglycan act as a potent immunostimulating agent on splenocyte, thymocyte and macrophagedependent immune systems [24,25] and also showed protective role on human lymphocytes [26]. Mushroom polysaccharides are protected the living body from oxidative damage causing by different reactive oxygen species (ROS) which are considered to be the most toxic substances [27]. Therefore, a detailed investigation was carried out on the polysaccharide of this mushroom and the results of structural studies are reported herein.

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2. Materials and methods

isolated and purified by the method described previously [28]. Finally, the crude polysaccharide (30 mg) was purified by gel permeation chromatography on column of Sepharose 6B using water as eluant by Redifrac fraction collector. A single fraction (test tube 20–32, Fig. 1a) was collected, and freeze-dried, yielding 20 mg of pure THPS. The same procedure was repeated in several lots to yield 100 mg of pure polysaccharide.

2.1. Isolation and purification of the crude polysaccharide Sixty gram dry mushroom powder, Termitomyces heimii were boiled with distilled water for 12 h. The aqueous extract was kept overnight at 4 °C and filtered through linen cloth. The crude polysaccharide was

a

0.5

Absorbance at 490 nm

0.4

0.3

0.2

0.1

0.0

0

10

20

30

40

50

60

No of test tubes

b

c

Fig. 1. a. Gel permeation chromatogram of crude polysaccharide isolated from the edible mushroom Termitomyces heimii using Sepharose 6B column. b. 1H NMR spectrum (500 MHz, D2O, 30 °C) of THPS isolated from the edible mushroom Termitomyces heimii. c. Combination of 13C NMR and DEPT-135 spectrum (125 MHz, D2O, 30 °C) of THPS isolated from an edible mushroom Termitomyces heimii.

P. Maity et al. / International Journal of Biological Macromolecules 151 (2020) 305–311

307

Table 1a GLC-MS analysis of methylated polysaccharide (THPS) isolated from mushroom Termitomyces heimii. Methylated sugars

Molar ratio

Linkage type

Major Mass Fragments (m/z)

3,4-Me2-Gal 2,4,6-Me3-Man 2,3,4-Me3-Fuc 2,3,4-Me3-Gal 2,4,6-Me3-Glc 2,4-Me2-Glc 2,3,4-Me3-Glc 2,3,4,6-Me4-Glc

1 2 1 1 3 1 1 1

→2,6)-α-D-Galp-(1→ →3)-α-D-Manp-(1→ α-L-Fucp-(1→ →6)-α-D-Galp-(1→ →3)-β-D-Glcp-(1→ →3,6)-β-D-Glcp-(1→ →6)-β-D-Glcp-(1→ β-D-Glcp-(1→

43,71,87,99,129,159,173,189,233 43,59,71,87,101,117,129,145,161,205 43,72,89,101,115,117,131,161,175 43,71,87,99,101,117,129,161,173,189,233 43,74,87,101,117,129,143,161,173,203,217,233 43,58,74,87,101,117,129,143,159,173,189,233 43,74,87,99,101,117,129,143,161,173,203,217,233 43,71,87,101,117,129,161,173,189,233

Table 1b GLC-MS analysis of methylated polysaccharide (THPS) after oxidation of sodium periodate isolated from mushroom Termitomyces heimii. Methylated sugars

Molar ratio

Linkage type

Major Mass Fragments (m/z)

2,4,6-Me3-Man 2,4-Me2-Glc 2,4,6-Me3-Glc

2 1 3

→3)-α-D-Manp-(1→ →3,6)-β-D-Glcp-(1→ →3)-β-D-Glcp-(1→

43,58,71,87,101,117,129,143,161,173,189,201,233 43,58,74,87,101,117,129,143,159,173,189,201,233 43,58,74,87,101,117,129,143,161,173,203,217,233

temperature of 200 °C. The NMR experiments were carried out as reported earlier [24,27].

2.2. General methods The molecular weight was measured by the method of Hara, Kiho, Tanaka, and Ukai [29]. The optical rotation was measured on a Jasco Polarimeter model P-1020 at 26 °C. For sugar analysis, the THPS (3.0 mg) was hydrolyzed with 2 M CF3COOH (2 mL) in a round-bottom flask at 100 °C for 18 h in a boiling water bath and the analysis was carried out as described in previous papers [24,27]. The absolute configuration of the monosaccharide constituents was determined by the method of Gerwig, Kamerling, and Vliegenthart [30]. The THPS was methylated according to the method of Ciucanu and Kerek method [31] where distilled DMSO and finely grounded NaOH were used and then converted to alditol acetates as reported earlier [24,27]. Periodate oxidation and Smith degradation experiment were carried out with this THPS as described in the earlier report [27]. A Gas-Liquid Chromatographic analysis (GLC) was done using Hewlett-Packard model 5730 A, having a flame ionization detector and glass columns (1.8 m × 6 mm) packed with 3% ECNSS-M (A) on Gas Chrom Q (100–120 mesh) and 1% OV225 (B) on Gas Chrom Q (100–120 mesh). All GLC analyses were performed at 170 °C. The Gas-liquid chromatography-mass spectrometric (GLC-MS) analysis was performed using Shimadzu GLC-MS Model QP2010 Plus automatic system, using ZB-5MS capillary column (30 m × 0.25 mm). The program was isothermal at 150 °C; hold time 5 min, with a temperature gradient of 2 °C min−1 up to a final

3. Results and discussion 3.1. Isolation, purification, and chemical analysis of the polysaccharide Sixty grams dry mushroom powder were boiled with distilled water for 12 h followed by centrifugation, precipitation in EtOH, and finally freeze dried to yield 1.5 g of crude polysaccharide. Fractionation of water soluble crude polysaccharide (30 mg) through Sepharose 6B column yielded single (test tubes 20–32, Fig. 1a) polysaccharide was collected and freeze dried. The pure THPS showed a specific rotation [α] 26 + 17.6 (c 0.1, water). The average molecular weight [29] of THPS D was estimated as ~1.98 × 105 Da from a calibration curve prepared with standard dextrans. GLC analysis of the alditol acetates of the hydrolyzed product of THPS revealed the presence of glucose, mannose, galactose, and fucose in a molar ratio of nearly 6:2:2:1. The absolute configuration of the monosaccharide was determined by the method of Gerwig et al. [30] and found that glucose, galactose, and mannose had the D configuration but fucose was present in the L configuration. The mode of linkages of the sugar moieties present in the THPS was determined by methylation analysis using the Ciucanu and Kerek method [31], followed by hydrolysis and preparation of alditol acetates. The

Table 1c The 1Ha and 13Cb NMR chemical shifts for the polysaccharide (THPS) isolated from mushroom Termitomyces heimiiin D2O at 30 °C. Glycosyl residue

H-1/C-1

H-2/C-2

H-3/C-3

H-4/C-4

H-5/C-5

H-6a,H-6b/C-6

→2,6)-α-D-Galp-(1→ A →3)-α-D-Manp-(1→ B(BI, BII) α-L-Fucp-(1→ C →6)-α-D-Galp-(1→ D →3)-β-D-Glcp-(1→ E →3,6)-β-D-Glcp-(1→ F →6)-β-D-Glcp-(1→ G β-D-Glcp-(1→ H

5.12 98.2 5.12×, 5.05y 101.8×,101.4y 5.05 97.6 4.98 97.9 4.77 102.0 4.73 102.4 4.53 102.4 4.50 102.4

3.84 76.9 4.00 70.8 3.80 68.7 3.85 69.7 3.35 72.8 3.50 72.9 3.50 73.1 3.30 73.2

3.98 69.8 3.86 78.2 4.07 69.9 4.03 69.8 3.74 84.7 3.71 84.5 3.46 75.5 3.48 75.5

4.13 69.5 3.79 68.0 4.05 70.9 4.13 69.2 3.44 69.6 3.46 69.6 3.40 69.6 3.37 69.6

4.21 68.4 3.83 74.0 4.17 68.0 4.21 68.4 3.62 75.2 3.65 75.0 3.64 75.0 3.42 76.0

3.67c, 3.67d 66.0 3.75c, 3.93d 61.0 1.21 15.5 3.64c, 3.64d 66.8 3.73c, 3.89d 60.5 3.85c, 4.20d 68.8 3.92c, 4.20d 68.9 3.68c, 3.83d 60.2

Interchangeable. x For residue BI. y For residue BII. Values of the 1H chemical shifts were recorded with respect to the HOD signal fixed at δ 4.70 at 30 °C. b Values of the 13C chemical shifts were recorded with reference to acetone as the internal standard and fixed at δ 31.05 at 30 °C.

c,d

a

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Fig. 2. The HSQC spectrum (D2O, 30 °C) of (a) anomeric part and (b) other than anomeric part of THPS isolated from an edible mushroom Termitomyces heimii. (c) The part of ROESY spectrum of THPS isolated an edible mushroom Termitomyces heimii. The ROESY mixing time was 300 ms.

P. Maity et al. / International Journal of Biological Macromolecules 151 (2020) 305–311 Table 2 ROE effects of THPS, observed in the ROESY spectrum recorded in D2O at 30 °C. Unit

A BI BII C D E F G H

ROE signals From

Intraunit

Interunit

AH-1 BI H-1 BII H-1 C H-1 D H-1 E H-1 F H-1 G H-1 H H-1

AH-2 BI H-2 BII H-2 C H-2 D H-2 E H-2, E H-3, E H-5 F H-2, F H-3, F H-5 G H-2, G H-3, G H-5 H H-2, H H-3, H H-5

G H-6a, G H-6b F H-3 D H-6a, D H-6b AH-2 A H-6a, A H-6b BI H-3 BII H-3 E H-3 F H-6a, F H-6b

GLC-MS analysis of the alditol acetates of methylated products have been shown in Table 1a. These linkages (Table 1a) were further confirmed by periodate oxidation experiment (Table 1b). GLC analysis of alditol acetates of the periodate-oxidized [32,33], NaBH4-reduced, and hydrolyzed products showed the presence of glucose and mannose, indicating that the galactose and fucose moieties were consumed during oxidation. GLC-MS analysis of periodate-oxidized, reduced, methylated [34] THPS showed the presence of 1,3,5-tri-O-acetyl-2,4,6-tri-O-methyl-D-glucitol, 1,3,5tri-O-acetyl-2,4,6-tri-O-methyl-D-mannitol, and 1,3,5,6-tetra-O-acetyl2,4-di-O-methyl-D-glucitol in a molar ratio of nearly 3:2:1. These results clearly indicated that the (1 → 3), (1 → 3,6)-linked glucopyranosyl and (1 → 3)-linked mannopyranosyl residues (Table 1b) remain unaffected whereas all other residues were consumed during oxidation, which further confirmed the mode of linkages present in the THPS.

3.2. NMR and structural analysis of the polysaccharide (THPS) The 1H NMR (500 MHz) spectrum (Fig. 1b, Table 1c) at 30 °C showed five signals in the anomeric region at δ 5.12, 5.05, 4.98, 4.77, 4.73, 4.53, and 4.50 in a ratio of nearly 2:2:1:3:1:1:1. The sugar residues were designated as A-H according to their decreasing anomeric proton chemical shifts. In the 13C (Fig. 1c, Table 1c) NMR (125 MHz) spectrum at 30 °C seven anomeric signals appeared at δ 102.4, 102.0, 101.8, 101.4, 98.2, 97.9, and 97.6 in a ratio of nearly 3:3:1:1:1:1:1.

309

Based on the result of the HSQC experiment (Fig. 2a), the anomeric carbon signals at δ 102.0, 101.8, 101.4, 98.2, 97.9, and 97.6 corresponded to the anomeric carbons of E, BI, BII, A, D and C residues, respectively while the peak at δ 102.4 was correlated to the anomeric carbon of F, G and H residues. All the 1H and 13C signals were assigned using DQF-COSY, TOCSY, HSQC (Fig. 2a & b) experiments [21,24,26,27,35–37,and]. The proton Coupling constants were measured from DQF-COSY spectrum and one-bond C\\H couplings were deduced from proton coupled 13C spectrum. Based on the coupling constant, JH-1,H-2–3.1 Hz and JC-1,H-1–171 Hz the residues A and D were established as α-anomer. A large JH-2,H-3 (~ 9 Hz) and small JH-3,H-4 (b5 Hz) indicated that those were D-galactosyl unit. In residue A, the downfield shift of C-2 (δ 76.9) and C-6 (δ 66.0) with respect to standard values of methyl glycosides [35] indicated that the moiety A was (1 → 2,6)-linked unit. On the other hand, in residue D, the downfield shift of C-6 (δ 66.8) with respect to standard values of methyl glycosides indicated that it was (1 → 6)-linked unit. The linking at C-6 of the both residue A and D were further confirmed from DEPT-135 spectrum (Fig. 1c). Hence, these observations confirmed that residue A was a (1 → 2,6)-linked-α-D-galactopyranosyl moiety and the residue D was a (1 → 6)-linked-α-D-galactopyranosyl moiety. The manno configuration of residues BI and BII were supported from large coupling constants of JH-3,H-4 (~7 Hz) and JH-4,H-5 (~9 Hz). The anomeric proton signals at δ 5.12 and 5.05 and the coupling constant values of JH-1,H-2 (~1.6 Hz), JH-2,H-3 (~3.5 Hz) and JC-1,H-1(~170 Hz) clearly indicated that both BI and BII residues were α-linked mannopyranosyl moieties. The downfield shift of C-3 (δ 78.2) with respect to standard values of methyl glycosides [35] indicated that the both BI and BII residues was (1 → 3)-linked-α-D-mannopyranosyl residue. Residue C was assigned to L-fucopyranosyl unit. This was strongly supported by the appearance of a proton signal at δ 1.21, carbon signal at δ 15.5 for a CH3 group, and the relatively small JH-3,H-4 (b3 Hz). The appearance of the anomeric proton and carbon signals for both residues at δ 5.05 and 97.6, respectively, as well as the coupling constant value JH-1, H-2–3.75 Hz clearly indicated that those were α-anomer. The anomeric configuration was further confirmed by 1H\\13C coupling constant JC-1, H-1–171 Hz. The carbon chemical shifts of residue C from C-1 to C-6 were assigned from HSQC experiment and corresponded nearly to the standard values of methyl glycoside of α-L-fucose. Thus, considering the results of chemical analysis and NMR experiments, it may be conclude that the residue C was a terminal α-L-fucopyranosyl moiety.

Fig. 3. The 13C NMR spectrum of the Smith-degraded glycerol-containing hexsaccharide of THPS isolated from isolated an edible mushroom Termitomyces heimii in D2O at 30 °C.

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Table 3 The 13C NMRa chemical shifts of Smith-degraded glycerol-containing hexasaccharide of the polysaccharide (THPS) isolated from mushroom Termitomyces heimii in D2O at 30 °C. Sugar residue

C-1

C-2

C-3

C-4

C-5

C-6

→3)-β-D-Glcp-(1→ I β-D-Glcp-(1→ J →3)-α-D-Manp-(1→ KI →3)-α-D-Manp-(1→ KII Gro-(3→ L

103.0

73.0

84.20

69.70

75.5

61.0

102.0

73.52

76.8

70.5

76.0

60.8

102.7

71.0

80.0

66.9

74.0

62.0

102.5

71.2

79.8

66.7

74.0

61.9

72.0

62.5

a

68.10

The values of chemical shifts were recorded with reference to acetone as internal standard and fixed at δ 31.05 ppm at 30 °C.

Residues E, F, G and H were established as β-anomer from coupling constant values JH-1,H-2 (~8 Hz), and JC-1,H-1 (~160 Hz) and the large coupling constant values JH-2,H-3 and JH-3,H-4 (~10 Hz) of the residues E, F, G and H confirmed their glucopyranosyl moiety. The downfield shift of C3 (δ 84.5) and C-6 (δ 68.8) with respect to standard values indicated that moiety F was linked at C-3 and C-6. These observations indicated that F was (1 → 3,6)-linked-β-D-glucopyranosyl moiety. On the other hand, the downfield shift of C-3 (δ 84.8) with respect to standard values of methyl glycosides [35] indicated that moiety E was linked at C-3. Thus it may be concluded that E was (1 → 3)-linked-β-Dglucopyranosyl moiety. The downfield shifts of C-6 (δ 68.9) of G residue supported the presence of (1 → 6)-linking in β-D-glucopyranosyl moiety. All the chemical shifts of residue H were nearly analogous with the standard values of methyl glycoside [28] of β-D-glucose. This observation clearly indicated that the residue H was non-reducing end β-Dglucopyranosyl moiety. The linking at C-6 of the residues F and G were further confirmed from DEPT-135 spectrum (Fig. 1c). The sequences of glycosyl moieties were determined from ROESY (Fig. 2c) experiments. In ROESY experiment, the inter-residual contacts along with some intra-residual contacts were observed (Fig. 2c, Table 2). The above ROESY connectivities established the following sequences: A (1 → 6) G; BI (1 → 3) F; BII (1 → 6) D; C (1 → 2) A; D (1 → 6) A; E (1 → 3) BI; F (1 → 3) BII; G (1 → 3) E; and H (1 → 6) F.

These data clearly indicated the positions of substitution and sequence of sugar residues in the polysaccharide. For further confirmation of the sequence of linkages in THPS, the Smith degraded material (SDPS) was prepared and NMR experiment was carried out. The 13C NMR (125 MHz) spectrum (Fig. 3, Table 3) at 30 °C of SDPS showed four anomeric carbon signals at δ 103.0, 102.7, 102.5, and 102.0 in a ratio of nearly 3:1:1:1, corresponding to →3)-β-DGlcp-(1 → (I), →3)-α-D-Manp-(1 → (KII), →3)-β-D-Manp-(1 → (KI) and β-D-Glcp-(1 → (J) residues respectively. The carbon signals C-1, C2, and C-3 of the glycerol moiety were assigned as δ 68.10, 72.0, and 62.50 respectively. The nonreducing β-D-Glcp-(1 → (J) was generated from one (1 → 3)-β-D-Glcp (E) due to complete oxidation of the →6)β-D-Glcp-(1 → (G) and also one (1 → 3)-β-D-Glcp (J) was produced from the (1 → 3,6)-β-D-Glcp (C) due to oxidation followed by Smith degradation of the β-D-Glcp-(1 → moiety (H) and the other two (1 → 3)-β-DGlcp (J) was retained from two (1 → 3)-β-D-Glcp (E) and also two (1 → 3)-α-D-Manp (KI and KII) was retained from two (1 → 3)-α-DManp (BI and BII) respectively. The glycerol (L) moiety was generated from (1 → 6)-α-D-Galp (D) after periodate oxidation followed by Smith degradation and be attached to (1 → 3)-α-D-Manp (KII) as a gro part. Hence, Smith degradation resulted in the formation of an oligosaccharide unit from the parent polysaccharide and the structure of which was established as:

So, considering all the results of chemical investigations and NMR spectroscopic evidences, the structure of repeating unit in the THPS was proposed as:

4. Conclusion A water soluble heteroglycan (THPS) was isolated from the aqueous extract of an edible mushroom Termitomyces heimii. The following structure was proposed by chemical analysis and 1D/2D NMR studies.

CRediT authorship contribution statement Author statement The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Prasenjit Maity: Conceptualization, Writing. Ashis K. Nandi: Methodology and analysis. Manabendra Pattanayak: Methodology and analysis. Dilip K. Manna: Methodology and analysis. Ipsita K. Sen: Writing-Reviewing and Editing. Indranil Chakraborty: Writing-

P. Maity et al. / International Journal of Biological Macromolecules 151 (2020) 305–311

Reviewing and Editing. Sunil K. Bhanja: Methodology. Atish K. Sahoo: Supervision. Nibha Gupta: Supervision. Syed S. Islam: Supervision. Acknowledgements The authors are grateful to Mr. Barun Majumdar, Bose Institute, Kolkata for preparing NMR spectra. The authors are also grateful to the Ministry of Science & Technology, Department of Biotechnology, and Government of India for sanctioning a project [No. BT/PR9175/NDB/ 39/315/2013]. References [1] S.P. Wasser, Medicinal mushrooms as a source of antitumor and immunomodulating polysaccharides, Appl. Microbiol. Biot. 60 (2002) 258–274. [2] S.P. Wasser, A.L. Weis, Medicinal properties of substances occurring in higher Basidiomycetes mushrooms: current perspectives (review), Int. J. Med. Mushrooms 1 (1999) 31–62. [3] K. Natarajan, South Indian Agaricales v: Termitomyces heimii, Mycologia 71 (4) (1979) 853–855, https://doi.org/10.2307/3759201. [4] N. Rahmad, J.R. Al-Obaidi, N.M.N. Rashid, N.B. Zean, M.H.Y.M. Yusoff, N.S. Shaharuddin, N.A.M. Jamil, N.M. Saleh, Comparative proteomic analysis of different developmental stages of the edible mushroom Termitomyces heimii, Biol. Res. 47 (30) (2014) 1–8. [5] S.S. Tripathy, A. Rajoriya, N. Gupta, Wild indigenous mushrooms as a source of food from different forest divisions of Odisha, J. Pharmaceut. Biol. 4 (3) (2014) 138–147. [6] B. Kumari, N.S. Atri, Evaluation of alkaloids of north Indian wild edible termitophilous mushrooms, Libyan Agric. Res. Cen. J. Intl. 3 (5) (2012) 229–232. [7] S.N.A. Malek, G. Kanagasabapathy, V. Sabaratnam, N. Abdullah, H. Yaacob, Lipid components of a malaysian edible mushroom, Termitomyces heimii Natarajan, Int. J. Food Prop. 15 (2012) 809–814. [8] N.G. Puttaraju, S.U. Venkateshaiah, S.M. Dharmesh, S.M.N. URS, R. Somasundaram, Antioxidant activity of indigenous edible mushrooms, Journal of Agriculture and Food Chemistry 54 (2006) 9764–9772. [9] P. Mitra, N.C. Mandal, K. Acharya, Polyphenolic extract of Termitomyces heimii: antioxidant activity and phytochemical constituents, Journal fur Verbraucherschutz und Lebensmittelsicherheit 11 (2016) 25–31. [10] T. Ao, C.R. Deb, Nutritional and antioxidant potential of some wild edible mushrooms of Nagaland, India, J. Food Sci. Technol. 56 (2) (2019) 1084–1089. [11] D.K. Manna, A.K. Nandi, M. Pattanayak, P. Maity, S. Tripathy, A.K. Mandal, S. Roy, S.S. Tripathy, N. Gupta, S.S. Islam, A water soluble β-glucan of an ediblemushroom Termitomyces heimii: structural and biological investigation, Carbohydr. Polym. 134 (2015) 375–384. [12] N. Abdullah, R. Abdulghani, S.M. Ismail, M.H.Z. Abidin, Immune-stimulatory potential of hot water extracts of selected edible mushrooms, Food Agric. Immunol. 28 (3) (2017) 374–387. [13] K.H. Shin, I. Nashwaan, J. Logis, T.W. Keat, W.K. Kit, L.C.S. Ying, T.Y. Hui, Charaterisation and decolorisation capacity of laccase from termitomyces Heimii, Malaysian Journal of Biochemistry and Molecular Biology 22 (1) (2019) 110–117. [14] K. Singha, B.R. Pati, K.C. Mondal, P.K.D. Mohapatra, Study of nutritional and antibacterial potential of some wild edible mushrooms from Gurguripal Ecoforest, West Bengal, India. Indian Journal of Biotechnology 16 (2) (2017) 222–227. [15] N.H.M. Fadzil, C.F. Misnan, N. Aminudin, N. Abdullah, Comparative SELDI-ToF/ms analysis of antihypertensive proteins from Termitomyces heimii natarajan and Ganoderma lucidum karst, Journal of the University of Malaya Medical Centre 16 (2013) 67. [16] S. Mondal, I. Chakraborty, M. Pramanik, D. Rout, S.S. Islam, Isolation and structural elucidation of a water-soluble polysaccharide (PS-I) of a wild edible mushroom, Termitomyces striatus, Carbohydr. Res. 341 (2006) 878–886.

311

[17] S. Mondal, I. Chakraborty, M. Pramanik, D. Rout, S.S. Islam, Structural studies of water-soluble polysaccharides of an edible mushroom, Termitomyces eurhizus. A reinvestigation, Carbohydr. Res. 339 (2004) 1135–1140. [18] K. Chandra, K. Ghosh, K.S. Roy, S. Mondal, D. Maiti, A.K. Ojha, D. Das, S. Mondal, S.S. Islam, A water-soluble glucan isolated from an edible mushroom Termitomyces microcarpus, Carbohydr. Res. 342 (2007) 2484–2489. [19] S. Mondal, K. Chandra, D. Maiti, A.K. Ojha, D. Das, S.K. Roy, K. Ghosh, I. Chakraborty, S.S. Islam, Chemical analysis of a new fucoglucan isolated from an edible mushroom, Termitomyces robustus, Carbohydr. Res. 343 (2008) 1062–1070. [20] S.K. Bhanja, C.K. Nandan, S. Mandal, B. Bhunia, T.K. Maiti, S. Mondal, S.S. Islam, Isolation and characterization of the immunostimulating β-glucans of an edible mushroom Termitomyces robustus var, Carbohydr. Res. 357 (2012) 83–89. [21] M. Pattanayak, S. Samanta, P. Maity, I.K. Sen, A.K. Nandi, D.K. Manna, P. Mitra, K. Acharya, S.S. Islam, Heteroglycan of an edible mushroom Termitomyces clypeatus: structure elucidation and antioxidant properties, Carbohydr. Res. 413 (2015) 30–36. [22] J.L. Jay, P.S. Gene, M.L. Catherine, The emerging importance of α-L-fucose in human breast cancer: a review, Am. J. Transl. Res. 3 (4) (2011) 292–322. [23] J.V. Jennifer, M.S. Jennifer, S.B. Barry, Hamster sperm-associated alpha-L fucosidase functions during fertilization, Biol. Reprod. 82 (2010) 572–579. [24] A.K. Nandi, S. Samanta, I.K. Sen, K.S.P. Devi, T.K. Maiti, K. Acharya, S.S. Islam, Structural elucidation of an immunoenhancing heteroglycan isolated from Russula albonigra (Krombh.) Fr, Carbohydr. Polym. 94 (2013) 918–926. [25] E.K. Mandal, K. Maity, S. Maity, S.K. Gantait, S. Maiti, T.K. Maiti, S.R. Sikdar, S.S. Islam, Structural characterization of an immunoenhancing cytotoxic heteroglycan isolated from an edible mushroom Calocybe indica var. APK2, Carbohydr. Res. 346 (2011) 2237–2243. [26] P. Maity, A.K. Nandi, I.K. Sen, M. Pattanayak, S. Chattopadhyay, S.K. Dash, S. Roy, K. Acharya, S.S. Islam, Heteroglycan of an edible mushroom Entoloma lividoalbum: structural characterization and study of its protective role for human lymphocytes, Carbohydr. Polym. 114 (2014) 157–165. [27] A.K. Nandi, S. Samanta, S. Maity, I.K. Sen, S. Khatua, K.S.P. Devi, K. Acharya, T.K. Maiti, S.S. Islam, Antioxidant and immunostimulant β-glucan from edible mushroom Russula albonigra (Krombh.) Fr, Carbohydr. Polym. 99 (2014) 774–782. [28] A.K. Nandi, I.K. Sen, S. Samanta, K. Maity, S.K.P. Devi, S. Mukherjee, T.K. Maiti, K. Acharya, S.S. Islam, Glucan from hot aqueous extract of an ectomycorrhizal edible mushroom, Russula albonigra (Krombh.) Fr.: structural characterization and study of immunoenhancing properties, Carbohydr. Res. 363 (2012) 43–50. [29] C. Hara, T. Kiho, Y. Tanaka, S. Ukai, Anti-inflammatory activity and conformational behavior of a branched (1→3)-β-D-glucan from an alkaline extract of Dictyophora indusiata Fisch, Carbohydr. Res. 110 (1982) 77–87. [30] G.J. Gerwig, J.P. Kamerling, J.F.G. Vliegenthart, Determination of the D and L configuration of neutral monosaccharides by high-resolution capillary g.l.c, Carbohydr. Res. 62 (1978) 349–357. [31] I. Ciucanu, F. Kerek, Simple and rapid method for the permethylation of carbohydrates, Carbohydr. Res. 131 (1984) 209–217. [32] I.J. Goldstein, G.W. Hay, B.A. Lewis, F. Smith, Controlled degradation of polysaccharides by periodate oxidation, reduction and hydrolysis, Methods Carbohydr. Chem. 5 (1965) 361–370. [33] G.W. Hay, B.A. Lewis, F. Smith, Periodate oxidation of polysaccharides: general procedures, Methods Carbohydr. Chem. 5 (1965) 357–361. [34] M. Abdel-Akher, F. Smith, Use of lithium aluminium hydride in the study of carbohydrates, Nature 166 (1950) 1037–1038. [35] P.K. Agrawal, NMR spectroscopy in the structural elucidation of oligosaccharides and glycosides, Phytochemistry 31 (1992) 3307–3330. [36] S. Mondal, I. Chakraborty, D. Rout, S.S. Islam, Isolation and structural elucidation of a water-soluble polysaccharide (PS-I) of a wild edible mushroom, Termitomyces striatus, Carbohydr. Res. 341 (2006) 878–886. [37] D. Maiti, K. Chandra, S. Mondal, A.K. Ojha, D. Das, S.K. Roy, K. Ghosh, I. Chakarborty, S.S. Islam, Isolation and characterization of a heteroglycan from fruit bodies of Astraeus hygrometricus, Carbohydr. Res. 343 (2008) 817–824.