Chinese Chemical Letters  2017, Vol. 28 Issue (5): 1052-1056   PDF    
Triterpenoids and phenolics from the fruiting bodies of Inonotus hispidus and their activations of melanogenesis and tyrosinase
Qing Rena,b, Xue-Ying Lua, Jian-Xin Hana,b, Haji Akber Aisaa, Tao Yuana     
a The Key Laboratory of Plant Resources and Chemistry of Arid Zone, Chinese Academy of Sciences, State Key Laboratory of Xinjiang Indigenous Medicinal Plants Resource Utilization, Xinjiang Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, Urumqi 830011, China;
b University of Chinese Academy of Sciences, Beijing 100049, China
Abstract: Two new 24-methyl lanostane triterpenoids, hispindic acids A and B (1 and 2), and a new phenolic compound, hispinine (7), along with nine known compounds (3-6, and 8-12), were isolated from the fruiting bodies of Inonotus hispidus. Their structures were elucidated based on the extensive analysis of spectroscopic data (NMR and HRMS). Hispindic acid A (1) possesses an unusual formyl group at C-30. Compounds 1, 3-4, and 8 showed stronger activate abilities of melanogenesis and tyrosinase in B16 melanoma cells than those of positive control, 8-methoxypsoralen, at 50 μmol/L.
Key words: Inonotus hispidus     Lanostane triterpenoids     Phenolic compound     Melanogenesis     Tyrosinase    
1. Introduction

Inonotus hispidus (Bull. Ex Fr.) Karst is a parasitic fungus in the family Hymenochaetaceae. It is preferably living on a variety of deciduous trees such as malus, fraxinus, sorbus and quercus. I. hispidus mainly distributed in the Northeast regions and Xinjiang province of China, and was used as a traditional medicine for the treatment of dyspepsia, cancer, diabetes and stomach problems in these regions [1]. Previous chemical investigations of this species have reported the presence of a considerable quantity of yellow brown pigments, e.g. hispidin, hispolon, and some hispidin derivatives dimmers, which exhibited antimicrobial, antioxidant, and anti-inflammatory activities [2-7]. In our interest in discover ing bioactive compounds from Xinjiang (China) indigenous medicinal fungus, three new compounds including two 24-methyl lanostane triterpenoids (1 and 2) and a phenolic compound (7), together with nine known compounds (3-6, and 8-12) (Fig. 1), were isolated and identified from the methanolic extract of the fruiting bodies of I. hispidus. Moreover, all of the isolates were evaluated for their activations of melanogenesis and tyrosinase, the related targets of vitiligo. Herein, the isolation and structural elucidation, as well as the evaluation of activating melanogenesis and tyrosinase, were present.

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Figure 1. Structures of compounds 1–12.

2. Results and discussion

The total of 12 compounds (1-12) including three new ones, were isolated from the fruiting bodies of I. hispidus. Herein, the structural elucidation of the new compounds is presented.

Compound 1 was obtained as white amorphous powder, its molecular formula was determined as C31H48O4 by HRESIMS at m/z 483.3489 [M-H]- (calcd. for C31H47O4, 483.3474). The IR absorptions showed the presence of hydroxyl (3445 cm-1) and carbonyl (1706 cm-1) functionalities. The 1H NMR spectrum of 1 (Table 1) showed six methyl groups signals at δH 0.71, 0.73, 0.87, 0.98 (each 3H, s), 0.95 (d, 3H, J = 7.5 Hz), 0.96 (d, 3H, J = 6.6 Hz), a terminal double bond signals at δH 4.72, 4.63 (s, each 1H), a formyl group signal at δH 9.36 (s, 1H) and an oxygen-bearing methine signal at δH 3.00 (m, 1H). The 13C NMR (Table 1) and HSQC spectra revealed the presence of 31 carbon resonances comprising six methyls, 11 methylenes, five methines and nine quaternary carbons (two of which are carbonyl and three olefinic carbons). The aforementioned data implied compound 1 to be a 24-methyl lanostane triterpenoid. Comparison of the 1H NMR and 13C NMR data of 1 with those of compound 3 (eburicoic acid) [8], indicated that they were structurally similar, the only difference being the group at C-30. The methyl group of position-30 was replaced by a formyl group in 1, which was supported by the NMR data of CHO-30 (δH 9.36, δC 199.8), and verified by the HMBC correlations (Fig. 2a) from H-30 to C-14 (δC 67.2) and C-15 (δC 22.3). The overall structure of compound 1 was further confirmed by 1H-1H COSY and HMBC data. The relative configurations of compound 1 were established by examination of the NOESY experiment (Fig. 2b) and coupling constant. The β-orientation of 3-OH was determined on the basis of coupling constant of H-3 (J = 10.3, 5.0 Hz) [9]. NOESY correlations from: H-3 to H-1α, H-5 and H3-28; H-30 to H-12α; H-12α/H-17, indicated that they were co-facial, and in an α-orienta tion. Consequently, the NOESY correlations from: H3-19 to H3-29, H-12β and H3-18; H3-18 to H-20, showed they were β-oriented. The above NOESY data indicated that compound 1 possessed the same relative configurations with eburicoic acid. Therefore, the structure of compound 1 was elucidated as 24-exomethylene-3β hydroxy-30-oxo-lanost-8-en-21-oic acid, assigned the trivial name hispindic acid A.

Table 1
1H NMR and 13C NMR Spectroscopic data of compounds 1, 2 and 7.a

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Figure 2. (a) Key 1H–1H COSY (—) and selected HMBC correlations (H → C) of 1; (b) key NOESY correlations (H ↔ H) of 1.

Compound 2 was obtained as a white, amorphous powder with a molecular formula of C31H50O4, as determined by HR-ESIMS at m/z 485.3661 [M-H]- (calcd. for C31H49O4, 485.3631). Analysis of 1H NMR and 13C NMR data of compound 2 suggested that the structure of 2 was also similar to that of compound 3 (eburicoic acid), except for the absence of a methyl signal, while the presence of an oxygenated methylene signal (δH 3.33, 3.10; δC 64.9) in compound 2. The HMBC correlations (Fig. 3a) from: H2-28 to C-3(δC 70.7), C-4 (δC 42.4), and C-5 (δC 42.3), H3-29 (δH 0.56) to C-3, C-4, C-5 and C-28 (δC 64.9), located the oxymethylene at C-28, and the NOESY correlation from H3-19 (δH 0.93) to H3-29 (δH 0.56), supported the assignment. The extensive analysis of 2D NMR data (1H-1H COSY, HSQC, HMBC) confirmed the determination of the structure of compound 2. Thus, the structure of compound 2 was elucidated as 24-exomethylene-3β, 28-dihydroxy-lanost-8-en-21-oic acid, gave the trivial name hispindic acid B.

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Figure 3. Key 1H–1H COSY (—) and selected HMBC correlations (H → C) of 2 (a) and 7 (b).

Compound 7 was obtained as yellow amorphous solid, displayed a molecular formula of C12H12O4 with 7° of unsaturation as determined by HRESIMS at m/z 221.0894 [M+H]+ (calcd. for C12H13O4, 221.0814). In the 1H NMR spectrum (Table 1), an ABX spin system signals at δH 6.77 (d, 1H, J = 8.5 Hz, H-5'), 6.86 (brs, 1H, H-2'), and 6.74 (brd, 1H, J = 8.5 Hz, H-6'), an olefinic proton at δH 5.40 (s, 1H, H-2), and a methyl signals at δH 2.05 (s, 3H, H-6) were identified. The 13C NMR (Table 1) and HSQC spectrum revealed the presence of 12 carbon resonances comprising one methyl, one methylenes, five methines (including four sp2 and one sp3 ones) and five quaternary carbons (one of which was characteristic of a carbonyl). Further analyses of the 1D and 2D NMR (including 1H-1H COSY, HSQC, HMBC) data allowed the establishment of the structure of 7. The 1, 3, 4-trisubstituted benzene ring was confirmed by the HMBC correlations showed in Fig. 3b. The 1H-1H COSY correlations between H-4 (δH 5.29) and H2-5 (δH 2.82, 2.47) established the linkage of C-4 and C-5. The HMBC correlations from: H3-6 to C-2 (δC 103.7), C-3 (δC 176.3); H-2 to C-5 (δC 41.3); and H-5 to C-1 (δC 194.4), determined the linkage of C-6-C-3-C-2-C-1-C-5. The C-4 was attached to C-1' of benzene ring by the HMBC correlations from H-2' and H-6' to C-4 (δC 80.9). The aforemen tioned functionalities accounted for 6° of unsaturation, the remaining one of unsaturation required the presence of an additional ring in 7. Analysis of MS and 13C NMR data of C-3 and C-4 indicated that the C-3 and C-4 were linked via ether bond. The small value of optical rotation and no Cotton effect (a straight line) showed in the CD spectrum (see Supporting information) suggested that compound 7 occurred as a racemate mixture. Therefore, the structure of compound 7 was elucidated as 2-(3, 4-dihydroxyphenyl)-6-methyl-2H-pyran-4(3H)-one, gave the trivial name hispinine.

Nine known compounds were identified on the basis of spectroscopic data (1H-, 13C NMR and MS) as eburicoic acid (3) [8], inotolactone B (4) [10], ergosterol (5) [11], ergosteryl-3-O-β-D glucopyranoside (6) [11], hispolon (8) [4], (E)-4-(3, 4-dihydrox yphenyl)but-3-en-2-one (9) [6], hispidin (10) [4], pinillidine (11) [12], 3, 14'-bihispidinyl (12) [13].

All of the isolates were evaluated for their abilities of activating melanogenesis and tyrosinase in B16 melanoma cells, 8-methox ypsoralen (8-MOP) was selected as positive control. Primary screening showed that all of the isolates could increase melanin content at 50 mmol/L except the compounds 5, 6 and 10. Compounds 1, 3-4, and 7-9 were then tested their effects of activating melanogenesis and tyrosinase at different concentra tions (50 mmol/L, 10 mmol/L and 2 mmol/L), as they showed stronger activities than those of 8-MOP at primary results. As shown in Fig. 4A, compounds 4, 7 and 9 showed the activities of increasing melanin content at all of the tested concentrations compared to the control. Notably, compounds 8 and 9 significantly increased the melanin levels by 66.9% and 62.3% at 10 mmol/L compared to the control, respectively, while 8-MOP increased the melanin level by 22.5% at 50 mmol/L. As shown in Fig. 4B, compounds 1, 3 and 4 remarkably increased tyrosinase activities at 10 mmol/L and 50 mmol/L, compound 8 could increase tyrosinase activity at 50 mmol/L, while suppressed tyrosinase activity at other two concentrations. Notably, compound 3 increased tyrosinase activity by 43.3% at 10 mmol/L compared to the control, which was higher than that of 8-MOP (by 38.1%) at 50 mmol/L. Combined analysis of melanogenesis and tyrosinase data, indicated that only compound 4 could stimulate melanogenesis and activate tyrosi nase at 10 mmol/L.

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Figure 4. The effects of compounds 1, 3, 4, 79, and 8-MOP on the melanin content (A) and tyrosinase activity (B) of B16 melanoma cells. Each value represents the mean ± SE (n = 3).

3. Conclusion

In summary, phytochemical investigation of the fruiting bodies of I. hispidus led to the isolation of 12 compounds (1-12) including three new compounds (1, 2 and 7). Compounds 1-4 are 24-methyl lanostane triterpenoids, 5 and 6 are steroids, and 7-12 are phenolic compounds. Biological evaluation revealed that compounds 1, 3-4, and 8 exhibited stronger activations of melanogenesis and tyrosinase in B16 melanoma cells than those of positive control, 8-MOP at 50 mmol/L, while only compound 4 could stimulate melanogenesis and activate tyrosinase at 10 mmol/L. As activations of melanogenesis and tyrosinase are beneficial to curing vitiligo, it suggested that compound 4 could serve as lead scaffold for the synthesis of structural analogs with more potent anti-vitiligo agents.

4. Experimental 4.1. General experiment procedures

The IR spectra were recorded on a Nicolet 6700 (Thermo Fisher Scientific) spectrometer. The UV spectra were measured on a Shimadzu UV-2550 UV-vis spectrophotometer. Optical rotations were measured on an Autopol Ⅵ automatic polarimeter (Rudolph Research Analytical) at room temperature. 1D and 2D NMR data were recorded on a Varian 600 MHz instrument with TMS as internal standard. HRESIMS data were acquired using a QSTAR Elite Q-TOF mass spectrometer (Applied Biosystems). Semi-preparative HPLC separations were performed on a Hitachi Chromaster system consisting of an 5110 pump, 5210 autosampler, 5310 column oven, 5430 diode array detector and a Phenomenex Luna C18 column (250 × 10 mm, S-5 mm), all operated by EZChrom Elite software. All solvents were of ACS or HPLC grade, and were obtained from Tianjin Zhiyuan Chemical (Tianjin, China), Sigma-Aldrich (St. Louis, MO), respectively. Silica gel (300-400 mesh), C18 reverse phased silica gel (150-200 mesh, Merck), and Sephadex LH-20 gel (Amersham Biosciences), were used for column chromatography. Pre-coated silica gel GF254 plates (Qingdao Marine Chemical Plant, Qingdao, People's Republic of China) were used for TLC.

4.2. Fungal materials

The fruiting bodies of I. hispidus were collected from Akesu, Xinjiang province, China, and identified by Prof. Xinping Yang (Institute of Applied Microbiology, Xinjiang Academy of Agricul tural Sciences). A voucher specimen (IH-201410) has been deposited in the Key Laboratory of Plant Resources and Chemistry of Arid Zone, Xinjiang Technical Institute of Physics and Chemistry, Chinese Academy of Sciences (Xinjiang, China).

4.3. Extraction and isolation

The air-dried fruiting bodies of I. hispidus (1.0 kg) were pulverized, and then extracted with methanol (5.0 L × 3) by maceration at room temperature (7 days each time) to afford 147.1 g of crude methanol extract. The extract was suspended in distilled H2O and then extracted successively with petroleum ether, EtOAc and n-BuOH. The EtOAc fraction (14.2 g) was subjected to silica gel column chromatography (CC) eluted with CHCl3:MeOH (100:1-2:1 v/v) gradient to yield eight fractions (A-H). Fraction B (129.4 mg) was chromatographed over a column of Sephadex LH-20 eluted with MeOH to yield four sub-fractions (B1-B4). Sub-fraction B1 (79.7 mg) was purified by semi-preparative HPLC, eluting with MeOH-H2O (0-30 min: 90:10-100:0; 30-32 min: 100:0; 32-33 min: 100:0-90:10; 33-40 min: 90:10; v/v, 3 mL/min), yielded compound 4 (4.0 mg). Fraction C (344.6 mg) was dissolved with MeOH, MeOH-insoluble part was filtered to get compound 3 (55.7 mg). Fraction D (230.0 mg) was applied to a column of Sephadex LH-20 eluted with MeOH to yield three sub-fractions (D1-D3). Sub-fraction D1 (99.6 mg) was subjected to silica gel CC eluted with CHCl2: MeOH (80:1-10:1 v/v) gradient to obtain three sub-fractions (D1a-D1c). Purification of sub-fraction D1a (70.4 mg) by semi-preparative HPLC, eluting with MeOH-H2O (0-35 min: 85:15-100:0; 36-37 min: 100:0; 37-38 min: 100:0-85:15; 38-45 min: 85:15; v/v, 3 mL/min), yielded compounds 1 (3.0 mg) and 2 (3.3 mg). Fraction F (2.66 g) was subjected to a column of Sephadex LH-20 eluted with MeOH to yield compound 10 (25.5 mg) and seven sub-fractions (F1 -F7). Sub-fraction F2 (100.2 mg) was purified by semi-preparative HPLC, eluting with MeOH-H2O (0-29 min: 95:5-100:0; 29-30 min: 100:0; 30-31 min: 100:0-95:5; 31-38 min: 95:5; v/v, 3 mL/min), to yield compounds 6 (7.6 mg). Sub-fraction F3 (456.8 mg) was subjected to RP-18 silica gel CC (MeOH-H2O, 20/80-70:30, v/v) to give four sub fractions (F3a-F3d). Purification of sub-fraction F3a (185.1 mg) by semi-preparative HPLC, eluting with MeOH-H2O (0-22 min: 37:63-38:62; 22-23 min: 38:62-100:0; 23-25 min: 100:0; 25-26 min: 100:0-37:63; 26-34 min: 37:63; v/v, 3 mL/min), yielded compounds 7 (3.6 mg) and 9 (8.5 mg). Sub-fraction F3b (152.4 mg) was subjected to a column of Sephadex LH-20 eluted with MeOH to yield six fractions (F3b1-F3b6). Purification of fraction F3b2 (71.7 mg) by semi-preparative HPLC, eluting with MeOH-H2O (0-40 min: 32:68-44:56; 40-41 min: 44:56-100:0; 41-42 min: 100:0; 42-43 min: 100:0-32:68; 43-50 min: 32:68; v/v, 3 mL/ min), yielded compound 11 (4.8 mg). Fraction G (1.42 g) was subjected to a column of Sephadex LH-20 eluted with MeOH to yield seven fractions (G1-G7). Fraction G1 (30.7 mg) was purified by semi-preparative HPLC, eluting with MeOH-H2O (0-24 min: 45:55-75:25; 24-25 min: 75:25-100:0; 25-26 min: 100:0; 26-27 min: 100:0-45:55; 27-34 min: 45:55; v/v, 3 mL/min), to yield compound 12 (2.5 mg). Fraction H (1.2 g) was subjected to RP-18 silica gel CC (MeOH-H2O, 30/70-80:20, v/v) to give five sub fractions (H1-H5). Sub-fraction H5 (289.5 mg) was purified by silica gel CC eluted with CHCl3:MeOH (80:1-30:1 v/v) gradient to yield compound 5 (2.5 mg).

The n-BuOH fraction (17.7 g) was subjected to silica gel column chromatography (CC) eluted with CHCl3:MeOH (90:1-5:1 v/v) gradient to yield nine fractions (a-i). Purification of fraction a (220.5 mg) by semi-preparative HPLC, eluting with MeOH-H2O (0-25 min: 54:46-56:44; 25-26 min: 56:44-100:0; 26-27 min: 100:0; 27-28min: 100:0-54:46; 27-35 min: 54:46; v/v, 3 mL/min), yielded compound 8 (3.7 mg).

Hispindic acid A (1): white amorphous powder; [α]D25 -217 (c 0.100, CHCl3/MeOH, 1:1); UV (MeOH) λmax 208, 240, 304 nm; IR νmax 3445, 2961, 1706, 1472, 1261, 1022, 802 cm-1; For 1H NMR and 13C NMR spectroscopic data, see Table 1; HR-ESIMS m/z 483.3489 [M-H]-(calcd. for C31H47O4, 483.3474).

Hispindic acid B (2): white amorphous powder; [α]D25 + 16 (c 0.100, CHCl3/MeOH, 1:1); UV (MeOH) λmax 208 nm; IR νmax 3422, 2924, 1716, 1653, 1466, 1261, 1023, 801 cm-1; For 1H NMR and 13C NMR spectroscopic data, see Table 1; HR-ESIMS m/z 485.3661 [M-H]- (calcd. for C31H49O4, 485.3631).

Hispinine (7): yellow amorphous solid; [α]D25 + 8 (c 0.100, MeOH); UV (MeOH) λmax 208, 270 nm; IR νmax 3448, 2923, 1690, 1590, 1501, 901, 802, 722 cm-1; For 1H NMR and 13C NMR spectroscopic data, see Table 1; HR-ESIMS m/z 221.0894 [M+H]+ (calcd. for C12H13O4, 221.0814).

4.4. Melanin content

The murine B16 melanoma cell line was purchased from the Type Culture Collection of the Chinese Academy of Sciences (Shanghai, China). The B16 melanoma cells were cultured in HG DMEM (Gibco), and then supplemented with 10% FBS (BI, Biological Industries), 100mg/mL streptomycin and 100 U/mL penicillin (Gibco). The cells were maintained in a humidified incubator with 5% CO2 at 37 ℃, which were sub-cultured every 2 days to maintain logarithmic growth.

B16 cells were seeded in a 6-well plate at a density of 1.8 × 105 cells/well. After incubation for 24 h, they were treated with different concentrations of the tested compounds for 48 h, 8-MOP treated with 50μmol/L as a positive control, and control group was added 2μL DMSO. After that, B16 cells were washed twice with PBS (PH 7.4), and lysed in 100μL RIPA buffer for 40 min at 4 ℃. The lysates were centrifuged at 12, 000 × g for 20 min at 4 ℃. The protein content of supernatants was valued by the BCA assay and after the pellets were traeted in 190 μL of 1 μmol/L NaOH (with 1% DMSO) for 2 h at 60 ℃, melanin content was detected at 405 nm by a multi-plate reader a multi-plate reader (SpectraMax M5/M5e, Molecular Devices, Sunnyvale, CA, USA). The melanin content was calculated and corrected for the concentrations of proteins, using control groups as 100%.

4.5. Tyrosinase activity

B16 cells were seeded in a 6-well plate at a density of 2.5 × 105 cells/well. After incubation for 24 h, they were treated with different concentrations of the tested compounds for 24 h, 8-MOP treated with 50 μmol/L as a positive control, and control group was added 2μL DMSO. After that, cells were lysed with PBS containing 1% Triton X-100 and 1% sodium deoxycholate for 30 min at -20 ℃. The lysates were centrifugated at 12, 000 × g for 15 min. A reaction mixture containing 90μL of each cell lysate, 10μL of 10 mmol/L L- DOPA was added to each well of a 96-well plate. Following a 20-60 min (according to the content of dopachrome formation) incubation at 37 ℃ in the dark, the product dopachrome was detected at 490 nm by a multi-plate reader (SpectraMax M5/M5e), the tyrosinase activity of each sample was calculated and corrected for the concentrations of proteins, using control groups as 100%.

Acknowledgments

This work was supported by the Recruitment Program of Global Experts (to Tao Yuan), China; and the Xinjiang Key Research and Development Program (No. 2016B03038-3).

Appendix A. Supplementary data

Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.cclet.2016.12.010.

References
[1] L.F. Zan, H.Y. Bao. Progress in Inonotus hispidus research. Acta Edulis Fungi 18 (2011) 78–82.
[2] R.L. Edwards, D.G. Lewis, D.V. Wilson. Constituents of the higher fungi. Part Ⅰ. hispidin, a new 4-hydroxy-6-styryl-2-pyrone from Polyporus hispidus (Bull.) Fr. J. Chem. Soc. (1961) 4995–5002. DOI:10.1039/jr9610004995
[3] J.L. Fiasson. Distribution of styrylpyrones in the basidiocarps of various Hymenochaetaceae. Biochem. Syst. Ecol. 10 (1982) 289–296. DOI:10.1016/0305-1978(82)90002-3
[4] N.A.A. Ali, R. Jansen, H. Pilgrim, K. Liberra, U. Lindequist. Hispolon, a yellow pigment from Inonotus hispidus. Phytochemistry 41 (1996) 927–929. DOI:10.1016/0031-9422(95)00717-2
[5] N.A.A. Ali, R.A.A. Mothana, A. Lesnau, H. Pilgrim, U. Lindequist. Antiviral activity of Inonotus hispidus. Fitoterapia 74 (2003) 483–485. DOI:10.1016/S0367-326X(03)00119-9
[6] L.F. Zan, J.C. Qin, Y.M. Zhang, et al., Antioxidant hispidin derivatives from medicinal mushroom Inonotus hispidus. Chem. Pharm. Bull. 59 (2011) 770–772. DOI:10.1248/cpb.59.770
[7] I.K. Lee, B.S. Yun. Highly oxygenated and unsaturated metabolites providing a diversity of hispidin class antioxidants in the medicinal mushrooms Inonotus and Phellinus. Bioorg. Med. Chem. 15 (2007) 3309–3314. DOI:10.1016/j.bmc.2007.03.039
[8] C.G. Anderson, W.W. Epstein. Metabolic intermediates in the biological oxidation of lanosterol to eburicoic acid. Phytochemistry 10 (1971) 2713–2717. DOI:10.1016/S0031-9422(00)97269-8
[9] T. Kikuchi, S. Kanomi, Y. Murai, et al., Constituents of the fungus Ganoderma lucidum (Fr.) Karst. Ⅱ.:structures of ganoderic acids F, G, and H, lucidenic acids D2 and E2 and related compounds. Chem. Pharm. Bull. 34 (1986) 4018–4029. DOI:10.1248/cpb.34.4018
[10] Y.M. Ying, L.Y. Zhang, X. Zhang, et al., Terpenoids with alpha-glucosidase inhibitory activity from the submerged culture of Inonotus obliquus. Phytochemistry 108 (2014) 171–176. DOI:10.1016/j.phytochem.2014.09.022
[11] J.W. Bok, L. Lermer, J. Chilton, H.G. Klingeman, G.H.N. Towers. Antitumor sterols from the mycelia of Cordyceps sinensis. Phytochemistry 51 (1999) 891–898. DOI:10.1016/S0031-9422(99)00128-4
[12] I.K. Lee, B.S. Yun. Peroxidase-mediated formation of the fungal polyphenol 3, 14'-bihispidinyl. J. Microbiol. Biotechnol. 18 (2008) 107–109.
[13] J.J. Han, L. Bao, L.W. He, et al., Phaeolschidins A-E, fivehispidin derivatives with antioxidant activity from the fruiting body of Phaeolus schweinitzii collected in the Tibetan plateau. J. Nat. Prod. 76 (2013) 1448–1453. DOI:10.1021/np400234u