Endertiins A-B, two lanostane triterpenoids from the fruit bodies of the mushoom Humphreya endertii

Abstract

Two new lanostane triterpnenoids named endertiins A and B (1 and 2) together with two known compounds as ganodecalone A (3) and ergosterol (4) have been successfully isolated from the cultivated fruit bodies of the mushroom Humphreya endertii Stey (Ganodermataceae). Their structures were elucidated by a combination of HR-MS and 2 D NMR spectroscopic analyses. In addition, endertiins A and B (1 and 2) were evaluated their cytotoxicity against two cancer cell lines, MCF7 (human breast carcinoma) and LU (human lung carcinoma). The result showed that endertiin A (1) could inhibit the growth of MCF-7 cells with its IC50 value of 71.16 ± 6.25 µg/ml.

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Endertiins A-B, two lanostane triterpenoids from
the fruit bodies of the mushoom Humphreya
endertii
Dang Ngoc Quang , Le Duc Long , Nguyen Quang Tung , Nguyen Ngoc Thanh
& Le Xuan Tham
To cite this article: Dang Ngoc Quang , Le Duc Long , Nguyen Quang Tung , Nguyen Ngoc Thanh
& Le Xuan Tham (2020): Endertiins A-B, two lanostane triterpenoids from the fruit bodies of the
mushoom Humphreya�endertii , Natural Product Research, DOI: 10.1080/14786419.2020.1800696
To link to this article: https://doi.org/10.1080/14786419.2020.1800696
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Published online: 03 Aug 2020.
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Endertiins A-B, two lanostane triterpenoids from the fruit
bodies of the mushoom Humphreya endertii
Dang Ngoc Quang
a
, Le Duc Long
b
, Nguyen Quang Tung
b
, Nguyen Ngoc
Thanh
b
and Le Xuan Tham
c
a
Faculty of Chemistry, Hanoi National University of Education, Hanoi, Vietnam;
b
Faculty of Chemical
Technology, Hanoi University of Industry, Hanoi, Vietnam;
c
Faculty of Biotechnology, Hongbang
International University, Hochiminh City, Vietnam
ABSTRACT
Two new lanostane triterpnenoids named endertiins A and B (1
and 2) together with two known compounds as ganodecalone A
(3) and ergosterol (4) have been successfully isolated from the
cultivated fruit bodies of the mushroom Humphreya endertii Stey
(Ganodermataceae). Their structures were elucidated by a combin-
ation of HR-MS and 2 D NMR spectroscopic analyses. In addition,
endertiins A and B (1and 2) were evaluated their cytotoxicity
against two cancer cell lines, MCF7 (human breast carcinoma) and
LU (human lung carcinoma). The result showed that endertiin A
(1) could inhibit the growth of MCF-7 cells with its IC
50
value of
71.16 ± 6.25 mg/ml.
ARTICLE HISTORY
Received 17 May 2020
Accepted 10 July 2020
KEYWORDS
Humphreya endertii;
lanostane triterpen-
oid; cytotoxic
1. Introduction
It is estimated that there are about 300 species of lingzhi mushrooms
(Ganodermataceae) globally and about 60 are described in Vietnam. They have been
used traditionally and are even being cultivated as food or for medicinal purposes for
a long time (Tham 2005). Previously, mushrooms from Ganodermataceae are proven
to be a good source of secondary metabolites with a broad spectrum in the biological
system, such as anti-cancer, anti-inflamatory, anti-microbial …(Tham 2005; Quang
et al. 2011; Phuc et al. 2014). In the course of our investigation on the biological active
compounds from Vietnamese mushroom belonging to Ganodermataceae, variuos
metabolites have been identified as lanostane trterpenoids, steroids, …(Quang et al.
CONTACT D. N. Quang quangdn@hnue.edu.vn
Supplemental data for this article can be accessed at https://doi.org/10.1080/14786419.2020.1800696.
ß2020 Informa UK Limited, trading as Taylor & Francis Group
NATURAL PRODUCT RESEARCH
https://doi.org/10.1080/14786419.2020.1800696

2011; Hien et al. 2013; Phuc et al. 2014). H. endertii Stey is a rare mushroom which
was only found in Indonesia and Malaysia. It was recently discovered in Cattien
National Park, Lamdong province, Vietnam. Then, its fruit bodies were successfully cul-
tivated in laboratory (Tham et al. 2018). The mushroom was then identified by not
only morphological analyses but also molecular biology. Interestingly, H. endertii has a
closed phylogenetic relationship with Ganoderma neo-japonicum (Tham et al. 2009).
However, its chemical composition and application remains unknown. This paper
describles the chemical constituents and anti-cancer activity of a new lingzhi
H. endertii.
2. Results and discussion
The HR-MS data for the molecular ion peak [M þH]
þ
(m/z 453.3355) of endertiin A (1)
supported a molecular formula of C
30
H
44
O
3
, with 9 degrees of unsaturation. Its
1
H
NMR spectrum displayed the presence of seven methyl signals, including one singlet
methyl at 1.75 ppm suggesting that it is connected with a double bond, five tertiary
methyls, one methyl doublet at 0.95 ppm (d, 6.5 Hz). In addition, one olefinic proton at
6.47 (1H, brs) and one aldehyde proton at low filed (9.40 ppm) are observed. The
13
C
NMR spectrum of compound 1revealed the presence of 30 carbon signals. They are
one saturated ketone (218.2 ppm), one conjugated ketone (198.9 ppm), one aldehyde
(195.6 ppm), two double bonds (164.2, 154.8, 139.3, 138.2 ppm) and others which are
listed in the section 3.3.1. Then, the structure of compound 1was deduced by the
detail analyses of its 2 D NMR (HSQC, HMBC and NOESY spectra). There were HMBC
correlations between i) H-27 (1.75 ppm) and the aldehyde (C-26); ii) H-27 and C-24, C-
25; iii) H-26 and C-24, C-25 and C-27, indicating the presence of unsaturated aldehyde
at C-26. The saturated ketone was determined at C-3 due to long-range correlations
between H-28, H-29 and C-3. Similarly, the unsaturated ketone was assigned at C-11
since it was coupled to H-12 and H-18 in its HMBC spectrum. Thus, compound 1was
predicted as a lanostane triterpenoid (Kikuchi et al. 1985; Gao et al. 2002). The relative
Figure 1. Structures of compounds 1–4.
2 D. N. QUANG ET AL.

configuration of compound 1was also deduced by its NOESY spectrum. There were
NOE correlation between H-24 and H-26 indicating the configuration of the double
bond C
24-25
was trans. H-5 was found to be a-face since it had NOE correlation with
H-28. From the above discussion, compound 1was assigned to be (24E)- 3,11-dioxo-
5a-lanosta-8,24-dien-26-al as shown in Fig. 1 and named endertiin A.
Endertiin B (2) was analyzed having the molecular formula of C
30
H
48
O
3
on the basis
of a [M þH]
þ
ion cluster at m/z 457.3646 (calcd for C
30
H
49
O
3
, 457.3682). Analysis of its
2 D NMR spectra and in comparison with those of compound 1suggested that com-
pound 2was also a lanostane triterpenoid but two alcohol signals were observed in 2
instead of two carbonyl groups. Detail interpretation of its 2 D NMR spectra indicated
that it contained one primary alcohol (4.0 ppm and 68.9 ppm in its
1
H NMR and
13
C
NMR spectra, respectively) and one secondary alcohol at C-3 [3.24 (dd, J¼11.0, 5.0, H-
3) and (78.7 C-3)]. Compound 2had similar structure with 3b,26-Dihydroxy-8,24E-
euphadien-11-one, which was previously isolated from Cassipourea lanceolata (Hou
et al. 2010). However, some chemical shifts of protons and carbons are different; sug-
gesting that compound 2had different stereochemistry at C-13, C-14 and C-17. Detail
analysis of its NOESY spectrum, H-3 was established as a-axial due to a big coupling
constant (J¼11.0, 5.0 Hz) due to being splitted by H-2a and H-2e, then 3-OH is
b-equatorial. In addition, H-5 was coupled to H-3, H-28 suggesting both of them are
a-face. The stereochemistry of H-18 and H-20 are b-face and H-30 is a-face due to
NOE correlations between i) H-12a-axial and H-17, H-30; ii) H-12b-equatorial and H-21.
Thus, compound 2possessed a lanostane triterpenoid skeleton (Huang et al. 2017).
Consequently, endertiin B (2) was found to be (24E)-3b,26-dihydroxy-5a-lanosta-11-
oxo-8,24-diene as shown in Figure 1.
In addition, two known compounds were determined as ganodecalone A (3) which
was purified from the fruiting body of Ganoderma calidophilum (Huang et al. 2017)
and ergosterol (4) (Quang et al. 2011; Phuc et al. 2014). Compound 4is a typical sec-
ondary metabolite which was found in almost all wood-rooting mushrooms, especially
in lingzhi mushrooms (Ganodermataceae) (Tham 2005; Quang et al. 2011; Phuc
et al. 2014).
Previously, ganodecalone A (3) and ergosterol (4) exhibited good anti-cancer activ-
ity against several cancer cells (Phuc et al. 2014; Kikuchi et al. 1985). Endertiins A and
B(1and 2) were tested their cytotoxic activity toward Lu and MCF-7 cell lines.
Accordingly, endertiin A (1) could inhibit the growth of MCF-7 cells (IC
50
value:
71.16 ± 6.25 mg/ml).
3. Experimental
3.1. General experimental procedures
Thin layer chromatography was carried out on pre-coated thin sheet Kieselgel 60 F254
of Merck, Germany. The spots were detected by ultraviolet lights with three wave-
lengths at 254, 302 and 366 nm or used a reagent of 10% H
2
SO
4
solution and heated
slowly until the color appeared. Column chromatography was carried out on a silica
gel (particle size 60-100 mM, Merck) and Sephadex LH-20 (GE Healthcare Life Sciences)
column chromatography. Nuclear magnetic resonance spectroscopy (NMR) was
NATURAL PRODUCT RESEARCH 3

measured on a Bruker AMX-500 (500 MHz for
1
H-NMR spectrum and 125 MHz for
13
C-
NMR spectrum) with an internal standard TMS. The chemical shifts (d) are expressed in
parts per million (ppm). HR-MS was recorded on a SCIEX X500 QTOF system.
3.2. Mushroom material
The fruit bodies of H. endertii Stey was cultivated in the laboratory as described in
(Tham et al. 2018). The sample was identified by Le Xuan Tham, Hongbang
International University. The specimen materials (LDL191) were deposited at Faculty of
Chemistry, Hanoi National University of Education, Hanoi, Vietnam.
3.3. Extraction and isolation
The powder of air-dried fruit bodies of H.endertii (600 g) were extracted in methanol
in a ultrasonic bath for 3 times, each 45 min, after filtration and all solvent removed
by rotary evaporator to give a crude extract (41.4 g), which was then chromato-
graphed on silica gel column, using hexane/EtOAc (10/1) to 100% EtOAc to afford 13
sub-fractions (1-13). Sub-fraction 3 (300 mg) was purified by silica gel column, solvent
system hexane/EtOAc (7/1, v/v) to obtain compound 4(23.9 mg). Sub-fraction 4
(185.0 mg) was a compound 1as a white crystal. Sub-fraction 7 (790 mg) was isolated
by silica gel column, using hexane/EtOAc (5/1, v/v) to give compound 3(72.8 mg).
Compound 2(5.0 mg) was obtained from sub-fraction 8 (332 mg) by silica gel column,
hexane/EtOAc (3/1, v/v) as a solvent system.
3.3.1. Endertiin A (1)
[a]20D þ64.0

(c0.10, CH
3
OH); IR (KBr) t
max
2928, 2859, 1709, 1655, 1589, 1462, 1381,
1281, 1177 cm
1
;
1
H-NMR (500 MHz, CDCl
3
)d
H
¼9.40 (s, H-26), 6.48 (t, J¼7.0 Hz, H-
24), 3.03 (m, H-1), 2.66 (d, J¼17.0 Hz, H-12a-axial), 2.52 (m, H-2), 2.51 (d, J¼17.0 Hz,
H-12b-equatorial), 2.46 (m, H-2), 2.45 (m, H-22), 2.42 (m, H-7), 2.40 (m, H-23), 2.38 (m,
H-7), 2.37 (m, H-23), 2.02 (m, H-16), 1.82 (m, H-15), 1.75 (s, H-27), 1.77 (m H-17), 1.67
(d, J¼11.5 Hz, H-5), 1.66 (m, H-6), 1.62 (m, H-1), 1.51 (m, H-6), 1.48 (m, H-20), 1.46 (m,
H-16), 1.32 (m, H-15), 1.28 (m, H-22), 1.15 (s, H-19, H-30), 1.12 (s, H-29), 1.08 (s, H-28),
0.95 (d, J¼6.5 Hz, H-21), 0.86 (s, H-18);
13
C-NMR (125 MHz, CDCl
3
)d
C
¼218.2 (C-3),
198.9 (C-11), 195.3 (C-26), 164.2 (C-8), 154.8 (C-24), 139.3 (C-9), 138.2 (C-25), 51.7 (C-5),
51.6 (C-14), 51.5 (C-12), 50.1 (C-17), 47.0 (C-13), 46.8 (C-4), 37.0 (C-10), 36.1 (C-20), 35.1
(C-1), 34.5 (C-22), 34.2 (C-2), 31.1 (C-15), 29.2 (C-7), 27.7 (C-29), 27.1 (C-16), 26.0 (C-23),
25.8 (C-30), 20.6 (C-28), 19.0 (C-19), 18.7 (C-6), 18.2 (C-21), 16.9 (C-18), 9.2 (C-27). HR-
MS: m/z 453.3355 [M þH]
þ
, calcd. for C
30
H
45
O
3
, 453.3369.
3.3.2. Endertiin B (2)
[a]20D 13.0

(c0.10, CH
3
OH); IR (KBr) t
max
3406, 2955, 2928, 2866, 1647, 1582, 1454,
1373, 1289, 1026 cm
1
;
1
H-NMR (500 MHz, CDCl
3
)d
H
¼5.39 (dt, J¼1.0, 7.0 Hz, H-24),
4.00 (s, H-26), 3.24 (dd, J¼5.0, 11.0 Hz, H-3), 3.01 (td, J¼3.5, 13.6 Hz, H-1), 2.65 (d,
J¼16.5 Hz, H-12), 2.47 (d, J¼16.5 Hz, H-12), 2.36 (dd, J¼5.5, 10.5 Hz, H-7), 2.27 (dd,
J¼7.0, 10.5 Hz, H-7), 2.10 (m, H-23), 1.99 (m, H-16), 1.95 (m, H-23), 1.77 (m, H-15), 1.73
4 D. N. QUANG ET AL.

(m, H-22), 1.72 (m, H-6, H-17), 1.71 (m, H-2), 1.66 (s, H-27), 1.62 (m, H-2), 1.46 (m, H-6),
1.36 (m, H-20), 1.35 (m, H-15), 1.34 (m, H-16), 1.13 (s, H-19), 1.12 (s, H-30), 1.07 (dd,
J¼4.5, 13.6 Hz, H-1), 1.06 (m, H-22), 1.02 (s, H-28), 0.91 (m, H-5), 0.90 (d, J¼6.5 Hz, H-
21), 0.83 (s, H-18, H-29);
13
C-NMR (125 MHz, CDCl
3
)d
C
¼199.4 (C-11), 164.3 (C-8), 139.6
(C-9), 134.6 (C-25), 126.5 (C-24), 78.7 (C-3), 68.9 (C-26), 51.9 (C-5), 51.8 (C-12), 51.6 (C-
14), 50.1 (C-17), 47.3 (C-13), 39.0 (C-4), 37.7 (C-10), 36.0 (C-20), 35.7 (C-22), 34.4 (C-1),
31.0 (C-15), 29.9 (C-7), 28.0 (C-2), 28.3 (C-28), 27.0 (C-16), 25.7 (C-30), 24.4 (C-23), 19.0
(C-19), 18.3 (C-21), 17.4 (C-6), 16.7 (C-18), 15.7 (C-29), 13.6 (C-27). HR-MS: m/z 457.3646
[M þH]
þ
, calcd. for C
30
H
49
O
3
457.3682.
3.4. Biological activity
Compounds 1and 2were tested for their cytotoxicity toward two cancer cells (Lu and
MCF-7) by the method described in (Scudiero et al. 1988).
4. Conclusion
Two new lanostane triterpenoids endertiins A-B (1,2) together with two known com-
pounds, ganodecalone A (3) and ergosterol (4) were purified from the cultivated
lingzhi Humphreya endertii in Vietnam. In addition, endertiin A (1) showed the cyto-
toxic activity against MCF-7 cells with IC
50
value of 71.16 ± 6.25 mg/ml. This is the first
report on the isolation, structural elucidation and anti-cancer activity of secondary
metabolites from Humphreya endertii.
Disclosure statement
No potential conflict of interest was reported by the authors.
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