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Chemical Analysis of the Edible Mushroom Tricholoma populinum:
Steroids and Sulfinyladenosine Compounds
Bernadett Kovácsa, Zoltán Bénib, Miklós Dékányb, Orsolya Orbán-Gyapaia, Izabella Sinkac,
István Zupkóc, Judit Hohmanna,d*and Attila Ványolósa*
aDepartment of Pharmacognosy, University of Szeged, Eötvös u. 6, 6720 Szeged, Hungary
bSpectroscopic Research, Gedeon Richter Plc., Gyömrői út 19–21, H-1103 Budapest, Hungary
cDepartment of Pharmacodynamics and Biopharmacy, University of Szeged, Eötvös u. 6, 6720 Szeged, Hungary
dInterdisciplinary Centre for Natural Products, University of Szeged, Eötvös u. 6, H-6720 Szeged, Hungary
hohmann@pharm.u-szeged.hu; vanyolosa@pharmacognosy.hu
Received: March 20th, 2017; Accepted: August 3rd, 2017
Ten compounds have been identified in the methanol extract of cottonwood mushroom (Tricholoma populinum J.E. Lange), 9 of them for the first time in this
species. Besides adenosine (8) and nicotinamide (7) the isolated compounds were ergostane type steroids (1-6) and rare sulfinyladenosine constituents (9 and
10). The chemical structures of these compounds were elucidated by means of extensive spectroscopic methods (NMR and MS). Compounds 3-6 were
evaluated for their potential antiproliferative activity against human cancer cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide
(MTT) assay. The xanthine oxidase (XO) inhibitory activity of 7-10 has been examined by spectrophotometric method. Cerevisterol (3), its methylated
derivative (5) and 3-glycoside of ergosterol peroxide (6) showed significant antiproliferative activity on human breast cancer cell lines.
Keywords: Mushroom, Tricholoma populinum, Antiproliferative, Sulfinyladenosine, Steroids.
For centuries Basidiomycetes mushrooms have been used as food
material and also for their various beneficial properties by several
traditional medicines around the world. Nowadays mushrooms are
recognized as functional foods with high nutritional value,
appreciable vitamin and mineral content, favorable essential amino
acid profile; furthermore there is a rising interest in exploring the
biologically active compounds of mushrooms, since they represent
a rich source of structurally diverse metabolites with potential
medical application [1].
In our ongoing research program, aiming the evaluation of
Hungarian macrofungi in terms of their chemical composition and
pharmacological activity, extracts of cottonwood mushroom
(Tricholoma populinum J.E. Lange) were found to exhibit XO
inhibitory property [2] and antiproliferative activity against several
human cancer cell lines [3]. T. populinum, member of the fairly
large genus of Tricholoma, is a less known relative of the
distinguished species of T. matsutake and T. magnivelare; it grows
on sandy soil under cottonwood (Populus sp.) trees near a source of
water. While T. matsutake and T. magnivelare are both highly
popular among consumers in Asia and America because of their
unique spicy odor and taste, T. populinum is less significant as food
material. Even if it has a lower culinary value than the above
mentioned species, T. populinum is however consumed in North
America by Salish Indian people from British Columbia [4] and the
Native American tribe of Taos Pueblo in New Mexico [5].
Furthermore, according to Lentini and Venza (2006) sporocarps of
cottonwood mushroom (“funci di chiuppu”) are eaten by locals in
Sicily, during the fall season [6]. Our literature survey on the
chemistry and pharmacology of cottonwood mushroom revealed
that little is known about this species; Kreisel et al. found that it
contains ergosterol peroxide, which demonstrated immuno-
suppressive activity [7]. The primary goals of the present study
were to explore the chemical profile of cottonwood mushroom and
to examine the pharmacological activity of the isolated compounds.
Figure 1: Structures of compounds 1-10 isolated from Tricholoma populinum.
The investigation of the methanol extract obtained from the
lyophilized fruiting bodies of cottonwood mushroom led to the
isolation of 10 compounds (Figure 1.). The fungal extract was
subjected to solvent-solvent partition between aqueous MeOH and
n-hexane, followed by extraction with chloroform. The obtained
n-hexane and chloroform extracts were purified using combined
chromatographic methods including column chromatography, flash
chromatography, rotational planar chromatography and reversed-
phase HPLC. Compounds 1 and 2 were identified by comparing
their chromatographic and spectroscopic data with those of
authentic samples. Compounds 3–10 were structurally characterized
on the basis of NMR and MS spectroscopic data and were
confirmed by comparing them to those reported earlier for these
compounds.
All compounds have been identified for the first time in T.
populinum, except nicotinamide (7), which has been detected by
Turner et al. [4] in cottonwood mushroom samples collected in
NPC Natural Product Communications 2017
Vol. 12
No. 10
1583 - 1584
1584 Natural Product Communications Vol. 12 (10) 2017 Kovács et al.
British Columbia (Canada). Among the isolated compounds six
constituents (1-6) belong to the group of triterpene steroids of
ergostane skeleton. Ergosterol (1) and 3β-hydroxyergosta-7,22-
diene (2) are common steroids with a widespread distribution
among fungal species, as well as cerevisterol (3), and 3β,5α,6β,9α-
tetrahydroxyergosta-7,22-diene (4), which are di- and trihydroxlated
derivatives of 3β-hydroxyergosta-7,22-diene (2). Compound 5, a
methylated derivative of 3, has been previously isolated from the
medicinal mushroom Agaricus blazei by Kawagishi et al. [8]. The
steroid derivative 6, identified by Takaishi et al. in Hericium
erinaceus is an ergosterol-3-glucoside, which has a peroxide
linkage between C5 and C8 [9]. Besides nicotinamide (7), the main
constituents of the chloroform extract proved to be the adenosine
type compounds of 8, 9 and 10.
Whilst adenosine (8) is commonly found in living organisms, the
methylsulfinyladenosine derivatives 9 and 10 represent a fairly rare
subclass of secondary metabolites. According to our literature
overview this type of compounds has only been detected in a single
fungal, a plant species, and two marine organisms so far.
Compounds 9 and 10 were first isolated as inseparable epimeric
mixture from Ganoderma lucidum, and have been characterized as
inhibitors of platelet aggregation [10]. In 1997 these compounds
were also identified in the ethanol extract of Sauropus androgynous,
a cultivated plant native to Southeast Asia [11]. The other known
sources of sulfinyladenosine compounds are the marine ascidians
Herdmania momus, which synthesizes four complex adenosides
with bromines in their structures [12], and Atriolum robustum,
which contains a metabolite with methoxyacrylic acid group linked
to the sulfinyladenosine moiety [13]. As compared to the findings
of Kawagishi et al. (1993) however, the fruiting bodies of T.
populinum contain approx. 80 times higher amount from these
sulfinyladenosines than G. lucidum [10].
The isolated steroids 3-6 were evaluated for their potential cytotoxic
activity by MTT method on three different human breast cancer cell
lines (Table 1). Compounds 3 and 6 proved to be the most active
against T47D cells, while the methylated derivative of cerevisterol
(5) demonstrated significant activity on MDA-MB-231 cells.
Table 1: Antiproliferative activity of compounds 3-6 on human cancer cell lines.
Compds. Cell growth inhibition (%) ± SEM
MCF7 cells T47D cells MDA-MB-231 cells
3 10 µM –* 38.4 ± 0.6 –
30 µM – 50.2 ± 1.6 –
4 10 µM – – –
30 µM – – –
5 10 µM – 20.1 ± 1.4 25.7 ± 1.5
30 µM – 23.7 ± 2.4 54.7 ± 1.6
6 10 µM – – –
30 µM 37.17 ± 2.49 46.0 ± 1.4 –
*Inhibition values less than 20% were considered unsubstantial and are not given numerically
for clarity
Among the three cell lines, MCF7 proved to be the least susceptible
against the isolated compounds; however 6 exerted moderate
activity on this cell line as well. Compounds 7-10 were evaluated
for their potential XO inhibitory activity, though our results indicate
that they do not possess inhibitory activity on this enzyme.
The present study is the first comprehensive chemical analysis of
the edible mushroom Tricholoma populinum. The detailed
investigation of the methanol extract of T. populinum led to the
isolation and identification of 10 compounds. Nine of these are
identified for the first time in this species. The low-molecule
chemical profile of cottonwood mushroom includes rare
sulfinyladenosine compounds (9 and 10) and ergostane type
triterpenoids (1-6). Biological evaluation of the isolated compounds
showed that cerevisterol (3) and its methylated derivative (5), as
well as 3-glycoside of ergosterol peroxide (6) have significant
antiproliferative activity on some human breast cancer cell lines.
Based on the results obtained the current study makes a valuable
contribution to the chemistry and pharmacology of the edible
mushroom T. populinum.
Supplementary data: The electronic supplementary material
contains information on isolation and structural determination of
compounds 3-10, and the biological assays performed.
Acknowledgments - The authors thank Éva Bata Hullmann
(Hungarian Mycological Society) for her help in the identification
and collection of mushroom material.
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