Aromatic glycosides from Eulophia andamanensis

Article abstract of DOI:10.1016/j.phytol.2021.01.008

Two new phenolic glycosides, eulophiosides A and B (3, 4), were isolated from Eulophia andamanensis in addition to six known compounds. The known compounds were identified as gastrodin (1), vitexnegheteroin A (2), grammatophylloside A (5), grammatophylloside B (6), pleionoside E (7), and pleionoside F (8). Their structures were determined based on the physical data and the spectroscopic evidence including 1D and 2D NMR experiments.

Full text of DOI:10.1016/j.phytol.2021.01.008

Phytochemistry Letters 42 (2021) 24–26
Contents lists available at ScienceDirect
Phytochemistry Letters
journal homepage: www.elsevier.com/locate/phytol
Aromatic glycosides from Eulophia andamanensis
Jedsada Maliwonga, Nitirat Chimnoia, Wassapol Thamniyoma, Somsak Ruchirawata,
,b,
Tripetch Kanchanapooma
*
a Chulabhorn Research Institute, Kamphaeng Phet 6, Talat Bang Khen, Lak Si, Bangkok 10210, Thailand
b Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen 40002, Thailand
A R T I C L E I N F O
A B S T R A C T
Keywords:
Two new phenolic glycosides, eulophiosides A and B (3, 4), were isolated from Eulophia andamanensis in addition
to six known compounds. The known compounds were identified as gastrodin (1), vitexnegheteroin A (2),
grammatophylloside A (5), grammatophylloside B (6), pleionoside E (7), and pleionoside F (8). Their structures
were determined based on the physical data and the spectroscopic evidence including 1D and 2D NMR
experiments.
Eulophia andamanensis
Orchidaceae
Aromatic glucoside
Glucosyloxybenzyl derivatives
Eulophiosides A-B
1. Introduction
formula, C20H22O9, was determined by using HR-ESI-TOF-MS (m/z:
405.1184 [Mꢀ H]ꢀ ). The 1H NMR spectroscopic data (Table 1) showed
Eulophia andamanensis Rchb.f. (Thai name: Chang-Pa-Som-Khong) is
a species of the family Orchidaceae, distributed in India and Southeast
Asia. In Thai traditional medicine, the dried pseudobulbs are externally
used to treat wounds for antiseptic purposes. The phytochemical
investigation of this species has not been carried out. However, some
species of Eulophia were reported to contain phenanthrenes (Tuchinda
et al., 1998, 1989; Blitzke et al., 2000; Temkitthawon et al., 2017). In
this article, we described the isolation and structural identification of
four simple aromatic glycosides (1–4, Fig. 1), of which compounds 3 and
4 were new, in addition to four known glucosyloxybenzyl succinate
derivatives (5–8) from the n-BuOH soluble fraction of this plant.
the presence of two sets of 1,4-disubstituted aromatic rings from the
chemical shifts at δH 7.16 and 7.04 (each 2H, d, J =8.5 Hz), and δH 7.90
and 6.85 (each 2H, d, J =8.7 Hz); a hydroxymethyl group at δδH 4.50
(2H, s) in addition to an anomeric proton signals at δ 4.91 (1H, d, J
H
=7.3 Hz) for a β-D-glucopananosyl moiety. Acid hydrolysis of 3 afforded
D-glucose. In the 13C NMR spectrum, the chemical shifts were similar to
those of gastrodin (1), except for the presence of the additional signals at
δC 116.2 (2C), 122.0 (1C), 132.9 (2C), 163.5 (1C) and 167.8 (1C). These
carbons belonged to the 4-hydroxybenzoyl moiety, related to the
structure part of vitexnegheteroin A (2) (Table 2). Comparison of the
chemical shifts of 3 with those of 1 indicated that the 4-hydroxybenzoyl
moiety was connected to C-6′ of the glucopyranosyl unit since this car-
2. Results and discussion
bon atom appeared downfield at δ 65.0. Also, the chemical shifts of H2-
C
6′ of the glucopyranosyl moiety were observed downfield at δH 4.65 (1H,
dd, J = 11.8, 1.8 Hz) and 4.35 (1H, dd, J =7.6 Hz). The assignment was
confirmed by the HMBC experiment, in which the correlation was
observed from H-6′ (δ 4.68 and 4.35) to C"-7 (δ 167.8) as shown in
The methanolic extract of the leaves of E. andamanensis was sus-
pended in water and partitioned with Et2O and n-BuOH. The n-BuOH
fraction fraction was separated by combination of chromatographic
methods to obtain two new aromatic glycosides (3, 4), and six known
compounds. The known compounds were identified as gastrodin (1)
(Sahakitpicham et al., 2013), vitexnegheteroin A (2) (Hu et al., 2015),
grammatophylloside A (5), grammatophylloside B (6) (Sahakitpicham
et al., 2013), pleionoside E (7), and pleionoside F (8) (Han et al., 2019)
by comparison of physical data with literature values and from spec-
troscopic evidence.
H
C
Fig. 2. Therefore, this compound was identified as 6′-O-4-hydrox-
ybenzoyl-gastrodin, and named eulophioside A.
Compound 4 was isolated as amorphous powder. The molecular
formula, C26H32O14, was determined by using HR-ESI-TOF-MS (m/z:
567.1705 [Mꢀ H]ꢀ ). Inspection of the 1H NMR and 13C NMR spectro-
scopic data (Tables 1 and 2) indicated that this compound was a de-
rivative compound of 1-3. In addition, the 13C NMR signals arising from
a β-D-glucopyranosyl moiety (δC 103.0, 75.1, 78.1, 71.7, 78.0 and 62.8)
Compound 3 was isolated as amorphous powder, and the molecular
* Corresponding author at: Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen 40002, Thailand.
E-mail address: trikan@kku.ac.th (T. Kanchanapoom).
https://doi.org/10.1016/j.phytol.2021.01.008
Received 2 December 2020; Received in revised form 21 January 2021; Accepted 25 January 2021
Available online 3 February 2021
1874-3900/© 2021 Phytochemical Society of Europe. Published by Elsevier Ltd. All rights reserved.

J. Maliwong et al.
Phytochemistry Letters 42 (2021) 24–26
Table 2
13C NMR spectroscopic data of compounds 1-4 (100 MHz for 13C NMR, in
CD3OD).
Position
1
2
3
4
Aglycone
1
136.5
129.4
117.5
158.3
117.5
129.4
64.7
137.7
112.5
150.8
147.0
117.8
120.5
64.9
136.5
129.3
117.6
158.1
117.6
129.3
64.8
132.9
130.6
117.5
158.5
117.5
130.6
71.3
2
3
4
5
6
7
Glc
1’
102.2
74.8
77.9
71.2
77.8
62.3
102.7
74.9
77.8
72.0
75.6
64.9
102.1
74.8
77.8
71.9
75.5
65.0
102.1
75.0
77.9
72.0
75.6
65.0
2’
3’
4’
5’
6’
Ester moiety
1"
122.1
133.0
116.2
163.7
167.9
122.0
132.9
116.2
163.5
167.8
122.1
133.0
116.3
163.7
167.9
2", 6"
3", 5"
4"
7"
Glc
1"’
2"’
3"’
4"’
5"’
6"’
103.0
75.1
78.1
71.7
78.0
62.8
Fig. 1. Structures of compounds 1-4.
Table 1
1H NMR spectroscopic data of compounds 3 and 4 (400 MHz for 1H NMR, in
CD3OD).
Position
3
4
2, 6
3, 5
7.16 (2H, d, J =8.5 Hz)
7.04 (2H, d, J =8.5 Hz)
7.24 (2H, d, J =8.6 Hz)
7.04 (2H, d, J =8.6 Hz)
4.85 (1H, d, J =11.5 Hz)
4.58 (1H, d, J =11.5 Hz)
7
4.50 (2H, s)
Glc
1’
4.91 (1H, d, J = 7.3 Hz)
3.51 (1H, dd, J = 9.2, 7.3 Hz)
3.53 (1H, dd, J = 9.2, 8.9 Hz)
3.44 (1H, dd, J = 9.4, 8.9 Hz)
3.78 (1H, ddd, J = 9.4, 7.6,
1.8 Hz)
4.92 (1H, d, J = 7.3 Hz)
3.50 (1 H)a
2’
3’
3.51 (1 H)a
4’
3.42 (1H, dd, J = 9.5, 8.9 Hz)
3.78 (1H, ddd, J = 9.5, 7.6,
2.1 Hz)
5’
4.68 (1H, dd, J = 11.8, 1.8 Hz)
4.35 (1H, dd, J = 11.8, 7.6)
4.68 (1H, dd, J = 11.8, 2.1 Hz)
4.35 (1H, dd, J = 11.8, 7.6)
6’
Ester
moiety
2", 6"
3", 5"
Glc
Fig. 2. HMBC correlations of compounds 3 and 4.
7.90 (2H, d, J =8.7 Hz)
6.85 (2H, d, J =8.7 Hz)
7.90 (2H, d, J =8.8 Hz)
6.87 (2H, d, J =8.8 Hz)
secondary metabolites obtaining from members of the subfamily Epi-
dendroideae of the orchid family, such as compounds 1, 5, and 6 from
Grammatophyllum speciosum; and compounds 5–8 from Pleione bulboco-
dioides (Mochizuki et al., 1992; Simmler et al., 2011; Yoshikawa et al.,
2013, 2014; Sahakitpicham et al., 2013; Han et al., 2019; Auberona
et al., 2019; Liu et al., 2019). From the chemotaxonomical point of view,
the occurrence of these specific constituents provided further confir-
mation of the typical profile of the secondary metabolites found in this
subfamily.
1"’
4.33 (1H, d, J =7.8 Hz)
3.23 (1H, dd, J = 9.0, 7.8 Hz)
3.28 (1 H)a
2"’
3"’
4"’
3.29 (1 H)a
5"’
3.32 (1H, m)
3.91 (1H, dd, J = 12.0, 2.1 Hz)
3.70 (1H, dd, J = 12.0, 5.6)
6"’
a
Signal was assigned by HSQC.
were observed in 4, as compared to 3. The additional glucopyranosyl
unit was assigned to be attached at C-7 (δC 71.3) of the aglycone moiety
because of the glycosylation shift effect to this carbon atom (Kasai et al.,
1977). Moreover, the HMBC spectrum provided further confirmation
with a significant correlation from H-7 (δH 4.85 and 4.58, each d, J
=11.5 Hz) of the aglycone moiety to C-1"’ (δC 103.0) of the additional
sugar unit. Besides, acid hydrolysis of 4 gave D-glucose. Consequently,
the structure of compound 4 was assigned as shown, named eulophio-
side B.
3. Experimental
3.1. General procedures
NMR spectra were recorded in CD3OD using Bruker AV-400 spec-
trometer. The MS data was obtained from the Bruker Micro TOF-LC mass
spectrometer. Optical rotations were measured using a Jasco P-1020
digital polarimeter. SiliaFlash® P60 (230–400 mesh, SiliCycle Inc.), and
RP-18 (50 μm, YMC) were used to create a column chromatography.
The phytochemical investigation of E. andamanensis isolated eight
compounds (1-8), which contained the glucosyloxybenzyl moiety as
part of the structure. These results were closely related to those of the
HPLC (Jasco PU-980 pump) was carried out for ODS columns (column
20 mm i.d. × 250 mm length, YMC ODS-AQ) with a Jasco UV-970
25

J. Maliwong et al.
Phytochemistry Letters 42 (2021) 24–26
detector at 210 nm and flow rates were 6 mL/min. The TLC spraying
reagent was 10 % H2SO4 in H2O-EtOH (1:1, v/v).
heated at 80 ◦C for 5 h. After cooling, the reaction was diluted with H2O
and extracted with EtOAc. The aqueous layer was neutralized with 2 N
KOH and concentrated to dryness affording the sugar fraction. This part
was dissolved with H2O (1 mL), analyzed by HPLC (Jasco OR-2090 plus
chiral detector; VertisepTM sugar LMP, 7.8 mm x 300 mm i.d.; mobile
phase: H2O; flow rate 0.4 mL/min; temperature: 80 ◦C) and comparison
with their retention times and optical rotations with an authentic sam-
ple. Hydrolysis of compounds 3 (2.1 mg) and 4 (0.8 mg) gave peaks
corresponding to D-glucose at 19.3 min with positive optical rotation.
3.2. Plant material
The whole plants of E. andamanensis Rchb.f. were collected from
Phitsanulok Province, Thailand, in July 2020. Plant specimen was
identified by one researcher (TK). Voucher specimens (TK-PSKKU-0094)
are deposited at the Herbarium of the Faculty of Pharmaceutical Sci-
ences, Khon Kaen University.
Declaration of Competing Interest
The authors report no declarations of interest.
Acknowledgements
3.3. Extraction and isolation
The air-dried whole plants of E. andamanensis (5.8 kg) were extrac-
ted three times with MeOH, and the extracted solution was then
concentrated to dryness. The residue (196.1 g) was suspended in H2O
and partitioned with Et2O and n-BuOH. The n-BuOH part (13.1 g) was
applied to a silica gel column using solvent systems of EtOAc (2.0 L);
EtOAc-MeOH (9:1, 6.0 L); EtOAc-MeOH-H2O (40:10:1, 4.0 L); EtOAc-
MeOH-H2O (70:30:3, 2.0 L); and EtOAc-MeOH-H2O (6:4:1, 2.0 L),
respectively to produce seven fractions (A to G). Fraction C (170.8 mg)
was applied to a RP-18 column using a gradient solvent system, H2O-
MeOH (90:10 → 20:80, v/v) to yield compound 3 (74.6 mg). Fraction D
(1.25 g) was separated on a RP-18 column using solvent system, H2O-
MeOH (90:10 → 20:80, v/v) to provide seven sub-fractions. Compound
1 (192.7 mg) was precipitated from sub-fraction D-1. Sub-fraction D-4
was purified by preparative HPLC-ODS with solvent system H2O-MeCN
(80:20, v/v) to afford compound 2 (7.9 mg). Fraction E (2.8 g) was
applied to a RP-18 column using a gradient solvent system, H2O-MeOH
(90:10 → 20:80, v/v) to provide eight sub-fractions. Sub-fraction E-3
was purified by preparative HPLC-ODS using solvent system H2O-MeCN
(85:15, v/v) to provide compounds 4 (0.8 mg) and 8 (20.4 mg). Sub-
fraction E-6 was purified by preparative HPLC-ODS using solvent sys-
tem H2O-MeCN (80:20, v/v) to produce compound 6 (40.2 mg). Frac-
tion F (1.78 g) was applies to a RP-18 column using a gradient solvent
system, H2O-MeOH (90:10 → 20:80, v/v) to provide eight sub-fractions.
Sub-fraction F-3 was purified by preparative HPLC-ODS using solvent
system H2O-MeCN (85:15, v/v) to afford compound 5 (1.2 mg). Finally,
sub-fraction F-5 was purified by preparative HPLC-ODS using solvent
system H2O-MeCN (80:20, v/v) to obtain compound 7 (14.4 mg).
This research project was supported by the grant from Chulabhorn
Research Institute, and Khon Kaen University.
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26
Amorphous powder,
[
α
]
ꢀ 35.2 (CH3OH, c 1.17); 1H NMR
D
(CD3OD): Table 1; 13C NMR (CD3OD): Table 2; Negative HRESITOFMS,
m/z: 405.1184 [Mꢀ H]ꢀ (calcd for C20H21O9, 405.1191).
3.5. Eulophioside B (4)
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constituents of Cymbidium Great Flower Marie Laurencin and their antioxidative
activity. J. Nat. Med. 67, 217–221.
26
Amorphous powder,
[α
]
ꢀ 80.1 (CH3OH, c 0.08); 1H NMR
D
(CD3OD): Table 1; 13C NMR (CD3OD): Table 2; Negative HRESITOFMS,
Yoshikawa, K., Okahuji, M., Iseki, K., Ito, T., Asakawa, Y., Kawano, S., Hashimoto, T.,
2014. Two novel aromatic glucosides, marylaurencinosides D and E, from the fresh
flowers of Cymbidium Great Flower ‘Marylaurencin’ Kazuko. J. Nat. Med. 68,
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m/z: 567.1705 [Mꢀ H]ꢀ (calcd for C26H31O14, 567.1719).
3.6. Determination of the absolute configuration of sugar
Each compound was dissolved in 2 N HCl–dioxane (6:1, 2.0 mL) and
26