Catomentosaponin, a new triterpene saponin from the roots of Catunaregam tomentosa

Article abstract of DOI:10.1080/14786419.2017.1385009

A new triterpene saponin, catomentosaponin (1) and 11 known analogues (2–12) were isolated from the roots of Catunaregam tomentosa. The structures of 1–12 were determined on the basis of extensive NMR and MS data analysis. The sugar residues were identified by co-TLC and HPLC analysis after hydrolysis. All isolated compounds were evaluated for their cytotoxicity against KB and HeLa cell lines. Compound 2 showed moderate cytotoxicity against KB cell with IC₅₀ value of 24.84?μM.

Full text of DOI:10.1080/14786419.2017.1385009

Natural Product Research
Formerly Natural Product Letters
ISSN: 1478-6419 (Print) 1478-6427 (Online) Journal homepage: http://www.tandfonline.com/loi/gnpl20
Catomentosaponin, a new triterpene saponin from
the roots of Catunaregam tomentosa
Sutin Kaennakam, Kitiya Rassamee, Pongpun Siripong & Santi Tip-pyang
To cite this article: Sutin Kaennakam, Kitiya Rassamee, Pongpun Siripong & Santi Tip-pyang
(2017): Catomentosaponin, a new triterpene saponin from the roots of Catunaregam tomentosa,
Natural Product Research, DOI: 10.1080/14786419.2017.1385009
To link to this article: http://dx.doi.org/10.1080/14786419.2017.1385009
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Date: 19 October 2017, At: 06:33

Natural Product research, 2017
ꢁꢁpꢂ://ꢃꢄi.ꢄꢅg/10.1080/14786419.2017.1385009
ꢀ
Catomentosaponin, a new triterpene saponin from the roots
of Catunaregam tomentosa
a
b
b
a
Sutin Kaennakam , Kitiya Rassamee , Pongpun Siripong and Santi Tip-pyang
ꢆ
Fꢆꢇꢈꢉꢁy ꢄf sꢇiꢊnꢇꢊ, dꢊpꢆꢅꢁmꢊnꢁ ꢄf cꢀꢊmiꢂꢁꢅy, Nꢆꢁꢈꢅꢆꢉ Pꢅꢄꢃꢈꢇꢁꢂ rꢊꢂꢊꢆꢅꢇꢀ uniꢁ, cꢊnꢁꢊꢅ ꢄf exꢇꢊꢉꢉꢊnꢇꢊ in Nꢆꢁꢈꢅꢆꢉ
b
Pꢅꢄꢃꢈꢇꢁꢂ cꢀꢊmiꢂꢁꢅy, cꢀꢈꢉꢆꢉꢄngkꢄꢅn univꢊꢅꢂiꢁy, Bꢆngkꢄk, tꢀꢆiꢉꢆnꢃ; Nꢆꢁꢈꢅꢆꢉ Pꢅꢄꢃꢈꢇꢁꢂ rꢊꢂꢊꢆꢅꢇꢀ sꢊꢇꢁiꢄn,
rꢊꢂꢊꢆꢅꢇꢀ diviꢂiꢄn, Nꢆꢁiꢄnꢆꢉ cꢆnꢇꢊꢅ Inꢂꢁiꢁꢈꢁꢊ, Bꢆngkꢄk, tꢀꢆiꢉꢆnꢃ
ABSTRACT
ARTICLE HISTORY
A new triterpene saponin, catomentosaponin (1) and 11 known
analogues (2–12) were isolated from the roots of Catunaregam
tomentosa. The structures of 1–12 were determined on the basis
of extensive NMR and MS data analysis. The sugar residues were
identified by co-TLC and HPLC analysis after hydrolysis. All isolated
compounds were evaluated for their cytotoxicity against KB and HeLa
cell lines. Compound 2 showed moderate cytotoxicity against KB cell
with IC value of 24.84 μM.
rꢊꢇꢊivꢊꢃ 19 Jꢈnꢊ 2017
aꢇꢇꢊpꢁꢊꢃ 13 sꢊpꢁꢊmbꢊꢅ 2017
KEYWORDS
Catunaregam
tomentosa; rꢈbiꢆꢇꢊꢆꢊ;
ꢇꢆꢁꢄmꢊnꢁꢄꢂꢆpꢄnin;
ꢁꢅiꢁꢊꢅpꢊnꢊ ꢂꢆpꢄnin;
ꢇyꢁꢄꢁꢄxiꢇiꢁy
5
0
1
. Introduction
Catunaregam tomentosa (Blume ex DC.) Tirveng (synonymous with Randia tomentosa), also
known as ‘Khet’in Thai is a tree widely distributed in the north-east region of Thailand. The
fruits are used in Thai for washing the hair and clothes, assuredly because of the presence
of saponins (Kanchanapoom et al. 2002). In previous study, iridoid glycosides have been
reported from the leaves and branches of this plant (Kanchanapoom et al. 2002). The genus
Catunaregam (Rubiaceae) comprises 12 species distributed over tropical and subtropical
regions of Southeast Asia and Africa. This genus has interesting pharmacological activities,
such as anti-inflammatory, antispasmodic, antidysenteric, antifertility, immunomodulatory
CONTACT sꢆnꢁi tip-pyꢆng
sꢆnꢁi.ti@ꢇꢀꢈꢉꢆ.ꢆꢇ.ꢁꢀ
sꢈppꢉꢊmꢊnꢁꢆꢉ ꢃꢆꢁꢆ fꢄꢅ ꢁꢀiꢂ ꢆꢅꢁiꢇꢉꢊ ꢇꢆn bꢊ ꢆꢇꢇꢊꢂꢂꢊꢃ ꢆꢁ ꢀꢁꢁpꢂ://ꢃꢄi.ꢄꢅg/10.1080/14786419.2017.1385009.
©
2017 Infꢄꢅmꢆ uK limiꢁꢊꢃ, ꢁꢅꢆꢃing ꢆꢂ tꢆyꢉꢄꢅ & Fꢅꢆnꢇiꢂ Gꢅꢄꢈp

2
S. KAENNAKAM ET AL.
and nephrotoxicity properties (Li et al. 2015; Adikay and Sravanthi 2016). Previous phyto-
chemical investigation on this genus revealed the presence of triterpene saponins, iridoid
glucosides, coumarin glucosides and lignans (Gao et al. 2011). In this report, we discuss the
isolation and structural elucidation of a new triterpene saponin, catomentosaponin (1) and
eleven known analogues (2–12) from the methanol extract of the roots of C. tomentosa. The
cytotoxic activities against KB (human epidermoid carcinoma) and Hela S–3 (human cervix
adenocarcinoma) cell lines were evaluated for all isolated compounds.
2
. Results and discussion
The methanol extract of the dried roots (10.0 kg) of C. tomentosa was subjected to multiple
chromatographic steps over silica gel, and Sephadex LH-20. Twelve triterpene saponins were
obtained, including a new compound, catomentosaponin (1) and 11 known compounds,
araliasaponin V (2), araliasaponin IV (3), araliasaponin VI (4); (Miyase et al. 1996), taibaienoside
VI (5) (Kong and Min 1996), elatoside K (6), elatoside A (7), elatoside C (8); (Yoshikawa et al.
1995), quinovic acid 3-O-α-L-rhamnoside (9) (Asase et al. 2008), gongganoside B (10) (Ohashi
et al. 1994), cinchonaglycoside C (11) (Li et al. 2007) and 3-O-β-D-glucopyranosyl quinovic
acid-28-β-D-glucopyranosyl ester (12) (Aquino et al. 2001). The structures of 1–12 (Figure 1)
were identified by physical data analyses, including 1D and 2D NMR, and HRESIMS. The sugar
residues were identified by co-TLC and HPLC analyses after hydrolysis.
Figure 1. cꢀꢊmiꢇꢆꢉ ꢂꢁꢅꢈꢇꢁꢈꢅꢊꢂ ꢄf 1–12.

NATURAL PRODUCT RESEARCH
3
2
0
Catomentosaponin (1) was obtained as white amorphous powder with [ꢀ] + 117.8
D
(
c 0.90, MeOH). Its molecular formula was determined as C H O from the positive HRESIMS
47 76 18
+
[M + Na] ion peak at m/z 951.4915 (calcd. for C H O Na, 951.4929). The IR spectrum showed
47 76 18
−1
absorption bands at 3425, 1731 and 1643 cm , suggesting the presence of hydroxyl, carbonyl
and olefinic groups, respectively. The H and C NMR data of 1 exhibited signals for seven
1
13
methyl groups at δ 0.75 (3H, s, H-26), 0.85 (3H, s, H-24), 0.95 (3H, s, H-29), 0.96 (3H, s, H-25),
H
0
.96 (3H, s, H-30), 1.06 (3H, s, H-23) and 1.30 (3H, s, H-27), one olefinic proton at δ 5.33 (1H,
H
br s, H-12) with two typical olefinic carbon signals at δ 125.0 (C-12) and 144.2 (C-13), indicative
C
of an olean-12-ene skeleton. The spectroscopic data also showed signals for two oxymethines
at δ 3.16 (1H, dd, J = 5.0, 10.7 Hz, H-3) with δ 91.0 (C-3) and δ 3.28 (1H, d, J = 3.3 Hz, H-19)
H
C
H
with δ 82.4 (C-19), and one carbon of carbonyl group at δ 178.6 (C-28). The NOESY correla-
C
C
tions (Figure S8) between H-3 and H-23, and between H-19 and H-30 indicated the α-orien-
tation of H-3 and β-orientation of H-19. Thus, the aglycone was identified as
3
β,19α-dihydroxyolean-12-ene-28-oic acid (siaresinolic acid) (Barton et al. 1952). The presence
of three sugar residues was confirmed by the observation of three anomeric protons at δH
.43 (1H, d, J = 7.7 Hz, H-1′), δ 4.53 (1H, d, J = 7.4 Hz, H-1″) and δ 5.39 (1H, d, J = 8.1 Hz, H-1‴),
4
H
H
which were correlated in the HSQC spectrum with anomeric carbons at δ 105.2, 106.2 and
C
95.8, respectively. After acid hydrolysis, the sugar units were confirmed to be D-glucose and
D-xylose, which were identified by co-TLC and HPLC analysis. The configuration of glucose
and xylose glycosidic bonds were established as β-glucose and β-xylose by the coupling
constants (J = 7.4–8.1 Hz) and the NOESY correlations (Figure S8) between the anomeric pro-
ton at H-1′ (δ 4.43) with the proton at H-23 (δ 1.06), between the anomeric proton at H-1″
H
H
(
δ 4.53) with the proton at H-3″ (δ 3.36) and between the anomeric proton at H-1‴ (δ 5.39)
H
H
H
with the proton H-5‴ (δ 3.37). The HMBC correlations (Figure S8) between the anomeric
H
proton at H-1′ (δ 4.43) with the carbon at C-3 (δ 91.0), between the anomeric proton at H-1″
H
C
(
δ 4.53) with the carbon at C-2′ (δ 83.2) and C-5″ (δ 67.1), between the anomeric proton
H C C
at H-1‴ (δ 5.39) with the carbon at C-28 (δ 178.6), and between the oxymethine proton at
H
C
H-2′ (δ 3.44) with the carbons at C-1′ (δ 105.2) and C-1″ (δ 106.2) confirmed the β-D-xy-
H
C
C
lopyranosyl-(1→2)-β-D-glucopyranosyl chain was linked at C-3 and the β-D-glucopyranosyl
1
13
was substituted at C-28 of aglycone. The comparison of the H and C NMR spectroscopic
data of 1 showed similar resonances to the known compound, 3-O-β-D-xylopyranosyl-(1→2)-
β-D-glucopyranosyl-28-O-β-D-glucopyranosyl-3-β-olean-12-en-28-oic acid (Jayasinghe et al.
2002), apart from the lack of a hydroxyl group at C-19. On the basis of the above results, the
structure of compound 1 was identified as 3-O-[β-D-xylopyranosyl-(1→2)-β-D-glucopyranosyl]-
28-O-(β-D-glucopyranosyl)-3β,19α-dihydroxyolean-12-en-28-oic acid.
The in vitro cytotoxic activities of all compounds are shown in Table 1, compound 2
showed moderate cytotoxicity against KB cell with IC value of 24.84 μM, compounds 1, 5
5
0
and 7 showed weak cytotoxicity against KB cell with IC50 values in the range of 51.10–
5.77 μM, compounds 1, 2, 7, 8 and 9 showed weak cytotoxicity against HeLa S–3 cell with
IC values in the range of 44.99–77.24 μM, and other compounds were inactive.
5
5
0
3
. Experimental
3
.1. General experimental procedures
For structural elucidation:The IR spectra were measured on a Nicolet 6700 FT-IR spectrometer
using KBr discs. Optical rotations were detected by a Jasco P-1010 polarimeter. The NMR

4
S. KAENNAKAM ET AL.
Table 1. In vitro ꢇyꢁꢄꢁꢄxiꢇiꢁy ꢄf ꢇꢄmpꢄꢈnꢃꢂ 1–7 ꢆgꢆinꢂꢁ KB ꢆnꢃ hꢊlꢆ s-3 ꢇꢊꢉꢉ ꢉinꢊꢂ.
IC50 (μM)
IC50 (μM)
HeLa S-3
Compound
KB
HeLa S-3
Compound
KB
1
51.44
24.84
> 100
> 100
51.10
> 100
0.15
58.58
52.57
> 100
> 100
> 100
> 100
0.02
7
8
55.77
> 100
> 100
> 100
> 100
> 100
77.24
71.39
44.99
> 100
> 100
> 100
2
3
9
4
10
11
12
5
6
dꢄxꢄꢅꢈbiꢇin
Nꢄꢁꢊꢂ: Ic50 ≤ 10 = gꢄꢄꢃ ꢆꢇꢁiviꢁy, 10 < Ic50 ≤ 30 = mꢄꢃꢊꢅꢆꢁꢊ ꢆꢇꢁiviꢁy, Ic50 > 100 = inꢆꢇꢁivꢊ.
1
spectra were recorded on a Bruker 400 AVANCE spectrometer (400 MHz for H and 100 MHz
1
3
for C). The HRESIMS was obtained using a Bruker MICROTOF model mass spectrometer.
HPLC analysis were obtained using a Alltech System equipped with model 626 binary gra-
dient pumps, select TM degasser, 580 autosampler, 2000 ES evaporative light scattering
detector (Alltech) and peak simple chromatography data system software. For isolation:
Column chromatography was performed by silica gel 60 (0.063–0.200 mm), and Sephadex
LH-20 (25–100 μm, GE Healthcare). For bioassay: Microplate reader TECAN infinite M200PRO.
3
.2. Plant material
The roots of C. tomentosa were collected from Sahatsakhan district, Kalasin province,
Thailand, in January 2016. The plant material was identified by Ms. Suttira Khumkratok, a
botanist at the Walai Rukhavej Botanical Research Institute, Mahasarakham University and
a specimen retained as a reference (Khumkratok No. 6–12).
3
.3. Extraction and isolation
The air-dried roots of C. tomentosa (10.0 kg) were extracted with MeOH over a period of
days at room temperature, respectively (2 × 25 L). Removal of the solvent under reduced
3
pressure provided MeOH (85.5 g) crude extract. The MeOH crude extract was further sepa-
rated by column chromatography (CC) over silica gel (40 × 10 cm, 2.5 kg) and eluted with a
gradient of EtOAc-MeOH (100% EtOAc, 80, 60, 40, 20 and 100% MeOH each 5 L, respectively)
to give four fractions (A–D). Fraction A (15 g) was purified by silica gel CC (45 × 5 cm, 0.5 kg)
and eluted with 80 and 60% CH Cl -MeOH (each 10L, respectively) to give two subfraction
2
2
(A1 and A2). Subfraction A1 (3 g) was separated by Sephadex LH-20 column (50 × 5 cm,
1
50 g) eluted with 50% CH Cl -MeOH (2 L) to yield 2 (20 mg) and 10 (15 mg). Subfraction
2
2
A2 (5 g) was subjected to silica gel CC (40 × 4 cm, 0.4 kg) and eluted with 80% CH Cl -MeOH
2
2
(
(
5 L) to provide 9 (12 mg) and 11 (15 mg). Fraction B (10 g) was fractionated by silica gel CC
45 × 5 cm, 0.5 kg) and eluted with 80 and 60% CH Cl -MeOH (each 10L, respectively) to
2
2
afford two subfraction (B1 and B2). Compound 5 (10 mg) was purified from subfraction
B1(2 g) by Sephadex LH-20 column (50 × 5 cm, 150 g) eluted with 50% CH Cl -MeOH (2 L).
2
2
Subfraction B2 (3 g) was separated by Sephadex LH-20 column (50 × 5 cm, 150 g) eluted
with 50% CH Cl -MeOH (2 L) to obtain 1 (15 mg) and 4 (10 mg). Fraction C (5 g) was separated
2
2
by Sephadex LH-20 column (50 × 3 cm, 150 g) eluted with 50% CH Cl -MeOH (2 L) to yield
2
2
5
(15 mg) and 7 (12 mg). Fraction D (7 g) was chromatographed over silica gel (40 × 4 cm,

NATURAL PRODUCT RESEARCH
5
0
.4 kg) and eluted with 70 and 50% CH Cl -MeOH (each 10L, respectively) to provide two
2 2
subfractions (D1 and D2). Compound 4 (13 mg) was purified from subfraction D1 (2 g) by
Sephadex LH-20 column (50 × 3 cm, 150 g) eluted with 20% CH Cl -MeOH (2 L). Finally,
2
2
Subfraction D2 (5 g) was separated by silica gel CC (40 × 4 cm, 0.4 kg) and eluted with 70%
CH Cl -MeOH (3 L) to yield 6 (11 mg) and 8 (12 mg).
2
2
3
.3.1. Catomentosaponin (12)0
White amorphous powder;[ꢀ] + 117.8 (c 0.90, MeOH); IR (KBr) ν 3425, 1731, 1643 cm−1;
D
max
1
H NMR (400 MHz, in MeOH-d ) δ : 1.00 (1H, m, H-1), 1.61 (1H, m, H-1), 1.72 (1H, m, H-2), 1.93
4
H
(
1H, m, H-2), 3.16 (1H, dd, J = 5.0, 10.7 Hz, H-3), 0.79 (1H, m, H-5), 1.42 (1H, m, H-6), 1.56 (1H,
m, H-6), 1.31 (1H, m, H-7), 1.50 (1H, m, H-7), 1.73 (1H, m, H-9), 1.95 (2H, m, H-11), 5.33 (1H, br
s, H-12), 1.02 (1H, m, H-15), 1.73 (1H, m, H-15), 2.33 (2H, m, H-16), 3.06 (1H, br s, H-18), 3.28
(
1H, d, J = 3.3 Hz, H-19), 1.68 (2H, m, H-21), 1.67 (1H, m, H-22), 1.79 (1H, m, H-22), 1.06 (3H, s,
H-23), 0.85 (3H, s, H-24), 0.96 (3H, s, H-25), 0.75 (3H, s, H-26), 1.30 (3H, s, H-27), 0.95 (3H, s,
H-29), 0.96 (3H, s, H-30), 4.43 (1H, d, J = 7.7 Hz, H-1′), 3.44 (1H, m, H-2′), 3.37 (1H, m, H-3′), 3.47
(
1H, m, H-4′), 3.56 (1H, m, H-5′), 3.69 (1H, m, H-6′), 3.84 (1H, m, H-6′), 4.53 (1H, d, J = 7.4 Hz,
H-1″), 3.26 (1H, m, H-2″), 3.37 (1H, m, H-3″), 3.35 (1H, m, H-4″), 3.15 (1H, m, H-5″), 3.82 (1H, m,
H-5″), 5.39 (1H, d, J = 8.1 Hz, H-1‴), 3.34 (1H, m, H-2‴), 3.44 (1H, m, H-3‴), 3.37 (1H, m, H-4‴),
1
3
3
.37 (1H, m, H-5‴), 3.69 (1H, m, H-6‴), 3.84 (1H, m, H-6‴); C NMR (100 MHz, in MeOH-d ) δ :
4 C
39.7 (C-1), 27.1 (C-2), 91.0 (C-3), 40.9 (C-4), 57.2 (C-5), 19.4 (C-6), 33.9 (C-7), 40.3 (C-8), 49.4
(C-9), 38.0 (C-10), 24.8 (C-11), 125.0 (C-12), 144.2 (C-13), 42.6 (C-14), 29.5 (C-15), 28.4 (C-16),
4
1
7.1 (C-17), 45.0 (C-18), 82.4 (C-19), 35.9 (C-20), 29.4 (C-21), 33.2 (C-22), 28.2 (C-23), 16.5 (C-24),
5.9 (C-25), 17.8 (C-26), 25.0 (C-27), 178.6 (C-28), 28.6 (C-29), 25.2 (C-30), 105.2 (C-1′), 83.2
(C-2′), 78.2 (C-3′), 71.1 (C-4′), 78.6 (C-5′), 62.4 (C-6′), 106.2 (C-1″), 76.2 (C-2″), 78.2 (C-3″), 71.4
(
C-4″), 67.1 (C-5″), 95.8 (C-1‴), 73.9 (C-2‴), 77.7 (C-3‴), 71.1 (C-4‴), 77.4 (C-5‴), 62.7 (C-6‴);
+
positive HRESIMS m/z 951.4915 [M + Na] (calcd. for C H O Na, 951.4929).
4
7
76 18
3
.4. Acidic hydrolysis
A solution of catomentosaponin (1) (5 mg) in 2 M HCl (1 mL) was heated at reflux for 24 h.
The reaction mixture was neutralised with 2 M NaOH and extracted by partition with EtOAc
(5 × 1 mL). The sugar residues were identified by co-TLC (TLC Silica gel 60 F254) by comparison
with standard sugar. The solvent system was CH Cl -MeOH-H O (2:1:0.2), and spots were
2
2
2
visualised by spraying with EtOH-H SO -anisaldehyde (9:0.5:0.5, v/v), then heated at 150 °C.
2
4
The R values of D-glucose and D-xylose by TLC was 0.30 and 0.44, respectively. In addition,
f
the sugar were identified by HPLC analysis (column: lichrocart-NH 250 × 4.0 mm, carrier:
2
8
2% ACN in H O 1.5 mL/min, retention time: 8.13 and 9.25 min) in comparison with the
2
authentic D-glucose and D-xylose, respectively. Actually, the natural occurring of glucose
and xylose are in the D-isomer.
3
.5. Cytotoxicity assay
All isolated compounds (1–12) were subjected to cytotoxic evaluation against KB (human
epidermoid carcinoma) and HeLa S–3 (human cervical carcinoma) cell lines employing the
colorimetric method (Skehan et al. 1990), (Kongkathip et al. 2003). Doxorubicin was used as
the reference substance which exhibits activity against five cancer cell lines. The

6
S. KAENNAKAM ET AL.
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (Sigma Chemical Co., USA)
was dissolved in saline to make a 5 mg/mL stock solution. Cancer cells (3 × 103 cells) sus-
pended in 100 μg/wells of MEM medium containing 10% fetal calf serum (FCS, Gibco BRL,
Life Technologies, NY, USA) were seeded onto a 96-well culture plate (Costar, Corning
Incorporated, NY 14831, USA). After 24 h pre-incubation at 37 °C in a humidified atmosphere
of 5% CO /95% air to allow cellular attachment, various concentrations of test solution (10
2
μL/well) were added and these were then incubated for 48 h under the above conditions.
At the end of the incubation, 10 μL of tetrazolium reagent was added into each well followed
by further incubation at 37 °C for 4 h. The supernatant was decanted, and DMSO (100 μL/
well) was added to allow formosan solubilisation. The optical density (OD) of each well was
detected using a Microplate reader at 550 nm and for correction at 595 nm. Each determi-
nation represented the average mean of six replicates. The 50% inhibition concentration
(
IC value) was determined by curve fitting.
50
4
. Conclusion
The MeOH crude extract from the roots of C.tomentosa comprise a new catomentosaponin
1) and eleven known triterpene saponins (2–12). Compound 2 showed moderate cytotox-
icity against KB cell with IC values of 24.84 μM, compounds 1, 5 and 7 showed weak cyto-
(
5
0
toxicity against KB cell with IC values in the range of 51.10–55.77 μM, compounds 1, 2, 7,
5
0
8
and 9 showed weak cytotoxicity against HeLa S-3 cell with IC50 values in the range of
44.99–77.24 μM, and other compounds were inactive. Therefore, we believe that this plant
is an important source for the diverse structure of triterpene saponins and should be further
investigated for other biological activities.
Disclosure statement
No potential conflict of interest was reported by the authors.
Funding
The authors are grateful to the Graduate School of Chulalongkorn University for a Postdoctoral
Fellowship (Ratchadaphiseksomphot Endowment Fund) to SK. is thankful for financial support from
the National Research University Project of Thailand (WCU-58-013-FW). We also thank Ms. Suttira
Khumkratok, Walai Rukhavej Botanical Research Institute, Mahasarakham University, Mahasarakham
44000, Thailand for identification and deposition of the plant material.
References
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