Antibacterial ellagic Acid derivatives and other constituents from pancovia pedicellaris

Article abstract of DOI:10.1515/znb-2009-0913

Two new ellagic acid derivatives, named panconosides A (1)and B (2) were isolated from Pancovia pedicellaris together with eleven known compounds (3-13). The structures of 1 and 2,as well as those of the known compounds were established by spectroscopic m

Full text of DOI:10.1515/znb-2009-0913

Antibacterial Ellagic Acid Derivatives and Other Constituents from
Pancovia pedicellaris
Rene´ F. Soha, Jean K. Bankeua, Bruno N. Lentab,c, Brice M. Mba’ninga, Silve`re Ngouelaa,
Etienne Tsamoa, and Norbert Sewaldc
a Department of Organic Chemistry, Faculty of Science, TWAS Research Unit (TRU) of the
University of Yaounde´ I, P. O. Box 812, Yaounde´, Cameroon
b Department of Chemistry, Higher Teacher’s Training College, University of Yaounde´ I,
P. O. Box 47, Yaounde´, Cameroon
c Department of Chemistry, Organic and Bioorganic Chemistry, Bielefeld University,
P. O. Box 100131, 33501 Bielefeld, Germany
Reprint requests to Prof. Etienne Tsamo. E-mail: tsamo@cm.refer.org or Silve`re Ngouela.
E-mail: sngouela@yahoo.fr
Z. Naturforsch. 2009, 64b, 1070 – 1076; received June 16, 2009
Two new ellagic acid derivatives, named panconosides A (1) and B (2) were isolated from Pancovia
pedicellaris together with eleven known compounds (3 – 13). The structures of 1 and 2, as well as
those of the known compounds were established by spectroscopic methods and by comparison with
previously reported data. Compounds 1 and 2 were tested in vitro for their antibacterial potential
against six strains of microorganisms: Micrococcus luteus, Streptococcus ferus, Streptococcus minor,
Escherichia coli, Bacillus subtilis, and Pseudomonas agarici. They were found to exhibit moderate
antibacterial activity against all the tested strains compared to standard drugs.
Key words: Pancovia pedicellaris, Sapindaceae, Ellagic Acid Derivatives, Panconosides A and B,
Antibacterial Activity
Introduction
known compounds identified as 3,3ꢀ,4ꢀ-tri-O-methyl-
ellagic acid 4-O-β-D-glucopyranoside (3), allantoin
(4), L-quebrachitol (5), stigmasterol (6), umbelliferone
(7), scopoletin (8), 8ꢀ-epi-cleomiscosin A (9), 3-oxo-
tirucalla-7,24-dien-21-oic acid (10), benjaminamide
(11), 3-O-β-D-glucopyranosylstigmasterol (12), and
7-oxostigmasteryl-3-O-β-D-glucopyranoside (13).
Plants of Pancovia genus (Sapindaceae) are trees
or shrubs generally found in tropical or subtropical
regions. In Cameroon, they are used in traditional
medicine for the treatment of several ailments such
as skin diseases, dysentery and rheumatism [1]. Pre-
vious phytochemical studies on plants of this family
reported the presence of various classes of secondary
metabolites including flavonoids, coumarins, ellagic
acid, and saponins as major constituents [2 – 5]. Some
of these compounds were shown to exhibit interesting
pharmacological properties including antiplasmodial,
haemolytic and antioxidant activities [5 – 7]. Pancovia
pedicellaris Radlk & Gilg is a tree or shrub of about
5 – 10 m high, found in Central Africa [1]. No previ-
ous reports on the chemical constituents or biological
properties of this species have been reported. As part
of our ongoing search for bioactive metabolites from
Cameroonian plants, we recently investigated extracts
from the leaves and the stem bark of P. pedicellaris.
Results and Discussion
The air-dried, powdered leaves and stem bark of
P. pedicellaris were extracted separately with MeOH.
The crude extract from the leaves obtained after con-
centration was further extracted with n-hexane, and the
MeOH soluble fraction subjected to successive col-
umn chromatography over silica gel to yield five com-
pounds, including the two new ellagic acid derivatives,
i. e. the panconosides A (1) and B (2), 3,3ꢀ,4ꢀ-tri-O-
methylellagic acid 4-O-β-D-glucopyranoside (3) [8],
allantoin (4) [9], and L-quebrachitol (5) [10].
The crude MeOH extract obtained from the stem
In this paper, we report the results of these investiga- bark was extracted with EtOAc, and the EtOAc-
tions which led to two new ellagic acid derivatives, soluble fraction subjected to repeated column chro-
panconosides A (1) and B (2), together with eleven matography over silica gel. This process yielded
c
0932–0776 / 09 / 0900–1070 $ 06.00 ꢀ 2009 Verlag der Zeitschrift fu¨r Naturforschung, Tu¨bingen · http://znaturforsch.com
Unauthenticated
Download Date | 10/8/15 12:07 AM

R. F. Soh et al. · Antibacterial Ellagic Acid Derivatives
1071
Table 1. 1H (500 MHz) and 13C NMR (125 MHz) data for 1
and 2 in [D6]DMSO.
Position
1
2
δc
δH [m, J (Hz)]
δc
δH [m, J (Hz)]
1
2
3
4
112.8
141.7
142.2
152.1
112.6
141.7
142.4
151.6
5
6
112.6 7.90 (s)
114.2
112.2 7.83 (s)
114.2
7
158.6
113.1
141.6
141.3
154.8
108.0 7.63 (s)
113.3
158.6
113.1
141.6
141.3
154.8
108.0 7.65 (s)
113.3
1ꢀ
Fig. 1. Selected HMBC (→) and NOESY (↔) correlations
in compound 1.
2ꢀ
3ꢀ
4ꢀ
tives, suggesting that this compound has an ellagic acid
skeleton [18, 19]. The IR spectrum displayed charac-
teristic absorptions for hydroxy groups (3420 cm−1),
lactone functions (1748 cm−1), aromatic C=C groups
(1611 cm−1), and glycosidic C–O bonds (1085 cm−1).
The broad band-decoupled 13C NMR spectrum (Ta-
ble 1) of compound 1 displayed only 28 carbon signals
which were sorted by DEPT and HSQC techniques
into eight methines including three aromatic methine
carbons and five oxymethine carbons, one oxymethy-
lene, fifteen quaternary aromatic carbons including two
carbonyls of α, β-unsaturated lactone groups at δ =
158.6 and 158.8 [8], one ester carbonyl group at δ =
165.9 and three methoxy carbons at δ = 57.2, 61.8, and
62.2, respectively. The fact that only 28 carbon signals
were observed in the 13C NMR spectrum of 1 com-
pared to the 30 carbons of the molecular formula was
very significant and suggested that it possessed a unit
of symmetry. The 1H and 13C NMR spectra (Table 1)
also indicated the presence of a sugar residue. The sig-
nals at δ = 101.3, 78.0, 77.4, 72.0, 67.9, and 60.6 in
the 13C NMR spectrum suggested that the sugar moi-
ety in 1 was a glucopyranoside [8]. The coupling con-
stant between the anomeric protons H-1ꢀꢀ [δ = 5.40 (d,
J = 8.2 Hz)] and H-2ꢀꢀ [δ = 3.63 (m)] corresponded to
two trans diaxial protons, suggesting a chair confor-
mation of the carbohydrate moiety with the C-1ꢀꢀ and
C-2ꢀꢀ substituents in equatorial position and indicating
5ꢀ
6ꢀꢀ
7ꢀꢀ
1ꢀꢀ
2ꢀꢀ
3ꢀꢀ
4ꢀꢀ
5ꢀꢀ
6 a
6ꢀꢀb
1ꢀꢀꢀ
2ꢀꢀꢀ
3ꢀꢀꢀ
4ꢀꢀꢀ
5ꢀꢀꢀ
6ꢀꢀꢀ
7ꢀꢀꢀ
3-OMe
158.8
158.8
101.3 5.40 (d, J = 8.2)
72.0 3.63 (m)
78.0 5.17 (t, J = 9.4)
67.9 3.55 (m)
77.4 3.65 (br s)
99.0 5.46 (d, J = 7.5)
76.7 3.63 (m)
77.5 3.46 (m)
70.1 3.24 (t, J = 8.8)
77.7 3.56 (br t, J = 8.8)
60.6 3.73 (dd, J = 11.3; 5.1) 60.8 3.65 (m)
3.57 (m)
3.48 (m)
120.4
109.3 7.04 (s)
145.9
138.6
145.9
109.3 7.04 (s)
165.9
62.2 4.10 (s)
100.7 5.26 (d, J = 1.3)
70.9 3.37 (br s)
70.8 3.72 (m)
72.3 3.18 (t, J = 9.4)
69.0 3.70 (m)
18.5 1.11 (d, J = 6.3)
62.2 4.11 (s)
61.8 4.05 (s)
57.2 4.01 (s)
3ꢀ-OMe 61.8 4.06 (s)
4ꢀ-OMe 57.2 4.01 (s)
the known compounds stigmasterol (6) [11], um-
belliferone (7) [12], scopoletin (8) [13], 8ꢀ-epi-
cleomiscosin A (9) [14], 3-oxotirucalla-7,24-dien-
21-oic acid (10) [15], benjaminamide (11) [16], 3-
O-β-D-glucopyranosylstigmasterol (12) [11], and 7-
oxostigmasteryl-3-O-β-D-glucopyranoside (13) [17].
The known compounds 3 – 13 were identified by anal-
ysis of their MS, 1H and 13C NMR data as well as by
comparison with previously reported data and authen-
tic samples.
Compound 1 was obtained as beige crystals and
gave a positive reaction with ferric chloride, indicat-
ing its phenolic nature. It also responded positively
to the Molish test, suggesting that it was a glyco-
side. Its elemental composition C30H26O17, with 18
degrees of unsaturation, was deduced from its HRMS
((+)-ESI) which exhibited a pseudo-molecular ion
peak [M+Na]+ at m/z = 681.10664 (calculated m/z =
681.10991). Its UV spectrum (λmax = 255, 280 and
355 nm) was similar to that of ellagic acid deriva-
1
the presence of a β-glucoside. Its H NMR spectrum
(Table 1) showed also four aromatic protons as sin-
glets at δ = 7.90 (s, H-5), 7.63 (s, H-5ꢀ), and 7.04 (s,
H-2ꢀꢀ/H-6ꢀꢀ) and three methoxy groups (δ = 4.01, 4.06
and 4.10). The 28 carbon signals observed in the 13C
NMR of compound 1 include, besides glucopyranosyl
and methoxy carbons, those for the two 7-carbon units
of an ellagic acid skeleton and those for another highly
oxygenated phenyl ring attributed to a gallic acid moi-
ety with the carbonyl group at δ = 165.9 [20]. The
Unauthenticated
Download Date | 10/8/15 12:07 AM

1072
R. F. Soh et al. · Antibacterial Ellagic Acid Derivatives
ellagic acid moiety was confirmed by its HMBC cor- experiments into twelve methine groups including two
relations, in particular those from H-5 (δ = 7.90) to aromatic methine carbons and ten oxymethine carbons,
C-3, C-4, C-6 and C-7 and from H-5ꢀ (δ = 7.63) to one oxymethylene, twelve quaternary aromatic car-
C-3ꢀ, C-4ꢀ, C-6ꢀ and C-7ꢀ. The location of the methoxy bons, including two lactone carbons and six oxygen-
groups at C-3ꢀ, C-4ꢀ and C-3 of the ellagic acid skele- bearing carbons, and three methoxy groups as well as
ton was also deduced from correlations observed be- one methyl group at δ = 18.5 (Table 1). The NMR
tween the methoxy protons at δ = 4.01 (OMe-4ꢀ), 4.06 spectroscopic data of 2 were similar to those of com-
(OMe-3ꢀ) and 4.10 (OMe-3) and C-4ꢀ (δ = 154.8), C-3ꢀ pound 1, but lacked signals for a gallic acid moiety.
(δ = 141.3) and C-3 (δ = 142.2), respectively. The Its 1H NMR spectrum exhibited in addition to signals
presence of the gallic acid moiety was confirmed by for an ellagic acid moiety [δ = 7.83 (s, H-5) and 7.65
the two-proton singlet signal observed at δ = 7.04 in- (s, H-5ꢀ)], and a glucopyranosyl group, five oxygen-
dicating a 3, 4, 5-trihydroxy substitution pattern in one bearing methine protons, along with a doublet methyl
of the aromatic rings in 1. The aromatic methine H-2ꢀꢀꢀ proton signal [δ = 1.11 (d, J = 6.3 Hz)] indicating the
(δ = 7.04) showed no HMBC correlations with the el- presence of a 6-deoxy sugar. The second sugar unit
lagic acid skeleton, but with C-1ꢀꢀꢀ, C-3ꢀꢀꢀ, C-4ꢀꢀꢀ and was easily determined as rhamnose through analysis
C-7ꢀꢀꢀ, supporting the existence of the gallic acid unit. of chemical shifts and coupling constant patterns of its
To establish the connectivity between the ellagic acid proton signals (Table 1) which were connected by a
moiety and the glucose on one hand and between the 1H-1H COSY spectrum. In addition, the set of chemi-
sugar and the gallic acid moiety on the other hand, cal shifts observed at δ = 100.7, 70.9, 70.8, 72.3, 69.0,
use was made of observed HMBC correlations in com- and 18.5 (C-1ꢀꢀꢀ to C-6ꢀꢀꢀ) resembled the correspond-
pound 1 (Fig. 1). The correlation observed from the ing signals of substituted rhamnose glycosides [18].
anomeric proton H-1ꢀꢀ (δ = 5.40) to C-4 (δ = 152.1) The anomeric protons (H-1ꢀꢀ, H-1ꢀꢀꢀ) of the sugar units
clearly indicates that the glucose unit was linked to C-4 were observed at δ = 5.46 (d, J = 7.5 Hz, H-1ꢀꢀ) and
of the aglycon (ellagic acid skeleton). Those from the 5.26 (d, J = 1.3 Hz, H-1ꢀꢀꢀ) and attributed to glucose
proton H-3ꢀꢀ (δ = 5.17) of the glucose moiety to C-7ꢀꢀꢀ and rhamnose, respectively. The values of the coupling
(δ = 165.9, carbonyl group), C-2ꢀꢀ (δ = 72.0) and C-4ꢀꢀ constants suggest the β configuration for glucose [8]
(δ = 67.9) suggest that the galloyl unit was linked to and α for rhamnose [18]. The junction between the
the glucose moiety across C-3ꢀꢀ (δ = 78.0). Addition- rhamnose and glucose and between the glucose and the
ally, the location of the glucose substituent was further ellagic acid moiety were made using the correlations
confirmed by the observation of an NOE between the observed in the HMBC spectrum (Fig. 2). In particular,
anomeric proton H-1ꢀꢀ (δ = 5.40) and H-5 (δ = 7.90) those observed from the anomeric protons [H-1ꢀꢀ (δ =
in the NOESY spectrum. Thus compound 1 was de- 5.46)] to C-4 (δ = 151.6) and [H-1ꢀꢀꢀ (δ = 5.26)] to C-2ꢀꢀ
termined as 3,3ꢀ,4ꢀ-tri-O-methylellagic acid 4-O-(3ꢀꢀ- (δ = 76.7) indicated that the glucose was linked to the
galloyl)-β-D-glucopyranoside, named panconoside A. aglycon at position C-4, and the rhamnose to glucose
at position C-2ꢀꢀ (C1ꢀꢀꢀ-O-C2ꢀꢀ junction), respectively. In
Compound 2 was obtained as yellow crystals. Its
the NOESY experiment, the observation of an NOE
between H-5 (δ = 7.83) and H-1ꢀꢀ (δ = 5.46) supported
the location of the glucose unit. Acid hydrolysis of
molecular formula C29H32O17 was determined from
HRMS ((+)-ESI) which showed the pseudo-molecular
ion peak at m/z = 675.15354 [M+Na]+ (calculated
m/z = 675.15686) corresponding to 14 degrees of
unsaturation. It responded positively to the Molish
test, suggesting its glycoside nature. Its UV spectrum
(λmax = 255, 360 nm) was similar to that of ellagic
acid derivatives, suggesting that compound 2 like com-
pound 1 had an ellagic acid skeleton [18, 19]. The
IR spectrum displayed characteristic absorptions for
hydroxy groups (3430 cm−1) and lactone functions
(1749 cm−1), aromatic C=C groups (1600 cm−1), and
glycosidic C–O bonds (1090 cm−1). The broad band-
decoupled 13C NMR of compound 2 showed 29 car-
bon signals which were sorted by DEPT and HSQC
Fig. 2. Selected HMBC (→) and NOESY (↔) correlations
in compound 2.
Unauthenticated
Download Date | 10/8/15 12:07 AM

R. F. Soh et al. · Antibacterial Ellagic Acid Derivatives
1073
Table 2. In vitro antibacterial
activity of compounds 1, 2 and
extracts.
Tested
compound
MIC (µg mL−1
)
Tested microorganisms
M. luteus
E. coli
P. agarici
S. ferus
B. subtilis
S. minor
Leavesa
Stema
1
156.25
19.53
78.12
na
0.01
nt
39.06
9.76
78.12
78.12
nt
78.12
9.76
39.06
78.12
nt
78.12
156.25
39.06
na
1.95
nt
78.12
78.12
78.12
na
3.90
nt
39.06
19.53
39.06
na
3.90
nt
2
a = extract; na = not active; nt = not
tested.
Ampicillin
Gentamicin
2.44
3.90
Fig. 3. Structures of com-
pounds 1, 1a, 2 and 2a.
compound 2 gave glucose, rhamnose and 3,3ꢀ,4ꢀ-tri-O- methanolic extract of the leaves exhibited antibacte-
methylellagic acid, which were identified on TLC by rial potency with MIC values of 39.06, 78.12 and
comparison with known samples of glucose and rham- 156.23 µg mL−1. The EtOAc extract of the stem bark
nose using n-BuOH-Me2CO-H2O (4 : 5 : 1), and NMR showed a remarkable antibacterial effect with the best
data for the aglycon. The three methoxy groups at δ = value of MIC against E. coli and P. agarici (MIC =
4.01 (OMe-4ꢀ), 4.05 (OMe-3ꢀ) and 4.11 (OMe-3) cor- 9.76 µg mL−1) close to that of gentamycin used as
relate with C-4ꢀ, C-3ꢀ and C-3 carbons of the aglycon. standard drug (Table 2). These results provide promis-
Thus, the structure of 2 was established as 3, 3ꢀ,4ꢀ- ing baseline information for the potential use of these
tri-O-methylellagic acid 4-O-[α-L-rhamnopyranosyl- crude extracts in the treatment of bacterial infections.
(1→2)]-β-D-glucopyranoside, named panconoside B.
Although ellagic acid has already been isolated from
the Sapindaceae family [4], this is the first report
of its derivatives in this family. Despite the moder-
ate antibacterial potency of compounds 1 and 2, sev-
eral ellagic acid derivatives isolated from plants ex-
hibit antiplasmodial, cytotoxic, antioxidant, and α-
glucosidase inhibitory activities [4, 20, 23, 24]. All
these results highlight the potency of this class of sec-
ondary metabolites and suggest potential uses of plants
from the genus Pancovia in medicine.
Acetylation of compounds 1 and 2 afforded the per-
acetylated derivatives 1a and 2a (Fig. 3).
The antibacterial activity of compounds 1 and 2 was
evaluated in an antibacterial assay against 6 strains
of microorganisms, Micrococcus luteus, Streptococcus
ferus, Streptococcus minor, Escherichia coli, Bacil-
lus subtilis, and Pseudomonas agarici (Table 2), by
the agar well diffusion method [21, 22]. Compound 1
displayed a moderate antibacterial effect expressed as
minimum inhibitory concentration (MIC) against all
strains. In particular, P. agarici, S. ferus and S. mi-
nor were the most inhibited (MIC = 39.06 µg mL−1).
Compound 2 also showed moderate activity against
Experimental Section
General
Melting points were determined on a Bu¨chi-540 melting
E. coli and P. agarici (MIC = 78.12 µg mL−1). The point apparatus. UV spectra were measured on a Spectronic
Unauthenticated
Download Date | 10/8/15 12:07 AM

1074
R. F. Soh et al. · Antibacterial Ellagic Acid Derivatives
Unicam Spectrophotometer. Optical rotations were measured
The crude MeOH extract from the leaves was dissolved in
in DMSO solution on a Jasco Digital Polarimeter (model MeOH (500 mL) and washed with n-hexane (5 × 500 mL).
DIP-360). IR spectra were determined on a JASCO FT/IR- After concentration, this yielded 30 g of a hexane-soluble
410 spectrometer. HRMS ((+)-ESI) were run on a Bruker fraction and 17 g of a MeOH fraction. This latter fraction was
1
FT-ICR: APEX III (7.0 T). H and 13C NMR spectra were subjected to silica gel (Merck, 230 – 400 mesh) flash chro-
run on a Bruker DRX spectrometer equipped with 5 mm 1H matography and eluted with gradient n-hexane/EtOAc and
and 13C probes operating at 500 and 125 MHz, respectively, EtOAc/MeOH mixtures in the order of increasing polarity.
with TMS as internal standard. Silica gels (Merck, 230 – 400 Thirty-five fractions, each containing 250 mL, were collected
and 70 – 230 mesh) were used for column chromatography, and combined according to their TLC profiles on pre-coated
while pre-coated aluminum silica gel 60 F254 sheets were silica gel 60 F254 plates developed with n-hexane/EtOAc and
used for TLC with different mixtures of n-hexane, EtOAc, CH2Cl2/MeOH to yield 3 main fractions (Fꢀ1−3). Fraction
CH2Cl2, and MeOH as eluents; spots were visualized under Fꢀ1 was subjected to CC over silica gel (Merck, 70 – 230
UV lamps (254 and 365 nm) or by heating after spraying with mesh) and eluted with a CH2Cl2/MeOH mixture (9.8 : 0.2
50 % H2SO4 reagent.
to 8 : 2) to yield panconoside A (1) (15 mg) and 3,3ꢀ,4ꢀ-tri-
O-methylellagic acid 4-O-β-D-glucopyranoside (3) (80 mg).
Fraction Fꢀ2 was subjected to CC over silica gel (Merck,
70 – 230 mesh) with a CH2Cl2/MeOH mixture (9.5 : 0.5 to
8 : 2) to yield panconoside B (2) (500 mg) and allantoin (4)
(855 mg). Fraction Fꢀ3 yielded L-quebrachitol (5) (725 mg)
by column chromatography separation over silica gel using
the mixture CH2Cl2/MeOH (7 : 3).
Plant material
The stem bark and leaves of P. pedicellaris were collected
in April 2008 from Mount Kala in the Central Region of
the Republic of Cameroon. The plant was identified by Mr.
Nana Victor, plant taxonomist at the National Herbarium of
Cameroon, where a voucher specimen (Nr. 2741 / SRFCam)
has been deposited.
Antibacterial assays
In vitro antibacterial activity screening of the crude ex-
tracts and of compounds 1 and 2 were determined by the
agar well diffusion method as described in previous reports
[22, 23].
Extraction and isolation
The air-dried and powdered stem bark (2.5 kg) and leaves
(2 kg) of P. pedicellaris were extracted separately with
MeOH (5 L) at r. t. for 48 h and filtered. The filtrate was
concentrated to dryness under vacuum to give 120 g and 50 g
of crude extract, respectively.
Acid hydrolysis of compound 2
A solution of 7 mg of compound 2 in 10 mL MeOH/H2O
The MeOH extract from the stem bark was extracted se-
lectively with EtOAc. The EtOAc-soluble fraction was evap-
orated to dryness under vacuum to give a dry residue (40 g)
which was then subjected to column chromatography over
silica gel (Merck, 230 – 400 mesh) and eluted with a gradient
mixture of n-hexane/EtOAc and EtOAc/MeOH of increasing
polarity. Ninety column fractions, each containing 200 mL,
were collected and combined according to their TLC pro-
files on pre-coated silica gel 60 F254 plates developed with
n-hexane/EtOAc mixtures to yield 3 fractions (F1−3). Frac-
tion F1 was subjected to CC over silica gel (Merck, 70 –
230 mesh) eluted with an n-hexane/EtOAc mixture (9.5 : 0.5
to 7 : 3). This resulted in the isolation of stigmasterol (6)
(15 mg), 3-oxotirucalla-7, 24-dien-21-oic acid (10) (16 mg)
and umbelliferone (7) (56 mg). Fraction F2 was also sub-
jected to successive CC (Merck, 70 – 230 mesh) and eluted
with a mixture of n-hexane and EtOAc (6 : 4) to give scopo-
letin (8) (903 mg) and benjaminamide (11) (32 mg). Frac-
tion F3 was eluted with an n-hexane/EtOAc mixture of in-
creasing polarity (5 : 5 to 1 : 9) to yield 8ꢀ-epi-cleomiscosin
(9) (10 mg), 3-O-β-D-glucopyranosylstigmasterol (12) (900
mg) and 7-oxostigmasteryl-3-O-β-D-glucopyranoside (13)
(11 mg).
◦
(1 : 1) mixed with 5 mL 2N HCl was refluxed at 100 C for
3 h. After evaporation of the organic solvent under vacuum
and threefold extraction of the aqueous phase with EtOAc,
the aglycon was identified as 3,3ꢀ,4ꢀ-tri-O-methylellagic acid
by NMR techniques. The neutralized aqueous portion re-
sulted in the detection of glucose and rhamnose by com-
parison with standard sugar samples on TLC plates using n-
BuOH/Me2CO/H2O (4 : 5 : 1) as solvents and using (NH4)6
Mo7O24Ce(SO4)2H2SO4 reagent for visualization.
Acetylation of Panconosides A (1) and B (2)
Panconoside A (1) (4 mg) or B (2) (8 mg) was dissolved
in dry pyridine (0.5 mL), and Ac2O (1.0 mL) was added. The
mixture was stirred overnight at r. t. After the usual work-up,
filtration on a short silica gel column yielded the acetylated
derivative 1a (5 mg) or 2a (10 mg).
Panconoside A [3,3ꢀ,4ꢀ-tri-O-methylellagic acid 4-O-(3ꢀꢀ-
galloyl)-β-D-glucopyranoside] (1)
Beige crystals, m. p. 247 – 249 ◦C. – UV/Vis (MeOH):
λmax (lgεmax) = 255 (3.42), 280 (3.43), 355 (3.44) nm. –
Unauthenticated
Download Date | 10/8/15 12:07 AM

R. F. Soh et al. · Antibacterial Ellagic Acid Derivatives
[α]2D0 = +66.6◦ (c = 0.3, DMSO). – IR (KBr): νmax
1075
=
158.7 (C-7ꢀ), 163.6 (C-7ꢀꢀꢀ), 166.4 (C=O), 167.5 (C=O),
3420 (O–H), 1748 (C=O), 1611 (C=C), 1356, 1085 (C – O) 169.3 (C=O), 169.5 (C=O), 170.9 (C=O).
cm−1. – 1H NMR ([D6]DMSO) and 13C NMR ([D6]DMSO)
spectroscopic data, see Table 1. – MS (EI, 70 eV): m/z (%) =
344 (100), 329 (17), 286 (12). – HRMS ((+)-ESI): m/z =
681.10664 (calcd. 681.10991 for C30H26O17Na, [M+Na]+).
Panconoside B peracetate (2a)
Yellow amorphous powder. – 1H NMR (500 MHz,
CDCl3): δ = 1.15 (d, J = 6.3 Hz, 3H, 6ꢀꢀꢀ-Me), 1.98, 2.00,
2.06, 2.09, 2.13, 2.16 (6 × s, 18 H, COMe), 3.89 (br t, J =
8.8 Hz, 1H, 5ꢀꢀ-H), 3.96 (m, 1H, 2ꢀꢀ-H), 4.06 (s, 3H, OMe),
4.10 (m, 1H, 6ꢀꢀb-H), 4.15 (m, 1H, 5ꢀꢀꢀ-H), 4.18 (m, 1H,
6ꢀꢀa-H), 4.21 (s, 1H, 2ꢀꢀꢀ-H), 4.23 (s, 3H, OMe), 4.28 (s, 3H,
OMe), 5.05 (m, 1H, 3ꢀꢀꢀ-H), 5.08 (m, 1H, 4ꢀꢀ-H), 5.10 (m, 1H,
3ꢀꢀ-H), 5.16 (d, J = 1.3 Hz, 1H, 1ꢀꢀꢀ-H), 5.30 (d, J = 7.5 Hz,
1H, 1ꢀꢀ-H), 5.40 (t. J = 9.4 Hz, 1H, 4ꢀꢀꢀ-H), 7.71 (s, 1H, 5ꢀ-H),
7.86 (s, 1H, 5-H). – 13C NMR (125 MHz, CDCl3): δ = 17.5
(C-6ꢀꢀꢀ), 20.6, 20.7, 20.8, 20.9 (all COMe), 56.9 (OMe), 61.9
(C-6ꢀꢀ), 62.0 (OMe), 62.2 (OMe), 67.2 (C-5ꢀꢀꢀ), 68.3 (C-4ꢀꢀ),
68.4 (C-3ꢀꢀꢀ), 70.1 (C-2ꢀꢀꢀ), 70.9 (C-4ꢀꢀꢀ), 72.4 (C-2ꢀꢀ), 74.2
(C-3ꢀꢀ), 76.4 (C-5ꢀꢀ), 97.7 (C-1ꢀꢀ), 99.3 (C-1ꢀꢀꢀ), 107.9 (C-5ꢀ),
112.2 (C-5), 112.6 (C-1), 113.1 (C-1ꢀ), 113.7 (C-6ꢀ), 115.2
(C-6), 141.2 (C-3ꢀ), 141.4 (C-2), 141.9 (C-2ꢀ), 143.3 (C-3),
150.2 (C-4), 154.8 (C-4ꢀ), 158.4 (C-7), 158.6 (C-7ꢀ), 169.7
(C=O), 169.8 (C=O), 169.9 (C=O), 170.1 (C=O), 170.2
(C=O), 170.7 (C=O).
Panconoside B [3,3ꢀ,4ꢀ-tri-O-methylellagic acid 4-O-[α-L-
rhamnopyranosyl-(1→2)]-β-D-glucopyranoside] (2)
◦
Yellow crystals, m. p. 257 – 259 C. – UV/Vis (MeOH):
λ
max (lgεmax) = 255 (3.71), 360 (2.47) nm. – [α]2D0 = −80.0◦
(c = 0.5, MeOH). – IR (KBr): νmax = 3430 (O–H), 1749
(C=O), 1600 (C=C), 1090 cm−1. – 1H NMR ([D6]DMSO)
and 13C NMR ([D6]DMSO) spectroscopic data, see Ta-
ble 1. – MS (EI, 70 eV): m/z (%) = 344 (100), 329 (20), 286
(17). – HRMS ((+)-ESI): m/z = 675.15354 (calcd. 675.15686
for C29H32O17Na, [M+Na]+).
Panconoside A peracetate (1a)
Yellow amorphous powder. – 1H NMR (500 MHz,
CDCl3): δ = 2.03, 2.07, 2.10, 2.21, 2.32 (6 × s, 18 H,
COMe), 3.50 (br s, 1H, 5ꢀꢀ-H), 4.06 (s, 3H, OMe), 4.09 (m,
1H, 6ꢀꢀb-H), 4.15 (s, 3H, OMe), 4.24 (s, 3H, OMe), 4.29
(m, 1H, 6ꢀꢀa-H), 5.27 (d, J = 7.6 Hz, 1H, 1ꢀꢀ-H), 5.31 (t,
J = 9.4 Hz, 1H, 3ꢀꢀ-H), 5.54 (m, 1H, 4ꢀꢀ-H), 5.57 (m, 1H,
2ꢀꢀ-H), 7.73 (s, 1H, 5ꢀ-H), 7.78 (s, 2H, 2ꢀꢀꢀ/6ꢀꢀꢀ-H), 7.96 (s,
1H, 5-H). – 13C NMR (125 MHz, CDCl3): δ = 20.2, 20.6
(all COMe), 56.9 (OMe), 62.0 (2 × OMe), 62.3 (C-6ꢀꢀ), 68.0
(C-4ꢀꢀ), 70.7 (C-2ꢀꢀ), 72.7 (C-5ꢀꢀ), 73.4 (C-3ꢀꢀ), 100.0 (C-1ꢀꢀ),
108.0 (C-5ꢀ), 112.5 (C-5), 112.7 (C-1), 113.0 (C-1ꢀ), 113.7
(C-6ꢀ), 115.4 (C-6), 122.6 (C-2ꢀꢀꢀ/6ꢀꢀꢀ), 126.7 (C-1ꢀꢀꢀ), 139.4
(C-4ꢀꢀꢀ), 141.4 (C-3ꢀ), 141.6 (C-2ꢀ), 141.9 (C-2), 143.3 (C-3),
143.6 (C-3ꢀꢀꢀ/5ꢀꢀꢀ), 151.5 (C-4), 154.9 (C-4ꢀ), 158.4 (C-7),
Acknowledgements
The authors wish to acknowledge the German Academic
Exchange Service (DAAD) for awarding a grant to Soh
Fongang Rene´, the European Commission for awarding a
Marie Curie post doctoral fellowship to Bruno N. Lenta
(MIF1-CT-2006-021591), both at Bielefeld University, and
The Academy of Science for the Developing World (TWAS)
for awarding a research grant Nr. 07-141 LDC/CHE/AF/AC-
UNESCO FR: 3240171776 to our TWAS Research Unit.
[1] A. Aubreville, J. F. Leroy, Flore du Cameroun: Les
Sapindaceae, Vol. 1, Muse´um National d’Histoire Na-
turelle, Paris, 1973, pp. 3 – 118.
[2] M. Abdou-Shoer, G. E. Ma, X. H. Li, M. N. Koon-
chanok, R. L. Geahlin, C. J. Chang, J. Nat. Prod. 1993,
56, 967 – 969.
M. Litaudon, C. Lavaud, Phytochemistry 2005, 66,
825 – 835.
[8] D. D. Khac, S. Tran-Van, A. M. Campos, J.-Y. Lalle-
mand, M. Fetizon, Phytochemistry 1990, 29, 251 – 256.
[9] S. E. Drewes, J. V. Staden, Phytochemistry 1975, 14,
751 – 753.
[3] Z. Li, X. Li, D. Li, L. Gao, J. Xu, Y. Wang, Fitoterapia
2007, 78, 605 – 606.
[4] N. Rangkadilok, L. Worasuttayangkum, R. N. Bennett,
J. Satayavivad, J. Agri. Food Chem. 2005, 53, 1387 –
1392.
[5] H.-C. Huang, M.-D. Wu, W.-J. Tsai, S.-C. Liao, C.-C.
Liaw, L.-C. Hsu, Y.-C. Wu, Y.-H. Kuo, Phytochemistry
2008, 69, 1609 – 1616.
[10] M. Diaz, A. Gonzalez, I. Castro-Gamboa, D. Gonzalez,
C. Rossini, Carbohydr. Res. 2008, 343, 2699 – 2700.
[11] M. S. Alam, N. Chopra, M. Ali, M. Niwa, Phytochem-
istry 1996, 41, 1197 – 1200.
[12] B. T. Ngadjui, S. M. Mouncherou, J. F. Ayafor, B. L.
Sondemgam, F. Tillequin, Phytochemistry 1991, 30,
2809 – 2811.
[13] L. Cardona, B. Garcia, J. R. Pedro, J. Pe´rez, Phyto-
chemistry 1992, 31, 3989 – 3991.
[14] V. U. Ahmad, F. Ullah, J. Hussain, U. Farooq,
M. Zubair, M. T. H. Khan, M. I. Choudhary, Chem.
Pharm. Bull. 2004, 52, 1458 – 1461.
[6] M. L. D. Mesquita, P. Grellier, A. Blond, J.-P. Brouard,
J. E. D. Paula, L. S. Espindola, L. Mambu, Bioorg.
Med. Chem. 2005, 13, 4499 – 4506.
[7] L. Voutquenne, P. Guinot, C. Froissard, O. Thoison,
Unauthenticated
Download Date | 10/8/15 12:07 AM

1076
R. F. Soh et al. · Antibacterial Ellagic Acid Derivatives
[15] C.-M. Ma, N. Nakamura, M. Hattori, H. Kakuda, J.-C.
Qiao, H.-L. Yu, J. Nat. Prod. 2000, 63, 238 – 242.
[16] F. C. C. Simo, S. F. Kouam, H. M. P. Poumale, I. K.
Simo, B. T. Ngadjui, I. R. Green, K. Karsten, Biochem.
Syst. Ecol. 2008, 36, 238 – 243.
[17] R. Lie-Ching, C. C. Ming, H. J. Ching, J. Chinese
Chem. Soc. 2003, 50, 1103 – 1107.
[18] S. A. A. El-Toumy, H. W. Rauwald, Planta Med. 2003,
69, 682 – 684.
mad, N. K. Shamsum, B. T. Ngadjui, B. Luu, Chem.
Pharm. Bull. 2008, 56, 847 – 850.
[21] J. Parekh, S. Chanda, Turk. J. Biol. 2007, 31, 3 – 58.
[22] C. Perez, M. Paul, P. Bazerque, Acta Biol. Med. Exp.
1990, 15, 113 – 115.
[23] L. Verotta, M. Dell’Agli, A. Giolito, M. Guerrini,
P. Cabalion, E. Bosisio, J. Nat. Prod. 2001, 64, 603 –
607.
[24] A. Ito, H.-B. Chai, D. Lee, L. B. S. Kardono, S. Riswan,
N. R. Farnsworth, J. M. P. Cordell, D. Kinghorn, Phyto-
chemistry 2002, 61, 171 – 174.
[19] G. Pakulski, J. Budzianowski, Phytochemistry 1996,
41, 775 – 778.
[20] T. K. Tabopda, J. Ngoupayo, L. Jiawei, S. A. Muham-
Unauthenticated
Download Date | 10/8/15 12:07 AM