Flavonoid galloyl glucosides from the pods of Acacia farnesiana

Article abstract of DOI:10.1016/S0031-9422(97)00010-1

Three new flavonoids: naringenin 7-O-β-(4_,6_- digalloylglucopyranoside), quercetin 7-O-β-(6_-galloylglucopyranoside) and myricetin 7-O-β-(6_-galloylglucopyranoside) were identified from the pods of Acacia farnesiana, together with naringenin and kaempferol 7-(6_- galloylglucoside). The structures were determined by conventional methods of analysis and confirmed by ESI-MS (negative mode) and NMR spectroscopy.

Full text of DOI:10.1016/S0031-9422(97)00010-1

\
PERGAMON
Phytochemistry 40 "0888# 028Ð031
Flavonoid galloyl glucosides from the pods of Acacia farnesiana
Heba H[ Barakat\ Ahmed M[ Souleman\ Sahar A[M[ Hussein\ Ola A[ Ibrahiem\
Mahmoud A[M[ Nawwarꢀ
National Research Centre\ El!Dokki\ Cairo\ Egypt
Received in revised form 3 November 0885
Abstract
Three new ~avonoids] naringenin 6!O!b!"3ý\5ý!digalloylglucopyranoside#\ quercetin 6!O!b!"5ý!galloylglucopyranoside# and myr!
icetin 6!O!b!"5ý!galloylglucopyranoside# were identi_ed from the pods of Acacia farnesiana\ together with naringenin and kaempferol
6!"5ý!galloylglucoside#[ The structures were determined by conventional methods of analysis and con_rmed by ESI!MS "negative
mode# and NMR spectroscopy[ Þ 0888 Elsevier Science Ltd[ All rights reserved[
Keywords] Acacia farnesiana^ Leguminoseae^ naringenin 6!"3ý\5ý!digalloylglucoside#^ quercetin and myricetin 6!"5ý!galloylglucoside#^ ESI!MS^ NMR
0[ Introduction
Mr of 475 "ðM!HŁ−]474[3# and 2 exhibited a Mr of 599
0
amu "ðM!HŁ−]488#[ H and 02C NMR of 1 and 2 were
recorded and assigned for the _rst time[
As a part of our continuing search among Egyptian
medicinal plants for novel phenolics\ which might possess
biological activity\ we report here the isolation and struc!
tural elucidation of three new ~avonoids "0\ 3 and 4# from
the aqueous ethanolic pod extract of Acacia farnesiana
Willd[ Previous studies "El Sissi\ El Ansari\ + El
Negoumy\ 0862^ El Negoumy + El Ansari\ 0870#\ proved
the presence of naringenin 6!"5ý!galloylglucoside# in pods
and kaempferol 6!"5ý!galloylglucoside# in ~owers of this
plant[ Both known compounds "1 and 2# were isolated
during the course of the present study and subjected to
ESI!MS\ 0H and 02C NMR analysis for the _rst time[
The 0H NMR spectrum of 1\ "DMSO!d5\ at room temp#
revealed two aliphatic resonances at d 1[64 "dd\ J ꢁ 06
and 2 Hz# and at d 2[21 "dd\ J ꢁ 06 and 02 Hz# from the
axial and equatorial methylene protons at C!2 as well as
a third aliphatic resonance at d 4[37 "dd\ J ꢁ 02 and 2
Hz# from the methine proton\ which bears an oxygen and
phenyl function at C!1 of the ~avanone moiety in 1[
The presence of a 5ý!O!galloylglucoside moiety at the 6!
hydroxyl group of naringenin followed from the low_eld
shift of the resonances of H!5 and H!7 protons to d 5[07
"d\ J ꢁ 1[4 Hz# and 5[19 "d\ J ꢁ 1[4 Hz#\ respectively\
as well as the down_eld shift of the glucose methylenic
protons H!5 and H!5? to d 3[11 "dd\ J ꢁ 01[4 and 4 Hz#
and 3[31 "d J ꢁ 01[4 Hz#[ The two equivalent galloyl
protons\ H!1 and H!5 appeared as a sharp singlet inte!
grated to two protons at d 5[86 ppm and the anomeric
glucose proton as a doublet of J ꢁ 7 Hz at d 4[09\ thus
1[ Results and discussion
The meal of the deseeded pods of A[ farnesiana was
exhaustively extracted with aqueous ethanol "2]0#[ Com!
pounds 0Ð4 were isolated and puri_ed by polyamide col!
umn chromatography\ followed by Sephadex LH!19
column chromatography and preparative paper chro!
matography[ The known compounds 1 and 2 gave Rf
similar to\ and UV spectral data and hydrolytic products
identical with those of naringenin 6!"5ý!galloylglucoside#
"1#\ "El Sissi et al[\ 0862# and kaempferol 6!"5ý!gal!
loylglucoside# "2#\ "El Negoumy + El Ansari\ 0870#\
respectively[ On negative ESI!MS analysis\ 1 exhibited a
0
proving a b!con_guration and C3 conformation for the
glucose moiety of 1[ The remaining resonances in this
spectrum\ "Table 0#\ agreed well with the proposed
structure of
1
as naringenin!6!O!b!"5ý!galloyl!
glucopyranoside#[
The 02C NMR spectrum of 1 showed the characteristic
04 distinct carbon resonances of a naringenin moiety[
Glucosidation at the 6!hydroxyl of naringenin was
deduced from the recognized up_eld shift "Dd ppm ꢁ 0[5#
of the C6 carbon resonance "Table 1# as well as b!glu!
copyranose resonances in which that for the anomeric
carbon appeared at d 88[10[ Galloylation of the C!5 glu!
ꢀ Corresponding author[
9920!8311:88:, ! see front matter Þ 0888 Elsevier Science Ltd[ All rights reserved[
PII] S9920!8311"86#99909!0

039
H[H[ Barakat et al[ : Phytochemistry 40 "0888# 028Ð031
Table 0
0H NMR spectral data for compounds 0Ð4
Protons
Compound
of
0
1
2
3
4
Flavonoid
H!1
4[37 dd "01[4 + 2#
4[4 dd "01[4 + 2#
H!2
axial
equatorial
H!5
H!7
H!1?
H!2?
H!4?
H!5?
2[1 m
2[2 dd "06 + 01[4#
1[61 dd "06 + 2#
5[07 d "1[4#
5[19 d "1[4#
6[29 d "6[4#
5[79 d "6[4#
5[79 d "6[4#
6[29 d "6[4#
1[67 dd "06 + 2#
5[07 d "1[4#
5[19 d "1[4#
6[27 d "6[4#
5[77 d "6[4#
5[77 d "6[4#
6[27 d "6[4#
5[33 d "1[4#
5[71 d "1[4#
7[92 d "6[4#
5[84 d "6[4#
5[84 d "6[4#
7[92 d "6[4#
5[31 d "1[4#
5[79 d "1[4#
6[29 d "1#
5[31 d "1[4#
5[61 d "1[4#
6[17 s
5[89 d "6[4#
6[41 dd "6[4 + 1#
6[17 s
Glucose
H!0
H!1
H!2
H!3
4[14 d "7#
4[09 d "7#
4[07 d "7#
4[19 d "7#
4[19 d "7#
2[09Ð2[59 m
2[09Ð2[59 m
2[09Ð2[59 m
2[74 m
2[14Ð2[59 m
2[14Ð2[59 m
4[11 t "7#
2[14Ð2[69 m
2[14Ð2[69 m
2[14Ð2[69 m
2[89 m
2[29Ð2[59 m
2[29Ð2[59 m
2[29Ð2[59 m
2[89 m
2[29Ð2[59 m
2[29Ð2[59 m
2[29Ð2[59 m
2[89 m
H!4
2[89 m
H!5
H!5?
3[44 dd "01[4 + 4#
3[54 d "01[4#
3[11 dd "01[4 + 4#
3[31 d "01[4#
3[29 dd "01[4 + 4#
3[35 d "01[4#
3[29 dd "01[4 + 4#
3[37 d "01[4#
3[25 m
3[25 m
Galloyl:s
H!1 + H!5
6[07 + 6[08
5[84
5[86
5[86
5[86
Coupling constants "J in Hz# in parentheses[
cose hydroxyl group was evidenced from the down_eld
shift "Dd ppm ꢁ 0[0# of this carbon resonance\ in com!
parison with the resonance of the corresponding carbon
in the spectrum of free b!glucopyranose "Kalinowski\
Berger\ + Braun\ 0873#[ The galloyl moiety gave a charac!
teristic pattern with _ve distinct resonances located at the
expected chemical shifts "Table 1#[ Esteri_cation of the
carboxyl group of this moiety followed from the up_eld
shift "Dd ppm ꢁ 1[11# of the resonance of the esteri_ed
carboxyl carbon[ Consequently the identity of 1 is con!
_rmed[
moiety to C!6 of the kaempferol moiety followed from
the up_eld shift of this carbon resonance and the
accompanying down_eld shift of the resonances of its
ortho related carbons "C!5 and C!7# due to the a! and b!
e}ect\ respectively "all in comparison with the chemical
shifts of the corresponding carbon resonances in the 02
C
NMR spectrum of kaempferol itself# "Nawwar\ Soule!
man\ Buddrus\ + Linscheid\ 0873#[ Galloylation at the
glucopyranose carbon C!5 followed from the down_eld
shift of its resonance to d 51[78 ppm "compared with 59[5
ppm in the spectrum of free b!glucopyranose "Kalinowski
et al[\ 0873## and from the up_eld shift of the resonance
of the galloyl carbonyl carbon to d 054[70 ppm "compared
with d 056[6 in free gallic acid "Nawwar et al[\ 0873##[
The new compound\ 0\ isolated as a light brown
amorphous powder was found to possess chro!
matographic and colour properties "dark purple spot on
PC and UV light\ intense blue FeCl2 colour reaction and
a positive rose colour with aqueous KIO2\ speci_c for
galloyl esters "Haddok\ Gupta\ Al!Sha_\ + Haslam\
0871## and UV absorption maxima consistent with gal!
loylated naringenin 6!O!glucoside[ It exhibited a Mr of
627 amu in negative ESI!MS\ "ðM!HŁ−]626[3#[ On com!
plete acid hydrolysis\ 0 yielded naringenin\ gallic acid
"CoPC\ UV and 0H NMR analysis# and glucose "CoPC#[
On controlled acid hydrolysis it yielded naringenin 6!O!b!
The 0H NMR spectrum of 2 "DMSO!d5\[ room temp#
showed the expected kaempferol 6!O!b!glucopyranoside
proton pattern of signals with the exception of the re!
cognizable down_eld shift of the two methylenic glucose
proton resonances to d 3[35 "d\ J ꢁ 01[4 Hz# and to 3[2
"dd\ J ꢁ 01[4 and 4 Hz#\ which proved esteri_cation of
their geminal hydroxyl group[ In addition\ a sharp singlet
at d 5[86 was attributed to the H!1 and H!5 galloyl
protons\ thus con_rming the structure of 2 as kaempferol
6!O!b!"5ý!galloylglucopyranoside#[
02C NMR spectral analysis further con_rmed the struc!
ture of 2[ Thus\ most of the chemical shift values "Table
1# were the same as for kaempferol 6!O!glucopyranoside
"Markham + Mohan Shari\ 0871# and 5ý!O!gal!
loylglucose in 1[ The attachment of the galloylglucose

H[H[ Barakat et al[ : Phytochemistry 40 "0888# 028Ð031
030
Table 1
Merfort\ 0883^ Nawwar + Hussein\ 0883#\ as well as by
consideration of the known a! and b!e}ect "Nawwar\
Souleman\ Buddrus\ + Linscheid\ 0873# caused by esteri!
_cation of the glucose hydroxyl groups[ The b!anomeric
carbon resonance was identi_ed from its characteristic
chemical shift value "099[91 ppm#\ while the further
up_eld glucose carbon resonance at d 52[1 ppm was
assigned to the methylenic carbon C!5 to which one of
the galloyl moieties is attached[ Attachment of the second
galloyl moiety to C!3 of glucose was evidenced by the b!
up_eld shift recognized for the vicinal carbons "C!2 + C!
4# resonances to d ppm 64[08 and 63[92\ respectively
"by comparison with the chemical shift values of the
corresponding carbon resonances in the spectrum of the
unsubstituted b!glucopyranose "Nawwar et al[\ 0873#[ C!
3 was found to resonate down_eld at d ppm 60[47 "a!
e}ect# thus con_rming galloylation of its hydroxyl group[
Other resonances in this spectrum exhibited chemical
shift values which were in accordance with the
structure of 0 as naringenin 6!O!b!"3ý\5ý!digalloyl!
glucopyranoside#\ a new natural product[ This represents
the _rst report of a ~avonoid glucoside digallate[
Compound 3 was isolated as yellow crystals "mp 104>#\
which appeared yellow in UV light turning yellow!orange
when fumed with ammonia vapour[ It gave quercetin\
gallic acid and glucose on complete acid hydrolysis and
a Mr of 505 amu in negative ESI!MS "ðM!HŁ−]504#[
These data\ together with Rf values and UV spectral
analysis\ indicated that 3 is the quercetin analogue of 2[
This was supported by controlled acid hydrolysis of 3
which gave 5!monogalloyl!"a:b#!glucopyranose as an
intermediate "CoPC\ UV spectral\ ESI!MS and 0H NMR
analysis "Nawwar et al[\ 0883##[ 0H NMR "Table 0# and
02C NMR "Table 1# of 3 con_rmed its structure as quer!
cetin 6!O!b!"5ý!galloylglucopyranoside#\ which is ano!
ther new natural product[
02C NMR spectral data for compounds 0Ð4
Carbons
of
Compound
0
1
2
3
4
Flavonoid
1
2
3
4
5
6
7
8
09
0?
1?
2?
3?
4?
5?
68[8
32[1
67[7 036[6 036[8 037[1
31[0 025[9 025[0 025[1
087[9
053[5
85[0
054[3
86[3
086[2 065[0 065[9 065[2
052[9 059[4 059[4 059[2
85[3
054[0 051[4 051[4 051[4
84[2 83[1 83[1 83[5
87[6
87[5
87[7
053[0
093[4
029[3
018[0
005[9
047[5
005[9
018[0
052[9 044[8 044[7 045[9
092[3 093[7 093[7 094[9
017[5 010[5 010[7 010[0
017[6 018[6 004[5 096[8
004[2 004[4 034[1 035[0
046[7 048[3 036[6 025[1
004[2 004[4 004[4 035[0
017[6 018[6 019[0 096[8
Glucose
0
1
2
3
4
5
099[9
62[1
64[1
60[5
63[9
52[1
88[1
62[9
65[0
58[2
63[7
52[9
88[6
62[0
65[0
58[0
62[7
51[7
88[6 099[9
62[1
65[0
58[0
62[7
51[7
62[1
65[3
58[1
62[4
52[9
Galloyl:s
0
1
2
3
4
010[9 + 010[3
098[9 + 098[3
034[8
027[7 + 027[8
034[8
008[3 008[2 008[3 008[3
097[6 097[5 097[6 097[7
034[5 034[4 034[5 034[3
027[5 027[5 027[4 027[5
034[5 034[4 034[4 034[4
097[6 097[5 097[6 097[7
054[4 054[7 054[7 055[9
5
098[9 + 098[3
055[3 + 055[4
C1O
The new compound 4 "yellow crystals mp 127>#\ was
identi_ed as the myricetin analogue of 2 and 3 from
chromatographic\ UV spectral\ hydrolytic\ and ESI!MS
"Mr ꢁ 521 amu\ ðM!HŁ−]520# data[ Its structure was con!
"5ý!galloylglucopyranoside#\ "1#\ among other products[
Compound 1 was separated from the concentrated aque!
ous hydrolysate by preparative paper chromatography
and fully characterized by CoPC\ UV absorption\ ESI!
MS and 0H NMR analysis[
0
_rmed by H NMR "Table 1# and 02C NMR "Table 1#
as myricetin 6!O!b!"5ý!galloylglucopyranoside#\ which is
The 0H NMR spectrum of 0 exhibited the characteristic
resonance pattern of naringenin 6!O!b!"5ý!gal!
loylglucopyranoside# "Table 0# with the exception of the
recognized glucose proton signal appearing as a down!
_eld shifted triblet "J ꢁ 7Hz# at d ppm ꢁ 4[11 and the
additional galloyl protons which appeared as a singlet at
d ppm ꢁ 6[07 "or 6[08\ see Table 0#[ These data indicated
the presence of two galloyl moieties in the molecule of 0[
This was con_rmed by the two distinct galloyl carboxylic
carbon resonances at d ppm 055[3 and 055[4 and the
characteristic pattern of galloyl carbon resonances in the
02C NMR spectrum[ Resonances of the glucose carbons
were assigned by comparison with the 02C NMR data
reported for similar galloyglucose "Nawwar\ Hussein\ +
the third new natural product[
2[ Experimental
0H NMR spectra were measured at 399 MHz[ 0H res!
onances were measured relative to TMS and 02C NMR
resonances to DMSO!d5 and converted to TMS scale by
adding 28[4[ Typical conditions] spectral width ꢁ 5999
Hz for 0H and 11 999 Hz for 02C\ 21 K data points and a
~ip angle of 34>[ ESI!MS "negative mode#] the direct ~ow
injection technique was applied\ sample in MeOH was
introduced "0[14 ml min−0# together with MeOH sheath!
liquid "4 ml min−0# by a Harvard in~usion pump 8 ml

031
H[H[ Barakat et al[ : Phytochemistry 40 "0888# 028Ð031
min−0 SF5 sheath gas into the ESI!ion source of a Fin!
nigan MAT 3599 spectrometer[ PC was carried out on
Whatman no[ 0 paper\ using solvent systems] "0# H1O^
"1# 04) HOAc^ "2# BAW "n!BuOHÐHOAcÐH1O\ 3]0]4\
upper layer#^ "3# C5H5Ðn!BuOHÐH1OÐpyridine "0]4]2]2\
upper layer#[ Solvents 1 and 2 were used for prep[ PC on
Whatman no[ 2MM paper and solvents 2 and 3 for sugar
analysis[
229 sh[ Normal acid hydrolysis gave glucose "CoPC#\
0
naringenin and gallic acid "CoPC\ UV and H NMR
spectral data#[ 0H] see Table 0^ 02C NMR] Table 1[
2[4[ Kaempferol 6!O!b!"5ý!`alloyl`lucoside# "2#
Mr 599\ −ve ESI!MS ðM!HŁ−] 488[ Rf !values] 9[90
MeOH
max
"H1O#\ 9[07 "HOAc#\ 9[34 "BAW#[ UV] l
nm] 156\
208\ 254[ Normal acid hydrolysis gave glucose "CoPC#\
0
2[0[ Plant material[
kaempferol and gallic acid "CoPC\ UV and H NMR
spectral data#[ 0H NMR] Table 0^ 02C NMR] Table 1[
Fresh pods of Acacia farnesiana Willd\ were collected
from a mature tree in the Nile Delta near Tanta city\
Egypt\ during October 0883 and authenticated by Dr L[
Boulos\ Professor of Botany\ NRC\ Cairo\ Egypt[
2[5[ Quercetin 6!O!b!"5ý!`alloyl`lucoside# "3#
Mr 505\ −ve ESI!MS ðM!HŁ−] 504[ Rf !values] 9[90
MeOH
max
"H1O#\ 9[01 "HOAc#\ 9[31 "BAW#[ UV] l
nm] 147\
2[1[ Isolation and identi_cation
157\ 260[ Normal acid hydrolysis gave glucose "CoPC#\
quercetin and gallic acid "CoPC\ UV and 0H NMR spec!
tral data#[ 0H NMR] Table 0^ 02C NMR Table 1[
Powdered deseeded pods were extracted with EtOHÐ
H1O "2]0#[ The concd extract was applied to a polyamide
5S CC "Riedel!De Haen\ Seelze Hanover\ Germany# and
Ã
2[6[ Myricetin 6!O!b!"5ý!`alloyl`lucoside# "4#
eluted with H1O followed by H1OÐEtOH mixts of
decreasing polarities to yield eight major frs "IÐVIII#[
Compounds 0 and 1 were isolated from fr[ V "eluted by
39) EtOH# and 2Ð4 from fr[ VI "eluted by 79) EtOH#
by CC on Sephadex LH!19 using EtOH and EtOH con!
taining 0]0 mixture of Me1COÐH1O "6]0#[ Compounds
0Ð4 were puri_ed by prep[ PC\ using BAW and 0 and 1
were repuri_ed by prep[ PC\ using 04) HOAc[
Mr 521\ −ve ESI!MS ðM!HŁ−] 520[ Rf !values] 9[90
MeOH
max
"H1O#\ 9[98 "HOAc#\ 9[27 "BAW#[ UV] l
nm] 141\
208\ 261[ Normal acid hydrolysis gave glucose "CoPC#\
myricetin and gallic acid "CoPC\ UV and 0H NMR spec!
tral data# 0H NMR] Table 0\ 02C NMR] Table 1[
2[2[ Narin`enin 6!O!b!"3ý\5ý!di`alloyl`lucopyranoside#
"0#
References
El Sissi\ H[ I[\ El Ansari\ M[ A[\ + El Negoumy\ S[ I[ "0862#[ Phy!
tochemistry\ 01\ 1292[
El Negoumy\ S[ I[\ + El Ansari\ M[ A[ "0870#[ E`ypt Journal of Chem!
istry\ 13\ 360[
Mr 627\ −ve ESI!MS ðM!HŁ−]626[ Rf !values] 9[36
MeOH
max
"H1O#\ 9[37 "HOAc#\ 9[63 "BAW#[ UV l
nm] 172\
221 sh[ Normal acid hydrolysis gave glucose "CoPC#\
naringenin ðCoPC\ UV spectral data ð1Ł\ 0H NMR] ppm
4[27 "dd\ J ꢁ 01[4 + 2 Hz\ H!1#\ 2[05 "dd\ J ꢁ 06[0 and
01[4 Hz\ H!2ax#\ 1[55 "dd\ J ꢁ 06[0 and 2 Hz\ H!2eq#\ 4[89
"s\ H!5#\ 4[80 "s\ H!7#\ 6[2 "d\ J ꢁ 7 Hz\ H!1? and H!5?#\
5[72 "d\ J ꢁ 7 Hz\ H!2? and H!4?#Ł and gallic acid "CoPC\
UV and 0H NMR spectral data#[ Controlled acid
hydrolysis gave 1[ 0H NMR] Table 0^ 02C NMR] Table 1[
Haddok\ A[\ Gupta\ R[ K[\ Al!Sha_\ S[ M[\ + Haslam[ E[ "0871#[
Journal of the Chemistry Society Perkin Transactions I\ 1404[
Kalinowski\ H[ O[\ Berger\ S[\ + Braun\ S[ "0873#[ 02C NMR Spek!
troskopie[ Georg Thieme\ Stuttgart[
Markham\ K[ R[\ + Mohan Shari\ V[ "0871#[ In J[ B[ Harborne + T[ J[
Mabry "Eds[#\ The Flavonoids Advances in Research "p[ 75#[ London]
Chapman and Hall[
Nawwar\ M[ A[ M[\ Hussein\ S[ A[ M[\ + Merfort\ I[ "0883#[ Phy!
tochemistry\ 25\ 687[
Nawwar\ M[ A[ M[\ + Hussein\ S[ A[ M[ "0883#[ Phytochemistry\ 25\
0924[
2[3[ Narin`enin 6!O!b!"5ý!`alloyl`lucoside# "1#
Nawwar\ M[ A[ M[\ Souleman\ A[ M[\ Buddrus\ J[\ + Linscheid\ M[
"0873#[ Phytochemistry\ 12\ 1236[
Nawwar\ M[ A[ M[\ Souleman\ A[ M[\ Buddrus\ J[\ + Linscheid\ M[
"0873#[ Tetrahedron Letters\ 14\ 38[
Mr 475\ −ve ESI!MS ðM!HŁ−] 474[ Rf !values] 9[40
MeOH
max
"H1O#\ 9[43 "HOAc#\ 9[69 "BAW#[ UV] l
nm] 171\