Free-radical-scavenging and xanthine oxidase inhibitory constituents from Stereospermum personatum

Article abstract of DOI:10.1021/np058036y

Bioassay-guided fractionation of different extracts of both stem and stem bark of Stereospermum personatum led to the isolation of free-radical-scavenging and xanthine oxidase inhibitory molecules along with three new anthraquinones, sterequinones F-H (1-3), a new naphthoquinone, sterequinone I (4), two new phenethyl esters, 2(4′-hydroxyphenyl)ethyl undecanoate (14) and 2(4′-hydroxyphenyl)ethyl nonacosanoate (15), and a new 3,4,5-trimethoxycinnamyl ether, 2-methoxy-4-[3′-(3″,4″, 5″trimethoxyphenyl)allyloxymethyl]phenol (16), together with known compounds. The antioxidant and xanthine oxidase inhibitory potentials of the isolated compounds are reported.

Full text of DOI:10.1021/np058036y

J. Nat. Prod. 2005, 68, 1615-1621
1615
Free-Radical-Scavenging and Xanthine Oxidase Inhibitory Constituents from
Stereospermum personatum§
U. Sampath Kumar,† Ashok K. Tiwari,‡ S. Venkat Reddy,† P. Aparna,† R. Jagadeeshwar Rao,† A. Zehra Ali,‡ and
J. Madhusudana Rao*,†
Natural Products Laboratory, Division of Organic Chemistry-I, and Division of Pharmacology, Indian Institute of Chemical
Technology, Hyderabad-500 007, India
Received March 10, 2005
Bioassay-guided fractionation of different extracts of both stem and stem bark of Stereospermum
personatum led to the isolation of free-radical-scavenging and xanthine oxidase inhibitory molecules along
with three new anthraquinones, sterequinones F-H (1-3), a new naphthoquinone, sterequinone I (4),
two new phenethyl esters, 2(4′-hydroxyphenyl)ethyl undecanoate (14) and 2(4′-hydroxyphenyl)ethyl
nonacosanoate (15), and a new 3,4,5-trimethoxycinnamyl ether, 2-methoxy-4-[3′-(3′′,4′′,5′′trimethoxyphe-
nyl)allyloxymethyl]phenol (16), together with known compounds. The antioxidant and xanthine oxidase
inhibitory potentials of the isolated compounds are reported.
Stereospermum personatum Hassk-Chatterjii (Bignoni-
aceae) is an Ayurvedic medicinal plant used in the tradi-
tional Indian System of medicine1-3 and is widely advo-
cated as a diuretic, antiinflammatory and in prepara-
tions for hemorrhoids, vesicle calculi and for cardiotonic,
diabetic, cancer, renal, and hyperacidity disease condi-
tions.4 In a continuation of our search to identify potential
antioxidants derived from plants5 we have tested the free-
radical-scavenging (DPPH) and xanthine oxidase inhibitory
potential of extracts of various parts of S. personatum.
Although the petroleum ether and CHCl3 extracts of the
stem bark displayed mild free-radical-scavenging activity,
the petroleum ether extract resulted in the isolation of
biogenetically related quinones.6 The CHCl3 extract of the
stem bark furnished three new anthraquinones, stere-
quinones F-H (1-3), a new naphthoquinone, sterequinone
I (4), norviburtinal (5) (Figure 1), vanillin, and ferulic acid.
The acetone extract displayed potent free-radical-scaveng-
ing and moderate xanthine oxidase inhibition and yielded
three known iridoid glycosides (6-8). The CHCl3 extract
of the stem also displayed antioxidant and xanthine oxidase
inhibitory potential and afforded two new esters (14, 15)
of 4-hydroxyphenylethanol, a new 3,4,5-trimethoxycin-
namyl ether (16) (Figure 1), and several known compounds.
The structure of 16 was confirmed by spectroscopic meth-
ods and synthesis. Lignans (25-28) that were isolated in
substantial amounts were evaluated for their antioxidant
and xanthine oxidase inhibition potentials.
ortho-coupled aromatic protons (H-4 and 3). Two two-proton
triplets at δ 3.06 and 2.75 accounted for the presence of
two adjacent methylene groups (H-11 and 12). Two three-
proton singlets at δ 2.78 and 2.18 are due to aromatic and
acetyl methyl groups, respectively. The data showed re-
semblance to those of anthrakunthone isolated from Ste-
reospermum kunthianum7 with a different aromatic sub-
stitution. The 13C NMR data (Table 1) indicated the
presence of 19 carbons, which were sorted by DEPT into
two methyls, two methylenes, six methines, and nine
quaternary carbons. The above data led to the structure
1, named sterequinone F, a new 7-deoxy analogue of
anthrakunthone.
Sterequinone G (2) was isolated as a pale yellow semi-
solid. Its molecular formula was deduced as C20H18O4 by
HREIMS [M+] (m/z 322.1196), which was consistent with
13C NMR and DEPT data. Its IR spectrum displayed
carbonyl bands at 1713 and 1665 cm-1. The 1H NMR
spectrum (Table 1) displayed features similar to those of
sterequinone-F (1) but with a different aromatic substitu-
tion pattern. Two one-proton doublets at δ 8.20 (J ) 8.5
Hz) and 7.68 (J ) 2.5 Hz) and a one-proton double doublet
at δ 7.22 (J ) 8.5, 2.5 Hz) suggested the presence of a 1,2,4-
1
trisubstituted aromatic ring. Comparison of H, 13C NMR
and MS data with those of 1 revealed that 2 had an
aromatic O-methyl group at δ 3.98 with a corresponding
signal at δ 55.8 in its 13C NMR spectrum (Table 1). The
position of the O-methyl group was established by HMBC
correlations (Figure 2) of C-10a with H-8 and H-6 and of
C-8a with H-5. The above data showed that sterequinone
G had a structure as presented in 2, a new 7-methoxy
analogue of anthrakunthone.
Sterequinone H (3) was isolated as a pale yellow solid.
It had a molecular formula of C19H18O3 by the analysis of
its HREIMS [M+] (m/z 294.1246). The IR spectrum showed
absorption bands at 1668 and 1651 cm-1 assignable to two
carbonyl groups. The 1H NMR spectrum (Table 1) indicated
the presence of aromatic substitution identical with that
in sterequinone F (1) but with a different side chain. A one-
proton multiplet at δ 3.85 was typical of an aliphatic
hydroxy methine (H-13). Two multiplets (2H) at δ 2.90 and
1.75 suggested the presence of two adjacent methylene
groups, the former being characteristic of a benzylic me-
thylene. A three-proton doublet at δ 1.25 (J ) 6.0 Hz)
Results and Discussion
Sterequinone F (1) was isolated as a pale yellow solid.
The molecular formula was deduced from HREIMS analy-
sis [M+] (m/z 292.1088), 13C NMR, and DEPT experiments
as C19H16O3. The IR spectrum displayed absorptions at
1716 and 1667 cm-1 due to carbonyl functionalities. The
1H NMR spectrum (Table 1) revealed two two-proton
multiplets, at δ 8.25 and 7.75, indicating a typical 1,2-
disubstituted aromatic ring. Two doublets at δ 8.18 (1H, J
) 8.3 Hz) and 7.58 (1H, J ) 8.3 Hz) were indicative of two
§ IICT Communication No. 050613.
* Corresponding author. Tel: +91-40-2719 3166. Fax: +91-40-2716 0512.
E-mail: janaswamy@iict.res.in.
† Natural Products Laboratory, Division of Organic Chemistry-I.
‡ Division of Pharmacology.
10.1021/np058036y CCC: $30.25
© 2005 American Chemical Society and American Society of Pharmacognosy
Published on Web 10/29/2005

1616 Journal of Natural Products, 2005, Vol. 68, No. 11
Kumar et al.
Figure 1.
a
Table 1. 1H and 13C NMR Data of Compounds 1-4 in CDCl3
1
2
3
position
δH
δC
δH
δC
δH
δC
1
139.8 C
139.8 C
147.2 C
134.4 CH
125.6 CH
134.1 C
129.0 CH
120.6 CH
164.3 C
109.9 CH
137.3 C
189.0 C
133.7 C
126.3 C
182.3 C
28.1 CH2
43.4 CH2
206.9 C
139.9 C
2
147.7 C
149.0 C
3
4
7.58 d (8.3)
8.18 d (8.3)
134.0 CH
125.6 CH
135.2 C
7.55 d (8.0)
8.18 d (8.0)
7.55 d (8.5)
8.15 d (8.5)
134.0 CH
125.5 CH
135.3 C
4a
5
8.25 m
7.75 m
7.75 m
8.25 m
126.4 CH
133.3 CH
134.3 CH
127.2 CH
133.5 C
8.20 d (8.5)
7.22 dd (8.5, 2.5)
8.25 m
7.75 m
7.75 m
8.25 m
126.4 CH
133.3 CH
134.4 CH
127.2 CH
133.1 C
6
7
8
7.68 d (2.5)
8a
9
185.6 C
185.9 C
9a
132.0 C
132.1 C
10a
10
132.6 C
132.7 C
183.5 C
183.6 C
11
3.06 t (7.2)
2.75 t (7.2)
29.6 CH2
43.3 CH2
206.8 C
3.10 t (7.2)
2.78 t (7.2)
2.90 m
1.75 m
3.85 m
2.80 s
29.6 CH2
39.3 CH2
67.5 CH
17.4 CH3
23.8 CH3
12
13
1-CH3
13-CH3
OCH3
2.78 s
2.18 s
17.4 CH3
30.0 CH3
2.75 s
2.20 s
3.98 s
17.4 C
29.3 CH3
55.8 CH3
1.25 d
a 1H and 13C NMR spectra recorded for 1 and 3 at 200 and 50 MHz, respectively, for 2 at 300 and 75.462 MHz, respectively. Chemical
shifts presented in ppm (δ) with TMS as internal standard; J values are given in Hz in parentheses.
methine carbon at δ 67.5. The above data were consistent
with structure 3 (Figure 1) for sterequinone H.
Sterequinone I (4) was isolated as a pale yellow solid.
The molecular formula was determined as C20H18O4 by
HRFABMS [M + H]+ (m/z 323.1296), which was consistent
with 13C NMR and DEPT data. The IR spectrum indicated
two sharp bands at 1717 and 1664 cm-1 accounting for two
1
carbonyl groups. The H NMR spectrum of compound 4
showed two one-proton doublets at δ 8.18 (J ) 8.6 Hz) and
7.60 (J ) 2.6 Hz) and a one-proton double doublet at δ 7.20
(J ) 8.6, 2.6 Hz), suggesting a typical 1,2,4-trisubstituted
aromatic ring. A sharp singlet at δ 3.98 (3H) accounted for
an aromatic O-methyl group. A double multiplet at δ 3.02
(2H) and a multiplet at δ 2.25 (2H) indicated the presence
Figure 2.
of two adjacent methylene groups (H-2 and H-1). Two
accounted for the terminal methyl group. The 13C NMR
spectrum (Table 1) indicated 19 carbons. The presence of
the carbinol function was evident by the presence of a
singlets at δ 2.70 (3H) and 1.65 (3H) were indicative of two
olefinic methyl groups. The downfield shift of the former
could be due to its proximity to both the carbonyl and

Constituents from Stereospermum personatum
Journal of Natural Products, 2005, Vol. 68, No. 11 1617
Figure 3.
1
hydroxyl groups. The above H NMR data showed resem-
at 1733 cm-1. The 1H NMR spectrum of compound 14
showed two two-proton doublets at δ 7.05 (2H, J ) 9.0 Hz)
and 6.72 (2H, J ) 9.0 Hz), suggesting the presence of a
1,4-disubstituted aromatic ring. Two triplets were observed
at δ 2.85 (2H) and 4.22 (2H), the former attributed to a
benzylic methylene and the latter to an oxymethylene
group of an ester (-CH2-O-). A broad singlet at δ 4.60,
which was exchanged with D2O, accounted for a phenolic
hydroxyl group. The above data suggested the presence of
a 4-hydroxyphenylethyl group, which was substantiated by
a fragment at m/z 121 in its EI MS spectrum.
blance to those of kigelinol isolated from Kigelia pinnata.8
The 13C NMR spectrum revealed the presence of 20
carbons. The position of the aromatic O-methyl group was
established by HMBC correlations (Figure 2), where C-6a
showed correlations with H-10 and H-8. C-10a and C-9
showed correlations with H-7. The above data established
the structure 4 for sterequinone I. Sterequinone I probably
represents the missing link in the biogenetic sequence of
sterequinone B to sterequinone D previously isolated by
us.6
Norviburtinal (5) was isolated as an orange powder and
was identified by comparing its spectroscopic data with
literature data.9 However, the 13C NMR data whose as-
signments are based on a DEPT experiment and in
comparison with halitunal10 are reported here for the first
time.
The acetone extract of the stem bark upon further
purification yielded substantial amounts of three iridoid
glycosides (Figure 3), which were identified as (-)-6-
feruloyl catalpol (6),11 (-)-specioside (7),12 and (-)-vermi-
noside (8)13 by comparison of spectroscopic data with
literature values.
The 1H NMR spectrum further displayed a three-proton
triplet at δ 0.90, which was characteristic of a terminal
methyl group of an aliphatic chain. Another triplet at δ
2.30 (2H) was characteristic of a methylene adjacent to a
carbonyl group. A broad multiplet at δ 1.30 integrating for
14 protons accounted for the presence of an aliphatic chain
comprising seven methylenes. 13C assignments were made
by comparing the data with that of bongardol (octacosanoic
acid ester of 4-hydroxyphenylethanol) isolated from Bon-
gardia chrysogonum.22 The above data suggested that
compound 14 is a new undecanoic acid ester of 4-hydrox-
yphenylethanol.
The powdered stem of S. personatum was first defatted
with petroleum ether followed by the extraction with
CHCl3. Further purification of the CHCl3 extract yielded
three new compounds (14-16) (Figure 1) along with several
known compounds. The known compounds were identified
as lapachol (9),14 dehydro-R-lapachone (10),14 coniferalde-
hyde (11),15 sinapaldehyde (12),16 3,4,5-trimethoxycinna-
maldehyde (13),17 pinoresinol (24),18 7′-hydroxydivanil-
lyltetrahydrofuran (25),19 (-)-secoisolariciresinol (26),20 (-)-
olivil (27),21 and (+)-cycloolivil (28)21 (Figure 3). Except
lapachol, all the remaining compounds were isolated for
the first time from this plant.
Compound 14 was obtained as a white solid. The
molecular formula was deduced from its positive ion
FABMS as C19H30O3, which showed an [M + Na]+ ion at
m/z 329. The IR spectrum revealed the presence of a
hydroxyl group at 3408 cm-1 and an ester carbonyl function
Compound 15 was also isolated as a white solid. The
molecular formula C37H66O3 was deduced from its FABMS
1
(M+ 558). The H NMR has indicated features similar to
compound 14 except for the multiplet at δ 1.30 integrating
for 50 protons that indicated the presence of 25 methylenes.
These data along with the established molecular mass
allowed the structure of compound 15 to be defined as a
new nonacosanoate ester of 4-hydroxyphenylethanol.
Compound 16 was isolated as a pale brown semisolid.
The molecular formula C20H24O6 was deduced from its
HREIMS analysis (m/z 360.1562), 13C NMR, and a DEPT
experiment. The IR spectrum indicated the presence of a
hydroxyl group at 3416 cm-1 and an aliphatic ether at 1034
cm-1. The 1H NMR spectrum showed a double triplet at δ
6.25 (1H) and a doublet at δ 6.50 (1H, J ) 15.0 Hz)
indicative of a trans double bond. A sharp singlet at δ 6.60
(2H) indicated the presence of two equivalent aromatic

1618 Journal of Natural Products, 2005, Vol. 68, No. 11
Kumar et al.
Scheme 1a
a Reagents and conditions: (a) TBDMSCl, Et3N, DCM, rt, 8 h; (b) NaBH4, MeOH, rt, 5 h; (c) PBr3, DCM, -10 °C, 30 min; (d) carbethoxymethylene
triphenyl phosphorane, benzene, reflux, 12 h; (e) DIBAL-H, DCM, -10 °C 1 h; (f) sodium hydride, THF, compound 19, 24 h; (f) TBAF, THF, 10 min.
protons. A two-proton doublet at δ 4.30 (J ) 5.0 Hz) was
indicative of the oxymethylene of an ether function. Its
deshielding may be attributed to its allylic nature. Two
singlets integrating for six and three protons respectively
resonated at δ 3.88 and 3.82, respectively, and suggested
tion of the cinnamyl alcohol with the benzyl bromide (19)
using sodium hydride followed by deprotection of the silyl
group using tetrabutylammonium fluoride yielded com-
pound 16 with spectroscopic data identical with the natural
product.
1
the presence of three aromatic O-methyl groups. The H
Iridoids are distributed widely in plants belonging to the
Bignoniaceae family24 and are well known for their broad
spectrum of biological properties.25 Some of the iridoid
glycosides are well documented for their free-radical-
scavenging and xanthine oxidase inhibitory properties.26,27
In our search for potential antioxidants derived from
plants5 we have screened the three iridoids (6-8) for their
antioxidant (DPPH) activity as well as potential for xan-
thine oxidase inhibition. Table 2 summarizes the SC50 for
free-radical DPPH scavenging and IC50 for xanthine oxi-
dase inhibitory concentration of extract as well as com-
pounds isolated from it. Among the three iridoids (-)-
verminoside (8) displayed strong free-radical-scavenging
properties, whereas (-)-specioside (7) displayed mild activ-
ity. The xanthine oxidase inhibition pattern of these three
compounds was in the reverse order with (-)-specioside (7)
presenting high inhibition followed by (-)-6-feruloylcatalpol
(6). (-)-Verminoside (8) did not display xanthine oxidase
inhibitory property.
Lignans possess a broad spectrum of biological activi-
ties.28 Since the lignans 7′-hydroxydivanillyltetrahydro-
furan (25), (-)-secoisolariciresinol (26), (-)-olivil (27), and
(+)-cycloolivil (28) (Figure 3) were obtained in substantial
quantities, they were also evaluated for their free-radical-
scavenging (DPPH) and xanthine oxidase inhibitory activi-
ties. Table 3 displays the free-radical (DPPH)-scavenging
and xanthine oxidase inhibitory concentration of extract
as well as lignans isolated from it. These compounds
NMR spectrum displayed a three-proton singlet at δ 3.92
accounting for one more aromatic O-methyl group. A broad
singlet at δ 5.52, which was exchanged with D2O, suggested
the presence of a phenolic hydroxyl group. A singlet at δ
4.60 (2H) was characteristic of the benzylic methylene
group adjacent to an ether functionality. A doublet at δ
6.85 (2H, J ) 2.5 Hz) and a singlet at δ 6.90 (1H) indicated
the presence of three more aromatic protons. The above
data suggested the presence of a 4-hydroxy-3-methoxyben-
zyl group, which was confirmed by the presence of a
fragment at m/z 136 in its EIMS spectrum. The 13C NMR
spectrum of the compound indicated the presence of 20
carbons, which are consistent with the proposed structure.
The above data suggested the structure of compound 16
as shown (Figure 1). The structure was further confirmed
via synthesis from vanillin (Scheme 1).
Vanillin was protected as TBDMS ether (17) followed by
reduction of the aldehyde group of 17 with sodium boro-
hydride to afford 4-tert-butyldimethylsilyloxyvanillyl alco-
hol (18) in quantitative yield. The resultant alcohol was
converted into the corresponding benzyl bromide (19) using
phosphorus tribromide under anhydrous conditions.23 Ethyl
3′,4′,5′-trimethoxycinnamate (21) was prepared by the
Wittig reaction of 3,4,5-trimethoxybenzaldehyde with car-
bethoxymethylene triphenylphosphorane in anhydrous
benzene. The cinnamic ester was reduced with DIBAL-H
to afford the corresponding cinnamyl alcohol (22). Alkyla-

Constituents from Stereospermum personatum
Journal of Natural Products, 2005, Vol. 68, No. 11 1619
Table 2. Antioxidant and Xanthine Oxidase Inhibitory Activity
of Extracts and Isolated Compounds from the Stem Bark of S.
personatuma
300 MHz spectrometers using TMS as internal standard. IR
spectra were recorded on a Nicolet-740 spectrometer. EIMS
was recorded on a VG 70-70 micromass instrument. FABMS
and HREIMS were recorded on a VG Auto Spec-M instrument.
Column chromatography was performed on silica gel (60-120
mesh). Solvents and reagents were purified according to
standard procedures.
Plant Material. Plant material was collected from the
forests of Tirumala, Andhra Pradesh, India, in January 2000.
It was identified by Dr. K. Madhava Chetty, Department of
Botany, Sri Venkateswara University, Tirupati. A voucher
specimen of the plant is deposited in Herbarium, Department
of Botany, with an accession number 547.
Extraction and Isolation. The shade-dried stem bark of
S. personatum (2 kg) was powdered and extracted with
petroleum ether in a Soxhlet apparatus for 20 h. The material
was then extracted with CHCl3 for 20 h followed by extraction
with acetone. The CHCl3 extract was evaporated under
reduced pressure to obtain a residue (4.0 g). The residue was
adsorbed on silica gel and subjected to column chromatography
eluted with petroleum ether and by mixtures containing
increasing amounts of EtOAc. The fractions eluted at 5, 10,
15, and 20% EtOAc in petroleum ether were collected sepa-
rately, concentrated, and rechromatographed using silica gel
(60-120 mesh) to obtain compounds 1 (20 mg), 2 (8 mg), 3 (20
mg), 4 (12 mg), and 5 (25 mg) in pure form.
The acetone extract (20 g) was purified using column
chromatography (silica gel, 60-120 mesh) with CHCl3 and
increasing amounts of MeOH. The fractions eluted with 9, 10,
and 11% MeOH were collected separately and concentrated
to obtain compounds 6 (2.5 g), 7 (5.5 g), and 8 (2.0 g) in pure
form.
Similarly, the shade-dried stem of S. personatum (2.5 kg)
was extracted with petroleum ether in a Soxhlet apparatus
followed by the extraction with CHCl3 for 20 h. The CHCl3
extract on evaporation yielded a residue of 22 g. The residue
was adsorbed on silica gel (60-120 mesh) and subjected to
chromatography by eluting first with CHCl3, followed by
CHCl3/acetone (9:1, 8.5:1.5) (I and II) and CHCl3/MeOH (9.5:
0.5) (III). Fraction I eluted at 10% acetone was subjected to
chromatography over silica gel using CHCl3 with increasing
percentage of acetone. The fractions eluted at 2%, 4%, 6%, 8%,
and 10% acetone yielded compounds 9 (65 mg), 10 (20 mg),
11 (8 mg), 12 (6 mg), and 13 (8 mg) in pure form. Fraction II
eluted at 15% acetone in CHCl3 was also subjected to column
chromatography over silica gel with CHCl3 with increasing
percentages of acetone. The fractions eluted at 5%, 10%, 15%,
and 20% acetone yielded compounds 14 (18 mg), 15 (20 mg),
16 (15 mg), and 24 (6 mg) in pure form. Fraction III was
subjected to column chromatography using CHCl3 and MeOH.
The fractions eluted at 2%, 3%, 4%, and 5% MeOH in CHCl3
yielded compounds 25 (50 mg), 26 (1 g), 27 (2.5 g), and 28 (2.2
g) in pure form.
antioxidant (DPPH) xanthine oxidase
activity
(SC50
inhibition
(IC50
compound
)
)
CHCl3 extract
sterequinone A-I
ferulic acid
12.14 µg/mL
NA
32.51 µM
6.78 µg/mL
40.49 µM
160.85 µM
7.88 µM
NA
NA
NA
acetone extract
6-feruloylcatalpol (6)
specioside (7)
verminocide (8)
allopurinol
54.2 µg/mL
150.4 µM
79.88 µM
NA
34.91 µM
trolox
15.94 µM
a NA, compounds displaying less than 10% activity at 50 µg/
mL concentration were not followed for IC50 or SC50 analysis. IC50
or SC50 values were determined by linear regression analysis using
at least five different concentrations in triplicate and represent
the mean of the experiment. SDM were within 10% in any case.
IC50 are concentrations of compounds or mixtures required to
inhibit 50% activity of xanthine oxidase enzyme, and SC50, the
concetration of extracts or compounds to scavenge 50% of the free-
radical (DPPH) under experimental conditions.
Table 3. Antioxidant and Xanthine Oxidase Inhibitory Activity
of Extract and Isolated Compounds from the Stem of S.
personatuma
antioxidant (DPPH) xanthine oxidase
compound
CHCl3 extract
activity (SC50
)
inhibition (IC50)
7.47 µg/mL
7.58 µM
12.5 µM
59.83 µg/mL
NA
213 µM
compound (16)
secoisolariciresinol (26)
cycloolivil (28)
7′-hydroxydivanillyl-
tetrahydrofuran (25)
olivil (27)
17.38 µM
15.16 µM
101.38 µM
110.88 µM
15.10 µM
15. 94 µM
23.63% at 132 µM
34.91 µM
trolox
allopurinol
a NA, compounds displaying less than 10% activity at 50 µg/
mL concentration were not followed for IC50 or SC50 analysis. IC50
or SC50 values were determined by linear regression analysis using
at least five different concentrations in triplicate and represent
the mean of the experiment. SDM were within 10% in any case.
IC50 are concentrations of compounds or mixtures required to
inhibit 50% activity of xanthine oxidase enzyme, and SC50, the
concetration of extracts or compounds to scavenge 50% of the free-
radical (DPPH) under experimental conditions.
displayed moderate xanthine inhibitory activity. As far as
their xanthine oxidase inhibitory potential is concerned,
(+)-cycloolivil (28) and 7′-hydroxydivanillyltetrahydrofuran
(25) were more potent than (-)-secoisolariciresinol (26) and
(-)-olivil (27). While the DPPH free-radical-scavenging
activities of (-)-olivil and (+)-cycloolivil are known,29 the
DPPH-scavenging activities for 7′-hydroxydivanillytetrahy-
drofuran (25) and secoisolarciresinol (26) are reported here
for the first time. Similarly the xanthine oxidase inhibitions
of these lignans are also reported for the first time.
The combined application of free-radical scavengers of
varied nature with multiple activities is increasingly being
advocated for the prevention and treatment of oxidative
stress related disorders.30,31 The presence in substantial
yield of free-radical-scavenging, xanthine oxidase inhibitory
iridoids and lignans in S. persontum, therefore, may
provide scientific basis for its use in Ayurvedic preparations
prescribed under a variety of disease conditions.
Sterequinone F (1): pale yellow solid; UV (MeOH) λmax
(log ꢀ) 259 (3.45), 252 (3.41) nm; IR(KBr) νmax 2916, 1716, 1667,
1
1330, 1287, 1169, 760 cm-1; H and 13C NMR data, see Table
1; EIMS m/z (rel int) 292 (4) [M+], 274 (100), 259 (46), 249
(30), 178 (27), 149 (18), 129(7), 97 (30); HREIMS m/z 292.1088
(calcd for C19H16O3, 292.1099).
Sterequinone G (2): pale yellow semisolid; UV (MeOH)
λmax (log ꢀ) 267 (3.6) nm; IR (KBr) νmax 2918, 2818, 1713, 1665,
1596,1282 cm-1; 1H and 13C NMR data, see Table 1; EIMS m/z
(rel int) 322 (M+) (10), 304 (85), 225 (35), 165 (30), 149 (50),
81 (65), 57 (100); HREIMS m/z 322.1196 (calcd for C20H18O4,
322.1205).
Sterequinone H (3): yellow solid; UV (MeOH) λmax (log ꢀ)
278 (3.50), 259 (3.9) nm; IR (KBr) νmax 3456, 2822, 1668, 1651,
1
1581, 1321, 1127, 759 cm-1; H and 13C NMR data, see Table
1; EIMS m/z (rel int) 294 [M+] (5), 279 (10), 199 (12), 125 (30),
106 (100); HREIMS m/z 294.1246 (calcd for C19H18O3, 294.1255).
Sterequinone I (4): pale yellow solid; UV (MeOH) λmax (log
ꢀ) 272 (3.92), 252 (3.78) nm; IR (KBr) νmax 3431, 2918, 1717,
Experimental Section
General Experimental Procedures. 1H NMR and 13C
NMR spectra were obtained on Varian 200 MHz and Bruker
1664, 1595, 1272, 1178 cm-1 1H NMR (300 MHz, CDCl3) δ
;
8.18 (1H, d, J ) 8.6 Hz, H-7), 8.13 (1H, s, H-5), 7.60 (1H, d, J

1620 Journal of Natural Products, 2005, Vol. 68, No. 11
Kumar et al.
) 2.6 Hz, H-10), 7.20 (1H, dd, J ) 8.6, 2.6 Hz, H-8), 3.98 (3H,
s, OCH3), 3.02 (2H, dm, H-2), 2.70 (3H, s, 12-CH3), 2.25 (2H,
m, H-1), 1.65 (3H, s, 3-CH3); 13C NMR (75.462 MHz) δ 182.8
(C, C-11), 181.7 (C, C-6), 162.0 (C, C-9), 151.2 (C, C-5a), 148.4
(C, C-4), 138.8 (C, C-10a), 133.2 (C, C-13), 132.9 (C, C-3), 129.4
(C, C-6a), 127.9 (CH, C-7), 126.8 (C, C-12), 119.6 (CH, C-8),
118.2 (CH, C-5), 107.3 (CH, C-10), 79.6 (C, C-11a), 54.2 (CH3,
OCH3), 39.4 (CH2, C-1), 28.0 (CH2, C-2), 27.8 (CH3, 3-CH3),
17.0 (CH3, 12-CH3); HRFABMS m/z 323.1296 (calcd for
C20H19O4, 323.1283).
(15 mL) was added NaBH4 (0.141 g, 0.003 mol) slowly at 0 °C,
and the mixture was stirred at room temperature for 5 h. The
solvent was evaporated under vacuum. The residue was
dissolved in EtOAc (50 mL), washed with H2O (50 mL), dried
over anhydrous Na2SO4, and concentrated to obtain 18 (1.2 g,
90%) as a colorless liquid: IR (KBr) νmax 2935, 1514, 1465,
1293, 899 cm-1; 1H NMR (200 MHz, CDCl3) δ 6.85 (1H, s, H-2),
6.75 (2H, s, H-5 and 6), 4.58 (2H, Ar-CH2), 3.80 (3H, OCH3),
1.40 (brs, OH), 1.0 (9H, (CH3)3-C), 0.18 (6H, Si-(CH3)2); EIMS
m/z 210 (M+ - (CH3)3C).
Norviburtinal (5): orange powder; IR (KBr) νmax 1645,
(c) 4-tert-Butyldimethylsilyloxy-3-methoxybenzyl bro-
mide (19). To a solution of 18 (0.350 g, 0.001 mol) in
anhydrous CH2Cl2 (5 mL) was added slowly PBr3 (0.11 g, 0.042
mol) in anhydrous CH2Cl2 (1 mL) at -10 °C under N2
atmosphere, and the mixture was stirred for 30 min. Solid
NaHCO3 (0.100 g) was added, and the organic layer was
decanted and washed with CH2Cl2 again. The combined
organic layers are concentrated to obtain 19 (0.380 g, 87%) as
a colorless liquid. The product was immediately used for the
1559, 1460, 1396, 1117, 1023, 965, 736 cm-1 1H NMR (200
;
MHz, CDCl3) δ 9.85 (1H, s, CHO), 9.21 (1H, s, H-1), 7.82 (1H,
d, J ) 3.5 Hz, H-3), 7.79 (1H, d, J ) 6.0 Hz, H-6), 7.30 (1H, d,
J ) 6.0 Hz, H-5), 6.59 (1H, d, J ) 3.5 Hz, H-4); 13C NMR (50
MHz, CDCl3) δ 184.7 (CH, CHO), 151.1 (CH, C-1), 146.1 (CH,
C-6), 143.2 (CH, C-3), 134.4 (C, C-9), 124.2 (C, C-8), 123.1 (C,
C-7), 110.5 (CH, C-5), 111.1 (CH, C-4); EIMS m/z (rel int) 146
[M+] (78), 145 (100), 117 (30), 89 (37.5), 63 (43.7), 43 (31.2).
2-(4′-Hydroxyphenyl)ethyl undecanoate (14): white
solid; UV (MeOH) λmax (log ꢀ) 277 (3.2) nm; IR (KBr) νmax 3408,
2918, 1733, 1517, 1464, 1175 cm-1; 1H NMR (200 MHz, CDCl3)
δ 7.05 (2H, d, J ) 9.0 Hz, H-2′ and 6′), 6.72 (2H, d, J ) 9.0
Hz, H-3′ and 5′), 4.60 (brs, ArOH), 4.22 (2H, t, J ) 6.9 Hz,
H-1), 2.85 (2H, t, J ) 6.9 Hz, H-2), 2.30 (2H, t, J ) 6.9 Hz,
H-2′′), 1.60 (2H, m, H-3′′), 1.30 (14H, s, H-4′′ to H-10′′), 0.90
(3H, t, J ) 6.2 Hz, H-11′′); 13C NMR (50 MHz, CDCl3) δ 173.8
(C, C-1′′), 154.4 (C, C-4′), 129.9 (CH, C-6′ and 2′), 129.8 (C,
C-1′), 115.3 (CH, C-3′ and 5′), 64.9 (CH2, C-1), 34.3 (CH2, C-2),
31.9 (CH2, C-2′′), 29.7-22.7 (CH2, C-3′′-10′′), 14.1 (CH3, C-11′′);
FABMS m/z 329 [M + Na]+.
1
subsequent reaction as it decomposed rapidly. H NMR (200
MHz, CDCl3) δ 6.70-6.82 (3H, m, H-2, 5, and 6), 4.42 (2H,
Ar-CH2), 3.80 (3H, s, OCH3), 1.0 (9H, (CH3)3-C), 0.18 (6H, Si-
(CH3)2).
(d) Ethyl 3′,4′,5′-trimethoxycinnamate (21). 3,4,5-Tri-
methoxybenzaldehyde (0.49 g, 0.002 mol) and carbethoxym-
ethylenetriphenyl phosphorane (0.869 g, 0.002 mol) were
dissolved in anhydrous benzene (15 mL). The mixture was
refluxed under nitrogen for 12 h, the solvent was removed
under vacuum, and the product was purified by column
chromatography (silica gel, 60-120 mesh). The fractions eluted
at 20% EtoAc in petroleum ether yielded 21 (0.55 g, 83%): IR
(KBr) νmax 2932, 1710, 1512, 1467, 1274, 1160 cm-1; 1H NMR
(200 MHz, CDCl3) δ 7.58 (1H, d, J ) 17.3 Hz, H-â), 6.70 (2H,
s, H-2′ and 6′), 6.30 (1H, d, J ) 17.3 Hz, H-R), 4.20 (2H, q,
H-2), 3.88 (6H, s, 3′-OCH3 and 5′-OCH3), 3.82 (3H, s, 4′-OCH3),
1.30 (3H, t, H-3); EIMS m/z 266 (M+).
2-(4′-Hydroxyphenyl)ethyl nonacosanoate (15): color-
less solid; UV (MeOH) λmax (log ꢀ) 278 (3.2) nm; IR (KBr) νmax
3406, 2917, 1733, 1517, 1464, 1175 cm-1; 1H NMR (400 MHz,
CDCl3) δ 7.08 (2H, d, J ) 9.0 Hz, H-2′ and 6′), 6.75 (2H, d, J
) 9.0 Hz, H-3′ and 5′), 4.50 (brs, ArOH), 4.25 (2H, t, J ) 6.9
Hz, H-1), 2.85 (2H, t, J ) 6.9 Hz, H-2), 2.30 (2H, t, J ) 6.9 Hz,
H-2′′), 1.60 (2H, m, H-3′′), 1.30 (50H, s, H-4′′ to H-28′′), 0.92
(3H, t, J ) 6.2 Hz, H-29′′); 13C NMR (75.462 MHz, CDCl3) δ
173.9 (C, C-1′′), 154.3 (C, C-4′), 129.9 (CH, C-2′ and 6′), 129.8
(C, C-1′), 115.3 (CH, C-3′ and 5′), 64.9 (CH2, C-1), 34.3 (CH2,
C-2), 31.9 (CH2, C-2′′), 29.6-22.6 (CH2, C-3′′-28′′), 14.0 (CH3,
C-29′′); FABMS m/z 558.
(e) 3′,4′,5′-Trimethoxycinnamyl alcohol (22). Ethyl
3′,4′,5′,-trimethoxycinnamate 21 (0.266 g, 0.001mol) was dis-
solved in anhydrous CH2Cl2 (10 mL), and the mixture was
cooled to -15 °C. To this mixture was added slowly 1.18 mL
(0.355 g, 0.002 mol) of a 30% solution of DIBAL-H in hexane.
The mixture was stirred for 1 h under nitrogen atmosphere.
MeOH (0.5 mL) was added slowly followed by the addition of
a saturated solution of sodium potassium tartarate (15 mL).
The mixture was stirred vigorously for 1 h, and the layers were
separated. The organic layer was dried over anhydrous Na2-
SO4 and concentrated to yield 22 (0.180 g, 81%) as a colorless
2-Methoxy-4-[3′-(3′′,4′′,5′′-trimethoxyphenyl)allyloxym-
ethyl]phenol (16): pale brown semisolid; UV (MeOH) λmax
(log ꢀ) 285 (2.92) nm; IR (KBr) νmax 3416, 1514, 1480, 1275,
1239, 1125, 1034, 757 cm-1; 1H NMR (200 MHz, CDCl3) δ 6.90
(1H, s, H-3), 6.85 (2H, d, J ) 2.5 Hz, H-5 and 6), 6.60 (2H, s,
H-2′′ and 6′′), 6.50 (1H, d, J ) 15.0 Hz, H-3′), 6.25 (1H, dt, J
) 15, 5.0 Hz, H-2′), 5.52 (brs, ArOH), 4.60 (2H, s, H-1′′′), 4.30
(2H, d, J ) 5.0 Hz, H-1′), 3.92 (3H, s, 2-OCH3), 3.88 (6H, s, 3′′
and 5′′-OCH3), 3.82 (3H, s, 4′′-OCH3); 13C NMR (50 MHz,
CDCl3) δ 153.3 (C, C-3′′ and 5′′), 146.5 (C, C-2), 143.8 (C, C-1),
138.2 (C, C-4′′), 133.0 (C, C-4), 132.4 (C, C-1′′), 131.1 (CH, C-3′),
128.0 (CH, C-2′), 120.2 (CH, C-5), 114.3 (CH, C-3), 110.0 (CH,
C-6), 103.8 (CH, C-2′′ and 6′′), 65.3 (CH2, C-1′′′), 63.5 (CH2,
C-1′), 60.8 (CH3, 4′′-OCH3), 56.1 (CH3, 3′′ and 5′′-OCH3), 55.9
(CH3, 2-OCH3); EIMS 360 (M+) (28), 193 (12), 136 (100), 82
(42); HREIMS m/z 360.1562 (calcd for C20H24O6, m/z 360.1572).
Synthesis of Compound 16 (Scheme 1). 4-tert-Bu-
tyldimethylsilyloxy-3-methoxybenzaldehyde (17). Vanil-
lin (1 g, 0.0065 mol) was dissolved in anhydrous CH2Cl2 (20
mL) and Et3N (0.796 g, 0.0078 mole). The mixture was cooled
to 0 °C. To this solution was added slowly t-BDMSiCl (0.99 g,
0.0065 mole), and the mixture was stirred under nitrogen for
8 h. After completion of the reaction the reaction mixture was
diluted with CH2Cl2 (50 mL), washed with H2O (50 mL), dried
over anhydrous Na2SO4, and concentrated under vacuum to
give 17 (1.50 g, 86%) as a liquid: IR (KBr) νmax 2931, 1686,
liquid: IR (KBr) νmax 3400, 1580, 1450 cm-1 1H NMR (200
;
MHz, CDCl3) δ 6.82 (2H, s, H-2′ and 6′), 6.55 (1H, d, J ) 14.4
Hz, H-â), 6.20 (1H, m, H-R), 4.30 (2H, d, J ) 6.4 Hz, H-1),
3.88 (6H, s, 3′ and 5′ OCH3), 3.82 (3H, s, 4′-OCH3); EIMS m/z
224 (M+).
(f) tert-Butyl-{2-methoxy-4-[3′-(3′′,4′′,5′′-trimethoxy-
phenyl)allyoxymethyl]phenoxy}dimethylsilane (23). To
a suspension of NaH (0.029 g, 0.0012 mol) in anhydrous THF
(3 mL) was added slowly 22 (0.180 g, 0.0008 mol) in anhydrous
THF (5 mL) under nitrogen atmosphere at room temperature,
and the mixture was stirred for 0.5 h. To this mixture was
added 18 (0.267 g, 0.008 mol) in anhydrous THF (3 mL), and
the mixture was stirred for 24 h at room temperature. The
reaction mixture was poured into ice and extracted with EtOAc
(2 × 50 mL). The organic layer was dried over Na2SO4 and
concentrated. The residue was purified by column chromatog-
raphy over silica gel (60-120 mesh). The fractions eluted at
20% EtOAc in petroleum ether gave 23 (0.050 g): IR (KBr)
νmax 2930, 1512, 1465, 1129, 913, 794 cm-1; 1H NMR (200 MHz,
CDCl3) δ 7.0 (1H, s, H-2), 6.78 (2H, m, H-5 and 6), 6.55 (2H,
s, H-2′′ and 6′′), 6.52 (1H, d, J ) 17.3 Hz, H-3′), 6.20 (1H, dt,
H-2′), 4.82 (2H, s, H-1′′′), 4.25 (2H, d, J ) 4.3 Hz, H-1′), 3.82
(9H, s, 3′′, 5′′, and 3-OCH3), 3.80 (3H, s, 4′′-OCH3), 1.0 (9H, s,
(CH3)3-C), 0.20 (6H, s, Si-(CH3)2); FABMS m/z 475 (M + H)+.
(g) 2-Methoxy-4-[3′-(3′′,4′′,5′′-trimethoxyphenyl)allyl-
oxymethyl]phenol (16). To a solution of compound 23 (0.020
g) in THF (2 mL) was added a 1 M solution of TBAF (0.05
mL) in THF. The reaction was completed immediately, as
1
1508, 900, 782 cm-1; H NMR (300 MHz, CDCl3) δ 9.80 (1H,
s, CHO), 7.35 (2H, m, H-2 and 6), 6.92 (1H, d, J ) 9.1 Hz,
H-3), 3.90 (3H, s, OCH3), 1.05 (9H, s, (CH3)3-C), 0.20 (6H, s,
Si-(CH3)2); EIMS m/z 264 (M+).
(b) 4-tert-Butyldimethylsilyloxy-3-methoxybenzyl al-
cohol (18). To a solution of 17 (1.32 g, 0.005 mol) in MeOH

Constituents from Stereospermum personatum
Journal of Natural Products, 2005, Vol. 68, No. 11 1621
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evident from TLC (solvent system: petroleum ether/EtOAc, 70:
30). To the mixture was added H2O (5 mL), and the product
was extracted with EtOAc to afford the target compound 16
(0.010 g), which was homogeneous by TLC and identical with
the natural product in all aspects.
Bioassay Procedures. Determination of free-radical DPPH
scavenging and xanthine oxidase activity was done as per the
method described earlier.32
Acknowledgment. We thank Dr. J. S. Yadav, Director,
IICT, for his invaluable suggestions. S.V.R. and R.J.R. thank
CSIR and UGC, respectively, for their fellowships.
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NP058036Y