Terpenoids of Syzygium formosanum

Article abstract of DOI:10.1021/np980313w

A new natural product, 4-epifriedelin (1), and 12 known terpenoids have been isolated from the leaves of Syzygium formosanum. The known compounds include caryophyllene oxide, friedelin, canophyllal, glutinol, α-terpineol, phytol, betulinic acid, uvaol, lupeol, betulin, ursolic acid, and oleanolic acid. All of these compounds are reported for the first time from S. formosanum.

Full text of DOI:10.1021/np980313w

J . Nat. Prod. 1999, 62, 327-328
327
Ter p en oid s of Syzygiu m for m osa n u m
†
‡
§
†
,†
Churng-Werng Chang, Tien-Shung Wu, Yih-Shou Hsieh, Sheng-Chu Kuo, and Pei-Dawn Lee Chao*
Institute of Pharmaceutical Chemistry, China Medical College, Taichung, Taiwan, Republic of China
Received J uly 17, 1998
A new natural product, 4-epifriedelin (1), and 12 known terpenoids have been isolated from the leaves of
Syzygium formosanum. The known compounds include caryophyllene oxide, friedelin, canophyllal, glutinol,
R-terpineol, phytol, betulinic acid, uvaol, lupeol, betulin, ursolic acid, and oleanolic acid. All of these
compounds are reported for the first time from S. formosanum.
Syzygium formosanum (Hay.) Mori (Myrtaceae) is en-
demic to Taiwan. In the course of our continuing search
for anti-HIV agents from natural products, a 70% acetone
extract of the leaves demonstrated promising inhibition of
Moloney murine leukemia virus (Mo-MuLV) reverse tran-
methyl, and one carbon at δ 82.7 was shifted downfield
from δ 30.0 in comparison to friedelin. The IR spectrum of
-
1
3 showed hydroxyl absorption at 3481 cm
. EIMS
+
exhibited a significant peak at 426 (M - H
2
O). Based on
the above data, 3 was determined as maytensifolin A.
Comparison of the 1H NMR, C NMR, and EIMS with
those of maytensifolin A in the literature confirmed this
13
scriptase. The acetone extract was partitioned between H
2
O
and CH Cl , and the CH Cl fraction still showed inhibitory
2
2
2
2
asssignment.3 As to the structure of 2, its H NMR and
-5
1
effects on Mo-MuLV reverse transcriptase. This study
reports the isolation of 13 terpenoids from this fraction,
all of them reported for the first time from this plant, of
which 1 is a new natural product.
MS spectral data were identical with those reported for
3
canophyllal. The spontaneous transformation of canophyl-
lal to maytensifolin A has not been reported previously,
and the mechanism remains to be elucidated.
Compound 1 was obtained as colorless needles. Its IR
-
1
spectrum exhibited a band for carbonyl (1717 cm ). The
1
3
C NMR spectrum of compound 1 revealed chemical shifts
of carbons on C, D, and E rings very similar to those of
friedelin, but two methyl carbons were found further
downfield than friedelin (δ 13.5, 23.1 vs δ 6.8, 14.6). HMQC
of 1 revealed that the proton attached to the carbon at δ
1
3.5 was a doublet (J ) 7.3 Hz), indicating this carbon was
assignable to C-23. Based on the above data, compound 1
appeared to be the C-4 epimer of friedelin. Upon standing
3
in CDCl for one month after NMR measurement, com-
1
3
pound 1 had its C NMR spectrum recorded again, it
indicated the coexistence of friedelin1 with compound 1
(ratio ca. 1:1). The mixture was then separated again by
HPLC to afford compound 1 and friedelin. Therefore, the
structure of 1 was determined to be 4-epifriedelin, a
compound not reported previously as a natural product.
1
1
The structure of 1 was further confirmed by H- H COSY,
HMBC, and NOESY techniques. The epimerization of 1 at
C-4 in solvent could be accounted for by keto-enol tau-
tomerism of the ketone group at C-3, and the equilibrium
favored the more stable configuration of friedelin. 4-Epi-
friedelin had been previously reported as a product ob-
tained by photoepimerization of friedelin.2
Canophyllal (2) was obtained as a yellow powder. It
showed 30 carbons, including two carbonyls (δ 213.4 and
1
3
2
09.1) in the C NMR spectrum and an aldehyde group
1
in H NMR spectum (δ 9.48, s, 1H). Upon standing in
1
3
CDCl
3
for four months, compound 2 was subjected to
C
Based on the H NMR, 13C NMR, and MS data, together
1
NMR again, and it was found to have transformed into a
different compound that showed only 29 carbons, including
one carbonyl at δ 213.4. HMQC and HMBC analyses
revealed that 3 was of the friedelane type. By comparison
with the 13C NMR spectrum of friedelin, 3 had one less
with their physical constants, the other isolates were
characterized as caryophyllene oxide, friedelin, glutinol,
6
1
1
7
8
5,9,10
5,11
5,12
R-terpineol, phytol, betulinic acid,
uvaol,
lupeol,
5,9,10
13
5,13,14
betulin,
ursolic acid, and oleanolic acid.
Bioactivity tests of the isolates against Mo-MuLV reverse
transcriptase indicated that betulinic acid which has
previously been reported to show an EC50 of 1.4 µM, was
a promising inhibitor of human immunodeficiency virus
(HIV).
†
China Medical College.
Cheng Kung University.
Chung-Shan Medical and Dental College.
To whom correspondence should be addressed. Fax: 886 4 2032146.
‡
15
§
*
E-mail: pdlee@cmce.cmc.edu.tw.
1
0.1021/np980313w CCC: $18.00
© 1999 American Chemical Society and American Society of Pharmacognosy
Published on Web 01/05/1999

3
28 J ournal of Natural Products, 1999, Vol. 62, No. 2
Notes
Exp er im en ta l Section
measured using a Packard 2200 CA, Tri-carb Liquid Scintil-
lation Counter.
Gen er a l Exp er im en ta l P r oced u r es. The preparative
HPLC apparatus was equipped with a Shimadzu LC-10S
pump and a Waters 410 differential refractometer. LiChro-
spher Si 60 column (10 µm, 25 cm × 1 cm) was used for
separations. Melting points were measured on a Yanaco MP-
4
-Ep ifr ied elin (1): colorless needles (n-hexane-EtOAc); mp
1
-
2
2
42-245 °C, [R]
944, 2863, 1717, 1460, 1381, 1206, 1193, 1058, 978, 953; H
D
+2.7° (c 0.19, CHCl ,), IR νmax (KBr) cm
1
3
NMR (CDCl , 400 MHz) δ 0.80 (3H, s, H-25), 0.85 (1H, m,
3
H-22a), 0.86 (3H, s, H-24), 0.88 (3H, s, H-29), 0.93 (3H, s,
5
00 and are uncorrected. UV spectra were obtained on a
H-30), 0.94 (3H, s, H-26), 0.98 (3H, s, H-27), 1.05 (3H, d, J )
Shimadzu UV-160A, whereas IR spectra were recorded on a
Nicolet Impact 400 FT-IR. EIMS were obtained on a Hewlett-
Packard 5995 GC-MS system by direct probe (70 ev). NMR
spectra were recorded on Bruker ARX-200, ARX-300 or AMX-
7
.3 Hz, H-23), 1.11 (3H, s, H-28), 1.14 (1H, m, H-6a), 1.16 (2H,
m, H-19), 1.23 (1H, m, H-15a), 1.27 (1H, m, H-8), 1.29 (1H, m,
H-12), 1.31 (1H, m, H-11a), 1.36 (1H, m, H-11b), 1.44 (1H, m,
H-22b), 1.45 (1H, m, H-15b), 1.47 (1H, m, H-18), 1.54 (2H, m,
4
00 FT-NMR spectrometers. Optical activities were measured
H-1a and H-10),1.55 (1H, m, H-6b), 1.77 (1H, m, H-1b), 1.84
on a J ASCO DIP-1000 polarimeter.
13
(
1H, m, H-4), 2.24 (1H, m, H-2a), 2.44 (1H, m, H-2b); C NMR
P la n t Ma ter ia l. The leaves of Syzygium formosanum (4
kg) were collected in Taipei in September 1994, and identified
by Dr. Chung-Chuan Chen of the Institute of Chinese Phar-
maceutical Sciences, China Medical College.
(
3
3
CDCl
3
, 100 MHz) C-1 21.7, C-2 37.1, C-3 216.6, C-4 58.7, C-5
9.9, C-6 37.4, C-7 17.7, C-8 53.5, C-9 37.0, C-10 49.4, C-11
5.7, C-12 30.5, C-13 39.7, C-14 38.3, C-15 32.4, C-16 36.0,
C-17 30.0, C-18 42.7, C-19 35.3, C-20 28.1, C-21 32.7, C-22
9.2, C-23 13.5, C-24 23.1, C-25 18.0, C-26 20.4, C-27 18.7,
Extr a ction a n d Isola tion . The air-dried leaves were
3
blended with 70% Me
was removed by concentration under reduced pressure. The
residue was suspended in H O and partitioned six times with
CH Cl to afford an aqueous fraction (700 g) and a CH Cl
fraction (105 g).
The CH Cl fraction was chromatographed on a Si gel
2
CO in a juicer. After filtration, Me
2
CO
+
C-28 32.1, C-29 35.0, C-30 31.7; EIMS (m/z, %) 426 [M ] (8),
3
1
5
41 (3), 302 (7), 273 (12), 246 (12), 205 (17), 179 (19), 163 (31),
37 (26), 125 (44), 123 (53), 109 (68), 95 (100), 81 (71), 69 (91),
5 (72).
2
2
2
2
2
2
2
Refer en ces a n d Notes
column and eluted with increasing amounts of EtOAc in
n-hexane. Further separation was carried out by using Si gel
HPLC, which employed isocratic elution with various percent-
ages of EtOAc in n-hexane as mobile phases. The chromato-
graphed fraction yielded caryophyllene oxide (15 mg), friedelin
(1) Akihisa, T.; Yamamnoto, K.; Tamura, T.; Kimura, Y.; Iida, T.;
Nambara, T.; Chang, C. F. Chem. Pharm. Bull. 1992, 40, 789-791.
2) Aoyaki, R.; Yamda, S.; Tsuyuki, T.; Takahashi, T. Bull. Chem. Soc.
(
J pn. 1973, 46, 959-963.
(3) Nozaki, H.; Suzuki, H.; Hirayama, T.; Kasai, R.; Wu, R. Y.; Lee, K.
(600 mg), 4-epifriedelin (1, 165 mg), canophyllal (2, 10 mg),
H. Phytochemistry 1986, 25, 479-485.
(
(
(
4) Patra, A.; Chaudhuri, S. K. Magn. Reson. Chem. 1987, 25, 95-100.
5) Mahato, S. B.; Kundu, A. P. Phytochemistry 1994, 37, 1517-1575.
6) Sashida, Y.; Itokawa, H.; Akita, Y.; Fujita, M. Yakugaku Zasshi 1976,
96, 218-222.
glutinol (35 mg), R-terpineol (15 mg), phytol (20 mg), betulinic
acid (35 mg), uvaol (25 mg), lupeol (30 mg), betulin (90 mg),
ursolic acid (20 mg), and oleanolic acid (15 mg).
Rever se Tr a n scr ip ta se Assa y. The aforementioned iso-
lates were subjected to an in vitro reverse transcriptase assay.
(7) Harada, R.; lwasaki, M. Phytochemistry 1982, 21, 2009-2011.
(
(
8) Suga, T.; Aoki, T. Phytochemistry 1974, 13, 1623-1624.
9) Salimuzzaman, S.; Farrukh, H.; Sabira, B.; Bina, S. S. J . Nat. Prod.
n
Both (γA) -(dT)15 and unlabeled TTP were obtained from
1
988, 51, 229-233.
3
Boehringer Mannheim GmbH, Penzber, Germany. H-TTP was
purchased from Amersham Co. Reverse transcriptase from
Moloney murine leukemia virus (Mo-MuLV) is the product of
Bethesda Research Laboratories Life Technologies, Inc., Gaith-
ersburg, MD. The assay reaction mixture (30 µL) contained
(10) Sholichin, M.; Yamasaki, K.; Kasai, R.; Tanaka, O. Chem. Pharm.
Bull. 1980, 28, 1006-1008.
(
11) Salimuzzaman, S.; Farrukh, H.; Sabira, B.; Bina, S. S. J . Nat. Prod.
1
986, 49, 1086-1090.
(12) William, F. R.; Stewart, M.; J anusz, P. Tetrahedron 1986, 42, 3419-
3428.
5
0 mM Tris-HCl buffer, pH 8.3; 75 mM KCl; 10 mM
dithiothreitol; 3 mM MgCl ; 0.5 µg of (γ A) -(dT)15; 0.5 µ Ci
H]-dTTP, and 5 units of Mo-MuLV reverse transcriptase
(13) Tsutomu, F.; Yutaka, O.; Chisato, H. Phytochemistry 1987, 26, 715-
19.
14) Maillard, M.; Adewunmi, C. O.; Hostettmann, K. A. Phytochemistry
7
2
n
(
3
[
1
992, 31, 1321-1323.
1
6
preparation. After incubation at 37 °C for 30 min, 5 µL of
.4 M EDTA was added to terminate the reaction. The reaction
mixture (15 µL, in triplicate) was spotted on Whatman DEAE
cellulose paper and washed three times each with 5% Na
HPO , H O, and EtOH. The radioactivity of the sample was
(15) Fujioka, T.; Kashiwada, Y.; Kilkuskie, R. E.; Cosentino, L. M.; Ballas,
0
L. M.; J iang, J . B.; J anzen, W. P.; Chen, I. S.; Lee, K. H. J . Nat. Prod.
1
994, 57, 243-247.
(
16) Chu, S. C.; Hsieh, Y. S.; Lin, J . Y. J . Nat. Prod. 1992, 55, 179-183.
2
-
4
2
NP980313W