Synthesis of 8-desoxythunberginol A and (±)-8-desoxy-3,4- dihydrothunberginol A

Article abstract of DOI:10.1002/jccs.200300046

A two step synthesis of title isocoumarin isolated from Homalium longifolium and its conversion into corresponding 3,4-dihydroisocoumarin has been described. 3,4-Dimethoxybenzoyl chloride on condensation with homophthalic acid afforded 3-(3′,4′-dimethoxyphenyl)isocoumarin which was demethylated to furnish the 8-desoxythunberginol A, whereas its sequential saponification, reduction and demethylation yielded the (±)-8-desoxy-3,4- dihydrothunberginol A. The synthesized compounds were examined in vitro for antibacterial activity.

Full text of DOI:10.1002/jccs.200300046

Journal of the Chinese Chemical Society, 2003, 50, 313-317
313
Synthesis of 8-Desoxythunberginol A and (±)-8-Desoxy-
3,4-dihydrothunberginol A
Aamer Saeed
Department of Chemistry, Quaid-i-Azam University, Islamabad 45320, Pakistan
A two step synthesis of title isocoumarin isolated from Homalium longifolium and its conversion into cor-
responding 3,4-dihydroisocoumarin has been described. 3,4-Dimethoxybenzoyl chloride on condensation
with homophthalic acid afforded 3-(3¢,4¢-dimethoxyphenyl)isocoumarin which was demethylated to furnish
the 8-desoxythunberginol A, whereas its sequential saponification, reduction and demethylation yielded the
(±)-8-desoxy-3,4-dihydrothunberginol A. The synthesized compounds were examined in vitro for antibacte-
rial activity.
Keywords: Homalium longifolium; Hydrangea macrophylla; Thunberginol A; Isocoumarins; Anti-
bacterial.
INTRODUCTION
In 1995, K. Shaari isolated 3-(3¢,4¢-dihydroxyphen-
mine release from rat peritoneal mast cells induced by com-
pound 48/80, calcium ionophore A23187, with IC50 values of
less than 100 mM. Structure-activity studies have shown that
isocoumarins such as thunberginol A and B are more potent
as histamine release inhibitors as compared to the dihydro-
isocoumarins such as hydrangenol, thunberginol D and E.6 S.
Ohta has reported a synthesis of thunberginol A&B.7
Condensation of acid chlorides with homopthhalic acid
is useful for the preparation of 3-substituted isocoumarin
skeleton.8,9 Herein a short and efficient synthesis of 8-deso-
xythunberginol A achieved by using this method and its con-
version into corresponding (±)-3,4-dihydroisocoumarin as a
model for thunberginols C, D, E, G, hydrangenol and phyl-
lodulcin, and the antibacterial activity of the synthesized
compounds are described.
yl)isocoumarin (1a; R=H) from the stem of Homalium
longifolium Benth., a rain forest tree found in the southern
part of the Malay Peninsula.1 Earlier in 1992, M. Yoshikawa
had isolated its 8-hydroxy derivative: thunberginol A (1b;
R=OH) along with thunberginol B, and 3,4-dihydro- deriva-
tives: thunberginols C, D, E and G from the Hydrangeae
Dulcis Folium (amacha or sweet tea) a natural medicine in-
digenous to Japan, prepared from the leaves of Hydrangea
macrophylla SERINGE var. thunbergii MAKINO (Saxi-
fragaceae).2,3 This natural medicine is listed in the Japanese
Pharmacopoeia XII and is extensively used in confectionary,
drinks and food as an oral refrigerant and a sweetener. The
methanolic extract showed potent antiallergic, antibacterial,
antioxidative, antiulcer and cholagoic activities. Two isocou-
marins (thunberginol A and B), four dihydroisocoumarins
(thunberginol C, D, E and G) and some related compounds
were recognized as the antiallergic and antibacterial princi-
ples of this natural medicine.4 Phyllodulcin and hydrangenol
had already been identified as its principal constituents and
sweetening agents.5 Thunberginol A, B inhibited the hista-
Reaction of commercial homophthalic acid with 3,4-
dimethoxybenzoyl chloride at elevated temperature afforded
the 3-(3¢,4¢-dimethoxyphenylisocoumarin (2) in 85% yield.
The isocoumarin showed the characteristic 1H singlet of
isocoumarin moiety (H-4) at d 6.74 in the 1H NMR and at d
100.46 (C-4) and 153.49 (C-3) in the 13C NMR spectrum. IR
spectrum showed the lactonic carbonyl absorption at 1720
cm-1.
Complete demethylation of (2) was proceeded using
BBr3 in dry CH2Cl2 in good yield to furnish 3-(3¢,4¢-dihydro-
xyphenyl)isocoumarin (1a; R=H). The dihydroxy isocou-
marin was characterized by complete absence of both MeO
singlets and the downfield shift of the characteristic H-4 sin-
OH
4
1'
3'
3
OH
2'
1
O
8
1a; R=H
1b; R=OH
glet at d 7.26 in the 1H NMR and at d 99.68 (C-4) in the 13
C
R
O
* Corresponding author. E-mail: aamersaeed@yahoo.com

314 J. Chin. Chem. Soc., Vol. 50, No. 2, 2003
Saeed et al.
NMR spectrum. IR spectrum showed the lactonic carbonyl
absorption at 1722 cm-1.
Biological Activity
The isocoumarins (1a), (2), keto acid (3) and the dihy-
droisocoumarins (4) and (5) were screened in vitro for anti-
bacterial activity13 against various bacterial strains and were
shown to exhibit weak or moderate activity compared to the
standard drug (Table 1). The activity was determined via the
growth inhibition of microorganisms i.e. the zone of inhibi-
tion was measured in millimeters. It is evident from the table
that the dihydroisocoumarin 4 and 5 show more bactericidal
activity compared to corresponding isocoumarin 1a and 2
and the 3¢ and 4¢ hydroxyl groups are important for activity as
the corresponding dimethyl ethers are less active.
Alkaline hydrolysis of the isocoumarin (2) to furnish
the 2-(3,4-dimethoxybenzoylmethyl)benzoic acid (3) was ac-
complished in 80% yield. The keto acid showed the charac-
teristic 2H singlet at d 4.73 (H-4, ArCH2) in the 1H NMR and
that at d 44.02 (C-4) in 13C NMR spectrum.10 DEPT 90° and
135° experiments confirmed these assignments. The ketonic
and carboxylic carbonyl absorptions were observed in IR
spectrum at 1716 and 1685 cm-1, respectively.
Sodium borohydride reduction of the keto acid (3) af-
forded the corresponding racemic hydroxy acid (4a) which
underwent spontaneous cyclodehydration on standing for a
few minutes (as monitored by TLC) to (±)-3-(3¢,4¢-dimeth-
oxyphenyl)-3,4-dihydroisocoumarin (4) without any dehy-
drating agent.11 The methylenes protons (C4) adjacent to the
newly generated chiral centre (C3) in dihydroisocoumarin (4)
showed the diastereotopic effect. Thus, typical ABX split-
ting12 (dddd) of the three 3,4-hydrogens, (ArCH2, AB) and
It is interesting to note that whereas d-phyllodulcin and
dl-hydrangenol possess antifungal activities,14 our compounds
were inactive as antifungal agents indicating that 8-hydroxyl
group is essential for such activity.
EXPERIMENTAL
1
H3 (X) in H NMR at d 3.08-3.39 ppm was observed. Each
hydrogen was split by the other nearly to the same extent and
unequally by the adjacent methine proton. The double dou-
blet of the hydrogen cis to phenyl ring is located slightly up-
field at 3.13-3.08 ppm (Jgem = 16.3 Hz, Jcis = 3.14 Hz) and that
of trans hydrogen is located slightly downfield at 3.39-3.32
ppm (Jgem = 16.54, Jtrans = 12.16 Hz). The H-3 appeared as a
double doublet at 5.52-5.48 ppm with vicinal coupling con-
stant to the trans H4 of 12.04 Hz and to the cis H4 of 3.26 Hz
due to coupling with each of the unequivalent C-4 protons.
13C NMR spectrum showed signals at d 80.03 and 35.76 for
C-3 and C-4, respectively. The d-lactonic carbonyl absorp-
tion appeared at 1721 cm-1 in the IR spectrum.
Melting points of the compounds were determined us-
ing a Gallenkemp melting point apparatus and are uncor-
rected. Commercial ethyl acetate and petroleum ether (60-80
°C) were distilled before use; CH2Cl2 was dried over CaH2 1H
NMR and 13C NMR spectra were recorded respectively on
400 MHz (Bruker, AM-400) and 100 MHz (Bruker, AM-100)
as CDCl3 or DMSO-d6 solutions. Coupling constants J were
measured in Hz. IR spectra were recorded on a Bruker Vector
22 and Mass Spectra (EI, 70 eV) on a MAT 312 instrument.
Flash column chromatography was carried out on Merck
Kieselgel 60.
The final deprotection to unveil the 8-desoxy-3,4-dihy-
drothunberginol A (5), was carried out using boron tribro-
mide. Absence of both MeO singlets and the slight downfield
shift of the characteristic signals in the 1H and 13C NMR spec-
tra confirmed the complete demethylation.
3-(3¢,4¢-Dimethoxyphenyl)isocoumarin (2)
A stirred mixture of homophthalic acid (0.5 g, 2.77
mmol) and 3,4-dimethoxybenzoyl chloride (2.19 g, 9.7
mmol) was heated in an oil bath at 200 °C for 4 h. Flash chro-
matography of the residue (petroleum ether: ethyl acetate
Table 1.
Standard drug
(Imipenem)
Microorganism
2
1a
3
4
5
Escherichia coli
11
00
00
00
10
00
13
12
10
12
13
11
10
8
8
10
13
9
12
00
00
12
12
00
13
10
8
11
17
8
30
31
35
45
29
40
Bacillus subtilis
Shigella flexenari
Staphlococcus aureus
Pseudomonas aeruginosa
Salmonella typhi
(Concentration Used 100 mg/100 mL of DMSO) Zone of Inhibition (mm)

Synthesis of 8-Desoxythunberginol A and Derivatives
J. Chin. Chem. Soc., Vol. 50, No. 2, 2003 315
Scheme I
OMe
OMe
4'
OMe
4
3
ii
i
COOH
3'
Cl
(1a;R=H)
2'
+
OMe
O
1
COOH
2
O
iii
O
OMe
OMe
OMe
OMe
H
4
iv
3
2'
OH
COOH
O
COOH
3
1
4a
OH
OMe
H
O
H
v
OH
OMe
O
O
5
O
4
Reagents and conditions:
(i) 200 °C, 4 h ( 85%); (ii) BBr3/CH2Cl2, -78 °C, R.T., overnight (70%); (iii) 5% KOH / EtOH, 4 h reflux (80%); (iv)
NaBH4 / EtOH, 2 h, R.T. (85%); (v) BBr3, CH2Cl2, -78 °, R.T., overnight (75%)
8:3) followed by recrystallization from MeOH-H2O gave the
isocoumarin 2 (0.66 g, 2.35 mmol, 85%) as orange yellow
needles. m.p. 110 °C; EIMS m/z (%): 282 (M+, 30), 165 (28),
137 (19.8), 117 (100); IR (film, n, cm-1) 2913, 2849, 1720,
1694, 1598, 1572, 1471, 1151, 832 cm-1; 1H NMR (CDCl3, d,
ppm) 3.85 (s, 3H, MeO-3¢) 3.90 (s, 3H, MeO-4¢), 6.74 (s, 1H,
H-4), 7.26 (d, J = 7.6, 1H, H-5), 7.36 (td, J = 2.12, 1H, H-7),
7.57-7.62 (dt, J = 2.12, 1H, H-6), 8.17 (d, J = 7.6, 1H, H-8);
13C NMR (CDCl3, d, ppm): 162.7 (C1, C=O), 153.4 (C3),
100.4 (C4), 137.7 (C4a), 125.6 (C5), 134.7 (C6), 127.6 (C7),
129.4 (C8), 119.9 (C8a), 124.6 (C1¢), 111.9 (C2¢), 149.0
(C3¢), 150.6 (C4¢), 108.0 (C5¢), 118.4 (C6¢), 56.0 (MeO-4),
55.9 (MeO-3). HREIMS: m/z 282.0902 (calcd. for C17H14O4
282.0892).
117 (100); IR (film, n, cm-1) 3538, 3278, 1697, 1631, 1618,
1605, 1524, 1297, 1175; 1H NMR (DMSO-d6, d, ppm): 6.92
(d, 1H, J = 7.8, H-5), 7.26 (s, 1H, H-4), 7.32 (dd, J = 8.4, 2.24,
1H, H-2¢), 7.37 (d, J = 2.24, 1H, H-6¢), 7.60 (dt, J = 1.84, 1.0,
1H, H-7¢), 7.71 (t, J = 7.8, 1H, H-5¢), 7.88 (dt, J = 6.4, 1.64,
1H, H-6¢), 8.19 (d, J = 7.76, 1H, H-8); 13C NMR (DMSO-d6,
d, ppm): 161.53 (C1, C=O), 153.2 (C3), 99.7 (C4), 137.9
(4a), 126.2 (C5), 135.2 (C6), 127.8 (C7), 128.7 (C8), 119.2
(C8a), 122.9 (C1¢), 112.2 (C2¢), 145.6 (C3¢), 147.6 (C4¢),
116.0 (C5¢), 116.9 (C6¢). HREIMS: m/z 254.0571 (calcd. for
C15H10O4 254.0579).
2-(3¢,4¢-Dimethoxybenzoylmethyl)benzoic acid (3)
A stirred solution of 3-(3¢,4¢-dimethoxyphenyl)iso-
coumarin 1a (0.4 g, 1.42 mmol) in ethanol (20 mL) was
treated with 5% KOH (40 mL) and the mixture refluxed for 4
hr. After cooling the reaction mixture, most of the ethanol
was rotary evaporated. Cold water (20 mL) was added and the
mixture acidified with dil. hydrochloric acid when the yellow
solid was precipitated. Filtration followed by drying under
vacuum afforded 3 as a yellow solid. Recrystallized from
MeOH (0.34 g, 1.13 mmol, 80%). m.p 182-184 °C; EIMS m/z
(%): 300 [M+] (11.5), 282 (37.0), 256 (11.6), 178 (100); IR
(film, n, cm-1): 2915, 2849, 1713, 1694, 1601, 1202, 1162;
(DMSO-d6, d, ppm): 3.81 (s, 3H, MeO-3¢), 3.85 (s, 3H,
MeO-4¢), 4.73 (s, 2H, Ar-CH2, H4), 7.11-7.0 (d, J = 8.28, 1H,
H-5), 7.34 (dd, J = 7.72, 2.0, 1H, H-4¢), 7.42 (dt, J = 7.28,
1.26, 1H, H-6), 7.52 (d, J = 7.21, 1H, H-5¢), 7.56 (dt, J = 7.4,
1H, H-7), 7.77 (dd, J = 8.52, 2.0, 1H, H-2¢), 7.95 (dd, J = 7.76,
3-(3¢,4¢-Dihydroxyphenyl)isocoumarin (1a; R=H)
A 1M solution of BBr3 in CH2Cl2 (3.52 mL, 3.52 mmol)
was injected to a stirred solution of 2 (0.25 g, 0.88 mmol) at
-78 °C in dry CH2Cl2 (40 mL) under Ar. The reaction mixture
was allowed to warm to room temperature and stirred over-
night. The reaction mixture was poured into ice-water (50
mL) and the layers separated. The aqueous layer was ex-
tracted with CH2Cl2 (2 ´ 50 mL) and then EtOAc (50 mL).
The combined organic phases were dried (MgSO4) and con-
centrated. Flash chromatography (Petroleum ether:ethyl ace-
tate 7:3) followed by recrystallization from MeOH-H2O af-
forded 1a as intense yellow needles (0.15 g, 0.61 mmol,
70%). m.p. 229-230 °C (lit1. 234-235 °C); EIMS: m/z (%):
254 (M+, 19.4), 252 (14.1), 226 (19), 170 (37), 137 (19.8),

316 J. Chin. Chem. Soc., Vol. 50, No. 2, 2003
Saeed et al.
1.38, 1H, H-8); 13C NMR (DMSO-d6, d, ppm): 168.3 (C1,
COOH), 195.5 (C=O, C3), 44.0 (ArCH2, C4), 148.5 (4a),
132.6 (C5), 131.8 (C6), 130.8 (C7), 130.4 (C8), 152.9 (C8a),
137.3 (C1¢), 129.8 (C2¢), 126.8 (C3¢), 122.5 (C4¢), 110.9
(C5¢), 110.3 (C6¢), 55.7 (MeO-4), 55.5 (MeO-3). For direct
comparison and to avoid confusion, the C/H numbering of
isocoumarins 1a, 2 was maintained in the keto acid 3)
HREIMS: m/z 300.502 (calcd. for C17H16O5 300.0998).
= 16.3 Hz, Jcis = 3.14 Hz, H-4), 3.23-3.21 (dd, 1H, Jgem =
16.54, Jtrans = 12.16, H-4), 5.52-5.48 (dd, J = 12.04, 3.26,
H-3), 6.88 (dd, J = 7.16, 1H, H-5), 6.99 (dd, J = 7.76, 1H,
H-5¢), 7.29 (m, 2H, H-6¢, H-2¢), 7.74 (dt, J = 7.4, J = 0.5, 1H,
H-6*), 7.58 (dt, J = 7.28, J = 2.28, 1H, H-7*), 8.15 (d, J = 7.28,
1H, H-8); 13C NMR (CDCl3, d, ppm): 169.2 (C1), 80.0 (C3),
35.6 (C4), 142.1 (4a), 131.3 (C5), 135.9 (C6), 129.5 (C7),
129.0 (C8), 118.8 (C8a), 139.0 (C1¢), 130.2 (C2¢), 126.9
(C3¢), 127.3 (C4¢) (*interchangeable assignments) HREIMS:
m/z 256.0534 (calcd. for C15H12O4 256.0736).
(±)-3-(3¢,4¢-Dimethoxyphenyl)-3,4-dihydroisocoumarin (4)
Sodium borohydride (0.67 g, 18 mmol) was added por-
tion wise to a stirred solution of 3 (0.2 g, 0.66 mmol) in etha-
nol (25 mL) and water (75 mL). The reaction mixture was
stirred for 2 h at room temperature, diluted with water (150
mL), acidified with conc. HCl and stirred for a further 2 h. It
was then saturated with ammonium sulfate, and extracted
with EtOAc (3 ´ 100 mL). The layers were separated and the
organic layer dried (MgSO4) and concentrated. Flash chro-
matography (Petroleum ether:ethyl acetate 7:2) afforded 4 as
yellow prisms (0.16 g, 0.56 mmol, 85%). m.p. 56-58 °C
EIMS m/z (%): 284 (M+, 56), 147 (14), 118 (100), 90 (59), 89
(15%); IR (film, n cm-1) 2933, 2849, 1684, 1606, 1517, 1460,
1263, 1117, 1026; 1H NMR (CDCl3, d, ppm): 3.13-3.08 (dd,
1H Jgem = 16.3 Hz, Jcis = 3.14 Hz, H-4), 3.39-3.32 (dd, 1H,
Jgem 16.54, Jtrans = 12.16, H-4), 3.91 (s, 3H, MeO-3¢), 3.89 (s,
3H, MeO-4¢), 5.52-5.48 (dd, J = 12.04, 3.26 H-3), 6.88 (dd, J
= 7.16, 1H, H-5), 6.99 (dd, J = 7.76, 1H, H-5¢), 7.29 (m, 2H,
H-6¢, H-2¢), 7.74 (dt, J = 7.4, 0.5, 1H, H-6*), 7.58 (dt, J = 7.28,
J = 2.28, 1H, H-7*), 8.15 (d, J = 7.28, 1H, H-8); 13C NMR
(CDCl3, d, ppm): 165.5 (C1), 80.0 (C3), 35.7 (C4), 149.5
(4a), 131.2 (C5), 133.9 (C6), 129.5 (C7), 129.0 (C8), 118.8
(C8a), 139.2 (C1¢), 130.6 (C2¢), 127.9 (C3¢), 125.3 (C4¢),
109.6 (C5¢), 111.1 (C6¢), 56.1 (MeO-4), 55.0 (MeO-3).
HREIMS: m/z 284.1031 (calcd. for C17H16O4 284.1049).
ACKNOWLEDGEMENT
The author gratefully acknowledges a Georg Forster
research fellowship from the Alexander von Humboldt Foun-
dation Germany. The author is indebted to the Instrumental
Research Services, Institut für Organishe Chemie, Universität
Hannover, Germany for the spectral analyses.
Received May 21, 2002.
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(±)-3-(3¢,4¢-Dihydroxyphenyl)-3,4-dihydroisocoumarin (5)
A 1M solution of BBr3 in CH2Cl2 (1.10 mL, 1.1 mmol)
was injected to a stirred solution of (0.1 g, 0.35 mmol) in at
-78 °C in dry CH2Cl2 (4 mL) under Ar. The reaction mixture
was allowed to warm to room temperature and stirred over-
night. The layers were separated and the aqueous layer was
extracted with EtOAc (3 ´ 50 mL). The combined organic
phases were dried (MgSO4) and concentrated. Flash chroma-
tography (Petroleum ether: ethyl acetate 7:3) followed by
recrystallization from MeOH afforded 5 as yellow prisms
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256 (M+), 254 (18.6), 138 (19.8), 118 (100), 90 (22.4); IR
(film, n, cm-1) 2913, 2849, 1720, 1694, 1598, 1572, 1471,
1151, 832; 1H NMR (CDCl3 d, ppm): 2.94-3.02 (dd, 1H, Jgem

Synthesis of 8-Desoxythunberginol A and Derivatives
J. Chin. Chem. Soc., Vol. 50, No. 2, 2003 317
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