Synthesis and biological evaluation of functionalized coumarins as acetylcholinesterase inhibitors

Article abstract of DOI:10.1016/j.ejmech.2005.07.014

Three series of functionalized coumarin compounds were designed and prepared as cholinesterase (AChE and BuChE) inhibitors. The biological profile against AChE and BuChE of the prepared compounds was determined. Compound 7b exhibited a mixed-type of AChE

Full text of DOI:10.1016/j.ejmech.2005.07.014

European Journal of Medicinal Chemistry 40 (2005) 1307–1315
www.elsevier.com/locate/ejmech
Original article
Synthesis and biological evaluation of functionalized coumarins as
acetylcholinesterase inhibitors
Qiong Shen a,b, Quan Peng a, Jialiang Shao a, Xufeng Liu a, Zhishu Huang b, Xianzhang Pu b,
Lin Ma a,*,Yue-Ming Li c,*, Albert S.C. Chan b,c, Lianquan Gu a,b,
*
a School of Chemistry and Chemical Engineering, Sun Yat-Sen University, Guangzhou 510275, China
b School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou 510275, China
c Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University, Kowloon, Hong Kong
Received 5 January 2005; received in revised form 27 June 2005; accepted 1 July 2005
Available online 21 September 2005
Abstract
Three series of functionalized coumarin compounds were designed and prepared as cholinesterase (AChE and BuChE) inhibitors. The
biological profile against AChE and BuChE of the prepared compounds was determined. Compound 7b exhibited a mixed-type of AChE
inhibitor with IC50 value for the AChE inhibition of 0.19 0.01 µM and a high selectivity for AChE/BuChE, and compound 6b acted as
non-competitive AChE inhibitor with IC50 value of 0.43 0.02 µM. Structure–activity relationships (SARs) of prepared compounds were
discussed.
© 2005 Elsevier SAS. All rights reserved.
Keywords: Functionalized coumarins; Cholinesterase inhibitor; Synthesis
1. Introduction
pidium and ensaculin (Fig. 1) have shown to induce a modest
improvement in memory and cognitive functions [6]. How-
ever, these compounds do not appear to prevent or slow down
the progressive neurodegeneration. The optimal approach is
also being followed in searching new agents which can be
used to treat AD [7].
Recent study showed thatAChE could also play a key role
in accelerating Ab plaques deposition [8,9]. It is likely that
AChE interacts with Ab and promotes amyloid fibril forma-
Alzheimer’s disease (AD) is a progressive, chronic, neu-
rodegenerative disorder that is characterized by a loss of
memory and cognition, severe behavioral abnormalities and
ultimately death. It is the fourth leading cause of death in
Western countries. As general health care improves, and the
proportion of the elderly increases, the number ofAD patients
is anticipated to increase dramatically [1]. The neuropatho-
logical hallmarks of the disease is the presence of numerous
senile amyloid b-peptide (Ab) plaques, neurofibrillary tangles
(NFT), the dramatic loss of synapses and degeneration or atro-
phy of the basal forebrain cholinergic neurons [2]. The loss
of basal forebrain cholinergic cells results in sharp reduction
in acetylcholine (ACh), which is believed to play an impor-
tant role in the cognitive impairment associated with AD
[3–5]. On this basis, the cholinergic hypothesis has become
the leading strategy for the development of anti-AD agents.
Several anti-AChE agents such as tacrine, donepezil, pro-
* Corresponding authors.
E-mail addresses: bcyli@polyu.edu.hk (Y.-M. Li), cedc42@zsu.edu.cn
(L. Gu).
Fig. 1. Structures of compounds which have shown anti-AChE activities.
0223-5234/$ - see front matter © 2005 Elsevier SAS. All rights reserved.
doi:10.1016/j.ejmech.2005.07.014

1308
Q. Shen et al. / European Journal of Medicinal Chemistry 40 (2005) 1307–1315
Scheme 1. Preparation of 6a–6n.
tion through a pool of amino acids located in proximity of the
peripheral anionic binding site (PAS) of the enzyme [10].
Moreover, it has been shown that molecules are able to inter-
act either exclusively with PAS or with both catalytic and
peripheral binding site, which can prevent the pro-aggregation
of AChE toward Ab [8]. Compounds showing peripheral and
dual binding with AChE are intriguing and could represent a
new type of therapeutic agents through the prevention of Ab
aggregation.
In view of the above reasons, we focused on the develop-
ment of more active and selective compounds which are able
both to reduce the cognitive deficiency and to decrease Ab
production or aggregation. To this aim, three series of new
inhibitors were designed and prepared, their anti-AChE and
BuChE activities were tested and the SARs were discussed.
The results of the in vitro assays compared favorably with
the corresponding data of propidium and tacrine, indicating
that 6b and 7b might be regarded as the prototype of a new
class of compounds endowed with AChE inhibitory profile
that may favor for the therapeutic application in the treat-
ment of AD.
pounds have demonstrated high anti-AChE potency [12,16].
Studies also showed that benzylamine substructure were
found in many potent AChE inhibitors [17].
3. Chemistry
In general, compounds of series 1 (compounds 6a–6n)
were synthesized via a six-step procedure as described in
Scheme 1. Resorcinol and ethylacetoacetate were first treated
under Pechmann conditions [18] to give substituted cou-
marin l, and compound 1 was reacted with 2-chloro-
cyclohexanone under Williamson conditions [19] to give the
corresponding oxo ether 2 under refluxed conditions.
Reagents and conditions: (1) CH3COCH2COOC2H5,
H2SO4, 0–5 °C; (b) dry K2CO3, dry acetone, KI,
reflux; (c) NaOH, reflux, 2 h; (d) DDQ, Pd/C, toluene, reflux,
8 h; (e) SeO2, xylene, reflux, 29 h; (f) 95% ethanol, ary-
lamine, reflux, 2 h.
Thereafter, cyclization of oxo ether 2 by heating in strong
alkaline solution afforded compound 3. This compound was
aromatized in toluene with 2,3-dichloro-5,6-dicyano-1,4-
benzoquinone (DDQ) and a catalytic amount of Pd/C to give
the corresponding compound 4. Selective oxidation of com-
pound 4 to 4-carbaldehyde 5 was achieved using selenium
dioxide as oxidant in refluxing xylene. Finally, then reaction
of 5 with arylamine in 95% ethanol gave target compounds
6a–n as pure E-isomers [20].
Compounds 6 were reduced with NaBH4, producing com-
pounds of series B (7a–c) (Scheme 2).
Reagents and conditions: (a) THF, methanol, NaBH4,
0–10 °C.
To further verify the above hypothesis, another new target
compound 11a was also prepared according Scheme 3. The
compound 11a was obtained using the method similar to that
as shown in Scheme 1. Intermediate 9 was treated with 1,3-
dibromo-5,5-dimethylhydantoin (BDH) and a catalytic
amount of benzoyl peroxide in refluxing CCl4, bromination
2. Inhibitor design
A set of molecules composed of three moieties optimal
for the binding at PAS, catalytic site and linked by an appro-
priate distance were designed to obtain selective nonhepato-
toxic AChE inhibitor. The three moieties were the benzy-
lamino or arylamino group, benzofuran group and coumarin
(2H-2-Chromenone) skeleton. Coumarins were selected as
the parent nucleus for several reasons. At first, coumarins are
members of the set of compounds called benzopyrones which
display a variety of pharmacological properties [11], and some
functionalized coumarins are presented as potentAChE inhibi-
tor [12,13]. The second reason is that certain functionalized
coumarin derivatives are monoamine oxidase B (MAO-B)
inhibitors and have also been proposed for the treatment of
AD [14,15]. The third reason is the fact that coumarin com-

Q. Shen et al. / European Journal of Medicinal Chemistry 40 (2005) 1307–1315
1309
Scheme 2. Preparation of compounds 7.
Scheme 3. Preparation of compound 11a.
of the methyl substituent on the furan ring gave
6-bromomethylfuro[3,2-g] coumarin (10). Bromination of
4,6-dimethylfuro[3,2-g] coumarin (compound 9) occurs only
in the 6-methyl group, and the 4-methyl group is intact dur-
ing the reaction [21]. Then the bromo-derivative 10 was con-
densed with aniline to afford the target compound 11a.
Reagents and conditions: (a) dry K2CO3, KI,
CH3COCH2Cl, reflux, 18 h; (b) NaOH, reflux, 2 h; (c) BDH,
(PhCOO)2, CCl4, 6 h; (d) aniline, toluene, CH3CN, pow-
dered KOH, reflux.
5. Discussion
According to the screening data, the analogues of com-
pound 6 which possessed strong electron-donating groups,
such as –OCH3 in positions 2′ and 4′, –NH2 in position 2′,
–OH in position 2′, exhibited anti-AChE activity, while com-
pound 6b with a strongest electron-donor group –OCH3 in
the 2′-position exhibited the highest activity against AChE.
Analogues of compounds 6 with weaker electron-donating
group -CH3 in position 2′ and 4′ showed a decreasing inhibi-
tory activity, which suggests that strong electron-donating
group in these compounds could help them to access to the
enzyme-binding site. Possess two substituent groups in 2′ and
4′ positions exhibited a decreasing in AChE inhibitory effect
compared to those bearing either one group in either 2′- or
4′-positions. The congeners with bulkier substituent (com-
pound 6j) led to a huge decrease in activity than that bearing
small ones (compound 6i), indicating the steric hindrance
could limit the access of compounds to the enzyme-binding
site. These phenomena implied that both the steric and elec-
tronic effects might play a significant role in influencing the
activity.
Compounds 7a, 7b and 7c showed significantly increased
inhibitory activity compared with their unsaturated counter-
parts 6a, 6b and 6c. These results suggested that the thus com-
pounds may have strong interaction with AChE contributing
from their better binding affinity to AChE. Interesting that
the compounds 7a, 7b and 7c were less potent in inhibitory
activity against BuChE comparing withAchE, indicating that
these compounds can serve as selective inhibition agents for
AChE over BuChE. Comparing the selectivity of 7a, 7b and
7c for AChE/BuChE, it is clear that the 2-methoxy group on
the aromatic ring is endowed with optimal electronic proper-
ties, that agreement with above SARs results.
4. Biology
The inhibitory activities of the thus prepared compounds
against AChE and BChE were studied for determining the
rate of hydrolysis of acetylthiochoine (ATCh) and butylthio-
choline (BuTCh) in comparison with reference compound
tacrine and propidium using the method of Ellman et al. [22].
The results were listed in Table 1 as IC50 values. The nature
of AChE inhibition caused by these compounds was investi-
gated by the graphical analysis of steady-state inhibition data
(only 6b and 7b were shown in Fig. 1), and the Ki values and
the type of inhibition of the selected compounds were reported
in Table 2 along with the data for tacrine and propidium. Struc-
tural features of the three types of compounds are shown in
Fig. 2.
Table 1
AChE and BuChE inhibitory Activities (IC50, µM) of prepared compounds
Compound
6b
7b
Ki S.E. (nM)
0.20 0.02
Type of inhibition
Non-competitive
Mixed
0.061 0.003
8.13 1.50
Propidium
Non-competitive
Mixed
Tacrine
0.15 0.016
Compound 11a showed less activity than compound 6b
and 7b as an AChE inhibitor, suggested that the C-type com-
a Not determined.

1310
Q. Shen et al. / European Journal of Medicinal Chemistry 40 (2005) 1307–1315
Table 2
Inhibition constant and type of inhibition of selected compoundsa
Compound
Structural Feature
R1
R2
IC50 SEAChE
(µM)
IC50 SEBuChE
(µM)
nda
Selectivity for
AChE/BChE
6a
A
H
H
1.08 0.1
0.43 0.02
0.75 0.02
1.30 0.04
1.87 0.1
2.53 0.1
0.67 0.02
3.01 0.2
2.01 0.1
3.21 0.2
1.88 0.1
4.85 0.2
7.47 0.3
0.88 0.02
0.49 0.02
0.19 0.01
0.21 0.01
3.02 0.2
15.30 0.3
0.29 0.03
6b
6c
6d
6e
6f
6g
6h
6i
6j
6k
6l
6m
6n
7a
7b
7c
OCH3
H
H
< 20
OCH3
H
< 20
CH3
H
nd
CH3
CH3
H
nd
CH3
OH
H
nd
< 20
OH
CH3
C(CH3)3
SH
nd
OH
OH
H
nd
nd
nd
COOH
H
H
nd
COOH
H
nd
NH2
H
< 20
B
C
H
58.6 4.5
185 13.2
37.8 3.3
25.30 2.2
nd
119.60
973.68
180.0
8.38
OCH3
H
H
OCH3
H
11a
Propidium
Tacrine
H
0.0568 0.2
0.20
a Inhibition constants were calculated from kinetic data.
pound have weaker interaction withAChE, due to their poorer
binding affinity to AChE.
The kinetic studies showed that the selected compound 7b
display the mixed-types of inhibition similar to that of tacrine
[23], and the compound 6b showed as non-competitive bind-
ing mode that similar to that of propidium [23,24].
Compounds 7a, 7b and 7c showed significantly increased
activity compared with their unsaturated counterparts 6a, 6b
and 6c. These results indicated that the B-type compounds
have strong interaction withAChE that may due to better bind-
ing affinity to AChE. Meanwhile, compounds 7a, 7b and 7c
were less potent of inhibitory activity against BuChE than
that of AchE, indicating that the B-type compounds exhibit a
good selectivity for AChE over BuChE.
Compound 11a showed slightly (two to threefold) less
activity than compound 6a as an AChE inhibitor, indicating
that the C-type compound have weaker binding affinity to
AchE comparing with type- A or B compounds.
tacrine [23], while the other compounds showed the binding
mode of non-competitive similar to that of propidium [23,24].
The results from SARs analysis, biological assay data and
kinetic studies that provided coherent information concern-
ing the inhibitory action on AChE or BuChE and the selec-
tivity for AChE/BuChE, those to be favor for design a new
type of the prototypes.
Interestingly, Inestrosa [8] and Bartolini [23] recently
pointed out a direct link between AChE and Ab aggregation.
AChE was shown to exert an Ab pro-aggregation mediated
by its peripheral binding site [10]. We found that compound
6b with the binding mode similar to that of propidium and
showed the potency inhibitory activity against AChE [25].
This finding is particular interesting in the connection to the
Alzheimer’s disease, because recent observations show that
peripheral AChE inhibitors might decrease b-amyloid depo-
sition [8,23].
In addition, compound 7b showed a strong potency inhibi-
tory activity against AChE which was comparable to that of
tacrine [8,23,25], and also demonstrated a high selectivity
for AChE over BChE. This remarkable selectivity features
may have some implication on the future design and prepa-
ration of AChE inhibitors [23]. Actually, a peculiar structural
difference between the two types of cholinesterases is the lack
Reciprocal plots of initial velocity and substrate concen-
tration are reported. Lines were derived from a weighted least-
squares analysis of data points. The plots show non-
competitive inhibition for 6b, mixed-type inhibition for 7b.
The kinetic analysis showed that compounds7a, 7b, 7c and
11a, display the mixed-types of inhibition similar to that of
Fig. 2. Structural features of three types of compounds.

Q. Shen et al. / European Journal of Medicinal Chemistry 40 (2005) 1307–1315
1311
of the PAS (in particular of Trp286) in BuChE [26], that seems
6. Conclusion
to prevent the interaction of BuChE with Ab, resulting inef-
fective on Ab aggregation [8]. For instance, tacrine, a non-
competitive mixed-type AChE inhibitor, binds more tightly
to BuChE and is almost inactive in inhibiting the AChE-
promotedAb aggregation [23,26]. This inhibitor mostly inter-
acted with AChE catalytic site [27], as it likely did with
BuChE [26]. In contrast, propidiun, a well-known AChE
inhibitor which binding exclusively at the PAS, was almost
inactive against BuChE and strongly inhibits the AChE-
inducedAb aggregation [23]. The results presented in Tables 1
and 2 suggested the phenomenological correlation between
AChE selectivity and inhibition of AChE-induced Ab aggre-
gating action. A inhibitors of AChE that to be strongly inter-
acting with the PAS, that would be shown highAChE/BuChE
selectivity, and eventually performed inhibitory effect against
AChE-induced Ab aggregation Fig. 3.
In conclusion, we have prepared three series of com-
pounds and studied the enzyme-inhibitor binding mode and
the structure–activity relationship. We hope that the current
work could shed light on further investigations on the poten-
tial AChE inhibitors with high inhibitory activity and good
selectivity for AchE over BuChE.
7. Experimental section
7.1. Chemistry
Melting points were measured using an electro-thermal
8103 apparatus and were uncorrected. IR spectra were taken
with Perkin-Elmer 398 spectrometers. 1H NMR and 13C NMR
spectra were recorded on a Varian Unity INOVA 500 MHz
spectrometer with TMS as internal standard. All chemicals
used were of analytical grade. TLC monitored the progress
of reactions using pre-coated 60 F254 silica gel plates
(0.25 mm, Merck). Elemental analyses (C, H, N) were per-
formed on a Perkin-Elmer 240 elemental analyzer, and the
results were within 0.4% of the theoretical values. ESI-MS
spectra were performed on a Thermo Finigan LCQ DECA
XP ion trap mass spectrometer.
7.2. Synthesis of A-type compounds (6a–n)
4-Methyl-7-hydroxycoumarin (1). Compound 1 was pre-
pared based on a literature method [18].Yield 80%, m.p. 190–
192 °C; IR(KBr) v˜
: 3159, 1679, 1652, 1600, 1451 cm–1;
FAB-MS m/z:177([M + H]+), 154([M–OH]+); 1H NM-
R(CDCl3)d: 9.31 (s, 1H, 7-OH), 7.61 (d, 1H, J = 8.5, 1.0 Hz,
5-H), 6.85 (m, 1H, 6-H), 6.74 (d, 1H, J = 2.5 Hz, 8-H), 6.07
(s, 1H, 3-H), 2.42 (s, 3H, 4-CH3), 13C NMR (CDCl3) d: 161.8
(C2), 160.9 (C7), 156.3 (C8a), 153.6 (C4), 113.4 (C4a), 111.8
(C5), 106.4 (C6), 103.3 (C8), 96.6 (C3), 18.5 (CH3).
4-Methyl-7-(2′-oxocyclohexyloxy)coumarin (2). Com-
pound 2 was prepared based on a literature method [19].Yield
80%, m.p. 164–166 °C; IR(KBr) v˜: 3050, 2922, 2853, 1716,
1605, 1574, 1432, 747, 717 cm–1; FAB-MS m/z: 273 ([M +
H]+), 154([M-C6H7H2O]+); 1H NMR (DMSO-d6) d: 7.62 (d,
1H, J = 8.5 Hz, 5-H), 6.88 (dd, 1H, J = 6.0, 2.5 Hz, 6-H),
6.75 (s, 1H, 8-H), 5.13 (m, 1H, 1′-H), 2.76–2.65 (m, 2H, 3′-H),
2.48–2.43 (m, 2H, 6′-H), 2.42–2.38 (m, 3H, 4-CH3), 2.12–
1.89 (m, 4H, 4’-H, 5’-H); 13C NMR (CD3OD) d: 200.8 (C2′),
162.2 (C2), 160.8 (C7), 156.1 (C8a), 153.6 (C4), 126.8 (C5),
114.4 (C4a), 113.6 (C6), 112.4 (C8), 102.7 (C3), 81.1 (C1′),
41.3 (C3′), 35.0 (C4′), 28.1 (C5′), 24.1 (C6′), 18.5 (CH3).
6,7,8,9-Tetrahydro-4-methyl-2H-benzofuro [3,2-g]-1-
benzopyran-2-one (3). A solution of 2 (10.0 mmol) in 0.8 M
NaOH (1L) was refluxed in the dark for 3 h under N2. The
solution was cooled, diluted with water (1 l), and acidified
with diluted HCl. The precipitate obtained was collected and
crystallized from MeOH to give 3. Yield 95%, m.p. 190–
Fig. 3. Steady state inhibition of 6b and 7b against AchE.

1312
Q. Shen et al. / European Journal of Medicinal Chemistry 40 (2005) 1307–1315
3067, 3001, 2955, 2906, 2832, 1718, 1643,1602, 1569, 1514,
192 °C; IR (KBr) ˜v: 3053, 2934, 2886, 1711, 1628, 1573,
1431, 1247, 1222, 1138, 781, 745 cm–1; FAB-MS m/z: 370
([M + H]+), 171 ([M-C6H4.C7H7O]+); 1H NMR (THF-d8) d:
9.77 (s, 1H, HC=N), 8.83 (s, 1H, 5-H), 8.02(d, 1H, J = 7.0 Hz,
9-H), 7.58(d, 1H, J = 8.5 Hz, 6-H), 7.55 (s, 1H, 11-H), 7.47
(t, 1H, J = 15.0 Hz, 7-H), 7.37(t, 1H, J = 15.0 Hz, 8-H), 7.28–
7.11 (m, 2H, 4′-H, 6′-H), 7.00 –6.86 (m, 2H, 3′H, 5′-H), 6.76
(s, 1H, 3-H), 3.97 (s, 3H, OCH3); 13C NMR (THF-d8) d: 160.2
(C2), 159.0 (HC=N), 158.6 (C10a), 158.2 (C2′), 155.6 (C9a),
154.1 (C11a), 146.6 (C1′), 141.1 (C5b), 129.2 (C3), 128.5 (C4′),
124.3 (C6), 122.2 (C4a), 121.7 (C6), 121.5 (C7), 121.2 (C5),
120.5 (C9), 119.2 (C6′), 117.9 (C5a), 117.4 (C4), 113.0 (C3′),
112.4 (C5′), 100.5 (C11), 56.3(OCH3). C, H, N for C23H15NO4.
4-((4-Methoxyphenylimino)methyl)-2H-benzofuro[3,2-
1458, 1292, 875, 804 cm–1; FAB-MS m/z: 255 ([M + H]+),
154([M-C6H9H2O]+); 1H NMR (CDCl3) d: 7.46 (s, 1H, 5-H),
7.28 (s, 1H, 11-H), 6.17(s, 1H, 3-H), 2.75–2.72 (m, 2H, 9-H),
2.63–2.61(m, 2H, 6-H), 2.45 (s, 3H, 4-CH3), 1.98–1.85 (m,
4H, 7-H, 8-H); 13C NMR (CDCl3) d: 161.2 (C2), 156.2 (C10a),
155.8 (C9a), 152.8 (C11a), 150.9 (C4a), 126.2 (C5b), 115.6
(C5a), 113.1 (C5), 112.6 (C11), 99.2 (C3), 23.3 (C6), 22.6 (C8),
22.3 (C7), 20.2 (C9), 19.1 (CH3).
4-methyl-2H-benzofuro[3,2-g]-1-benzopyran-2-one (4).A
mixture of 3 (10 mmol), 2,3-dichloro-5,6-dicyano-1,4-
benzoquinone (25 mmol) and a catalytic amount of Pd/C in
anhydrous toluene (500 ml) was refluxed for 6 h. After cool-
ing, the solid was filtered off and the solvent evaporated under
reduced pressure. The residue was purified by column chro-
matography and crystallized from MeOH to give 4.Yield 98%,
g]chromen-2-one(6c). m.p. 244–245 °C; IR(KBr) v˜: 3064,
2951, 2842, 1705, 1643, 1604, 1554, 1503, 1436, 1250, 1197,
1133, 875 cm–1; FAB-MS m/z: 370 ([M + H]+),154 ([M-
C6H4OC7H7NO]+); 1H NMR (THF-d8) d: 9.46 (s, 1H,
HC=N), 8.93 (s, 1H, 5-H), 8.08 (d, 1H, J = 8.5 Hz, 6-H), 7.60
(s, 1H, 11-H), 7.51–7.47 (m, 3H, 8-H, 2′-H, 6′-H), 7.38(t,
1H, J = 7.5 Hz, 7-H), 7.03 (dd, 2H, J = 6.5, 7.0 Hz, 3′-H,
5′-H), 6.83 (s, 1H, 3-H), 3.84 (s, 3H, OCH3); 13C NMR (THF-
d8) d: 161.2 (C2), 160.3 (C10a), 158.8 (C9a), 158.3 (C11a), 155.8
(C4′), 155.5 (HC=N), 146.7 (C1′), 144.5 (C5b), 128.5 (C3),
124.6 (C4a), 124.3 (C6), 123.9 (C2′,C6′), 122.3 (C4), 121.7
(C8), 119.8 (C7), 118.3 (C5), 115.4 (C3′,C5′), 114.4 (C5a),
112.5 (C9), 100.7 (C11). C, H, N for C23H15NO4:
m.p. 228–230 °C; IR(KBr) v˜: 3099, 2940, 2870, 1715, 1617,
1553, 1513, 1437, 1294, 1265, 1206, 1148, 1082, 842 cm–1;
1
FAB-MS m/z: 251 ([M + H]+),154 ([M-C6H92H2O]+); H
NMR (CDCl3) d: 8.11 (s, 1H, 5-H), 7.98 (dd, 1H, J = 7.0,
1.0 Hz, 6-H), 7.58 (d, 1H, J = 8.0 Hz, 9-H), 7.51 (t, 1H,
J = 7.5 Hz, 7-H), 7.50 (s, 1H, 11-H), 7.40 (t, 1H, J = 15.0 Hz,
8-H), 6.29 (s, 1H, 3-H), 2.57 (s, 3H, 4-CH3); 13C NMR
(CDCl3) d: 160.8 (C2), 157.9 (C10a), 157.2 (C9a), 153.4 (C11a),
152.5 (C4), 127.8 (C6), 123.5 (C5b), 123.2 (C8), 121.6 (C4a),
120.6 (C7), 116.2 (C5a), 116.0 (C3), 113.5 (C5), 111.9 (C9),
100.2 (C11), 19.2 (CH3).
4-((o-Tolylimino)methyl)-2H-benzofuro[3,2-g]chromen-
2-one (6d). m.p. 260–262 °C; IR(KBr) v˜: 3059, 3025, 2914,
4-Carbaldhyde-2H-benzofuro [3,2-g]-1-benzopyran-2-
one (5). Powdered SeO2 (3.30 g, 30 mmol) was added to a
solution of 4 (20 mmol) in 20 ml of hot dry xylene and the
mixture were refluxed for 24 h with vigorous stirring. The
reaction mixture was filtered to remove black Se, and the deep
orange filtrate was allowed to stand overnight. Almost pure
crystals of 5 could be separated from the solution.Yield 90%,
1703, 1644, 1604, 1560, 1504, 1433, 1266, 1200, 1137,
742 cm–1; FAB-MS m/z: 354([M + H]+), 154([M-
C6H4O.C7H7N]+); 1H NMR (CDCl3) d: 9.66 (s, 1H, HC=N),
9.08 (s, 1H, 5-H), 7.68 (s, 1H, 11-H), 7.65 (d, 1H, J = 9.5 Hz,
9-H), 7.56–7.49 (m, 2H, 7-H, 8-H), 7.49–7.39 (m, 2H, 4′-H,
6′-H), 7.31(m, 2H, 3′-H, 5′-H), 6.97 (s, 1H, 3-H), 2.52–
2.51(m, 3H, CH3); 13C NMR (THF-d8) d: 160.8 (C2), 158.1
(C10a), 157.3 (HC=N), 155.5 (C9a), 151.0 (C11a), 149.0 (C1′),
147.0 (C2′), 138.9(C5b), 130.8 (C4′), 130.0 (C4a), 128.9 (C3),
124.6 (C5′), 124.4 (C5a), 122.7 (C6), 122.4 (C4), 122.0 (C8),
120.0 (C3′), 119.9 (C6′), 117.9 (C7), 112.6 (C5), 100.8 (C9),
88.3 (C11), 21.3 (CH3). C, H, N for C23H15NO3:
m.p. 259–261 °C; IR(KBr)˜v: 3056, 2854, 2717, 1730, 1639,
1600, 1586, 1453, 1433, 1402, 1380, 1339, 1302, 1267, 1212,
1048, 850, 800 cm–1; FAB-MS m/z: 265 ([M + H]+), 168
([M-C6H8O]+); 1H NMR(CDCl3) d: 10.17 (s, 1H, CHO), 9.22
(s, 1H, 5-H), 8.04 (d, 1H, J = 7.5 Hz, 6-H), 7.61–7.22 (m, 4H,
7-H, 8-H, 9-H, 11-H), 6.88 (s, 1H, 3-H); 13C NMR (CDCl3)
d: 191.9 (CHO), 160.3 (C2), 158.2 (C10a), 157.3 (C9a), 154.3
(C11a), 143.9 (C4), 128.2 (C3), 124.3 (C5b), 123.8 (C4a), 123.1
(C5a), 122.7 (C6), 121.1 (C8), 118.4 (C7), 111.9 (C5), 110.6
(C9), 100.5 (C11).
General procedure for preparation of compounds 6.A mix-
ture of 5 (5 mmol) and arylamine (15 mmol) in 95% ethanol
(25 ml) was refluxed for 2 h. After cooling, the precipitate
was collected, washed with 95% ethanol, and recrystallized
to give 6 in the yield of 35–90%.
4-((Phenylimino)methyl)-2H-benzofuro[3,2-g]chromen-
2-one(6a). m.p. 238–240 °C; FAB-MS m/z: 340 ([M + H]+);
1H NMR(THF-d8) d: 9.57 (s, 1H, HC=N), 8.88(s, 1H, 5-H),
8.05(d, 1H, J = 9.5 Hz, 9-H), 7.58 (m, 2H, 6-H, 11-H), 7.50–
7.42 (m, 5H, 2′-H, 3′-H, 4′-H, 5′-H, 6′-H), 7.40–7.32 (m, 2H,
7-H,8-H), 6.85 (s, 1H, 3-H). C, H, N for C22H13NO3:
4-((2-Methoxyphenylimino)methyl)-2H-benzofuro[3,2-
4-((p-Tolylimino)methyl)-2H-benzofuro[3,2-g]chromen-
2-one (6e). m.p. 226–228 °C; IR (KBr) ˜v:3055, 1706, 1642,
1604, 1564, 1486, 1267, 1138, 884, 845 cm–1. FAB-MS m/z:
354([M + H]+),154([M-C6H4O.C7H7N]+); 1H NMR (CDCl3)
d: 9.70 (s, 1H, HC=N), 8.98 (s, 1H, 5-H), 8.05 (d, 1H,
J = 8.0 Hz, 9-H), 7.84 (s, 1H, 11-H), 7.74 (d, 1H, J = 8.0 Hz,
9-H), 7.57 (t, 1H, J = 15.0 Hz, 8-H), 7.46 (t, 1H, J = 15.0 Hz,
7-H), 7.38–7.27 (m, 4H, 2′-H, 5′-H, 6′-H), 7.02 (s, 1H, 3-H),
2.50 (s, 3H, CH3); 13C NMR (CH3OD) d: 166.5(C2), 165.9
(C10a), 163.2 (C9a), 158.4(C11a), 155.2 (C1′), 142.2 (HC=N),
139.9 (C4′), 137.5 (C1a), 136.9(C3), 136.3 (C5b), 133.3 (C4a),
132.2 (C5a), 130.4 (C3′, C5′), 128.4 (C6), 127.0 (C2′, C5′),
126.3 (C8), 122.5 (C7), 121.2 (C5), 109.6 (C9), 104.8 (C11),
27.3(CH3). C, H, N for C23H15NO3.
4-((2,4-Dimethylphenylimino)methyl)-2H-benzofuro[3,2-
g]chromen-2-one(6b). m.p. 172–174 °C; IR(KBr)˜v: 3331,
g]chromen-2-one (6f). m.p. 247–249 °C; 1H NMR (THF-d8)

Q. Shen et al. / European Journal of Medicinal Chemistry 40 (2005) 1307–1315
1313
d: 9.78 (s, 1H, CH = N), 8.84 (s, 1H, 5-H), 8.04 (d, 1H,
J = 7.2 Hz, 9-H), 7.64 (s, 1H, 6′-H), 7.62 (d, 1H, J = 5.0 Hz,
6-H), 7.50–7.05 (m, 5H, 7-H, 8-H, 11-H, 3′-H, 5′-H), 6.84 (s,
1H, 3-H), 2.53 (s, 3H, 2′-CH3), 2.40 (s, 3H, 4′-CH3). C, H
and N for C24H17NO3.
(2-Hydroxyphenylimino)methyl)-2H-benzofuro[3,2-
g]chromen-2-one (6g). m.p. 258–259 °C; IR (KBr) ˜v: 3446,
3473, 3307, 3077, 1716, 1643, 1611, 1574, 1488, 1437,1288,
1215, 1143, 792, 750 cm–1; FAB-MS m/z: 384 ([M + H]+),
154 ([M-C6H4O.C7H5O2N]+); 1H NMR (Pyr-d5) d: 10.32 (s,
1H, COOH), 9.34 (s, 1H, HC=N), 8.97 (s, 1H, 5-H), 8.40 (d,
1H, J = 8.0 Hz, 6-H), 8.56 (s, 1H, 11-H), 8.25 (d, 1H,
J = 8.0 Hz, 3′-H), 7.98 (d, 1H, J = 7.5 Hz, 9-H), 7.92 (d, 1H,
J = 8.0 Hz, 6′-H), 7.66–7.57 (m, 2H, 7-H, 8-H), 7.52–7.44
(m, 2H, 4′-H, 5′-H), 7.20 (s, 1H, 3-H); 13C NMR (Pyr-d5) d:
171.6 (COOH), 158.9 (C2), 157.5 (C10a), 154.8 (HC=N),
3064, 1711, 1644, 1593, 1550, 1483, 1454, 1249, 1200, 1139,
1105, 799, 744.5 cm–1; FAB-MS m/z: 356([M + H]+), 171
1
([M-C6H4.C6H5O]+); H NMR (THF-d8) d: 9.41 (s, 1H,
152.7 (C9a), 150.1 (C1), 149.1 (C11a), 148.8 (C5b), 144.1 (C4a)
,
134.1 (C5′), 132.5 (C4′), 131.1 (C2′), 128.5 (C3′), 128.4 (C5a),
124.2 (C3), 124.1 (C6), 123.9 (C1′), 122.3 (C4), 121.5 (C8),
118.7 (C6′), 117.0 (C7), 115.5 (C9), 112.2 (C5), 100.6 (C5′),
96.5 (C11). C, H and N for C23H13NO5.
4-((2-Oxo-2H-benzofuro [3,2-g] chromen-4-yl) methyl-
eneamino)benzoic acid (6m). m.p. 260–262 °C; IR (KBr) v˜:
3067, 2979, 2886, 2665, 2545, 1708, 1644, 1603, 1562, 1429,
1324, 1289, 1139, 1108, 857, 817 cm–1. ESI-MS m/z: 382
([M-H]-); 1H NMR (Pyr-d5) d: 11.74(s, 1H, COOH), 9.64 (s,
1H, HC=N), 8.99 (s, 1H, 5-H), 8.53 (d, 1H, J = 12.0 Hz, 6-H),
8.51(s, 1H, 11-H), 8.43 (d, 2H, J = 20.0 Hz, 2′-H, 6′-H), 8.06
(d, 1H, J = 7.5 Hz, 9-H), 7.72–7.62(m, 2H, 3′-H, 5′-H), 7.58
(dd, 1H, J = 8.5, 2.0 Hz, 8-H), 7.50 (ddd, 1H, J = 1.5, 7.0,
1.0 Hz, 7-H), 7.15(s, 1H, 3-H). 13C NMR (Pyr-d5) d: 176.2
(COOH), 165.2 (C2), 158.4 (C10a), 156.8 (HC=N), 154.7 (C1′),
153.8 (C9a), 150.9 (C11a), 149.2 (C5b), 148.8 (C4a), 131.8 (C3′,
C5′), 130.7 (C4′), 129.4 (C3), 127.7 (C5a), 124.4 (C4), 123.6
(C6), 122.4 (C2′, C6′), 121.3 (C8), 120.2 (C7), 116.7 (C5), 113.6
(C9), 100.8 (C11). C, H and N for C23H13NO5.
HC=N), 9.20 (s, 1H, OH), 8.42 (s, 1H, 5-H), 8.09 (dd, 1H,
J = 7.5, 2 Hz, 6-H), 7.63 (s, 1H, 11-H), 7.62 (d, 1H, J = 3.5 Hz,
9-H), 7.51 (t, 1H, J = 7.5 Hz, 7-H), 7.47 (t, 1H, J = 12.0 Hz,
8-H), 7.39 (t, 1H, J = 10.0 Hz, 6′-H), 7.22 (t, 1H, J = 15.0 Hz,
3′-H), 7.06 (s, 1H, 3-H), 6.97 (d, 1H, J = 8.0 Hz, 5′-H), 6.91
(t, 1H, J = 15.0 Hz, 4′-H); 13C NMR (THF-d8) d: 160.3 (C2),
158.8 (C10a), 158.2 (C9a), 155.6 (C11a), 154.2 (HC=N), 146.7
(C2′), 137.6 (C1′), 130.7 (C3), 128.6 (C4′), 124.5 (C5b), 124.4
(C6), 122.3 (C4a), 121.7 (C8), 120.5 (C4), 119.2 (C5), 119.1
(C9), 117.4 (C6′), 116.7 (C5′), 114.6 (C5a), 112.5 (C3′), 100.7
(C11). C, H, N for C22H13NO4.
4-((4-Hydroxyphenylimino)methyl)-2H-benzofuro[3,2-
g]chromen-2-one (6h). m.p. 254–255 °C; IR (KBr) ˜v: 3601,
3380, 3343, 3094, 3028, 1692, 1642, 1602, 1574, 1506, 1475,
1456, 1387, 1341, 1266, 1218, 1139, 1099, 839, 810 cm–1;
FAB-MS m/z: 356 ([M + H]+),154 ([M-C6H4O.C6H6NO]+);
1H NMR (THF-d8) d: 9.64 (s, 1H, HC=N), 8.88 (s, 1H, 5-H),
8.50(s, 1H, OH), 8.07–8.05(m, 1H, 6-H), 7.84 (s, 1H, 11-H),
7.62–7.48 (m, 2H, 8-H, 7-H), 7.47–7.40 (m, 2H, 2′-H, 6′-H),
6.89–6.88 (m, 2H, 3′-H, 5′-H), 6.80(s, 1H, 3-H); 13C NMR
(THF-d8) d: 160.3 (C2), 159.4 (C10a), 158.7 (C9a), 158.2 (C4′),
155.7 (C11a), 154.3 (HC=N), 150.3 (C1′), 143.3 (C5b), 128.5
(C3), 124.6 (C4), 124.2 (C2′, C6′), 122.2 (C4a), 121.6 (C6),
119.8 (C8), 117.9(C7), 116.8 (C3′, C5′), 116.2 (C5), 114.5
(C5a), 112.5 (C9), 100.6 (C11). C, H, N for C22H13NO4.
4-((2-Hydroxy-4-methylphenylimino)methyl)-2H-
benzofuro[3,2-g]chromen-2-one (6i). m.p. 208–210 °C;
FAB-MS m/z: 370 ([M + H]+). 1H NMR (THF-d8) d: 10.05
(s, 1H, OH), 9.27 (s, 1H, HC=N), 9.02 (s, 1H, 5-H), 7.96 (d,
1H, J = 7.8 Hz, 9-H), 7.94 (d, 1H, J = 7.5 Hz, 6-H), 7.53 (t,
2H, J = 12.0 Hz, 4′-H, 6′-H), 7.40 (t, 1H, J = 6.9 Hz, 7-H),
7.29 (t, 1H, J = 6.9 Hz, 8-H), 7.14 (t, 1H, J = 6.9 Hz, 7-H),
6.94 (t, 1H, J = 2.9 Hz, 3′-H), 6.75(s, 1H, 3-H), 3.97(s, 1H,
3-H). C, H, N for C23H15NO4.
4-((2-Aminophenylimino)methyl)-2H-benzofuro [3,2-g]
chromen-2-one (6n). m.p. 276–278 °C; IR (KBr) v˜: 3270,
3066, 1674, 1643, 1605, 1564, 1492, 1428, 1391, 1366, 1274,
1251, 1230, 1104, 1029, 998, 886, 851, 762 cm–1; FAB-MS
1
m/z: 353 ([M-H]+), 154 ([M-C6H7O·C6H4N2]+); H NMR
(THF-d8) d: 12.36(s, 1H, CH = N), 10.01(s, 1H, 5-H), 8.17
(d, 1H, J = 8.0 Hz, 6-H), 7.92(d, 1H, J = 7.5 Hz, 9-H), 7.63(s,
1H, 11-H), 7.62–7.58(m, 2H, 3′-H, 6′-H), 7.50(t, 1H, J = 8.5,
7-H), 7.41–7.30(m, 3H, 8-H, 4′-H, 5′-H), 6.88 (s, 1H, 3-H).
C, H and N for C22H14N2O3.
7.3. Synthesis of B-type compounds (7a–c)
A suspension of NaBH4 (2.2 mmol) in 3 ml of tetrahydro-
furan was added to a methanolic solution of compound 6
(10 mmol) and the reaction mixture was stirred at 0 °C for
10 h. The methanol and tetrahydrofuran were evaporated and
the residue was diluted with water (20 ml), acidified with
diluted HCl. The precipitate obtained was collected and crys-
tallized to give compound 7 in the yield of 75–85%.
4-((Phenylamino)methyl)-2H-benzofuro[3,2-g]chromen-
2-one (7a). m.p. 296–298 °C; FAB-MS m/z: 342 ([M + H]+);
1H NMR (THF-d8) d: 8.32 (s, 1H, 5-H), 7.96 (d, 1H,
J = 7.8 Hz, 9-H), 7.81 (d, 1H, J = 8.2 Hz, 6-H), 7.73 (t,
J = 12.5 Hz, 7-H), 7.55 (t, J = 12.5 Hz, 8-H), 7.43 (s, 1H,
11-H), 7.18 (m, 2H, 3′-H, 5′-H), 7.11 (s, 1H, 4′-H), 6.87 (m,
2H, 2′-H, 6′-H), 6.62 (s, 1H, 3-H), 4.81 (t, 1H, NH), 3.85 (d,
J = 12.0 Hz, CH2N). C, H, N for C22H15NO3.
4-((4-tert-Butyl-2-hydroxyphenylimino)methyl)-2H-
1
benzofuro[3,2-g]chromen-2-one (6j). m.p. 243–245 °C; H
NMR (THF-d8) d: 10.56 (s, 1H, OH), 9.42 (s, 1H, CH = N),
9.02 (s, 1H, 5-H), 8.06 (t, 2H, J = 1.2 Hz, 6-H, 9-H), 7.68 (s,
1H, 11-H), 7.57–7.21 (m, 4H, 7-H, 8-H, 6′-H, 5′-H), 6.82 (s,
1H, 3-H), 3.61 (s, 9H). C, H, N for C26H21NO4.
4-((4-Mercaptophenylimino) methyl)-2H-benzofuro[3,2-g]
1
chromen-2-one (6k). m.p. 191–193 °C; H NMR (THF-d8)
d: 10.12 (s, 1H, SH), 9.23 (s, 1H, CH = N), 8.95 (s, 1H, 5-H),
8.17–6.92 (m, 9H), 6.71 (s, 1H, 3-H). C, H, N for
C22H13NO3S.
2-((2-Oxo-2H-benzofuro [3,2-g] chromen-4-yl) methyl-
eneamino)benzoic acid (6l). m.p. 256–258 °C; IR(KBr) ˜v:

1314
Q. Shen et al. / European Journal of Medicinal Chemistry 40 (2005) 1307–1315
4-((2-Methoxyphenylamino)methyl)-2H-benzofuro[3,2-
8. Biology
g]chromen-2-one (7b). m.p. 291–293 °C; FAB-MS m/z: 372
([M + H]+), 1H NMR (THF-d8) d: 8.27 (s, 1H, 5-H), 8.05 (d,
1H, J = 7.2 Hz, 9-H), 7.96 (d, 1H, J = 7.6 Hz, 6-H), 7.87 (t,
J = 10.5 Hz, 8-H), 7.71 (t, J = 10.5 Hz, 7-H), 7.65 (s, 1H,
11-H), 7.12–6.86 (m, 4H, 3′-H, 4′-H, 5′-H, 6′-H), 6.67 (s,
1H, 3-H), 3.86 (s, 3H, OCH3). C, H and N for C23H19NO4.
4-((4-Methoxyphenylamino)methyl)-2H-benzofuro[3,2-
g]chromen-2-one (7c). m.p. 325–327 °C; FAB-MS m/z: 372
([M + H]+), 1H NMR (THF-d8) d: 8.31 (s, 1H, 5-H), 8.13 (d,
1H, J = 7.8 Hz, 9-H), 7.91 (d, 1H, J = 7.6 Hz, 6-H), 7.78 (t,
J = 11.5 Hz, 8-H), 7.67 (t, J = 11.5 Hz, 7-H), 7.55 (s, 1H,
11-H), 7.15–6.86 (m, 4H, 2′-H, 3′-H, 5′-H, 6′-H), 6.71 (s,
1H, 3-H), 3.76 (s, 3H, OCH3). C, H and N for C23H19NO4.
8.1. Inhibition of AChE and BuChE
The inhibitory activities against AChE and BuChE of the
prepared compounds were carried out using a reported method
[22]. Five different concentrations of each compound were
used to measure the inhibitory activity forAChE and BuChE.
The assay solution consisted of 0.1 M phosphate buffer (pH
8.0), containing of 340 µM of 5,5′-dithio-bis(2-nitrobenzoic
acid), 0.035 unit ml–1 AChE or BuChE (0.5 and 3.4 UI mg–1,
respectively; Sigma Chemical), and 550 µM acetylthicoline
iodide or butylthicoline iodide. Test compound were added
to the assay solution and pre-incubated with the enzyme for
20 min followed by the addition of substrate. Assays were
carried out with a blank containing all components except
AChE or BuChE in order to account for non-enzymatic reac-
tion. The reaction rates were compared, and the percent inhi-
bition due to the presence of test compounds was calculated.
Each concentration was analyzed in triplicate, and IC50 val-
ues were determined graphically from log concentration-
inhibition curves.
7.4. Synthesis of C-type compound (11a)
7-(2-Oxo-propoxy)-4-methyl-2H-chromen-2-one (8). A
mixture of 1 (60.0 mmol), chloroacetone (120.0 mmol), and
anhydrous K2CO3 (60.0 g) in dry acetone (1000 ml) was
refluxed until the disappearance of 1 (72 h, monitored by
TLC). After cooling, the solid was filtered off and washed
with fresh acetone. The solvent was evaporated from the com-
bined filtrate and washings, and the residue was crystallized
form MeOH to give 8 as a white crystal. Yield 67%, m.p.
8.2. Kinetic characterization of AChE inhibition
Kinetic characterization of AChE were performed using a
reported method [29]. Five different concentrations of AChE
and inhibitors (6b, 7b, propidium and tacrine) were mixed in
the assay buffer (pH 8.0), containing of 340 µM of 5,5′-dithio-
bis(2-nitrobenzoic acid), 0.035 unit ml–1 AChE, and 550 µM
acetylthicoline iodide. Test compound were added to the assay
solution and pre-incubated with the enzyme at 37 °C for
20 min, followed by the addition of substrate. The determi-
nation of kinetic characterization of the AChE-catalyzed
hydrolysis of acetylthiocholine was taken by spectrometri-
cally at 412 nm. A parallel control with no inhibitor in the
mixture, allowed adjusting activities measured at various time.
1
161–163 °C; H NMR (CDCl3) d: 7.66–7.40 (m, 3H, 5-H,
6-H, 8-H), 6.25 (s, 1H, 3-H), 3.47 (s, 2H, 1′-H), 2.52 (s, 3H,
4-CH3), 2.30 (s, 3H, 3′-H).
3,5-Dimethyl-7H-furo[3,2-g]chromen-7-one (9). Prepara-
tion of 9 followed the procedure described by MacLeod [28].
1
Yield 83%, m.p. 220–222 °C; H NMR (CDCl3) d: 7.67(s,
1H, 5-H), 7.47 (s, 1H, 7-H), 7.40 (s, 1H, 8-H), 6.26 (s, 1H,
3-H), 2.51 (s, 3H, 6-CH3), 2.27 (s, 3H, 4-CH3).
3-(Bromomethyl)-5-methyl-7H-furo[3,2-g]chromen-7-
one (10). A mixture of 9 (25.0 mmol), 1,3-dibrom-5,5-
dimethylhydantoin, catalytic amount of benzoyl peroxide, and
carbon tetrachloride was refluxed for 8 h. After cooling, the
solvent was evaporated under reduced pressure. The residue
was washed with water, purified by column chromatography
(toluene/EtOAc, 100:1) and crystallized from MeOH to give
10.Yield 25%, m.p. 183–185 °C; 1H NMR (CDCl3) d: 7.58–
7.30 (m, 3H, 5-H, 7-H, 8-H), 6.27 (s, 1H, 3-H), 3.10 (s, 2H,
6-CH2Br), 2.51 (s, 3H, 4-CH3).
Acknowledgements
We thank the National Natural Science Foundation of
China (Grant No 20272085), Guangzhou City Science Foun-
dation and The Hong Kong Polytechnic UniversityASD fund
for financial support of this study.
5-Methyl-2-(phenylamino)-7H-furo[3,2-g]chromen-7-
one (11a).A solution of 10 (10.0 mmol) in CH3CN was mixed
with powdered KOH (11.0 mmol) under N2. Aniline was
added to the mixture at 50 °C and the mixture was vigorously
stirred.After 10 h, the resulting mixture was poured into water
and extracted with EtOAc (20 ml × 3). The organic layer was
dried over MgSO4, concentrated and purified by column chro-
matography (hexane/EtOAc/TEA, 16:2:1) to give 11a. Yield
References
[1] W.W. Pendlebury, P.R. Solomon, Neurobiol. Aging 15 (1994) 287–
289.
[2] M.R. Roberson, L.E. Harrell, Brain Res. Rev. 25 (1997) 50–69.
[3] R.T. Bartus, L.D. Dean, B. Beer, A.S. Lippa, Science 217 (1982)
408–414.
[4] M.J. Weinstock, Neural Transm, Suppl. 49 (1997) 93–102.
[5] S.B. Dunnett, H.C. Fobiger, Prog. Brain Res. 98 (1993) 413–420.
[6] M. Weinstock, CNS Drugs 12 (1999) 307–323.
1
72%, m.p. 203–205 °C; H NMR (DMSO-d6) d: 7.53–7.21
(m, 3H, 5-H, 7-H, 8-H), 7.12–6.86 (m, 5H, 2′-H, 3′-H, 4′-H,
5′-H, 6′-H), 6.25 (s, 1H, 3-H), 2.85 (d, 2H, J = 8.0 Hz, 4-CH2),
2.38 (s, 3H, 4-CH3). C, H, N for C19H15NO3.

Q. Shen et al. / European Journal of Medicinal Chemistry 40 (2005) 1307–1315
1315
[7] R. Diaz Brinton, R.S. Yamazaki, Pharm. Res. 15 (1998) 386–398.
[18] S.M. Sethna, N.M. Shah, Chem. Rev. 36 (1945) 1–62.
[8] N.C. Inestrosa, A. Alvarez, C.A. Perez, R.D. Moreno, M. Vicente,
C. Linker, O.I. Casanueva, C. Soto, J. Garrido, Neuron 16 (1996)
881–891.
[19] R.H. Baker, W.B. Martin, J. Org. Chem. 25 (1960) 1496–1498.
[20] The geometrical isomer of compound 6 constitution was determined
by 500 MHz ROESY and MM-2.
[9] J. Hardy, D.J. Selkoe, Science 297 (2002) 353–356.
[10] G.V. De Ferrari, M.A. Canales, I. Shin, L.M. Weiner, I. Silman,
N.C. Inestrosa, Biochem. 40 (2001) 10447–10457.
[21] J. Lecocq, N.P. Buu-Hoi, Compt. Rend. 224 (1947) 937–939.
[22] G.L. Ellmam, K.D. Courtney, V. Andres, R.M. Featherstone, Bio-
chem. Pharmacol. 7 (1961) 88–95.
[11] J.R.S. Hoult, M. Payd, Gen. Pharmacol. 27 (1996) 713–722.
[12] A. Rampa, A. Bisi, P. Valenti, M. Recanatini, A. Cavalli, V. Andrisano,
L. Fin, A. Buriani, P. Giusti, J. Med. Chem. 41 (1998) 3976–3986.
[13] C. Bruhlmann, F. Ooms, P.A. Carrupt, B. Testa, M. Catto, F. Leonetti,
C. Altomare, A. Carotti, J. Med. Chem. 44 (2001) 3195–3198.
[14] C. Gnerre, M. Catoo, F. Leonetti, P. Weber, P.A. Carrupt, C. Altomare,
A. Carotti, B. Testa, J. Med. Chem. 43 (2000) 4747–4758.
[15] B. Boyd, Drug News Perspect. 13 (2000) 425–438.
[16] L. Piazzi, A. Rampa, A. Bisi, S. Gobbi, F. Belluti, A. Cavalli, M. Bar-
tolini, V. Andrisamo, J. Med. Chem. 46 (2003) 2279–2283.
[17] H. Sugimoto, Y. Yamanishi, Y. Iimura, Y. Kawakami, Curr. Med.
Chem. 7 (2000) 303–339.
[23] M. Bartolini, C. Bertucci, V. Cavrini, V. Andrisano, Biochem. Phar-
macol. 65 (2003) 407–416.
[24] A. Fersht, Enzyme Structure and Mechanism, W. H. Freeman and Co.,
New York, 1985 (Chapter 3).
[25] Z. Radic, E. Reiner, P. Taylor, Mol. Pharmacol. 39 (1991) 98–104.
[26] J. Massoulie, S. Bon, I. Silman, Prog. Brain Res. 98 (1993) 139–146.
[27] J.L. Sussman, M. Harel, I. Silman, Chem. Biol. Interact. 87 (1993)
187–197.
[28] J.K. MacLeod, B.R. Worth, R.J. Well, Aust. J. Chem. 31 (1978)
1545–1552.
[29] S.R. Feaster, D.M. Quinn, Methods Enzymol. 286 (1997) 231–252.