3D-QSAR study of adamantyl N-benzylbenzamides as melanogenesis inhibitors

Article abstract of DOI:10.1016/j.bmcl.2013.11.056

Three-dimensional quantitative structure-activity relationship (3D-QSAR) modeling, comparative molecular field analysis (CoMFA), and comparative molecular similarity indices analysis (CoMSIA) of polyhydroxylated N-benzylbenzamide derivatives containing an

Full text of DOI:10.1016/j.bmcl.2013.11.056

Bioorganic & Medicinal Chemistry Letters 24 (2014) 667–673
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
3D-QSAR study of adamantyl N-benzylbenzamides as melanogenesis
inhibitors
a
b
c
a
a
Yong Deog Hong a, , Heung Soo Baek , Haelim Cho , Soo Mi Ahn , Ho Sik Rho , Young-Ho Park ,
⇑
Yung Hyup Joo a, Song Seok Shin a
a Materials Science, Medical Beauty Research Division, AmorePacific Corporation R&D Center, 314-1 Bora-Dong, Kiheung-Gu, Yongin-si, Kyounggi-do 446-729, South Korea
b T&J Tech Inc., Kasan-Dong, Keumchun-Gu, Seoul 153-770, South Korea
c Kyung Hee University Skin Biotechnology Center, Suwon 443-759, South Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Three-dimensional quantitative structure–activity relationship (3D-QSAR) modeling, comparative
molecular field analysis (CoMFA), and comparative molecular similarity indices analysis (CoMSIA) of
polyhydroxylated N-benzylbenzamide derivatives containing an adamantyl moiety were performed to
understand the mechanism of action and structure–activity relationship of these compounds. Contour
map analysis indicated that steric contributions of the adamantyl moiety and electrostatic contributions
of the hydroxyl group at the 3-position are important in the activity. Activities of the training set and test
sets predicted by CoMFA fit well with actual activities, demonstrating that CoMFA, along with the best
calculated q2 value, has the best predictive ability.
Received 2 September 2013
Revised 4 November 2013
Accepted 22 November 2013
Available online 7 December 2013
Keywords:
CoMFA
CoMSIA
3D-QSAR
Ó 2013 Elsevier Ltd. All rights reserved.
Melanogenesis
N-Benzylbenzamide
Melanin plays a beneficial role in protecting the skin from var-
ious external stimuli, such as ultraviolet light, free radicals, and
oxidative stresses. Melanogenesis is the biochemical process of
melanin formation and its distribution within the skin. This, com-
bined with the amount of melanin formed, determines skin color.
Through the actions of various harmful stimuli, over-production
of melanin from melanocytes leads to hyperpigmentation.1,2
Therefore, the control of melanin synthesis via attenuation or inhi-
length. We tried to find the optimized hydroxyl group position for
melanogenesis inhibition; modified hydroxyl substituents such as
3,4-dihydroxy, 2,5-dihydroxy, and 4-hydroxy groups on B ring
showed the same tendency as in the case of 2,4-dihydroxy substi-
tution, but were less effective toward melanogenesis inhibition.
Also, the chain length between parts A and B appeared to have a
relatively weak influence on the activity.
As an extension of this study, here we have prepared additional
benzamide derivatives with various substituents and chain
lengths. On the basis of this training set, we have performed stud-
ies to model three-dimensional quantitative structure–activity
relationships (3D-QSARs) by comparative molecular field analysis
(CoMFA) and comparative similarity indices analysis (CoMSIA)—
methods which may provide further insight into the mechanism
of action and optimal structure–activity relationships (SARs) of
these compounds.
An initial set of 48 molecules was used for the 3D-QSAR model-
ing studies.7 Both the training and test sets were designed to con-
tain wide structural diversity and activity ranges. An extended
dissimilarity selection method was used for finding a diverse rep-
resentative subset using the SYBYL OpSim method; the test set
molecules were selected with a diverse structure and activity
pIC50 range (4.2596 to 5.9586).8 The training set comprised 39
molecules containing an adamantyl or non-adamantyl moiety.
The test set, which comprised 9 molecules considering their struc-
tural diversity and activity ranges, was used to validate the
bition of melanogenesis has received attention as
a target
treatment.3,4
Among the known depigmenting agents, we were interested in
kazinols and broussonins, which contain a 1,3-diphenylpropane
skeleton. These agents, which have strong depigmenting activity,
have been used to design noble 1,3-diaryl polyhydroxy deriva-
tives.5 In an earlier study, we investigated polyhydroxylated
N-benzylbenzamide derivatives that contain an adamantyl moiety,
a new series of depigmenting agents with anti-melanogenic and
tyrosinase-inhibiting activities.6 We found that the depigmenting
activities of these analogues could be modulated by altering their
physicochemical properties via introduction of lipophilic or hydro-
philic functionalities, such as variation of hydroxyl substituents,
addition of a lipophilic adamantyl group, and modification of chain
⇑
Corresponding author.
E-mail
addresses:
hydhong@amorepacific.com,
hydhong@hanmail.com
(S.S. Shin).
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.11.056

668
Y. D. Hong et al. / Bioorg. Med. Chem. Lett. 24 (2014) 667–673
developed CoMFA/CoMSIA model (Table 2). Melanogenesis inhibi-
tion data were obtained from a murine melanoma cell line (B16).
The IC50 values of the 39 molecules in the training set, which were
CoMSIA models displayed values slightly lower than those of the
CoMFA model, e.g., for actual activities (r2 = 0.827, slope of 0.827
for the training set; r2 = 0.808, slope 0.794 for the test and training
sets). Therefore, these results indicate that the CoMFA model has
reliable predictive ability among the 3-D QSAR models. Analysis
of the CoMFA model was provided by the following results.
From the data given in Table 1, the contributions of the steric
and electrostatic fields of the CoMFA model are 56.6% and 43.4%,
respectively. The steric fraction of molecules was also determined
to play a slightly more significant role than the electrostatic frac-
tion in predicting biological activity.
distributed over a wide range from 0.5 to 100 lM with respect to
melanogenesis-inhibiting activity, were converted to pIC50 values
for the 3D-QSAR studies. The calculated pIC50 values ranged from
4.1249 to 6.1367.
Compound 9 was used as the template for alignment in these
studies because it showed the lowest IC50 values for biological
activity. To align template molecules, the minimum energy confor-
mation of 9 was sought by simulated annealing (heating the mol-
ecule at 700 K for 1000 fs and then annealing it to 200 K for
1000 fs). Subsequently, each molecule was structurally aligned
on the basis of this conformation.9 Following alignment, the steric
and electrostatic potential fields for CoMFA/CoMSIA were calcu-
lated with a grid spacing of 2.0 Å by default and an attenuation fac-
tor of 0.3 (Fig. 1).
A comparison of the CoMFA model and each of the CoMSIA
models is given in Table 1. Among the latter models, CoMSIA_all,
which used all descriptors including steric, electrostatic, hydro-
phobic, H-donor, and H-acceptor, provided the best q2 value of
0.691. Among all models, the CoMFA model had the highest q2 va-
lue of 0.708. On the basis of this finding, we assumed that CoMFA
was the best model in this study. The predicted and residual values
for the CoMFA/CoMSIA models are described in Table 1, and the
predicted versus actual pIC50 values are shown in Figure 2.
In Figure 2, the activities of both the training and test sets pre-
dicted by CoMFA fit well with the actual activities (r2 = 0.839, slope
of 0.839 for the training set; r2 = 0.817, slope of 0.808 for the test
and training sets). The blue data points correspond to the training
set and red shows the test set. As illustrated in Figure 2B, the
Table 3 provides a summary of the scrambling stability test re-
sults for the CoMFA model. The confidential Q2 values should be
high and near the 95% statistical confidence limit of 0.3,10and the
calculated cross-validated standard error of prediction (cSDEP)
should be low and similar to standard deviation of error of predic-
2
0
2
0
tion (SDEP). In general, the effective slope (dq /dr yy ) is the most
important value in the QSAR models.11 Of these models, unstable
ones are characterized by slopes greater than 1.20, whereas stable
models have slopes near unity.12 According to the summarized re-
sults, five components showed an acceptable range of slope values
2
2
0
0
(dq /dr yy ). Of these five, component model 3 was optimal be-
cause it showed the highest Q2 value of 0.597 and a dq /dr
va-
2
2
0
yy0
lue nearest to unity. This was also in accordance with the partial
least squares (PLS) results.
Contour maps of compound 9 in accord with these contribu-
tions are shown in Figure 3. Steric interactions are represented
by the green and yellow contours, whereas electrostatic interac-
tions are denoted by the red and blue contours. The steric contour
maps (A) are shown in green when the activity increases with a
large or bulky moiety. They are shown in yellow in the opposite
case, that is, when the activity decreases.
Figure 3A depicts a sterically favored green contour located at
the 3-position of 9, which suggests that biological activity in-
creases with bulky adamantyl substitution. This trend is observed
for all benzamide derivatives that possess an adamantyl substitu-
ent at the same position. Thus, compounds in the series, such as
15 with no substituent, exhibited reduced melanogenesis-
inhibiting activities. The para-position (R2) of
9 exhibited a
sterically unfavored small yellow contour, suggesting that activity
increases with the smaller OH group. However, more bulky substi-
tution (methoxy) and smaller substituent H yielded reduced
biological activity. Additionally, the B ring showed a yellow con-
tour, and the small OH group increased melanogenesis-activity.
The electrostatic contour maps in Figure 3B are indicated in red
when activity increases with negative atomic charges and in blue
when activity increases with positive atomic charges. In general,
Figure 1. Alignment of molecules used in the training set.
Table 1
Result of PLS (partial least square) analysis
Model
q2
N
r2
SEE
F
r2
Fraction
pred
Steric
Electrostatic
Hydrophobic
H-donor
0.182
H-acceptor
0.122
CoMFA
0.708
0.691
0.683
0.690
0.692
0.038
3
3
3
3
4
5
0.839
0.827
0.814
0.818
0.851
0.550
0.193
0.200
0.207
0.205
0.188
0.332
60.608
55.805
51.156
52.487
48.734
8.062
0.739
0.737
0.711
0.710
0.744
ꢀ0.562
0.566
0.181
0.370
0.223
0.434
0.258
0.630
0.451
CoMSIA_all
CoMSIA_ES
CoMSIA_ESH
CoMSIA_DAH
CoMSIA_DA
0.257
0.325
0.513
0.294
0.653
0.192
0.347
q2: Leave-One-Out cross-validated correlation coefficient.
N: optimum number of components.
r2: non-cross-validated correlation coefficient.
SEE: standard error of estimate.
F: F-test value.
r2pred: predictive r2 for the test set.
Fraction: contribution rate of each CoMFA field.
S: steric, E: electrostatic, H: hydrophobic, D: H-donor, A: H-acceptor.

Y. D. Hong et al. / Bioorg. Med. Chem. Lett. 24 (2014) 667–673
669
Table 2
Actual and CoMFA/CoMSIA-predicted pIC50 values of molecules in training and test sets
A ring
B ring
R1
O
R4
2
5
1
3
4
N
H
n
R2
6
R3
Compound
n
R1
R2
R3
R4
Actual pIC50
CoMFA
Pred. pIC50
CoMSIA_all
Pred. pIC50 Residual
Residual
Training set
3
4
5
6
7
8
9
1
1
1
2
1
1
1
1
1
1
1
2
2
1
1
2
2
2
1
1
2
2
1
1
1
1
2
1
1
2
1
1
1
1
1
2
1
2
1
OH
OH
OH
OH
OH
OH
OH
OMe
H
OH
OH
OH
OH
OH
OH
OH
H
2,4-OH
2,4-OH
3,4-OH
3,4-OH
2,5-OH
2,5-OMe
2,5-OH
3,4-OH
3,4-OH
3,4-OH
3,4-OH
4-OH
3,4-OH
2,4-OH
3,4-OH
3,4-OH
4-OH
3,4-OH
2,4-OH
3,4-OH
3,4-OH
3,4-OH
2,4-OH
2-OMe
4-OH
2,4-OMe
3,4-OMe
3,4,5-OMe
3,4-OMe
3,4-OMe
3,4-OMe
2,5-OMe
3,4-OMe
3,5-OMe
2,5-OMe
3,4-OH
3,5-OMe
4-OH
4.574
5.921
5.602
5.364
4.616
5.387
6.137
5.046
5.000
5.652
5.097
5.921
4.125
5.959
5.889
5.903
5.717
4.959
4.559
5.590
5.633
4.638
5.854
5.636
5.910
5.433
5.389
5.532
5.419
5.347
5.413
5.267
5.301
5.299
5.503
5.680
5.590
5.301
6.046
4.714
5.801
5.809
5.546
4.694
5.441
5.782
5.020
4.933
5.922
4.979
5.855
4.455
5.956
5.987
5.890
5.830
4.759
4.930
5.737
5.617
4.666
5.878
5.715
5.790
5.588
5.374
5.323
5.392
5.485
5.251
5.343
5.413
5.465
5.516
5.449
5.399
4.811
5.695
ꢀ0.141
0.120
4.761
5.824
5.846
5.553
4.696
5.435
5.756
5.019
4.920
5.917
4.972
5.843
4.465
6.013
5.976
5.834
5.788
4.742
4.944
5.760
5.626
4.640
5.881
5.539
5.711
5.505
5.452
5.361
5.367
5.547
5.301
5.325
5.476
5.483
5.507
5.482
5.427
4.780
5.733
ꢀ0.188
0.097
Ada
Ad
Ad
H
ꢀ0.207
ꢀ0.183
ꢀ0.078
ꢀ0.054
0.355
ꢀ0.244
ꢀ0.190
ꢀ0.080
ꢀ0.048
0.381
Ad
Ad
H
H
Ad
H
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
OMe
OMe
OMe
OMe
OMe
OH
OMe
OMe
OMe
OMe
OMe
OMe
OH
OMe
OMe
OMe
OH
OH
OH
0.026
0.067
0.027
0.080
H
ꢀ0.270
ꢀ0.265
OH
OMe
H
OMe
OH
OH
OH
OMe
OH
H
H
H
H
OH
OH
OH
OH
OH
OH
H
H
H
H
H
H
H
OH
OH
H
0.118
0.125
Ad
H
0.066
0.078
ꢀ0.330
ꢀ0.340
ꢀ0.054
ꢀ0.087
0.069
Ad
Ad
Ad
Ad
H
0.003
ꢀ0.098
0.013
ꢀ0.113
ꢀ0.071
0.200
0.217
H
ꢀ0.371
ꢀ0.147
0.016
ꢀ0.028
ꢀ0.024
ꢀ0.079
0.120
ꢀ0.385
ꢀ0.170
0.007
ꢀ0.002
ꢀ0.027
0.097
Ad
Ad
H
Ad
Ad
Ad
Ad
Ad
Ad
Ad
Ad
Ad
Ad
Ad
Ad
Ad
Ad
Ad
H
0.199
ꢀ0.155
ꢀ0.072
ꢀ0.063
0.171
OH
OH
OH
OMe
OH
0.015
0.209
0.027
0.052
ꢀ0.138
ꢀ0.200
0.162
0.112
OH
ꢀ0.076
ꢀ0.112
ꢀ0.166
ꢀ0.013
0.231
0.191
0.490
0.351
ꢀ0.058
ꢀ0.175
ꢀ0.184
ꢀ0.004
0.198
0.163
0.521
0.313
OMe
OMe
OMe
OH
OH
OMe
OH
Ad
2,4-OH
Test set
42
43
44
45
46
47
48
49
2
1
1
2
1
2
1
1
1
OH
OH
OMe
OMe
H
H
H
OH
OH
OH
Ad
Ad
H
H
H
Ad
Ad
H
4-OH
5.959
5.745
4.848
4.569
4.260
5.467
5.353
5.097
5.243
5.666
5.925
4.964
4.860
4.693
5.749
5.303
4.729
5.544
0.293
ꢀ0.180
ꢀ0.116
ꢀ0.291
ꢀ0.433
ꢀ0.282
0.050
5.662
5.977
5.004
4.840
4.741
5.714
5.305
4.786
5.538
0.297
ꢀ0.232
ꢀ0.156
ꢀ0.271
ꢀ0.481
ꢀ0.247
0.048
OMe
OMe
OMe
OH
OMe
OH
2,4-OH
2,4-OH
4-OH
3,4-OH
4-OH
3,5-OMe
3,4-OH
2-OH,4-OMe
OH
OH
0.368
ꢀ0.301
0.311
ꢀ0.295
50
Ad
a
1-Adamantyl.
red contour maps are assigned to heteroatoms such as nitrogen
and oxygen, whose partial atomic charges are highly negative.
The two large red contours indicated significant negative charges
corresponding to OH groups located on the phenyl ring in 9, which
are required for biological activity. The largely red contours located
at the para-position (R2) and 3-position of the B ring indicated that
biological activity increases with the negative atomic charge of OH
groups. Therefore, favorable steric and electrostatic contours pro-
vide an explanation for why compound 9 shows the best inhibitory
activity among the series of adamantyl-substituted compounds.
CoMFA and CoMSIA studies were used to derive 3D-QSAR mod-
els for a training set of 39 benzamide derivatives. On the basis of
the q2 values, we chose CoMFA as the best validated 3D-QSAR
model. Analysis of CoMFA studies yielded the following results:

670
Y. D. Hong et al. / Bioorg. Med. Chem. Lett. 24 (2014) 667–673
Figure 2. Plot of actual versus predicted activities for CoMFA (A) and CoMSIA (B).
Table 3
melanogenesis-inhibiting activity. Among compounds 3, 6, 9, 16,
Result of scrambling stability test for CoMFA
18, 27 and 41, which were shown to possess good anti-melanogen-
ic activities, compound 16 (in-house library code: AP-736), which
is also a good tyrosinase inhibitor, was selected as a promising can-
didate depigmenting agent with respect to large-scale preparation.
Further evaluation of this compound is currently underway
(Scheme 1).
Q2
cSDEP
dq20/dr2
yy0
#Component
1
2
3
4
5
0.486
0.585
0.597
0.543
0.497
0.335
0.304
0.304
0.326
0.351
0.535
0.731
0.973
0.695
0.715
Experimental section: Chemistry: 1H NMR spectra were re-
corded on a Varian Mercury 300 spectrometer with DMSO-d6 as
solvent. Chemical shifts are reported in values (ppm) relative to
the internal standard, and J values are reported in Hz. Mass spectra
were recorded on an LCQ Deca XP plus LC/MS mass spectrometer.
Reagents and solvents were purchased from common commercial
suppliers and used without further purification.
Synthesis of substituted N-benzylbenzamide or N-phenethylb-
enzamide: General procedures: Substituted benzoic acid
(2.1 mmol), N-hydroxysuccinimide (2.1 mmol), and N,N0-dicyclo-
hexylcarbodiimide (2.1 mmol) were dissolved in 1,4-dioxane
(5 mL) under a nitrogen atmosphere. The solution was stirred at
ambient temperature for 12 h. The precipitate was filtered off
and the filtrate added to a solution of substituted benzylamine
HCl salt (2.3 mmol) and NaHCO3 (2.3 mmol) in H2O (2 mL). The
reaction mixture was heated to 60 °C under nitrogen and stirred
for 2–3 h. After cooling to room temperature, 10% HCl solution
was added and the mixture was extracted with EtOAc. The com-
bined organic layers were washed with brine, dried over anhy-
drous MgSO4, and filtered, after which the filtrate was
Q2; 1-(cSDEP)2: predictive ability of the model after potential effects of redundancy
have been removed; that is, the expected value of q2 at the specified critical point.
cSDEP: scaled cross-validated standard error, SDEP normalized by the standard
deviation of the dependent variables.
2
dq20/dr yy : the slope of q2 with respect to the correlation of the original dependent
0
variables versus the perturbed dependent variables.
leave-one-out cross-validated q2, 0.708; non cross-validated r2,
0.839; F value, 60.608; standard error of estimation, 0.193; and
predictive r2 of test set, 0.739. The fractional contributions of the
steric and electrostatic fields were 56.6% and 43.4%, respectively,
in predicting biological activity. Additionally, the contour maps
provided a viable explanation for the biological activities of our
compounds. Accordingly, compound 42 in the test set, containing
a 2,4-dihydroxy group on the B ring and lipophilic adamantyl
group, showed the best correlation between actual biological
activity and predictive activity. Therefore, our CoMFA results
demonstrate an excellent correlation between steric/electrostatic
fields and biological activity, and provide a deeper insight into
the chemical structure design of new molecules with high
Figure 3. Contour maps of compound 10. (A) Steric contour maps: green and yellow contours indicate regions where steric bulky groups increase and decrease activity,
respectively. (B) Electrostatic contour maps: red and blue contours indicate regions where negative charge increases activity and positive charge increases activity,
respectively.

Y. D. Hong et al. / Bioorg. Med. Chem. Lett. 24 (2014) 667–673
671
A ring
R1
B ring
R4
R1
O
O
R1
O
a
b,c
N
H
OH
n
OH
R2
R2
R2
R3
R3
3~51
1
2
R1=H, OH, OMe
R2=OH, OMe
R3=H, 1-adamantyl
R4,R5=H, OH, OMe
n=1,2
Scheme 1. General reagents and conditions: (a) 1-adamantanol, c-H2SO4/AcOH, CH2Cl2, rt; (b) HOSu, DCC, dioxane, rt; (c) Amine(salt), NaHCO3, H2O/50–60 °C, 3–4 h.
evaporated in vacuo. The crude product was purified by column
chromatography to obtain the compound.
N-(3,4-Dihydroxybenzyl)-2-hydroxy-4-methoxybenzamide (13):
1H NMR (300 MHz, DMSO-d6): d 13.11 (s, 1H), 9.05 (br s, 1H),
8.86 (s, 1H), 8.74 (s, 1H), 7.83–7.80 (m, 2H), 6.70–6.41 (m, 5H),
4.30 (d, 2H, J = 5.7 Hz), 3.76 (s, 3H); MS (EI): 289 (M+).
5-Adamantan-1-yl-N-[2-(4-hydroxyphenyl)ethyl]-2,4-dimethoxy-
benzamide (14): 1H NMR (300 MHz, DMSO-d6): d 9.20 (s, 1H), 7.94
(br s, 1H), 7.73 (s, 1H), 7.04 (d, 2H, J = 8.1 Hz), 6.71 (d, 2H,
J = 8.1 Hz), 6.63 (s, 1H), 3.87 (s, 3H), 3.84 (s, 3H), 3.45–3.43 (m,
2H), 2.71–2.66 (m, 2H), 1.99 (s, 9H), 1.71 (s, 6H); MS (EI): 435 (M+).
N-[2-(3,4-Dihydroxyphenyl)ethyl]-4-hydroxybenzamide (15): 1H
NMR (300 MHz, DMSO-d6): d 9.89 (br s, 1H), 8.68 (br s, 1H), 8.22
(br s, 1H), 7.68 (d, 2H, J = 8.4 Hz), 6.77 (d, 2H, J = 8.4 Hz), 6.63–
6.43 (m, 3H), 3.40–3.35 (m, 2H), 2.63–2.58 (m, 2H); MS (EI): 273
(M+).
5-Adamantan-1-yl-N-(2,4-dihydroxybenzyl)-2,4-dimethoxybenza-
mide (16): 1H NMR (300 MHz, DMSO-d6): d 9.67 (s, 1H), 9.13 (s,
1H), 8.51 (m, 1H), 7.78 (m, 1H), 6.92 (d, 1H, J = 8.1 Hz), 6.66 (s,
1H), 6.27 (s, 1H), 6.16 (d, 1H, J = 8.1 Hz), 4.30 (d, 2H, J = 5.4 Hz),
3.93 (s, 3H), 3.88 (s, 3H), 1.98 (s, 9H), 1.71 (s, 6H); MS (EI): 437
(M+).
5-Adamantan-1-yl-N-(3,4-dihydroxybenzyl)-2-hydroxy-4-methoxy-
benzamide (17): 1H NMR (300 MHz, DMSO-d6): d 12.99 (s, 1H), 9.09 (br
s, 1H), 8.85 (s,1H), 8.72 (s, 1H), 7.48 (s, 1H), 6.71–6.43 (m, 4H), 4.30 (d,
2H, J = 5.4 Hz), 3.79 (s, 3H), 3.88 (s, 3H), 2.00 (s, 9H), 1.71 (s, 6H); MS
(EI): 423 (M+).
5-Adamantan-1-yl-N-[2-(3,4-dihydroxyphenyl)-ethyl]-2-hydroxy-
4-methoxybenzamide (18): 1H NMR (300 MHz, DMSO-d6): d 12.89
(s, 1H), 8.76–8.65 (m, 3H), 7.43 (s, 1H), 6.65–6.42 (m, 4H), 3.79
(s, 3H), 3.40–3.36 (m, 2H), 2.66–2.62 (m, 2H), 2.01 (s, 9H), 1.72
(s, 6H); MS (EI): 437 (M+).
N-(2,4-Dihydroxybenzyl)-2,4-dihydroxybenzamide (3): 1H NMR
(300 MHz, DMSO-d6): d 12.71 (s, 1H), 10.00 (s, 1H), 9.39 (s, 1H),
9.08 (s, 1H), 8.75 (m, 1H), 7.73 (d, 1H, J = 8.4 Hz), 6.89 (d, 1H,
J = 8.7 Hz), 6.24 (m, 3H), 6.15 (d, 1H, J = 8.7 Hz), 4.29 (d, 2H,
J = 5.4 Hz); MS (EI): 275 (M+).
5-Adamantan-1-yl-N-(2,4-dihydroxybenzyl)-2,4-dihydroxybenza-
mide (4): 1H NMR (300 MHz, DMSO-d6): d 12.41 (s, 1H), 9.91 (s,
1H), 9.40 (s, 1H), 9.09 (s, 1H), 8.83 (m, 1H), 7.47 (s, 1H), 6.89 (d,
1H, J = 8.1 Hz), 6.26 (s, 2H), 6.16 (d, 1H, J = 8.1 Hz), 4.29 (m, 2H),
2.02 (s, 9H), 1.70 (s, 6H); MS (EI): 409 (M+).
5-Adamantan-1-yl-N-(3,4-dihydroxybenzyl)-2,4-dihydroxybenza-
mide (5): 1H NMR (300 MHz, DMSO-d6): d 12.74 (s, 1H), 9.95 (s,
1H), 8.98 (br s, 1H), 8.85 (s, 1H), 8.71 (s, 1H), 7.44 (s, 1H), 6.70
(s, 1H), 6.67–6.43 (m, 2H), 6.56–6.54 (m, 1H), 6.26 (s, 1H), 4.28
(d, 2H, J = 5.4 Hz), 2.03 (s, 9H), 1.71 (s, 6H); MS (EI): 409 (M+).
5-Adamantan-1-yl-N-[2-(3,4-dihydroxyphenyl)ethyl]-2,4-dihydr-
oxybenzamide (6): 1H NMR (300 MHz, DMSO-d6): d 12.63 (s, 1H),
9.91 (s, 1H), 8.74 (s, 1H), 8.63 (s, 1H), 8.58 (br s, 1H), 7.38 (s, 1H),
6.65–6.61 (m, 2H), 6.48–6.45 (m, 1H), 6.25 (s, 1H), 3.45–3.40 (m,
2H), 2.65–2.60 (m, 2H), 2.03 (s, 9H), 1.72 (s, 6H); MS (EI): 423 (M+).
N-(2,5-Dihydroxybenzyl)-2,4-dihydroxybenzamide (7): 1H NMR
(300 MHz, DMSO-d6): d 12.50 (s, 1H), 10.05 (s, 1H), 8.85 (br s,
1H), 8.80 (s, 1H), 8.60 (s, 1H), 7.80–7.75 (m, 1H), 6.65–6.25 (m,
5H), 4.38–4.30 (m, 2H); MS (EI): 275 (M+).
5-Adamantan-1-yl-N-(2,5-dimethoxybenzyl)-2,4-dihydroxybenza-
mide (8): 1H NMR (300 MHz, DMSO-d6): d 12.39 (s, 1H), 9.94 (s,
1H), 8.90 (br s, 1H), 7.50 (s, 1H), 6.93–6.72 (m, 3H), 6.28 (s, 1H),
4.40 (d, 2H, J = 5.7 Hz), 3.77 (s, 3H), 3.65 (s, 3H), 2.04 (s, 9H), 1.71
(s, 6H); MS (EI): 437 (M+).
5-Adamantan-1-yl-2-hydroxy-N-[2-(4-hydroxyphenyl)ethyl]-4-methoxy-
1
5-Adamantan-1-yl-N-(2,5-dihydroxybenzyl)-2,4-dihydroxybenza-
mide (9): 1H NMR (300 MHz, DMSO-d6): d 12.50 (s, 1H), 9.94 (s,
1H), 8.92 (br s, 1H), 8.80 (s, 1H), 8.59 (s, 1H), 7.50 (s, 1H), 6.16–
6.43 (m, 3H), 6.28 (s, 1H), 4.33 (d, 2H, J = 4.8 Hz), 2.04 (s, 9H),
1.71 (s, 6H); MS (EI): 409 (M+).
benzamide (19): H NMR (300 MHz, DMSO-d6): d 12.85 (s, 1H), 9.17 (s,
1H), 8.70 (br s, 1H), 7.42 (s, 1H), 7.02 (d, 2H, J = 8.1 Hz), 6.68 (d, 2H,
J = 8.1 Hz), 6.41 (s, 1H), 3.78 (s, 3H), 3.50–3.39 (m, 2H), 2.73–2.71 (m,
2H), 2.00 (s, 9H), 1.72 (s, 6H); MS (EI): 421 (M+).
N-[2-(3,4-Dihydroxyphenyl)ethyl]-2,4-dimethoxybenzamide (20):
1H NMR (300 MHz, DMSO-d6): d 8.77 (br s, 1H), 8.67 (br s, 1H),
7.94 (br s, 1H), 7.84–7.81 (m, 2H), 6.67–6.47 (m, 5H), 3.81 (s,
3H), 3.80 (s, 3H), 3.50–3.41 (m, 2H), 2.64–2.60 (m, 2H); MS (EI):
317 (M+).
N-(2,4-Dihydroxybenzyl)-2-hydroxy-4-methoxybenzamide (21):
1H NMR (300 MHz, DMSO-d6): d 12.91 (s, 1H), 9.40 (s, 1H), 9.12
(s, 1H), 8.86 (m, 1H), 7.85 (d, 1H, J = 8.7 Hz), 6.90 (d, 1H,
J = 8.7 Hz), 6.44 (m, 2H), 6.28 (s, 1H), 6.16 (d, 1H, J = 8.1 Hz), 4.31
(d, 2H, J = 5.1 Hz), 3.76 (s, 3H); MS (EI): 289 (M+).
3-Adamantan-1-yl-N-(3,4-dihydroxybenzyl)-4-hydroxybenzamide
(22): 1H NMR (300 MHz, DMSO-d6): d 9.82 (br s, 1H), 8.80–8.60 (m,
3H), 7.63 (s, 1H), 7.56 (d, 1H, J = 8.1 Hz), 6.77 (d, 1H, J = 8.4 Hz),
N-(3,4-Dihydroxybenzyl)-2,4-dimethoxybenzamide (10): 1H NMR
(300 MHz, DMSO-d6): d 8.73 (br s, 1H), 8.34 (br s, 1H), 7.83 (d,
2H, J = 8.4 Hz), 6.70–6.54 (m, 5H), 4.31 (d, 2H, J = 5.7 Hz), 3.88 (s,
3H), 3.81 (s, 3H), 2.04 (s, 9H), 1.71 (s, 6H); MS (EI): 303 (M+).
N-(3,4-Dihydroxybenzyl)-4-methoxybenzamide (11): 1H NMR
(300 MHz, DMSO-d6):
d 8.72–8.59 (m, 3H), 7.77 (d, 2H,
J = 8.4 Hz), 6.89 (d, 2H, J = 8.7 Hz), 6.61–6.44 (m, 3H), 4.19 (d, 2H,
J = 5.7 Hz), 3.71 (s, 3H); MS (EI): 273 (M+).
3-Adamantan-1-yl-N-(3,4-dihydroxybenzyl)-4-methoxybenzam-
ide (12): 1H NMR (300 MHz, DMSO-d6): d 8.74 (br s, 1H), 7.76–7.70
(m, 2H), 7.03–7.00 (m, 1H), 6.70–6.53 (m, 3H), 4.27 (d, 2H, J = 6.0 Hz),
3.84 (s, 3H), 2.05 (s, 9H), 1.73 (s, 6H); MS (EI): 407 (M+).

672
Y. D. Hong et al. / Bioorg. Med. Chem. Lett. 24 (2014) 667–673
6.69 (s, 1H), 6.64 (d, 1H, J = 8.1 Hz), 6.53 (d, 1H, J = 8.1 Hz), 4.26 (d,
2H, J = 6.0 Hz), 2.07 (s, 9H), 1.72 (s, 6H); MS (EI): 393 (M+).
3-Adamantan-1-yl-N-[2-(3,4-dihydroxyphenyl)ethyl]-4-methoxy-
benzamide (23): 1H NMR (300 MHz, DMSO-d6): d 8.73 (br s, 1H),
8.63 (br s, 1H), 8.34 (br s, 1H), 7.68–7.63 (m, 2H), 6.99 (d, 1H,
J = 8.7 Hz), 6.64–6.60 (m, 2H), 6.46 (d, 1H, J = 8.1 Hz), 3.83 (s, 3H),
3.40–3.35 (m, 2H), 2.64–2.59 (m, 2H), 2.05 (s, 9H), 1.73 (s, 6H);
MS (EI): 421 (M+).
2H), 7.03–6.80 (m, 4H), 4.39 (d, 2H, J = 6.0 Hz), 3.84 (s, 3H),
3.72 (s, 3H), 3.71 (s, 3H), 2.05 (s, 9H), 1.73 (s, 6H); MS (EI):
435 (M+).
3-Adamantan-1-yl-N-(3,5-dimethoxybenzyl)-4-methoxybenzamide
(36): 1H NMR (300 MHz, DMSO-d6): d 8.83 (br s, 1H), 7.77–7.70 (m,
2H), 7.03 (d, 1H, J = 8.4 Hz), 6.46 (s, 2H), 6.36 (s, 1H), 4.38 (d, 2H,
J = 5.4 Hz), 3.84 (s, 3H), 3.70 (s, 6H), 2.05 (s, 9H), 1.73 (s, 6H); MS
(EI): 435 (M+).
3-Adamantan-1-yl-N-(2,5-dimethoxybenzyl)-4-methoxybenzamide
(37): 1H NMR (300 MHz, DMSO-d6): d 8.70 (br s, 1H), 7.78–7.71 (m,
2H), 7.03 (d, 1H, J = 8.7 Hz), 6.90 (d, 1H, J = 8.7 Hz), 6.79–6.72 (m,
2H), 4.39 (d, 2H, J = 6.0 Hz), 3.85 (s, 3H), 3.77 (s, 3H), 3.64 (s, 3H),
2.05 (s, 9H), 1.73 (s, 6H); MS (EI): 435 (M+).
N-[2-(3,4-Dihydroxyphenyl)ethyl]-4-methoxybenzamide (24): 1H
NMR (300 MHz, DMSO-d6): d 8.71 (br s, 1H), 8.63 (br s, 1H), 8.34
(br s, 1H), 7.79 (d, 2H, J = 8.7 Hz), 6.97 (d, 2H, J = 8.7 Hz), 6.64–
6.61 (m, 2H), 6.46 (d, 1H, J = 8.1 Hz), 3.80 (s, 3H), 3.45–3.35 (m,
2H), 2.65–2.60 (m, 2H); MS (EI): 287 (M+).
3-Adamantan-1-yl-N-(2,4-dihydroxybenzyl)-4-methoxybenzamide
(25): 1H NMR (300 MHz, DMSO-d6): d 9.58 (s, 1H), 9.09 (s, 1H), 8.81
(t, 1H, J = 5.4 Hz), 7.73 (m, 2H), 7.02 (d, 1H, J = 8.4 Hz), 6.90 (d, 1H,
J = 8.1 Hz), 6.18 (m, 2H), 4.26 (d, 2H, J = 5.7 Hz), 3.84 (s, 3H), 2.05 (s,
9H), 1.73 (s, 6H); MS (EI): 407 (M+).
3-Adamantan-1-yl-N-[2-(3,4-dihydroxyphenyl)ethyl]-4-hydroxy-
benzamide (38): 1H NMR (300 MHz, DMSO-d6): d 9.80 (br s, 1H),
8.70 (br s, 1H), 8.22 (br s, 1H), 7.57–7.48 (m, 2H), 6.77–6.43 (m,
4H), 3.45–3.35 (m, 2H), 2.62–2.58 (m, 2H), 2.07 (s, 9H), 1.73 (s,
6H); MS (EI): 407 (M+).
5-Adamantan-1-yl-2,4-dihydroxy-N-(2-methoxybenzyl)benzamide
(26): 1H NMR (300 MHz, DMSO-d6): d 12.46 (s, 1H), 9.93 (s, 1H),
8.93 (br s, 1H), 7.51 (s, 1H), 7.26–6.87 (m, 4H), 6.28 (s, 1H), 4.43
(d, 2H, J = 5.4 Hz), 3.82 (s, 3H), 2.04 (s, 9H), 1.71 (s, 6H); MS (EI):
407 (M+).
5-Adamantan-1-yl-N-(3,5-dimethoxybenzyl)-2,4-dihydroxybenza-
mide (39): 1H NMR (300 M Hz, DMSO-d6): d 12.6 (br s, 1H), 9.96 (s,
1H), 9.04 (br s, 1H), 7.46 (s, 1H), 6.46 (s, 2H), 6.38 (s, 1H), 6.27 (s,
1H), 4.39 (d, 2H, J = 5.4 Hz), 3.71 (s, 6H), 2.04 (s, 9H), 1.71 (s, 6H);
MS (EI): 437 (M+).
5-Adamantan-1-yl-2,4-dihydroxy-N-(4-hydroxybenzyl)benzamide
(27): 1H NMR (300 MHz, DMSO-d6): d 12.68 (s, 1H), 9.94 (s, 1H),
9.26 (s, 1H), 8.98 (m, 1H), 7.42 (s, 1H), 7.11 (d, 2H, J = 8.1 Hz),
6.70 (d, 2H, J = 8.1 Hz), 6.24 (s, 1H), 4.33 (m, 2H), 2.02 (s, 9H),
1.70 (s, 6H); MS (EI): 393 (M+).
2-Hydroxy-N-[2-(4-hydroxyphenyl)ethyl]-4-methoxybenzamide
(40): 1H NMR (300 MHz, DMSO-d6): d 13.0 (br s, 1H), 9.17 (s, 1H),
8.68 (br s, 1H), 7.77 (d, 1H, J = 8.7 Hz), 7.02 (d, 2H, J = 7.5 Hz),
6.67 (d, 2H, J = 7.5 Hz), 6.46–6.40 (m, 2H), 3.76 (s, 3H), 3.45–
3.38 (m, 2H), 2.74–2.69 (m, 2H); MS (EI): 287 (M+).
5-Adamantan-1-yl-N-(2,4-dimethoxybenzyl)-2,4-dihydroxybenza-
mide (28): 1H NMR (300 MHz, DMSO-d6): d 12.48 (s, 1H), 9.92 (s,
1H), 8.86 (m, 1H), 7.48 (s, 1H), 7.07 (d, 1H, J = 8.1 Hz), 6.56 (m,
1H), 6.48 (m, 1H), 6.27 (s, 1H), 6.18 (m, 2H),, 4.35 (d, 2H,
J = 5.4 Hz), 3.80 (s, 3H), 3.73 (s, 3H), 2.03 (s, 9H), 1.71 (s, 6H); MS
(EI): 437 (M+).
5-Adamantan-1-yl-N-[2-(3,4-dimethoxyphenyl)ethyl]-2,4-dihydr-
oxybenzamide (29): 1H NMR (300 MHz, DMSO-d6): d 12.58 (s, 1H),
9.90 (s, 1H), 8.56 (br s, 1H), 7.37 (s, 1H), 6.87–6.75 (m, 3H), 6.24 (s,
1H), 3.70 (s, 6H), 3.50–3.35 (m, 2H), 2.78–2.76 (m, 2H), 2.02 (s,
9H), 1.72 (s, 6H); MS (EI): 451 (M+).
5-Adamantan-1-yl-2,4-dihydroxy-N-(3,4,5-trimethoxybenzyl)benz-
amide (30): 1H NMR (300 MHz, DMSO-d6): d 12.51 (s, 1H), 9.95 (s,
1H), 8.99 (br s, 1H), 7.46 (s, 1H), 6.67–6.64 (m, 2H), 6.27 (s, 1H),
4.39 (d, 2H, J = 5.4 Hz), 3.74 (s, 6H), 3.62 (s, 3H), 2.03 (s, 9H), 1.71
(s, 6H); MS (EI): 467 (M+).
5-Adamantan-1-yl-N-(3,4-dimethoxy-benzyl)-2,4-dihydroxy-benz-
amide (31): 1H NMR (300 MHz, DMSO-d6): d 12.62 (s, 1H), 9.96 (s,
1H), 9.01 (br s, 1H), 7.44 (s, 1H), 6.93–6.84 (m, 3H), 6.26 (s, 1H),
4.38 (d, 2H, J = 5.4 Hz), 3.73 (s, 3H), 3.71 (s, 3H), 2.03 (s, 9H), 1.71
(s, 6H); MS (EI): 437 (M+).
3-Adamantan-1-yl-N-[2-(3,4-dimethoxyphenyl)ethyl]-4-methoxy-
benzamide (32): 1H NMR (300 MHz, DMSO-d6): d 8.34 (s, 1H), 7.68–
7.62 (m, 2H), 7.00 (d, 1H, J = 8.4 Hz), 6.86–6.72 (m, 3H), 3.83 (s, 3H),
3.70 (s, 6H), 3.50–3.35 (m, 2H), 2.77–2.73 (m, 2H), 2.04 (s, 9H), 1.73
(s, 6H); MS (EI): 449 (M+).
3-Adamantan-1-yl-N-(3,4-dimethoxybenzyl)-4-hydroxybenzam-
ide (33): 1H NMR (300 MHz, DMSO-d6): d 9.83 (s, 1H), 8.67 (br s,
1H), 7.63–7.55 (m, 2H), 6.92–6.76 (m, 4H), 4.36 (d, 2H,
J = 5.7 Hz), 3.72 (s, 3H), 3.71 (s, 3H), 2.07 (s, 9H), 1.72 (s, 6H); MS
(EI): 421 (M+).
3-Adamantan-1-yl-N-(2,5-dimethoxybenzyl)-4-hydroxybenzamide
(34): 1H NMR (300 MHz, DMSO-d6): d 9.85 (br s, 1H), 8.57 (br s, 1H),
7.65–7.57 (m, 2H), 6.90–6.71 (m, 4H), 4.37 (d, 2H, J = 5.7 Hz),
3.76 (s, 3H), 3.64 (s, 3H), 2.08 (s, 9H), 1.73 (s, 6H); MS (EI): 421
(M+).
3-Adamantan-1-yl-N-(2,4-dihydroxybenzyl)-4-hydroxybenzamide
(41): 1H NMR (300 MHz, DMSO-d6): d 9.87 (br s, 1H), 9.64 (s, 1H),
9.09 (s, 1H), 8.73 (br s, 1H), 7.63–7.55 (m, 2H), 6.89 (d, 1H,
J = 8.1 Hz), 6.78 (d, 1H, J = 7.8 Hz), 6.23–6.15 (m, 2H), 4.25 (m,
2H), 2.07 (s, 9H), 1.72 (s, 6H); MS (EI): 393 (M+).
5-Adamantan-1-yl-2,4-dihydroxy-N-[2-(4-hydroxyphenyl)ethyl]benz-
amide (42): 1H NMR (300 MHz, DMSO-d6): d 12.59 (s, 1H), 9.91 (s, 1H),
9.16 (s, 1H), 8.60 (br s, 1H), 7.38 (s, 1H), 7.03–7.00 (m, 2H), 6.69–6.66
(m, 2H), 6.25 (s, 1H), 3.40–3.35 (m, 2H), 2.72–2.67 (m, 2H), 2.03 (s,
9H), 1.72 (s, 6H); MS (EI): 407 (M+).
5-Adamantan-1-yl-N-(2,4-dihydroxybenzyl)-2-hydroxy-4-methoxy-
benzamide (43): 1H NMR (300 MHz, DMSO-d6): d 12.73 (s, 1H), 9.40
(s, 1H), 9.11 (s, 1H), 8.95 (m, 1H), 7.52 (s, 1H), 6.90 (d, 1H, J = 8.4 Hz),
6.43 (s, 1H), 6.28 (s, 1H), 6.17 (d, 1H, J = 8.4 Hz), 4.31 (d, 2H,
J = 5.4 Hz), 3.79 (s, 3H), 2.00 (s, 9H), 1.71 (s, 6H); MS (EI): 423 (M+).
N-(2,4-Dihydroxybenzyl)-2,4-dimethoxybenzamide (44): 1H NMR
(300 MHz, DMSO-d6): d 9.65 (s, 1H), 9.13 (s, 1H), 8.52 (m, 1H),
7.87 (d, 1H, J = 8.4 Hz), 6.93 (d, 1H, J = 8.7 Hz), 6.63 (m, 2H), 6.28
(s, 1H), 6.16 (d, 1H, J = 8.1 Hz), 4.30 (d, 2H, J = 5.7 Hz), 3.90 (s,
3H), 3.81 (s, 3H); MS (EI): 303 (M+).
N-[2-(4-Hydroxyphenyl)ethyl]-2,4-dimethoxybenzamide (45): 1H
NMR (300 MHz, DMSO-d6): d 9.19 (s, 1H), 7.95 (br s, 1H), 7.82 (d,
1H, J = 9.3 Hz), 7.04 (d, 2H, J = 7.5 Hz), 6.70 (d, 2H, J = 7.5 Hz),
6.62 (m, 2H), 3.81 (s, 3H), 3.80 (s, 3H), 3.48–3.41 (m, 2H), 2.72–
2.67 (m, 2H); MS (EI): 301 (M+).
N-(3,4-Dihydroxybenzyl)-4-hydroxybenzamide (46): 1H NMR
(300 MHz, DMSO-d6): d 9.92 (br s, 1H), 8.79 (br s, 1H), 8.66–8.61
(m, 2H), 7.75 (d, 2H, J = 8.7 Hz), 6.78 (d, 2H, J = 8.4 Hz), 6.69 (s,
1H), 6.64 (d, 1H, J = 7.8 Hz), 6.54 (d, 1H, J = 7.8 Hz), 4.26 (d, 2H,
J = 8.4 Hz); MS (EI): 259 (M+).
3-Adamantan-1-yl-N-[2-(4-hydroxyphenyl)ethyl]-4-methoxybenz-
amide (47): 1H NMR (300 MHz, DMSO-d6): d 9.14 (s, 1H), 8.33 (br s,
1H), 7.67–7.63 (m, 2H), 7.02–6.98 (m, 3H), 6.67 (d, 2H, J = 8.1 Hz),
3.83 (s, 3H), 3.40–3.35 (m, 2H), 2.72–2.67 (m, 2H), 2.05 (s, 9H),
1.74 (s, 6H); MS (EI): 405 (M+).
3-Adamantan-1-yl-N-(3,5-dimethoxybenzyl)-4-hydroxybenzamide
(48): 1H NMR (300 MHz, DMSO-d6): d 9.85 (s, 1H), 8.71 (br s, 1H),
7.64–7.56 (m, 2H), 6.78 (d, 1H, J = 7.8 Hz), 6.45 (s, 2H), 6.36 (s,
3-Adamantan-1-yl-N-(3,4-dimethoxybenzyl)-4-methoxybenzamide
(35): 1H NMR (300 MHz, DMSO-d6): d 8.79 (br s, 1H), 7.75–7.69 (m,

Y. D. Hong et al. / Bioorg. Med. Chem. Lett. 24 (2014) 667–673
673
1H), 4.36 (d, 2H, J = 5.4 Hz), 3.71 (s, 6H), 2.08 (s, 9H), 1.73 (s, 6H);
MS (EI): 421 (M+).
mol. Their charges were calculated by the Gasteiger–Huckel
method.
N-(3,4-Dihydroxybenzyl)-2,4-dihydroxybenzamide (49): 1H NMR
(300 MHz, DMSO-d6): d 12.5 (br s, 1H), 10.07 (br s, 1H), 8.90–
8.72 (m, 3H), 7.70 (d, 1H, J = 8.7 Hz), 6.69–6.21 (m, 5H), 4.28 (d,
2H, J = 5.4 Hz); MS (EI): 275 (M+).
References and notes
1. Gupta, A. K.; Gover, M. D.; Nouri, K. N.; Taylor, S. J. Am. Acad. Dermatol. 2006, 55,
1048.
5-Adamantan-1-yl-2,4-dihydroxy-N-(2-hydroxy-4-methoxybenzyl)benz-
amide (50): 1H NMR (300 MHz, DMSO-d6): d 12.39 (br s, 1H), 9.92 (s,
1H), 9.65 (s, 1H), 8.89 (br s, 1H), 7.47 (s, 1H), 7.02 (d, 1H, J = 8.4 Hz),
6.37–6.26 (m, 3H), 4.32 (s, 2H), 3.66 (s, 3H), 2.02 (s, 9H), 1.70 (s, 6H);
MS (EI): 423 (M+).
Biological evaluation: Measurement of melanin content: The cells
(2 ꢁ 105 cells/mL) were seeded into 24-well plates and derivatives
were added in triplicate. The medium was changed daily and after
4 d of culturing, the cells were lysed with 1 mL of 1 N NaOH. Sub-
sequently, 200 ll of each crude cell extract was transferred into
96-well plates. The relative melanin content was measured at
400 nm with a microplate reader (Molecular Devices).
3D QSAR analysis: 3D-QSAR studies were performed using SYB-
YL-X 2.0 running on a Windows 7 Home Premium K 64 bit OS plat-
form (http://www.tripos.com (St. Louis, MO, 2011)). Each structure
of a reasonably low-energy conformation was fully geometry-
optimized using a standard Tripos molecular mechanics force field
and conjugate gradient with a convergence criterion of 0.05 kcal/
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