Novel hydroxyphenylurea dual inhibitor against Acyl-CoA: Cholesterol acyltransferase (ACAT) and low density lipoprotein (LDL) oxidation as antiatherosclerotic agent

Article abstract of DOI:10.1016/S0968-0896(00)00303-5

Novel hydroxyphenylurea derivatives were synthesized and their inhibitory potency evaluated against acyl-CoA: cholesterol acyltransferase ACAT. Quantitative structure-activity relationship analysis revealed that their ACAT inhibitory activities were controlled by the hydrophobicity of the whole molecule, the substitution pattern of urea moiety, and the existence of carboxylic acid. The derivatives with strong activities inhibited foam cell formations. Moreover, these compounds showed antioxidative effects against low density lipoprotein LDL, owing to their characteristic 3-tert-butyl-2-hydroxy-5-methoxyphenyl substructure. Based on the mechanism of atherosclerosis generation, this hydroxyphenylurea-type dual inhibitor against both ACAT and LDL oxidation is expected to be a promising drug for atherosclerosis. Copyright

Full text of DOI:10.1016/S0968-0896(00)00303-5

Bioorganic & Medicinal Chemistry 9 (2001) 853±861
Novel Hydroxyphenylurea Dual Inhibitor Against Acyl-CoA:
Cholesterol Acyltransferase (ACAT) and Low Density Lipoprotein
(LDL) Oxidation as Antiatherosclerotic Agent
Kazuya Nakao,a Hitoshi Kubota,a Mikiko Yasuhara,b Keiko Saito,b Toshikazu Suzuki,b
Hiroshi Ohmizua and Ryo Shimizua,*
aDiscovery Research Laboratory, Tanabe Seiyaku Co., Ltd, 3-16-89 Kashima, Yodogawa, Osaka 532-8505, Japan
bDiscovery Research Laboratory, Tanabe Seiyaku Co., Ltd, 2-2-50 Kawagishi, Toda, Saitama 335-8505, Japan
Received 21 July 2000; accepted 28 October 2000
AbstractÐNovel hydroxyphenylurea derivatives were synthesized and their inhibitory potency evaluated against acyl-CoA: cho-
lesterol acyltransferase (ACAT). Quantitative structure±activity relationship analysis revealed that their ACAT inhibitory activities
were controlled by the hydrophobicity of the whole molecule, the substitution pattern of urea moiety, and the existence of carboxylic
acid. The derivatives with strong activities inhibited foam cell formations. Moreover, these compounds showed antioxidative e€ects
against low density lipoprotein (LDL), owing to their characteristic 3-tert-butyl-2-hydroxy-5-methoxyphenyl substructure. Based on
the mechanism of atherosclerosis generation, this hydroxyphenylurea-type dual inhibitor against both ACAT and LDL oxidation is
expected to be a promising drug for atherosclerosis. # 2001 Elsevier Science Ltd. All rights reserved.
Introduction
the intestine.6 Since the inhibition of ACAT can lower
plasma cholesterol level by the prevention of cholesterol
absorption, the ACAT inhibitors are expected to have
therapeutic e€ects for hypercholesterolemia. Moreover,
the inhibitors can reduce the extent of atherosclerotic
lesions by preventing macrophages from converting to
foam cells. Various ACAT inhibitors having urea sub-
structure (Fig. 2) are now under development for the
treatment of hypercholesterolemia and atherosclerosis.7,8
Atherosclerosis causes heart attack, stroke, and gang-
rene of the extremities.1 In the early stage of athero-
sclerosis, the massive accumulation of foam cells is
observed within the innermost layer of the artery wall.
The foam cells are mainly developed from monocytes/
macrophages, and ®lled with lipid droplets consisting of
cholesteryl esters. The hypothetical mechanism of the
formation of foam cells in the atherosclerotic lesions is
illustrated in Figure 1.2 Oxidized low density lipoprotein
(ox-LDL), viruses, and toxins can injure the endothelial
cells, which leads the adhesion of macrophages to the
artery wall.3 Then the macrophages migrate into the
intima and become foam cells owing to the unregulated
uptake of ox-LDL via the scavenger receptor.4 The ox-
LDL is hydrolyzed in the macrophages and the resulting
cholesterol is acylated again to form cholesteryl ester by
acyl-CoA: cholesterol acyltransferase (ACAT).5 Chol-
esteryl ester forms lipid droplets in macrophages which
®nally turn into mature foam cells.
Antioxidants are also promising as therapeutic agents
for atherosclerosis.9 The ox-LDL can play an important
role in atherosclerosis by injuring endothelial cells of the
artery wall and supplying cholesterol to macrophages.
Antioxidants could decrease the amount of the ox-LDL
production, which leads to the treatment for athero-
sclerosis. Vitamin E, a natural antioxidant, and probucol,
the well-known drug for hypercholesterolemia (Fig. 3),
have been known to reduce the progression of athero-
sclerosis by the suppression of oxidative stress.10
In our previous paper, we reported the antioxidantive
activities of hydroxyphenylureas.11 It was revealed that
hydroxyphenylureas with strong antioxidative activity
had electron-donating substituents on their phenol rings,
bulky substituents at the ortho-positions of the phenolic
ACAT is the key enzyme in both the foam cell forma-
tion of macrophages and the cholesterol absorption in
*Corresponding author. Tel.: +81-6-6300-2568; fax: +81-6-6300-2590
0968-0896/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(00)00303-5

854
K. Nakao et al. / Bioorg. Med. Chem. 9 (2001) 853±861
Figure 1. Hypothetical mechanism of the foam cell formation.
Figure 2. ACAT inhibitors.
hydroxyl group, and no carboxylate in the molecule. In
the course of our random screening programs searching
for useful biological e€ects of these hydroxyphenylureas,
compound 15 was found to lower the amount of plasma
cholesterol in rats. Because the hydroxyphenylurea 15
has a phenylurea moiety which is common to some
ACAT inhibitors,7,8 we assumed that it exhibited the
biological activity by inhibiting ACAT in the intestine.
Thus, we examined the inhibitory activity of the
hydroxyphenylurea 15 against ACAT and found it to
show moderate inhibition (14% inhibition at 1 mM).
Then we obtained various hydroxyphenylurea deriva-
tives and evaluated their ACAT inhibitory activities in
order to get a dual inhibitor against both ACAT activity
and LDL oxidation, which could be a promising drug
for hypercholesterolemia and atherosclerosis. In this
paper, we report the lead generation and optimization
of hydroxyphenylureas as dual inhibitors against ACAT
and LDL oxidation, and discuss their structure±activity
relationships.

K. Nakao et al. / Bioorg. Med. Chem. 9 (2001) 853±861
855
ortho-hydroxyaniline 31 with the protected hydroxy
group and the secondary amines 30 were combined by
triphosgene to give the methoxymethoxyphenylureas 32.
By removing the protecting group by treatment with
methanolic hydrogen chloride, trisubstituted ureas 6±26
were obtained in satisfactory yields. More detailed infor-
mation is shown in the Experimental.
Figure 3. Antioxidants.
Chemistry
Results and Discussion
Our principles of molecular design of dual inhibitors
against ACAT and LDL oxidation are shown in Figure 4.
First, in order to maintain antioxidative activity, we selec-
ted the 3-tert-butyl-2-hydroxy-5-methoxyaniline group as
the optimized substructure at the phenol ring (A-ring)
position. In the previous article, we showed that it was
able to modify hydroxyphenylureas around N-sub-
stituents on the opposite side of phenol ring (B-ring) part
without any decrease in antioxidative activity.11 Conse-
quently, various substituents were introduced in order
to obtain compounds with strong ACAT inhibitory
potency. The syntheses of compounds 1±5 in Table 1 have
been reported in the previous paper. The derivatives 6±26
were synthesized according to the procedure in Scheme 1.
Secondary amines 30 were prepared by alkylation of 28,
followed by removal of the protecting group. Then, the
Inhibitory activity against ACAT
The ACAT inhibitory activities were shown by the
decrease percentage (% inhibition) of the cholesteryl
ester production by ACAT at 1 mM dosing. Compounds
with strong inhibitory activity against ACAT had high
percentage values. The inhibitory potencies of the 1,3-
disubstituted ureas were very weak except for 1, the
logP value of which was comparatively high (4.48). CI-
976 (27),12 a well-known ACAT inhibitor, has also high
logP value (6.17). The disubstituted ureas 2±8 might show
low inhibitory activities because of their low hydro-
phobicity. In order to examine structural e€ects on ACAT
inhibition, we analyzed quantitative structure±activity
relationship (QSAR) of both di- and tri-substituted
ureas. The ACAT inhibition potencies were transformed
into their logit {logit=log[% inhibition/(100À% inhibi-
tion)]}.13 By this transformation, the value of % inhibi-
tion could correspond to the amount of compounds
interacting with the enzymes. The ratio of % inhibition to
(100À% inhibition) could be proportional to the binding
constants of compounds with the enzymes, and it was
shown that logit could be successfully applicable as the
indices of biological activity in QSAR studies.13À15 Com-
pounds with strong inhibitory potency had high logit
values and low 50% inhibition concentration values
(IC50). The % inhibition value and logit of every com-
pound, and moreover the IC50 values of rather high
potent compounds, are shown in Table 2, as well as every
compound's physicochemical parameters for QSAR ana-
lysis. The logP value, which was calculated based on the
summation of hydrophobicity of substituents and the
electronic and steric e€ects between substituents,11 indi-
cated the hydrophobicity of the whole molecule. The
ITRI and ICOOH terms were indicator variables, which
take a value of unity for tri-substituted ureas and
carboxylic acids, respectively, and zero for others. By
the correlation analysis of these parameters, eq 1 was
formulated.
Figure 4. Principles of molecular design.
Scheme 1. (a) (1) NaH, DMF, (2) H2, Pd-C, EtOH; (b) triphosgene, Et3N, CH2Cl2; (c) concd HCl, MeOH.

856
K. Nakao et al. / Bioorg. Med. Chem. 9 (2001) 853±861
Table 1. Structures and chemical data of hydroxyphenylureas
Substituents
No.
X
Y
Formula
mp (ꢀC)
Anal.a
1b
2b
3b
4b
5b
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
2,6-Diisopropylphenyl
2,4-Di¯uorophenyl
Cyclohexyl
H
H
H
H
H
H
H
H
C6H5
C6H5
C6H5
C6H5
C6H5
C6H5
C6H5
C6H5
C6H5
C24H34N2O3
C18H20N2O3F2
C18H28N2O3
C18H28N3O3
193±195
153±154
147±148
160±162
165±167
69±71
137±139
169±170
68±70
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHN
CHNF
CHN
CHN
CHN
CHN
CHNCl
CHNCl
CHN
Anilino
.
3-Pyridylmethyl
(CH2)3CO2C2H5
C(CH3)(CH2OH)2
C(CH2OH)3
C18H23N3O3 HCl
C18H28N2O5
C16H26N2O5
C16H26N2O6
C23H30N2O5
(CH2)2CO2C2H5
(CH2)3CO2C2H5
(CH2)3CONHC2H5
(CH2)3CON(C2H5)2
(CH2)3CO-Morpholino
(CH2)3CONHCH2CH(OH)CH2OH
(CH2)3CO2H
.
C24H32N2O5 1/4CHCl3
Syrup
C24H33N3O4
C26H37N3O4
C26H35N3O5
C25H35N3O6
C22H28N2O5
C23H30N2O5
C25H34N2O5
C26H37N3O5
C24H30N2O5F2
C28H34N2O5
C24H38N2O5
C25H40N2O5
C25H34N2O3
156±158
131±133
129±131
102±105
127±128
132±133
86±89
(CH2)4CO2H
(CH2)6CO2H
(CH2)3CO2C2H5
(CH2)3CO2C2H5
(CH2)3CO2C2H5
(CH2)3CO2C2H5
(CH2)3CO2C2H5
Cyclohexyl
Cycloheptyl
CH2C6H5
2-Cyclohexylethyl
4-Dimethylaminophenyl
2,4-Di¯uorophenyl
1-Naphthyl
Cyclohexyl
Cycloheptyl
CH2C6H5
4-Dimethylaminophenyl
4-Dimethylaminophenyl
4-Dimethylaminophenyl
77±79
115±116
104±105
113±115
99±101
140±141
140±142
183±185
124±125
. .
C27H39N3O3 HCl C2H5OH
.
C27H33N3O3 HCl
.
C28H41N3O3 1/4H2O
aDeviation between calculated and observed values were less than 0.4%.
bReported in our previous paper, ref 11.
logit ˆ 0:30 ꢀÆ0:11† logP ‡ 0:62 ꢀÆ0:39† ITRI
tri-substituted ureas inhibited the ACAT activity stronger
than did the di-substituted derivatives.18 The e€ect of
the substitution pattern shown quantitatively by ITRI
term coincides with the results by DeVries et al. How-
ever, it remains unclear at the molecular level why the
number of substituents can a€ect the anity between
the enzyme and the inhibitor.
À 1:28 ꢀÆ0:51† ICOOH À 0:98 ꢀÆ0:38†
n ˆ 26; s ˆ 0:39; r ˆ 0:90; F3;22 ˆ 31:90
ꢀ1†
In this equation, n is the number of tested compounds, s
the standard deviation, r the correlation coecient, and F
the ratio between regression and residual variances. The
®gures in parentheses are the 95% con®dence intervals.
The coecient of the ICOOH term in eq 1 was negative.
The hydroxyphenylurea with a carboxylic acid moiety
would exhibit 20 times lower activity than was expected.
The negative charge or the local hydrophilicity of car-
boxylate might prevent the inhibitor from interacting
with the enzyme.
The coecient of the logP term was positive, indicating
that compounds with higher hydrophobicity showed
stronger inhibitory activities against ACAT. In the assay
system used in this study, ACAT was obtained as the
hydrophobic microsome fraction from the homogenate of
rabbit intestinal mucosal scrapings by sequential cen-
trifugation. Cholesterol, which is highly hydrophobic
(logP=9.52 estimated by CLOGP software16,17), is acy-
lated by ACAT under a hydrophobic environment like
microsome fractions. The hydrophobic character of
hydroxyphenylurea ACAT inhibitors is thought to be
necessary so that inhibitor molecules could approach
the enzyme in the hydrophobic micrsomes.
Inhibition of the foam cell formation
Compounds with strong inhibitory activities against
ACAT were examined for the inhibitory potency of the
foam cell formation of macrophages. Compound 26
inhibited the transformation of macrophages to foam
cells stronger than CI-976. Compound 12 and 24 were
as potent as CI-976. They had the proper hydrophobic
character (their logP values were more than 3 as shown
in Table 2), and could permeate into macrophage cells.
Compounds with weak inhibitory activity against the
foam cell formation had weak ACAT inhibitory activity,
which might indicate that the foam cell formation of
macrophages was mainly controlled by ACAT activity.
The coecient of the ITRI term was positive, and it was
revealed that tri-substituted ureas showed about four
times stronger activity than di-substituted ones on the
average. DeVries et al. reported the similar results: in the
case of their ACAT inhibitors having phenylurea moiety,

K. Nakao et al. / Bioorg. Med. Chem. 9 (2001) 853±861
Table 2. Biological activities and physicochemical parameters of hydroxyphynylureas
857
No.
ACAT Inhibitory activity
Physicochemical parameters
Logita
Calcdb
Foam cell formation
inhibition
LDL oxidation
inhibition
IC50 (mM)
logPc
ITRI
ICOOH
d
e
% Inhibition
at 1 mM
Obsd.
Dev.
IC50 (mM)
IC50 (mM)
1
2
3
4
5
6
7
8
92
42
43
42
23
19
7
13
62
96
83
85
84
22
14
4
60
80
76
92
84
88
96
97
90
95
1.06
À0.14
À0.12
À0.14
À0.52
À0.63
À1.12
À0.83
0.21
1.38
0.69
0.75
0.72
À0.55
À0.79
À1.38
0.18
0.60
0.50
1.06
0.72
0.87
1.38
0.39
0.10
0.10
À0.12
À0.45
À0.36
À0.87
À1.23
0.80
0.70
0.30
0.56
0.30
À0.22
À0.88
À0.72
À0.39
0.76
0.91
1.07
0.78
0.96
1.22
1.47
1.19
1.55
0.67
À0.24
À0.22
À0.02
À0.07
À0.27
À0.25
0.40
0.06
Ð
Ð
Ð
Ð
Ð
Ð
Ð
Ð
Ð
Ð
0.08
0.27
Ð
Ð
Ð
Ð
0.20
Ð
0.05
0.26
0.17
0.04
0.05
0.10
0.07
4.4
Ð
Ð
Ð
Ð
Ð
Ð
Ð
0.09
Ð
Ð
Ð
Ð
Ð
Ð
Ð
Ð
Ð
Ð
0.11
0.20
Ð
Ð
Ð
Ð
Ð
Ð
0.06
Ð
Ð
0.12
0.22
0.12
0.79
4.48
3.55
3.56
2.84
1.80
2.08
0.46
À0.69
3.82
3.48
2.22
3.03
2.20
0.54
2.54
3.07
4.12
3.68
4.16
4.66
3.74
4.30
5.13
5.96
5.06
6.39
0
0
0
0
0
0
0
0
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
0
0
0
0
0
0
0
0
0
0
0
0
0
0
1
1
1
0
0
0
0
0
0
0
0
0
9
À0.59
Ð
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
0.68
0.39
>10
>10
1.1
4.4
Ð
Ð
Ð
Ð
>10
Ð
7.6
>10
>10
3.9
1.5
3.0
0.44
0.19
0.42
À0.33
0.09
À0.66
0.57
À0.16
À0.41
À0.01
À0.06
À0.09
0.16
1.51
0.95
1.28
0.04
À0.24
À0.27
27
87
0.83
Ð
Ð
0.09
1.4
Ð
6.17
Ð
±
alogit=log{% inhibition/(100À% inhibition)}.
bCalculated by eq 1.
cCalculated by our method, ref 11.
dIndicator variable for trisubstituted ureas.
eIndicator variable for carboxylic acids.
LDL oxidation inhibition
pounds were also potent in the inhibition of the foam cell
formation. Owing to the characteristic phenolic sub-
structure, hydroxyphenylureas could potently inhibit
LDL oxidation. These hydroxyphenylureas were dual
inhibitors of ACAT and LDL oxidation, which are
important for the treatment of atherosclerosis. Further
improvement and development of hydroxyphenylureas
are now being undertaken.19,20
All compounds in this report have a 3-tert-butyl-2-
hydroxy-5-methoxyphenylurea moiety which was opti-
mized for the strong antioxidative activity. Therefore,
those hydroxyphenylureas were expected to inhibit LDL
oxidation. Compouds with strong inhibitory activities
against the foam cell formation (1, 13, 14, 20, and 23±
26) were examined for their LDL oxidation inhibitory
potency. As we expected, all of the eight derivatives
inhibited the endothelial cell-induced oxidation of LDL
at the submicromolar level. Thus, we obtained dual
inhibitors against ACAT and LDL oxidation by the
rational structural evolution of the hydroxyphenylurea
antioxidant.
Experimental
Inhibitory activities of ACAT and the foam cell
formation of macrophages
ACAT enzyme was prepared from the intestines of
male Japanese White rabbits (Oriental Yeast, Japan).
Intestinal mucosal scrapings were suspended in 0.25 M
sucrose solution, and the suspension was homogenized
in 0.154 M potassium phosphate bu€er (pH 6.2) with
a Te¯on Potter homogenizer (Iuchi, Japan). The
microsomes were obtained from the mucosal homo-
genate by sequential centrifugation, and used for
ACAT assay. ACAT activity was determined by
measuring the incorporation of [14C]oleoyl-CoA into
cholesteryl oleate according to the methods of Heider
et al.6
Conclusion
We obtained hydroxyphenylureas with strong inhibitory
activities against both ACAT and LDL oxidation.
According to the result of QSAR analysis, it was revealed
that the ACAT inhibition depended on the hydrophobicity
of the whole molecule, the number of substituents at the
urea nitrogens, and the existence of carboxylates. Some of
these compounds exhibited comparable ACAT inhibition
with CI-976, a well-known ACAT inhibitors. These com-

858
K. Nakao et al. / Bioorg. Med. Chem. 9 (2001) 853±861
The foam cell formation was evaluated by measuring the
incorporation of [14C]oleic acid into cholesteryl ester in
the whole cells by the method of Tabas et al.21 The murine
macrophage cell line J774A.1 cells were obtained from
America Type Tissue Culture Collection (ATCC; Cam-
den, NJ, USA), grown in a 6-well plate (Iwaki Glass,
Japan), maintained in Dulbecco's modi®ed Eagle's med-
ium (DMEM; Nissui Pharmaceutical Co., Ltd, Tokyo),
and supplemented with 10% fetal bovine serum (FBS;
CSL Ltd, Rarville, Australia), penicillin (100 IU/mL),
streptomycin (100 mg/mL), and glutamine (292 mg/mL)
at 37 ꢀC in a 5% CO2 humidi®ed atmosphere. Mono-
layers of J774A.1 cells and mouse peritoneal macro-
phages were preincubated for 2 h in DMEM containing
10% lipoprotein de®cient serum and then incubated for
18 h in the presence of 30 mg/mL of oxidized LDL with
[14C]oleate-albumin complex, and the lipid extracts of
these cells were assayed for cholesteryl [14C]oleate.
Tested compounds were added with [14C]oleate-albumin
complex.
zyloxycarbonyl-N-(1-naphthyl)amine (28, 20.0 g, 72.1
mmol) at 0 ꢀC under nitrogen atmosphere and the mix-
ture was stirred for 90 min at rt. To the suspension was
added slowly a solution of ethyl 4-bromobutyrate (29,
14.8 g, 75.7 mmol) in DMF (50 mL) at 0 ꢀC under
nitrogen atmosphere and the reaction mixture was stir-
red for 90 min at rt. After the mixture was concentrated
in vacuo, the residue was extracted with ethyl acetate
(AcOEt), the organic layer was washed with brine, dried
(MgSO4), ®ltered and concentrated in vacuo. The resi-
due was puri®ed on silica gel chromatography using
hexane/AcOEt (3:1) to a€ord ethyl 4 - [N - benzyloxy-
carbonyl-N-(1-naphthyl)amino]butyrate (25.5 g, 90%):
1
syrup; IR (®lm) 2981, 1706, 1597 cmÀ1; H NMR (d in
CDCl3) 1.17 (t, 3H, J=7 Hz, CH3), 1.93 (m, 2H, CH2),
2.29±2.36 (m, 2H, CH2), 4.00±4.11 (m, 4H, CH2), 5.03
(s, 2H, CH2), 6.95±7.52 (m, 9H, ArH, NapH-3, -4, -6,
and -7), 7.73±7.89 (m, 3H, NapH-2, -5, and -8); EIMS
m/z 391 (M+). A mixture of ethyl 4-[N - benzyloxy-
carbonyl - N - (1 - naphthyl)amino]butyrate (25.5 g, 65.1
mmol), 10% palladium on carbon (5.0g) and EtOH
(400mL) was subjected to hydrogenation using Parr
apparatus (H2, 3.5 atm) for 6 h at rt. After removal of
catalyst by ®ltration, the ®ltrate was concentrated in
vacuo. The residue was puri®ed on silica gel chromato-
graphy using hexane/AcOEt (3:1) to a€ord 30 (X=(CH2)3
CO2C2H5, Y=1-naphthyl, 11.1 g, 66%): syrup; IR (®lm)
Inhibition against LDL oxidation
The e€ects of hydroxyphenylureas on the endothelial cell-
induced oxidation of LDL were assessed by the increase in
the thiobarbituric acid reactive substance (TBARS)
production.22À24 Bovine aortic endothelial cells were iso-
lated from the aorta with a scrape and cultured in Eagle's
minimum essential medium (Nissui Pharmaceutical Co.,
Ltd, Tokyo) a supplemented with 10% FBS, penicillin
(100 IU/mL) and streptomycin (100 mg/mL) at 37 ꢀC in a
5% CO2 humidi®ed atmosphere. The endothelial cell-
induced oxidation of LDL was achieved by incubating
100 mg/mL protein of LDL for 18h at 37 ꢀC in the pre-
sence of con¯uent endothelial cells in serum-free Ham's
F10 medium. Tested compounds were dissolved in
DMSO and added at the indicated concentration simul-
taneously with the LDL. The amounts of TBARS were
calculated as nanomole equivalents of malondialdehyde.
1
2981, 1732, 1583 cmÀ1; H NMR (d in CDCl3) 1.24 (t,
3H, J=7 Hz, CH3), 2.04±2.17 (m, 2H, CH2), 2.51 (t, 2H,
J=7 Hz, CH2), 3.33 (t, 2H, J=6.6 Hz, CH2), 4.14 (q,
2H, J=7 Hz, CH2), 6.58 (d, 1H, J=7.2 Hz, NH), 7.19±
7.51 (m, 4H, ArH, NapH-3, -4, -6, and -7), 7.75±7.87
(m, 3H, NapH-2, -5, and -8); EIMS m/z 257 (M+).
In the same manner, the other disubstituted amines
were obtained.
Urea formation and removal of the methoxymethyl
protecting group
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-(3-ethoxy-
carbonylpropyl)-1-(1-naphthyl)urea (20). To a solution
of triphosgene (2.63 g) in CH2Cl2 (150 mL), the solution
of 3-tert-butyl-5-methoxy-2-methoxymethoxyaniline 31
(6.1 g, 25.3 mmol) and triethylamine (10.6 mL) in CH2Cl2
(100 mL) were added dropwise at À78 ꢀC and the reac-
tion mixture was warmed up to rt for 1 h. After removal
of the solvent, the residue was dissolved in CH2Cl2
(150mL) and the solution of ethyl 4-(1-naphthylamino)-
butyrate (30; X=(CH2) 3CO2C2H5, Y=1-naphthyl, 6.5 g,
25.3mmol) and triethylamine (5.3mL) in CH2Cl2
(100mL) was added dropwise at rt. After stirring at the
same temperature for 10h, the reaction mixture was
washed with brine, dried (MgSO4), ®ltered and con-
centrated in vacuo, and the residue was chromatographed
on silica gel using hexane/AcOEt (3:1) to yield 3.52 g
(27%) of 3-(3-tert-butyl-5-methoxy-2-methoxymethoxy-
phenyl)-1-(3-ethoxycarbonylpropyl)-1-(1-naphthyl)urea
(32; X=(CH2) 3CO2C2H5, Y=1-naphthyl): syrup; IR
(®lm) 2957, 1736, 1677, 1592cmÀ1; 1H NMR (d in CDCl3)
1.77 (t, 3H, J=7 Hz, CH3), 1.22 (s, 9H, CH3), 1.93±2.04
(m, 2H, CH2), 2.27±2.47 (m, 2H, CH2), 2.63 (s, 3H, CH3),
3.82 (s, 3H, CH3), 4.01±4.44 (m, 6H, CH2), 6.50 (d, 1H,
Synthesis
Melting points were determined on Yamato melting point
apparatus and are uncorrected. Infrared spectra were
recorded on a Perkin±Elmer 1640 IR spectrophotometer.
Proton nuclear magnetic resonance (1H NMR) spectra
were recorded on a Bruker AC-200 (200MHz) spectro-
meter with tetramethylsilane as the internal standard.
Mass spectra were recorded on a Hitachi M-2000A
spectrometer. Elemental analyses were performed on a
Perkin±Elmer 2400 II. Column chromatography was
accomplished by using Kieselgel 60 (230±400 mesh, E.
Merck) with the indicated solvent system.
The procedures of syntheses and the spectral data of
compounds 1±5 were described in our previous article.11
Preparation of disubstituted amines 30
Ethyl 4-(1-naphthylamino)butyrate (30; X=(CH2)3CO2
C2H5, Y=1-naphthyl). To a suspension of NaH (3.17 g,
79.3 mmol, 60% mineral oil suspended) in N,N-dimethyl-
formamide (DMF, 100 mL) was added dropwise N-ben-

K. Nakao et al. / Bioorg. Med. Chem. 9 (2001) 853±861
859
J=3 Hz, ArH), 7.12 (s, 1H, NH), 7.54±7.58 (m, 4H,
NapH-3, -4, -6, and -7), 7.83 (d, 1H, J=3 Hz, ArH),
7.93±7.98 (m, 3H, NapH-2, -5, and -8); EIMS m/z 522
(M+). To a solution of 32 (R=3-C(CH3)3-5-OCH3,
X=(CH2)3 CO2C2H5, Y=1-naphthyl, 3.52 g, 6.74
mmol) in MeOH (30 mL) was added concd HCl
(3.0 mL) and the mixture was stirred at rt for 1 h. The
resulting mixture was concentrated in vacuo and the
crude crystals were recrystallized from AcOEt-diisopro-
pylether to give 20 (940 mg, 29%): IR (KBr) 3350, 2951,
1732 cmÀ1; 1H NMR (d in CDCl3) 1.21 (t, 3H, J=7 Hz,
CH3), 1.40 (s, 9H, CH3), 1.96±2.03 (m, 2H, CH2), 2.36±
2.45 (m, 2H, CH2), 3.46±3.53 (m, 1H, CH2), 3.57 (s, 3H,
CH3), 4.09 (q, 2H, J= 7 Hz, CH2), 4.16±4.30 (m, 1H,
CH2), 5.77 (d, 1H, J=3 Hz, ArH), 6.00 (s, 1H, NH or
OH), 6.66 (d, 1H, J=3 Hz, ArH), 7.56±7.67 (m, 4H,
NapH-3, -4, -6, and -7), 7.90±7.99 (m, 3H, NapH-2, -5,
and -8), 8.75 (s, 1H, NH or OH); EIMS m/z 478 (M+).
Anal. calcd for C28H34 N2O5: C 70.27, H 7.16, N 5.85.
Found: C 69.98, H 7.16, N 5.73.
6.22 (s, 1H, NH or OH), 6.70 (d, 1H, J=3 Hz, ArH),
7.34±7.57 (m, 5H, ArH), 8.55 (s, 1H, NH or OH); EIMS
m/z 414 (M+). Anal. calcd for C23H30N2O5: C 66.65, H
7.30, N 6.76. Found: C 66.53, H 7.28, N 6.69.
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-(3-ethoxy-
carbonylpropyl) - 1-phenylurea (10). IR (®lm) 2955,
;
1732 cmÀ1 1H NMR (d in CDCl3) 1.22 (t, 3H, J=
7.2 Hz, CH3), 1.41 (s, 9H, CH3), 1.84±1.98 (m, 2H,
CH2), 2.38 (t, 2H, J=7.4 Hz, CH2), 3.66 (s, 3H, CH3),
3.79 (t, 2H, J=7.4 Hz, CH2), 4.05±4.16 (q, 2H, J=
7.2 Hz, CH2), 6.07 (d, 1H, J=3 Hz, ArH), 6.26 (s, 1H,
NH or OH), 6.70 (d, 1H, J=3 Hz, ArH), 7.34±7.56 (m,
5H, ArH), 8.71 (s, 1H, NH or OH); EIMS m/z 428
+
.
(M ). Anal. calcd for C24H32N2O5 1/4CHCl3: C 63.54,
H 7.09, N 6.11. Found: C 63.38, H 7.11, N 6.04.
4-[3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-phenyl-
ureido]butanoic acid (15). IR (KBr) 2955, 1734 cmÀ1; 1H
NMR (d in CDCl3) 1.41 (s, 9H, CH3), 1.84±1.98 (m, 2H,
CH2), 2.44 (t, 2H, J=7.4 Hz, CH2), 3.67 (s, 3H, CH3),
3.81 (t, 2H, J=7.2 Hz, CH2), 6.08 (d, 1H, J=3 Hz, ArH),
6.17 (s, 1H, NH or OH), 6.70 (d, 1H, J=3 Hz, ArH),
7.34±7.57 (m, 5H, ArH), 8.50 (s, 1H, NH or OH); SIMS
m/z 401 (M+H)+. Anal. calcd for C22H28N2O5: C
65.98, H 7.05, N 7.00. Found: C 65.95, H 6.97, N 6.79.
In the same manner, the following compounds were
obtained.
1-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-3-(3-ethoxy-
carbonylpropyl)urea (6). IR (KBr) 3386, 2954, 1735,
1
1578 cmÀ1; H NMR (d in CDCl3) 1.25 (t, 3H, J=7Hz,
CH3), 1.41 (s, 9H, CH3), 1.77Ð1.91 (m, 2H, CH2), 2.37 (t,
2H, J=7Hz, CH2), 3.22±3.32 (dd, 2H, J=6.1, 6.7 Hz,
CH2), 3.72 (s, 3H, CH3), 4.07±4.18 (q, 2H, J=7 Hz, CH2),
5.30 (m, 1H, NH or OH), 6.40 (d, 1H, J=3 Hz, ArH),
6.76 (d, 1H, J=3 Hz, ArH), 6.76 (s, 1H, NH or OH),
8.15 (s, 1H, NH or OH); EIMS m/z 352 (M+). Anal.
calcd for C18H28N2O5: C 61.35, H 8.01, N 7.95. Found:
C 61.50, H 8.14, N 8.13.
5-[3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-phenyl-
ureido]pentanoic acid (16). IR (KBr) 2955, 1714 cmÀ1
;
1H NMR (d in CDCl3) 1.41 (s, 9H, CH3), 1.64±1.68 (m,
4H, CH2), 2.38 (m, 2H, CH2), 3.67 (s, 3H, CH3), 3.74±
3.81 (m, 2H, CH2), 6.03 (d, 1H, J=3 Hz, ArH), 6.08 (s,
1H, NH or OH), 6.70 (d, 1H, J=3 Hz, ArH), 7.34±7.57
(m, 5H, ArH), 8.68 (s, 1H, NH or OH); SIMS m/z 415
(M+H)+. Anal. calcd for C23H30N2O5: C 66.65, H
7.30, N 6.76. Found: C 66.99, H 7.36, N 6.63.
1-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-3-[1,1-bis
(hydroxymethyl)ethyl]urea (7). IR (KBr) 3384, 2949,
7-[3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-phenyl-
ureido]heptanoic acid (17). IR (KBr) 3415, 2940,
1651, 1558 cmÀ1 1H NMR (d in CDCl3+DMSO-d6)
;
1
1.24 (s, 3H, CH3), 1.41 (s, 9H, CH3), 2.32 (br, 1H, OH),
3.64±3.78 (m, 4H, CH2), 3.72 (s, 3H, CH3), 4.34 (br, 1H,
OH), 6.34 (s, 1H, NH or OH), 6.40 (d, 1H, J=3 Hz,
ArH), 6.68 (d, 1H, J=3 Hz, ArH), 8.01 (s, 1H, NH or
OH), 8.84 (s, 1H, NH or OH); SIMS m/z 327 (M+H)+.
Anal. calcd for C16H26N2O5: C 58.88, H 8.03, N 8.58.
Found: C 58.73, H 8.13, N 8.71.
1705 cmÀ1; H NMR (d in CDCl3) 1.34±1.61 (m, 8H,
CH2), 1.41 (s, 9H, CH3), 2.32 (t, 2H, J=7.4 Hz, CH2),
3.66 (s, 3H, CH3), 3.73 (t, 2H, J=7.2Hz, CH2), 6.03 (d,
1H, J=3 Hz, ArH), 6.07 (s, 1H, NH or OH), 6.70 (d, 1H,
J=3 Hz, ArH), 7.33±7.56 (m, 5H, ArH), 8.77 (s, 1H, NH or
OH); EIMS m/z 442 (M+). Anal. calcd for C25H34N2O5:
C 67.85, H 7.74, N 6.33. Found: C 67.76, H 7.76, N 6.13.
1-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-3-[1,1-bis
(hydroxymethyl)-2-hydroxyethyl]urea (8). IR (KBr)
3349, 2955, 1732, 1567 cmÀ1; 1H NMR (d in DMSO-d6)
1.34 (s, 9H, CH3), 3.59 (d, 6H, J=5.5 Hz, CH2), 3.64 (s,
3H, CH3), 4.73 (t, 3H, J=5.5 Hz, OH), 6.36 (s, 1H, NH or
OH), 6.47 (d, 1H, J=3 Hz, ArH), 6.57 (d, 1H, J=3 Hz,
ArH), 8.62 (s, 1H, NH or OH), 9.23 (s, 1H, NH or OH);
SIMS m/z 343 (M+H)+. Anal. calcd for C16H26 N2O6:
C 56.13, H 7.66, N 8.18. Found: C 56.08, H 7.71, N 8.20.
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-(4-N, N-di-
methylaminophenyl)-1-(3-ethoxycarbonylpropyl)urea (18).
IR (KBr) 3336, 2944, 1724 cmÀ1 1H NMR (d in
;
CDCl3) 1.23 (t, 3H, J=7.2 Hz, CH3), 1.41 (s, 9H, CH3),
1.82±1.97 (m, 2H, CH2), 2.37 (t, 2H, J=7.4Hz, CH2),
3.02 (s, 6H, CH3), 3.67 (s, 3H, CH3), 3.72 (t, 2H,
J=7.4Hz, CH2), 4.05±4.16 (q, 2H, J=7.2 Hz, CH2), 6.02
(d, 1H, J=3 Hz, ArH), 6.22 (s, 1H, NH or OH), 6.69 (d,
1H, J=3 Hz, ArH), 6.78 (d, 2H, J=9.2 Hz, ArH), 7.18 (d,
2H, J=9.2 Hz, ArH), 9.03 (s, 1H, NH or OH); EIMS
m/z 471 (M+). Anal. calcd for C26H37N3O5: C 66.22, H
7.91, N 8.91. Found: C 66.18, H 7.92, N 8.79.
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-(2-ethoxy-
carbonylethyl)-1-phenylurea (9). IR (KBr) 3358, 2954,
1729 cmÀ1; 1H NMR (d in CDCl3) 1.21 (t, 3H, J=7 Hz,
CH3), 1.41 (s, 9H, CH3), 2.66 (t, 2H, J=7.2 Hz, CH2),
3.67 (s, 3H, CH3), 4.02±4.13 (q, 2H, J=7 Hz, CH2), 4.02±
4.09 (t, 2H, J=7.2 Hz, CH2), 6.06 (d, 1H, J=3 Hz, ArH),
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-(3-ethoxy-
carbonylpropyl)-1-(2,4-di¯uorophenyl)urea (19). IR
1
(KBr) 3324, 2952, 1732 cmÀ1; H NMR (d in CDCl3)

860
K. Nakao et al. / Bioorg. Med. Chem. 9 (2001) 853±861
1.24 (t, 3H, J=7 Hz, CH3), 1.41 (s, 9H, CH3), 1.85±1.99
(m, 2H, CH2), 2.40 (t, 2H, J=7 Hz, CH2), 3.71 (s, 3H,
CH3), 3.71±3.77 (m, 2H, CH2), 4.08±4.18 (q, 2H,
J=7 Hz, CH2), 6.25 (d, 1H, J=3 Hz, ArH), 6.59 (br,
1H, NH or OH), 6.73 (d, 1H, J=3 Hz, ArH), 6.98±7.08
(m, 2H, ArH), 7.34±7.45 (m, 1H, ArH), 8.24 (s, 1H, NH
or OH); EIMS m/z 464 (M+). Anal. calcd for
C24H30N2O5F2: C 62.06, H 6.51, N 6.03, F 8.18. Found:
C 62.14, H 6.51, N 6.02, F 8.02.
N 8.68, Cl 7.32. Found: C 67.08, H 7.04, N 8.59, Cl
7.41.
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-(2-cyclo-
hexylethyl)-1-(4-N,N-dimethylaminophenyl)urea (26). IR
(KBr) 3404, 2922, 1639 cmÀ1; 1H NMR (d in DMSO-d6)
0.80±0.95 (m, 2H, CH2), 1.00±1.40 (m, 6H, CH2), 1.28
(s, 9H, CH3), 1.55±1.65 (m, 5H, CH2 and CH), 2.95 (s,
6H, CH3), 3.55±3.65 (m, 2H, CH2), 3.63 (s, 3H, CH3),
6.41 (d, 1H, J=3 Hz, ArH), 6.79 (d, 2H, J=9 Hz, ArH),
7.05 (d, 1H, J=3 Hz, ArH), 7.15 (d, 2H, J=9 Hz, ArH),
7.24 (s, 1H, NH or OH), 8.02 (s, 1H, NH or OH);
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-cyclo-
hexyl-1-(3-ethoxycarbonylpropyl) urea (21). IR (KBr)
1
+
3341, 2936, 1712 cmÀ1; H NMR (d in CDCl3) 1.27 (t,
EIMS m/z 467 (M ). Anal. calcd for C28H41N3O3 1/
4H2O: C 71.23, H 8.86, N 8.90. Found: C 71.01, H
8.49, N 8.88.
.
3H, J=7.2 Hz, CH3), 1.43 (s, 9H, CH3), 1.12±2.00 (m,
12H, CH2), 2.39±2.45 (m, 2H, CH2), 3.17±3.25 (m, 2H,
CH2), 3.74 (s, 3H, CH3), 4.12±4.23 (q, 2H, J=7.2 Hz,
CH2), 4.12±4.23 (m, 1H, CH), 6.58 (d, 1H, J=3 Hz,
ArH), 6.74 (d, 1H, J=3 Hz, ArH), 8.07 (s, 1H, NH or
OH), 9.35 (s, 1H, NH or OH); SIMS m/z 435 (M+H)+.
Anal. calcd for C24H38N2O5: C 66.33, H 8.81, N 6.45.
Found: C 66.29, H 8.82, N 6.34.
Synthesis of the derivatives including amide substructure
11±14
Compounds 11±14 were obtained from 3-(3-tert-butyl-
5-methoxy-2-methoxymethoxyphenyl)-1-(3-ethox-
ycarbonylpropyl)-1-phenylurea 32 (R=3-C(CH3)3-5-
OCH3, X=(CH2)3CO2C2H5, Y=phenyl). They were
hydrolyzed with 10% NaOH aq, made amide formation
with the corresponding amines by 1, 3-dicyclohexyl-
carbodiimide and N-hydroxysuccinimide, and removed
their methoxymethyl protecting group on the phenolic
hydroxyl with methanolic hydrogen chloride.
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-cyclohep-
tyl-1-(3-ethoxycarbonylpropyl) urea (22). IR (KBr)
1
3347, 2928, 1712 cmÀ1; H NMR (d in CDCl3) 1.27 (t,
3H, J=7.2 Hz, CH3), 1.43 (s, 9H, CH3), 1.43±2.04 (m,
14H, CH2), 2.39±2.45 (m, 2H, CH2), 3.17±3.25 (m, 2H,
CH2), 3.74 (s, 3H, CH3), 4.12±4.23 (q, 2H, J=7.2 Hz,
CH2), 4.33 (m, 1H, CH), 6.57 (d, 1H, J=3 Hz, ArH),
6.74 (d, 1H, J=3 Hz, ArH), 8.04 (s, 1H, NH or OH),
9.38 (s, 1H, NH or OH); EIMS m/z 448 (M+). Anal.
calcd for C25H40N2O5: C 66.94, H 8.99, N 6.25. Found:
C 67.07, H 9.03, N 6.19.
N-Ethyl-4-[3-(3-tert-butyl-2-hydroxy-5-methoxyphenyl)-
1-phenylureido]-butyramide (11). IR (KBr) 3303, 2940,
;
1631 cmÀ1 1H NMR (d in CDCl3) 1.13 (t, 3H, J=
7.2 Hz, CH3), 1.41 (s, 9H, CH3), 1.84±1.97 (m, 2H, CH2),
2.27 (d, 2H, J=6.8 Hz, CH2), 3.21±3.35 (m, 2H, CH2),
3.68 (s, 3H, CH3), 3.81 (t, 2H, J=6.8 Hz, CH2), 6.12 (d,
1H, J=3 Hz, ArH), 6.13 (br, 1H, NH or OH), 6.43 (br,
1H, NH or OH), 6.72 (d, 1H, J=3 Hz, ArH), 7.33±7.57
(m, 5H, ArH), 8.67 (s, 1H, NH or OH); SIMS m/z 428
(M+H)+. Anal. calcd for C24H33N3O4: C 67.42, H
7.78, N 9.83. Found: C 67.52, H 7.95, N 9.64.
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-benzyl-1-
cyclohexylurea (23). IR (KBr) 3375, 2939, 1621 cmÀ1
;
1H NMR (d in CDCl3) 1.39 (s, 9H, CH3), 1.39±1.88 (m,
10H, CH2), 3.58 (s, 3H, CH3), 4.38 (m, 1H, CH), 4.52 (s,
2H, CH2), 5.57 (d, 1H, J=3 Hz, ArH), 6.14 (s, 1H, NH
or OH), 6.66 (d, 1H, J=3 Hz, ArH), 7.34±7.47 (m, 5H,
ArH), 8.90 (s, 1H, NH or OH); SIMS m/z 411 (M+H)+.
Anal. calcd for C25H34N2O3: C 73.14, H 8.35, N 6.82.
Found: C 73.37, H 8.42, N 6.82.
N,N-Diethyl-4-[3-(3-tert-butyl-2-hydroxy-5-methoxy-
phenyl)-1-phenylureido]-butyramide (12). IR (KBr) 2964,
1
1615 cmÀ1; H NMR (d in CDCl3) 1.06±1.26 (m, 6H,
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-cyclohep-
tyl-1-(4-N,N-dimethylaminophenyl)urea (24). IR (KBr)
CH3), 1.41 (s, 9H, CH3), 1.93±2.07 (m, 2H, CH2), 2.38 (t,
2H, J=6.8 Hz, CH2), 3.24±3.42 (m, 4H, CH2), 3.70 (s,
3H, CH3), 3.75±3.82 (m, 2H, CH2), 6.32 (d, 1H, J=3 Hz,
ArH), 6.71 (d, 1H, J=3 Hz, ArH), 7.33±7.51 (m, 6H,
ArH, NH or OH), 8.85 (s, 1H, NH or OH); SIMS m/z
456 (M+H)+. Anal. calcd for C26H37N3O4: C 68.54, H
8.19, N 9.22. Found: C 68.51, H 8.30, N 9.31.
1
3397, 2927, 2430, 1657 cmÀ1; H NMR (d in CDCl3)
1.39 (s, 9H, CH3), 1.39±2.01 (m, 12H, CH2), 3.18 (s, 6H,
CH3), 3.68 (s, 3H, CH3), 4.53 (m, 1H, CH), 6.01 (s, 1H,
NH or OH), 6.25 (d, 1H, J=2.4 Hz, ArH), 6.68 (d, 1H,
J=2.4Hz, ArH), 7.47 (d, 2H, J=8 Hz, ArH), 7.87 (d, 2H,
J=8Hz, ArH); SIMS m/z 454 (M+H)+. Anal. calcd for
.
.
C27H39N3O3 HCl C2H5OH: C 64.97, H 8.65, N 7.84, Cl
6.61. Found: C 64.76, H 8.66, N 7.90, Cl 6.94.
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-(3-mor-
pholinocarbonylpropyl)-1-phenylurea (13). IR (KBr)
1
2946, 1642 cmÀ1; H NMR (d in CDCl3) 1.41 (s, 9H,
3-(3-tert-Butyl-2-hydroxy-5-methoxyphenyl)-1-benzyl-
1-(4-N,N-dimethylaminophenyl)urea (25). IR (KBr)
3385, 3059, 2376, 1607 cmÀ1; 1H NMR (d in DMSO-d6)
1.31 (s, 9H, CH3), 3.02 (s, 6H, CH3), 3.65 (s, 3H,
CH3), 4.93 (s, 2H, CH2), 6.48 (d, 1H, J=3 Hz, ArH),
7.01 (d, 1H, J=3 Hz, ArH), 7.19±7.34 (m, 9H, ArH),
7.67 (s, 1H, NH or OH); EIMS m/z 447 (M+).
CH3), 1.94±2.04 (m, 2H, CH2), 2.40 (t, 2H, J=7 Hz,
CH2), 3.43±3.47 (m, 2H, CH2), 3.62±3.68 (m, 6H, CH2),
3.68 (s, 3H, CH3), 3.81 (d, 2H, J=7 Hz, CH2), 6.17 (d,
1H, J=3 Hz, ArH), 6.71 (d, 1H, J=3 Hz, ArH), 6.72
(br, 1H, NH or OH), 7.34±7.54 (m, 5H, ArH), 8.77 (s,
1H, NH or OH); SIMS m/z 470 (M+H)+. Anal. calcd
for C26H35N3O5: C 66.50, H 7.51, N 8.95. Found: C
66.80, H 7.69, N 9.18.
.
Anal. calcd for C27H33 N3O3 HCl: C 67.00, H 7.08,

K. Nakao et al. / Bioorg. Med. Chem. 9 (2001) 853±861
861
N-(2,3-Dihydroxypropyl)-4-[3-(3-tert-butyl-2-hydroxy-5-
methoxyphenyl)-1-phenylureido]-butyramide (14). IR
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14. Yamagishi, M.; Yamada, Y.; Ozaki, K.; Asao, M.; Shimizu,
R.; Suzuki, M.; Matsumoto, M.; Matsuoka, Y.; Matsumoto, K.
J. Med. Chem. 1992, 35, 2085.
1
(KBr) 3322, 2952, 1638 cmÀ1; H NMR (d in CDCl3)
1.40 (s, 9H, CH3), 1.67 (br, 2H, OH), 1.86±1.93 (m, 2H,
CH2), 2.35 (t, 2H, J=6.8 Hz, CH2), 3.03 (m, 1H, CH),
3.35±3.41 (m, 2H, CH2), 3.51±3.53 (m, 2H, CH2), 3.68
(s, 3H, CH3), 3.81 (t, 2H, J=6.8 Hz, CH2), 6.18 (d, 1H,
J=3 Hz, ArH), 6.30 (s, 1H, NH or OH), 6.70 (d, 1H,
J=3 Hz, ArH), 6.72 (br, 1H, NH or OH), 7.35±7.58 (m,
5H, ArH), 8.25 (s, 1H, NH or OH); SIMS m/z 474
(M+H)+. Anal. calcd for C25H35N3O6: C 63.41, H
7.45, N 8.87. Found: C 63.39, H 7.58, N 9.01.
Acknowledgements
15. Watanabe, H.; Miyamoto, S.; Yoshimoto, M.; Kamioka,
T.; Nakayama, I.; Kobayashi, T.; Honda, T. Chem. Pharm.
Bull. 1987, 35, 1452.
The authors are grateful to Dr. Kimiaki Hayashi for
useful suggestions and comments. They with to thank
Katsuya Fujikawa, Teruhito Nakamine, Kiyoshi Saitoh
and Tomoko Wada for their skillfull experiments.
16. Leo, A. J. Chem. Rev. 1993, 93, 1281.
17. CLOGP for Windows, version 2.0.0, BioByte Corp. 201
W. Fourth St., Suite #204, Claremont, CA 91711 U. S. A.
18. DeVries, V. G.; Bloom, J. D.; Dutia, M. D.; Katocs, A. S.,
Jr.; Largis, E. E. J. Med. Chem. 1989, 32, 2318.
19. Yasuhara, M.; Saito, K.; Kubota, H.; Ohmizu, H.;
Suzuki, T. Biol. Pharm. Bull. 1997, 20, 1056.
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