Design and synthesis of 4-O-methylhonokiol analogs as inhibitors of cyclooxygenase-2 (COX-2) and PGF1 production

Article abstract of DOI:10.1016/j.bmc.2012.03.028

A series of novel 4-O-methylhonokiol analogs were synthesized in light of revealing structure-activity relationship for inhibitory effect of COX-2 enzyme. The key strategy of the molecular design was oriented towards modification of the potential metabolic soft spots (e.g., phenol and olefin) or by altering the polar surface area via incorporating heterocycles such as isoxazole and triazole. Most of all exhibited the inhibitory effects on COX-2 and PGF ₁ production but not macrophage NO production. Especially, aryl carbamates 10 and 11 exhibited more potent inhibitory activity against COX-2 and PGF₁ production.

Full text of DOI:10.1016/j.bmc.2012.03.028

Bioorganic & Medicinal Chemistry 20 (2012) 2860–2868
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Design and synthesis of 4-O-methylhonokiol analogs as inhibitors
of cyclooxygenase-2 (COX-2) and PGF1 production
Bit Lee a, , Jae-Hwan Kwak b, , Shin-Won Huang a, Jae-Yong Jang b, Sanglae Lim a, Young-Shin Kwak c,
,
⇑
Kiho Lee c,d, Hyung Sook Kim b, Sang-Bae Han b, Jin-Tae Hong b, Heesoon Lee b, Sukgil Song b,
b,
Seung-Yong Seo a, , Jae-Kyung Jung
⇑
⇑
a College of Pharmacy, Woosuk University, Wanju, Jeonbuk 565-701, South Korea
b College of Pharmacy, and Medicinal Research Center (MRC), Chungbuk National University, Cheongju 361-763, South Korea
c Korea Research Institute of Bioscience and Biotechnology, Ochang 363-883, South Korea
d College of Pharmacy, Korea University, Chochiwon 339-700, South Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of novel 4-O-methylhonokiol analogs were synthesized in light of revealing structure–activity
relationship for inhibitory effect of COX-2 enzyme. The key strategy of the molecular design was oriented
towards modification of the potential metabolic soft spots (e.g., phenol and olefin) or by altering the polar
surface area via incorporating heterocycles such as isoxazole and triazole. Most of all exhibited the inhib-
itory effects on COX-2 and PGF1 production but not macrophage NO production. Especially, aryl carba-
mates 10 and 11 exhibited more potent inhibitory activity against COX-2 and PGF1 production.
Crown Copyright Ó 2012 Published by Elsevier Ltd. All rights reserved.
Received 3 February 2012
Revised 9 March 2012
Accepted 10 March 2012
Available online 24 March 2012
This paper is dedicated to Professor
Young-Ger Suh on the occasion of his 60th
birthday
Keywords:
4-O-Methylhonokiol
Cyclooxygenase-2 (COX-2)
Aryl carbamate
Isoxazole
Triazole
1. Introduction
Several COX-2 selective inhibitors such as celecoxib have shown
excellent efficacy in humans with few side effects.2 In spite of this
Nonsteroidal anti-inflammatory drugs (NSAIDs) exhibit their
anti-inflammatory effect by inhibiting cyclooxygenases (COX)
which catalyzes initially in arachidonic acid metabolic cascade.1
There are two human enzymes, COX-1 and COX-2, which catalyze
the same biochemical reaction but they are clearly different in
terms of amino acids sequence, tissue distribution, and physiolog-
ical function. The chronic use of NSAIDs to treat pain and
inflammation is often accompanied by side effects such as gastro-
intestinal (GI) toxicities, bleeding, and suppressed renal function.
These common side effects of NSAIDs are ascribed to their ability
of inhibiting COX-1 and thus developing COX-2-selective inhibitors
is the logical strategy for designing NSAIDS out the adverse events.
initial success after the launch of selective COX-2 inhibitors, con-
cerns were raised regarding the adverse thrombotic cardiovascular
events. There has been a continuous progress made in the develop-
ment of novel anti-inflammatory agents having reduced GI and
cardiovascular risks.
The bark of the root and stem of Magnolia family has been used
in traditional medicine to treat various diseases.3,4 Among the bio-
active compounds isolated from Magnolia family, biphenyl-
neolignans such as magnolol (1), honokiol (2), 4-O-methylhonokiol
(3) and obovatol (4) have been reported to have anti-inflammatory,
anti-allergic, anti-bacterial and anti-depressant activities (Fig. 1).5
4-O-Methylhonokiol was isolated mainly from the bark of the root
and stem of Magnolia species and exhibits anti-inflammatory, anti-
oxidative and neurotropic activity. In a recent report, 4-O-methyl-
honokiol was capable of decreasing b-amyloid-induced memory
impairment by reduction of oxidative damages in Ab1-42-infused
Alzheimer’s disease mouse model.5a,f,g Moreover, 4-O-methylhon-
okiol was shown to have the higher anti-inflammatory activity
⇑
Corresponding authors. Tel.: +82 43 240 6017 (Y.-S.K.); tel.: +82 63 290 1566;
fax: +82 63 290 1812 (S.-Y.S.); tel.: +82 43 261 2635; fax: +82 43 268 2732 (J.-K.J.).
E-mail addresses: yskwak@kribb.re.kr (Y.-S. Kwak), syseo@woosuk.ac.kr
(S.-Y. Seo), orgjkjung@chungbuk.ac.kr (J.-K. Jung).
These authors have contributed equally to the preparation of this manuscript.
0968-0896/$ - see front matter Crown Copyright Ó 2012 Published by Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2012.03.028

B. Lee et al. / Bioorg. Med. Chem. 20 (2012) 2860–2868
2861
OH
OH
OMe
R
OR
OH
O
R-Br
K2CO3, DMF
OH
HO
O
5
6
(R = Me)
(R = iPr)
7 (R = prenyl)
3
O
Ac2O, DMAP
1
2
4
Obovatol ( )
Magnolol ( )
Honokiol ( , R = H)
OMe
Et3N, CH2Cl2
(for 8)
3
R
O
4-O-Methylhonokiol ( , R = Me)
or
8
9
(R = Me)
Figure 1. The structure of representative natural neolignans from Magnolia family.
Triphosgene
pyridine, CH2Cl2
R-NH2, (for
(R = NHCH2CH=CH2)
10 (R = NHCH2C6H5)
(R = N(CH3)CH2C6H5)
11
9 11
)
~
(e.g., IC50 of 0.06 lM for COX-2) than honokiol (e.g., IC50 of 2.1 lM
for COX-2) and a variety of honokiol analogs.6 As 4-O-methylhon-
okiol is the attractive research candidate for the drug discovery
against Alzheimer’s disease and other inflammatory conditions
such as rheumatoid arthritis (RA) and osteoarthritis (OA), we re-
cently reported the efficient synthesis of 4-O-methylhonokiol.7
Now this article deals with the structure–activity relationship
(SAR) study aiming at identifying novel COX-2 selective 4-methyl-
honokiol analogs via modifying the potential metabolic soft spots8
and increasing the polar surface area for the future assessment of
structure–property relationship.
Scheme 2. Synthesis of alkyl ethers, acetate and aryl carbamates analogs.
followed by the addition of allylamine, benzylamine, and N-ben-
zymethylamine to give the corresponding aryl carbamates,
respectively.10
Chloride 12 and bromide 13 were obtained by the ortho-haloge-
nation of phenol in which DCDMH (1,3-dichloro-5,5-dimethyl-
hydantoin) and DBDMH (1,3-dibromo-5,5-dimethyl-hydantoin)
were used to serve as chloride and bromide source after the phen-
oxide of 4-O-methylhonokiol was generated with iPrMgCl
(Scheme 3).11 Two double bonds of 3 could be modified in terms
of increasing the biological activity as well as the metabolic stabil-
ity and/or the proper polarity (Scheme 3, compounds 14–17). Thus,
compounds 14 and 15 were obtained by hydrogenation and m-
CPBA epoxidation, respectively. For the synthesis of alcohol 16
we performed hydroboration and subsequent oxidation to give
two regioisomers. For diprenyl compound 17, two allyl groups
were smoothly converted to prenyl groups by cross metathesis
using 2nd generation Grubbs catalyst and 2-methyl-2-butene in
good yield.12
We chose five-member heterocyclic ring, especially isoxazole
and triazole in replace of a phenyl ring of 4-O-methylhonokiol to
be endowed with higher solubility and less reactivity of phenol
in A ring through hydrogen bond with nitrogen of azole ring. In this
connection, isoxazoles were synthesized from commercially avail-
able 5-bromo-2-methoxy-benzaldehyde 18 which was treated
with hydroxylamine to give oxime 19 (Scheme 4). Isoxazoles 20a
and 20b were prepared by cyclization of either 1-pentyne or
2. Result and discussion
4-O-Methylhonokiol consists of a 5,30-diallyl-biphenyl skeleton
bearing hydroxy group in C2 of A ring and methoxy group in C4 of
B ring. The major route of metabolism in vivo for both 2 and 3 is via
glucuronidation of the phenol group.8 Thus we were interested in
modification of the phenol group on A ring as well as the introduc-
tion of electron-withdrawing substituents at ortho-position of phe-
nol group, which could reduce phase II metabolism (Scheme 1).
Both of allyl groups would be replaced with appropriate surrogates
in order to slow metabolic degradation while retaining potency at
the same time. In addition, hydrogen bond of the phenol group on
A ring with neighboring moiety was expected to lower a metabolic
change. Consideration of other potential substituents that would fit
the structural criteria required for biological activity led to the pos-
sibility of employing azole moieties, such as isoxazole and triazole.
Our synthesis of 4-O-methylhonokiol analogs commenced with
the modification in three different ways; (1) the alkylation and
acylation of phenol group in A ring, (2) ortho-halogenation of phe-
nol group, and (3) olefin modification for the synthesis of more
polar compounds. Thus alkyl phenyl ether (5–7) was obtained by
treatment of the alkyl halide and potassium carbonate in DMF in
good yield (Scheme 2).9 Acetate 8 was prepared by acetic anhy-
dride, Et3N and DMAP.9 For the synthesis of aryl carbamates
9–11, 3 was treated with triphosgene and pyridine in CH2Cl2,
OMe
Br
OMe
OMe
OH
OH
OH
Cl
12
DCDMH
iPrMgCl, THF
-78 o
13
14
DBDMH
H2, Pd/C
MeOH
iPrMgCl, THF
-78 o
Phenol substituents
ortho-Position
modification
C
C
OMe
OH
R
3
4-O-methylhonokiol ( )
3
modification of
4-O-methylhonikiol
2nd Grubbs
cat.
mCPBA
NaHCO3
CH2Cl2
BH3-THF
then H2O2,
NaOH
Olefin modification
4'
3'
OMe
OH
B
2
OMe
OMe
OH
OMe
H
A
X
OH
OH
N
O
Y
5
Introduction of
heterocycles
Azole analogs
X = O, Y = C: isoxazole
X, Y = N: triazole
4-O-Methylhonokiol (3)
O
OH
15
16
17
O
Scheme 1. Structure of 4-O-methylhonokiol (3) and schematic design of its
analogs.
Scheme 3. Ortho-Halogenation of phenol group and olefin modification.

2862
B. Lee et al. / Bioorg. Med. Chem. 20 (2012) 2860–2868
Table 1
O
N
OMe
OMe
Br
OMe
Inhibitory activity of COX-2 enzyme and PGF1 production in macrophages by 100 nM
of the alkylated derivatives (5–7)
R
CHO
OH
N
NH2OH
NCS
R1
OMe
then Et3N,
R
O
Br
Br
18
19
20a (R = CH2CH2CH3)
20b
(R = CH2OH)
O
O
N
N
OMe
OH
Pd(PPh3)4
BCl3
R
R
R
COX-2 enzyme (%)
PGF1 production (%)
CH2Cl2
SnBu3
toluene
Celecoxib
—
H
60
57
46
23
44
75
42
48
24
19
-78 o
C
21a,b
22a,b
3
5
6
7
CH3
CH(CH3)2
CH2CH@C(CH3)2
O
N
OH
Br
BCl3
20a
CH2Cl2
-78 oC
Table 2
23
Inhibitory activity of COX-2 enzyme and PGF1 production in macrophages by 100 nM
of the acylated derivatives (8–11)
Scheme 4. Synthesis of isoxazole analogs of 4-O-methylhonokiol.
O
OMe
R
O
propargyl alcohol with oxime 19 in the presence of NCS and Et3N,
respectively.13 Stille cross-coupling of compounds 20a and 20b
with allyltributyltin and Pd(PPh3)4, followed by demethylation
with BCl3 provided isoxazole analogs 22a and 22b. In a similar
manner, 20a was treated with BCl3 to give the resulting phenol 23.
For the synthesis of 1,2,3-triazole analogs, one-pot method of
Cu(I)-catalyzed azide–alkyne 1,3-dipolar cycloaddition was uti-
lized14 and thus 2-ethynylphenol (for A ring) and alkyl azide (for
B ring) are required to access diverse 1,2,3-triazoles compounds
(Scheme 5). 4-Propylphenol 24 could readily be converted into 2-
ethynyl-4-propylphenyl acetate 26 by ortho-bromination and acet-
ylation of phenol, followed by Sonogashira coupling reaction with
ethynyltrimethylsilane, Pd(PPh3)4, and CuI. The formation of 1,2,3-
triazole from compound 26 was accomplished by using CuSO4, so-
dium ascorbate and an alkyl azide which was prepared in situ by
the treatment of either allyl bromide or (bromomethyl)cyclobu-
tane with NaN3. Finally, the acetates 27a and 27b were hydrolyzed
to afford the resulting 1,2,3-triazole compounds 28a and 28b,
respectively.
R
COX-2 enzyme (%)
PGF1 production (%)
8
9
10
11
CH3
39
48
65
63
45
41
63
41
NHCH2CH@CH2
NHCH2C6H5
N(CH3)CH2C6H5
Table 3
Inhibitory activity of COX-2 enzyme and PGF1 production in macrophages by 100 nM
of the halogenated derivatives (3–7)
OMe
OH
R
The in vitro inhibitory activities of cyclooxygenase-2 (COX-2)
enzyme and PGF1 production were listed in Tables 1–5. Celecoxib
and 4-O-methylhonokiol (3) were used as reference compounds.
Inhibition of COX-2 enzyme by 100 nM of synthesized compounds
was evaluated by using COX inhibitor screening kit. All of com-
pounds showed the direct inhibition of COX-2 enzyme with the
range of 22–65% inhibition at 100 nM treatment. Especially, mod-
R
COX-2 enzyme (%)
PGF1 production (%)
12
13
Cl
Br
NDa
52
52
54
a
ND, not determined
Table 4
Inhibitory activity of COX-2 enzyme and PGF1 production in macrophages by 100 nM
of the olefin-modified derivatives (14–17)
OH
OAc
OAc
1. Pd(PPh3)4
CuI, Et3N
OMe
OH
Br
1. Br2, CHCl3
2. Ac2O
TMS
R2
2. CsCO3
24
25
26
R1
N
N
N
R1 and R2
1 = CH2CH3
R2 = CH2CH3
R1 = oxiranyl
COX-2 enzyme (%)
46
PGF1 production (%)
55
OAc
N
OH
R
Br
N
N
R
R
NaN3, CuSO4
K2CO3
MeOH
14
15
16
17
R
sod. ascorbate
DMSO / H2O
58
ND
41
51
61
42
R
2 = oxiranyl
R1 = CH@CH2
27a
28a
(R = CH2CH=CH2)
(R = CH2CH=CH2)
R2 = CH2CH2OH
R1 = CH@C(CH3)2
R2 = CH@C(CH3)2
27b (R =
)
28b (R =
)
a ND, not determined
Scheme 5. Synthesis of 1,2,3-triazole analogs 4-O-methylhonokiol.

B. Lee et al. / Bioorg. Med. Chem. 20 (2012) 2860–2868
2863
Table 5
the IC50 value of 10–100
l
M, while they have the cytotoxicity at
Inhibitory activity of COX-2 enzyme and PGF1 production in macrophages by 100 nM
of the isoxazole and triazole analogs
the similar concentration (Table 6). Thus, it was noted that 4-O-
methylhonokiol analogs were not able to inhibit NO production
selectively without cytotoxicity in macrophage. In terms of struc-
ture–cytotoxicity relationship, 4-O-methylhonokiol analogs having
the free phenol group showed more cytotoxicity than alkylated or
acylated compounds.
O
N
N
OH
N
OH
N
R1
R3
R2
R1/R2 or R3
1 = CH2CH2CH3
R2 = CH2CH@CH2
R1 = CH2OH
COX-2 enzyme (%)
39
PGF1 production (%)
54
3. Materials and methods
3.1. Synthetic procedures
22a
22b
23
R
38
35
49
50
R
2 = CH2CH@CH2
R1 = CH2CH2CH3
R2 = Br
All starting materials and reagents were obtained from com-
mercial suppliers and were used without further purification. Air
and moisture sensitive reactions were performed under an argon
atmosphere. Flash column chromatography was performed using
Silica Gel 60 (230–400 mesh, Merck) with the indicated solvents.
Thin-layer chromatography was performed using 0.25 mm silica
gel plates (Merck). 1H and 13C NMR spectra were recorded on
either a Bruker AVANCE III 400 MHz, Bruker AVANCE 500 MHz,
or a JEOL JNM-EX400 (400 MHz) spectrometer as solutions in deu-
teriochloroform (CDCl3). 1H NMR data were reported in the order
of chemical shift, multiplicity (s, singlet; d, doublet; t, triplet; m,
multiplet and/or multiple resonances), number of protons, and
coupling constant (J) in hertz (Hz).
28a
28b
R3 = CH2CH@CH2
22
33
29
43
R
3 = cyclobutylmethyl
ifications of 4-O-methylhonokiol by introducing methyl, isopropyl
and prenyl group at phenol slightly decrease the inhibiting activity
of COX-2 enzyme and PGF1 production (Table 1). Both of 10 and 11
which contain a benzyl carbamate group in A ring showed slightly
more potent inhibitory activity (65% and 63% inhibition, respec-
tively) than celecoxib (60%) and 4-O-methylhonokiol (57%) (Table
2). When RAW 264.7 cells were treated with LPS in the presence
of 100 nM analogs for 24 h and the concentrations of PGF1 were
measured, all of compounds showed the inhibitory activities with
the range of 19–63% inhibition. There is no marked difference be-
tween COX-2 and PGF1 production inhibition.
Replacing phenyl ring (B ring) with azole rings as a hydrogen
bond acceptor to the potential metabolic soft spots phenol group
(A ring) had the similar activity to 4-O-methyl-honokiol (Table
4). As the results, the identification of lead compounds for new
anti-inflammatory agents from the current synthesized analogs
would be achieved through in vitro/in vivo DMPK (drug metabo-
lism and pharmacokinetic) studies.
3.1.1. 30,5-Diallyl-2,40-dimethoxy-1,10-biphenyl (5)
To a DMF solution of 4-O-methylhonokiol (50 mg, 0.18 mmol)
and potassium carbonate (73 mg, 0.53 mmol) was added at ambi-
ent temperature. After stirring for 30 min, methyl iodide (75 mg,
0.53 mmol) was added. After stirring for 3 h, the reaction mixture
was diluted with EtOAc and washed with 1 N HCl and brine, dried
over MgSO4 and concentrated under reduced pressure. The residue
was purified by flash column chromatography on silica gel (ethyl
acetate/hexanes = 1:8) to afford the methyl ether 5 (51 mg, 98%);
1H NMR (CDCl3, 400 MHz) d 7.37 (dd, 1H, J = 8.3, 2.2 Hz), 7.31
(ds, 1H, J = 2.2 Hz), 7.12 (ds, 1H, J = 2.1 Hz), 7.10 (dd, 1H, J = 8.2,
2.3 Hz), 6.90 (d, 2H, J = 8.3 Hz), 6.08–5.93 (m, 2H), 5.12–5.02 (m,
4H), 3.86 (s, 3H), 3.79 (s, 3H), 3.43 (d, 2H, J = 6.6 Hz), 3.37 (d, 2H,
J = 6.7 Hz); the compound 5 was reported. See Ref. 7.
The in vitro inhibitory activities of nitric oxide (NO) production
and the cell viability were listed in Table 6. The synthesized ana-
logs were evaluated for their ability to inhibit LPS-activated NO
production by RAW264.7 macrophage cell line. Cell viability was
examined by the propidium iodide (PI) nuclear staining method.
Most of all show the inhibition of macrophage NO production at
3.1.2. 30,5-Diallyl-2-isopropoxy-40-methoxy-1,10-biphenyl (6)
To a DMF solution of 4-O-methylhonokiol (30 mg, 0.11 mmol)
potassium carbonate (44 mg, 0.32 mmol) was added at ambient
temperature. After stirring for 30 min, 2-bromopropane (220 mg,
0.16 mmol) was added. After stirring for 4 h, the reaction mixture
was diluted with EtOAc and washed with 1 N HCl and brine, dried
over MgSO4 and concentrated under reduced pressure. The residue
was purified by flash column chromatography on silica gel (ethyl
acetate/hexanes = 1:25) to afford the isopropyl ether product 6
Table 6
Inhibitory activity of compounds on macrophage NO production (IC50, lM) and the
cell viability (IC50
, lM)
Compounds
NO production
Viability
Celecoxib
3
5
6
7
8
9
10
11
12
13
14
15
16
17
22a
22b
23
28a
28b
12,090
18
40
78
>100
24
>100
>100
33
40
28
11
24
41
13
61
36
63
>50
33
11,130
27
>100
>100
>100
29
>100
>100
44
81
33
13
64
(21 mg, 60%).; 1H NMR (400 MHz, CDCl3)
d 7.41 (d, 1H,
J = 2.3 Hz), 7.38 (dd, 1H, J = 8.3 and 2.3 Hz), 7.14 (ds, 1H,
J = 2.3 Hz), 7.05 (dd, 1H, J = 8.3 and 2.3 Hz), 6.90 (d, 1H,
J = 8.3 Hz), 6.88 (d, 1H, J = 8.3 Hz), 6.10–5.93 (m, 2H), 5.13–5.03
(m, 4H), 4.45–4.34 (m, 1H), 3.87 (s, 3H), 3.43 (d, 2H, J = 6.7 Hz),
3.37 (d, 2H, J = 6.7 Hz), 1.24 (d, 6H, J = 6.1 Hz); 13C NMR
(100 MHz, CDCl3) d 156.3, 153.3, 138.0, 137.2, 132.7, 131.9,
131.4, 131.3, 131.2, 128.3, 127.9, 127.8, 115.8, 115.6, 115.5,
110.0, 71.1, 55.6, 39.6, 34.4, 22.3; IR (neat): 2975, 1638, 1488,
1242, 811 cmꢀ1; LRMS (ESI) m/z 323.1 (M+H+).
53
15
81
>50
63
3.1.3. 30,5-Diallyl-40-methoxy-2-((3-methylbut-2-en-1-yl)oxy)-
1,10-biphenyl (7)
To a DMF solution of 4-O-methylhonokiol (56 mg, 0.20 mmol)
potassium carbonate (55 mg, 0.40 mmol) was added at ambient
>50
>50

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B. Lee et al. / Bioorg. Med. Chem. 20 (2012) 2860–2868
temperature. After stirring for 30 min, 3,3-dimethylallyl bromide
(47 mg, 0.32 mmol) was added. After stirring for 5 h, the reaction
mixture was diluted with EtOAc and washed with 1 N HCl and
brine, dried over MgSO4 and concentrated under reduced pressure.
The residue was purified by flash column chromatography on silica
gel (ethyl acetate/hexanes = 1:10) to afford the 3,3-dimethylallyl
ether 7 (60 mg, 86%); 1H NMR (400 MHz, CDCl3) d 7.40 (m, 2H),
7.15 (d, 1H, J = 2.4 Hz), 7.09 (dd, 1H, J = 8.3 and 2.4 Hz), 6.93 (m,
2H), 6.05 (m, 2H), 5.45 (t, 1H, J = 2.3 Hz), 5.08 (m, 4H), 4.50 (d,
2H, J = 2.3 Hz), 3.87 (s, 3H), 3.44 (d, 2H, J = 6.3), 3.39 (d, 2H,
J = 6.8), 1.81 (s, 3H), 1.75 (s, 3H); 13C NMR (100 MHz, CDCl3) d
156.2, 154.1, 137.8, 137.0, 136.6, 132.3, 131.1 130.88, 130.84,
130.7, 128.1, 127.77, 127.72, 120.3, 115.4, 115.2, 113.3, 109.8,
65.7, 55.4, 39.4, 34.2, 25.6, 18.1.
154.7, 146.1, 138.1, 137.7, 137.1, 136.9, 134.6, 130.7, 130.4, 129.9,
128.5, 128.2, 127.9, 127.8, 127.5, 127.4, 123.0, 116.0, 115.3, 110.1,
55.3, 45.1, 39.6, 34.2. IR (KBr) 3629, 3346, 2931, 1710, 1529, 1490,
1261, 1219 cmꢀ1; LRMS (ESI) m/z 436.1 (M+Na+).
3.1.7. 30,5-Diallyl-40-methoxy-[1,10-biphenyl]-2-yl
benzyl(methyl)carbamate (11)
To a CH2Cl2 solution (1 mL) of 4-O-methylhonokiol (42 mg,
0.15 mmol) and pyridine (80 mg, 1 mmol) was added triphosgene
(89 mg, 0.3 mmol) at 0 °C. After stirring for 2 h at ambient temper-
ature, N-benzylmethylamine (36 mg, 0.3 mmol) was added to the
reaction mixture. After stirring for 12 h at ambient temperature,
the reaction mixture was diluted with CH2Cl2 and washed with
aqueous NH4Cl solution and water, dried over MgSO4 and concen-
trated under reduced pressure. The residue was purified by flash
column chromatography on silica gel (ethyl acetate/hexanes = 1:3)
to afford the benzylmethylcarbamate 11 (38 mg, 59%). 1H NMR
(400 MHz, CDCl3) d 7.23 (m, 5H), 7.13 (m, 3H), 7.00 (m, 2H), 6.80
(m, 1H), 5.99 (m, 2H), 5.09 (m, 4H), 4.41 and 4.35 (S6, 2H), (S6,
2H), 3.82 (S, 3H), 3.38 (m, 4H), 2.79 (s, 3H); 13C NMR (100 MHz,
CDCl3) d 156.5, 155.0, 146.7, 137.5, 137.3, 137.2, 137.0, 136.8,
136.7, 130.8, 130.7, 130.4, 130.1, 128.4, 128.0, 127.9, 127.6,
127.4, 127.3, 123.0, 115.9, 115.4, 109.9, 55.3, 52.6, 39.5, 34.2,
33.8; IR (neat) 3341, 3074, 2911, 2836, 1719, 1504, 1484, 1245,
1200 cmꢀ1; LRMS (ESI) m/z 450.2 (M+Na+).
3.1.4. 30,5-Diallyl-40-methoxy-[1,10-biphenyl]-2-yl acetate (8)
To a CH2Cl2 solution of 4-O-methylhonokiol (47 mg, 0.17 mmol)
4-dimethylaminopyridine (5 mg, 34 lmol), triethylamine (86 mg,
0.85 mmol), and acetic anhydride (20 mg, 0.20 mmol) was added
at ambient temperature. After stirring for 20 min, the reaction mix-
ture was treated with sodium bicarbonate, diluted with ethyl ace-
tate and dried over MgSO4 and concentrated under reduced
pressure. The residue was purified by flash column chromatogra-
phy (ethyl acetate/hexanes = 1:10) to afford the acetate (54 mg,
93%); 1H NMR (CDCl3, 400 MHz) d 7.24 (dd, 1H, J = 8.3, 2.2 Hz),
7.21 (ds, 1H, J = 2.2 Hz), 7.20 (ds, 1H, J = 2.1 Hz), 7.14 (dd, 1H,
J = 8.2, 2.2 Hz), 7.02 (d, 1H, J = 8.2 Hz), 6.89 (d, 1H, J = 8.3 Hz),
6.06–5.93 (m, 2H), 5.14–5.02 (m, 4H), 3.86 (s, 3H), 3.42–3.39 (m,
4H), 2.09 (s, 3H); The compound 8 was reported. See Ref. 7.
3.1.8. 30,5-Diallyl-3-chloro-40-methoxy-[1,10-biphenyl]-2-ol (12)
To a THF solution of 4-O-methylhonokiol (48 mg, 0.17 mmol)
isopropyl magnesium chloride (2.0 M solution in diethyl ether,
0.20 mL, 0.10 mmol) was added at ꢀ78 °C dropwise. After stirring
for 30 min at ꢀ78 °C, the reaction mixture was heated to ambient
temperature for 10 min. DCDMH (27 mg, 0.14 mmol) was added to
the reaction mixture. After stirring for 2 h, the reaction mixture
was treated with aqueous NH4Cl, extracted with ethyl acetate
and dried over MgSO4 and concentrated under reduced pressure.
The residue was purified by flash column chromatography (ethyl
acetate/hexanes = 1:10) to afford the chloride 12 (17 mg, 32%);
1H NMR (400 MHz, CDCl3) d 7.37 (dd, 1H, J = 8.4 and 2.3 Hz), 7.31
(d, 1H, J = 2.3 Hz), 7.13 (d, 1H, J = 2.2 Hz), 7.01 (d, 1H, J = 2.2 Hz),
6.94 (d, 1H, J = 8.4 Hz), 6.08–5.89 (m, 2H), 5.57 (s, OH), 5.14–5.04
(m, 4H), 3.88 (s, 3H), 3.44 (d, 2H, J = 6.7 Hz), 3.33 (d, 2H,
J = 6.7 Hz); 13C NMR (100 MHz, CDCl3) d 157.1, 146.8, 137.1,
136.8, 132.9, 130.7, 129.5, 129.4, 129.3, 129.0, 128.2, 127.9,
120.5, 116.3, 115.8, 110.5, 55.7, 39.3, 34.4; IR (neat) 3511, 1470,
1244, 814, 675 cmꢀ1; LRMS (ESI) m/z 313.2 (M–H+)
3.1.5. 3’,5-Diallyl-40-methoxy-[1,10-biphenyl]-2-yl
allylcarbamate (9)
To a CH2Cl2 solution (1 mL) of 4-O-methylhonokiol (42 mg,
0.15 mmol) and pyridine (80 mg, 1 mmol) was added triphosgene
(89 mg, 0.3 mmol) at 0 °C. After stirring for 2 h at ambient temper-
ature, allylamine (20 mg, 0.35 mmol) was added to the reaction
mixture. After stirring for 12 h at ambient temperature, the reac-
tion mixture was diluted with CH2Cl2 and washed with aqueous
NH4Cl solution and water, dried over MgSO4 and concentrated un-
der reduced pressure. The residue was purified by flash column
chromatography on silica gel (ethyl acetate/hexanes = 1:6) to af-
ford the allylcarbamate 9 (35 mg, 65%). 1H NMR (400 MHz, CDCl3)
d 7.22 (m, 2H), 7.13 (S, 1H), 7.07 (m, 2H), 6.85 (d, 1H, J = 8.7 Hz),
6.01 (m, 2H), 5.77 (m, 1H), 5.09 (m, 6H), 4.87 (b, 1H), 3.81 (S,
3H), 3.74 (t, 2H, J = 5.8) 3.37 (d, 4H, J = 6.3); 13C NMR (100 MHz,
CDCl3) d 156.6, 154.5, 146.1, 137.6, 137.1, 137.0, 134.4, 134.0,
130.7, 130.5, 129.9, 128.2, 127.9, 127.8, 123.0, 116.1, 116.0,
115.3, 110.1, 55.4, 43.5, 39.6, 34.2; IR (neat) 2931, 1720, 1200 cm
ꢀ1; LRMS (ESI) m/z 386.1 (M+Na+).
3.1.9. 30,5-Diallyl-3-bromo-40-methoxy-[1,10-biphenyl]-2-ol (13)
To a THF solution of 4-O-methylhonokiol (50 mg, 0.18) isopro-
pyl magnesium chloride (2.0 M solution in diethyl ether, 0.11 mL,
0.22 mmol) was added at ꢀ78 °C dropwise. After stirring for
30 min at ꢀ78 °C, the reaction mixture was heated to ambient tem-
perature for 10 min. DBDMH (41 mg, 0.14 mmol) was added to the
reaction mixture. After stirring for 2 h, the reaction mixture was
treated with aqueous NH4Cl, extracted with ethyl acetate and dried
over MgSO4 and concentrated under reduced pressure. The residue
was purified by flash column chromatography (ethyl acetate/hex-
anes = 1:15) to afford the bromide 13 (42 mg, 66%); 1H NMR
(400 MHz, CDCl3) d 7.34 (dd, 1H, J = 8.4 and 2.3 Hz), 7.29 (d, 1H,
J = 2.3 Hz), 7.26 (d, 1H, J = 2.1 Hz), 7.03 (d, 1H, J = 2.1 Hz), 6.93 (d,
1H, J = 8.4 Hz), 6.07–5.88 (m, 2H), 5.55 (s, OH), 5.13–5.02 (m,
4H), 3.87 (s, 3H), 3.42 (d, 2H, J = 6.7 Hz), 3.32 (d, 2H, J = 6.7 Hz);
13C NMR (100 MHz, CDCl3) d 157.2, 147.7, 137.1, 136.8, 133.5,
131.0, 130.7, 130.3, 129.4, 129.3, 129.1, 128.2, 116.4, 115.8,
110.7, 110.5, 55.7, 39.2, 34.4; IR (neat) 3506, 2922, 1465, 1245,
815, 614 cmꢀ1; LRMS (ESI) m/z 357.1 (M–H+)
3.1.6. 30,5-Diallyl-40-methoxy-[1,10-biphenyl]-2-yl
benzylcarbamate (10)
To a CH2Cl2 solution (1 mL) of 4-O-methylhonokiol (42 mg,
0.15 mmol) and pyridine (80 mg, 1.0 mmol) was added triphos-
gene (89 mg, 0.30 mmol) at 0 °C. After stirring for 2 h at ambient
temperature, benzylamine (32 mg, 0.30 mmol) was added to the
reaction mixture. After stirring for 12 h at ambient temperature,
the reaction mixture was diluted with CH2Cl2 and washed with
aqueous NH4Cl solution and water, dried over MgSO4 and concen-
trated under reduced pressure. The residue was purified by flash
column chromatography on silica gel (ethyl acetate/hexanes = 1:4)
to afford the benzylcarbamate 10 (50 mg, 81%). 1H NMR (400 MHz,
CDCl3) d 7.30 (m, 6H), 7.13 (S, 1H), 7.11 (m, 4H), 6.80 (d, 1H, J = 8.2),
5.98 (m, 2H), 5.12 (m, 4H), 4.29 (d, 2H, J = 6.3), 3.81 (S, 3H), 3.37(d,
2H, J = 6.8), 3.33 (d, 2H, J = 6.3); 13C NMR (100 MHz, CDCl3) d 156.6,

B. Lee et al. / Bioorg. Med. Chem. 20 (2012) 2860–2868
2865
3.1.10. 40-Methoxy-30,5-dipropyl-[1,10-biphenyl]-2-ol (14)
A solution of 4-O-methylhonokiol (30 mg, 0.11 mmol) and 10%
Pd/C (10 mg) in anhydrous MeOH was placed under an atmosphere
of hydrogen. After stirring for 1 h, the reaction mixture was diluted
with EtOAc, filtered through Celite pad and concentrated under re-
duced pressure. The residue was purified by flash column chroma-
tography on silica gel (ethyl acetate/hexanes = 1:13) to afford the
hydrogenated compound 14 (30 mg, 98%) as a colorless oil.; 1H
NMR (500 MHz, CDCl3) d 7.27 (dd, 1H, J = 8.3 and 2.3 Hz), 7.23 (d,
1H, J = 2.2 Hz), 7.05–7.03 (m, 2H), 6.95 (d, 1H, J = 8.3 Hz), 6.89(d,
1H, J = 7.9 Hz), 5.16 (s, 1H), 3.87 (s, 3H), 2.64 (t, 2H, J = 7.5 Hz),
2.55 (t, 2H, J = 7.5 Hz), 1.68–1.61 (m, 4H), 0.99–0.94 (m, 6H); The
compound 14 was reported. See Ref. 7.
(764 mg, 11 mmol) was added. After the reaction mixture was re-
fluxed for 12 h, it was concentrated to a half of volume under re-
duced pressure. The mixture was poured to ice water and filtered
using glass filter to afford the resulting oxime as a yellow solid.
1H NMR (400 MHz, CDCl3) d 8.34 (s, 1H), 7.76 (d, 1H, J = 2.4 Hz),
7.55 (br s, 1H), 7.37 (dd, 1H, J = 2.4 and 8.8 Hz), 6.72 (d, 1H,
J = 8.8 Hz), 3.77 (s, 3H); 13C NMR (100 MHz, CD3OD) d 157.6,
144.5, 134.1, 129.1, 124.4, 114.1, 113.8, 56.3. IR (KBr) 3345,
1487, 1460, 1258, 968, 807 cmꢀ1; LRMS (ESI) m/z 229.9 (M+H+)
3.1.15. 3-(5-Bromo-2-methoxyphenyl)-5-propylisoxazole (20a)
To a CHCl3 (1 mL) solution of the crude aldoxime 19 (92 mg,
0.4 mmol) NCS (58 mg, 0.44 mmol) was added at 0 °C. After stirring
for 3 h, pentyne (40 mg, 0.6 mmol) and triethylamine (0.11 mL,
0.8 mmol) was added to the solution and heated to 50 °C. After stir-
ring for 12 h, the reaction mixture was diluted with ethyl acetate
and the organic phase was washed with water and brine, dried
over MgSO4 and concentrated under reduced pressure. The residue
was purified by flash column chromatography on silica gel (ethyl
acetate/hexanes = 1:10) to afford the isoxazole 20a (75 mg, 63%).
1H NMR (400 MHz, CDCl3) d 7.97 (d, 1H, J = 2.4 Hz), 7.44 (dd, 1H,
J = 8.8 and 2.9 Hz), 8.83 (d, 1H, J = 8.8 Hz), 6.43 (s, 1H), 3.84 (s,
3H), 2.75 (t, 2H, J = 7.8 Hz), 1.79 (m, 2H), 1.00 (t, 3H, J = 7.8 Hz);
13C NMR (100 MHz, CDCl3) d 173.1, 158.7, 156.2, 133.3, 131.8,
120.2, 113.1, 112.9, 102.0, 55.8, 28.6, 20.9, 13.6; IR (neat) 3742,
3680, 3621, 2371, 2317, 1741, 1691, 1514 cmꢀ1; LRMS (ESI) m/z
318.0 (M+Na+).
3.1.11. 40-Methoxy-30,5-bis(oxiran-2-ylmethyl)-[1,10-biphenyl]-
2-ol (15)
To
a
CH2Cl2 solution of 4-O-methylhonokiol (51 mg,
0.18 mmol), m-chloroperoxybenzoic acid (94 mg, 0.54 mmol) and
NaHCO3 (76 mg, 0.90 mmol) were added at ꢀ78 °C dropwise. After
stirring for 1 day at ambient temperature, the reaction mixture
was diluted with water, extracted with ethyl acetate, dried over
MgSO4 and concentrated under reduced pressure. The residue
was purified by flash column chromatography (ethyl acetate/hex-
anes = 1:2) to afford the epoxide 15 (30 mg, 53%); 1H NMR
(500 MHz, CDCl3) d 7.33 (dd, 1H, J = 8.3 and 2.2 Hz), 7.30 (d, 1H,
J = 2.2 Hz), 7.12–7.09 (m, 2H), 6.97 (d, 1H, J = 8.4 Hz), 6.91 (d, 1H,
J = 8.0 Hz); The compound 15 was reported. See Ref. 7.
3.1.12. 5-Allyl-30-(3-hydroxypropyl)-40-methoxy-[1,10-
biphenyl]-2-ol (16)
3.1.16. (3-(5-Bromo-2-methoxyphenyl)isoxazol-5-yl)methanol
(20b)
To an anhydrous THF solution of BH3–THF (1.0 M solution in
THF, 0.68 mL, 0.68 mmol) a THF solution of 4-O-methylhonokiol
(190 mg, 0.68 mmol) was added during 5 min at 0 °C. After stirring
for 2 h at ambient temperature, the reaction mixture was treated
with 3 N NaOH and H2O2 at ꢀ10 °C. After heated to 70 °C for 2 h,
the reaction mixture was extracted with diethyl ether three times
and dried over MgSO4 and concentrated under reduced pressure.
The residue was purified by flash column chromatography (ethyl
acetate/hexanes = 1:1) to afford the alcohol 16 (77 mg, 36%); 1H
To a CHCl3 (2 mL) solution of aldoxime 19 (164 mg, 0.76 mmol)
NCS (120 mg, 0.9 mmol) was added at 0 °C. After stirring for 3 h,
propargyl alcohol (84 mg, 1.5 mmol) and triethylamine (0.26 mL,
1.9 mmol) was added to the solution and heated to 50 °C. After
stirring for 12 h, the reaction mixture was diluted with ethyl ace-
tate and the organic phase was washed with water and brine, dried
over MgSO4 and concentrated under reduced pressure. The residue
was purified by flash column chromatography on silica gel (ethyl
acetate/hexanes = 1:2) to afford the isoxazole 20b (90 mg, 44%).
1H NMR (400 MHz, CDCl3) d 7.98 (d, 1H, J = 2.4 Hz), 7.49 (dd, 1H,
J = 9.3, 2.9 Hz), 6.86 (d, 1H, J = 9.3 Hz), 6.71 (s, 1H), 4.80 (d, 2H,
J = 5.4 Hz), 3.85 (s, 3H), 2.20 (br s, 1H); 13C NMR (100 MHz, CDCl3)
d 170.7, 158.9, 156.2, 133.7, 131.9, 119.6, 113.1, 103.3, 56.6, 55.8,
29.6. IR (KBr) 3394, 2940, 2850, 1605, 1504, 1460, 1268, 1030,
813 cmꢀ1; LRMS (ESI) m/z 305.9 (M+Na+).
NMR (500 MHz, CDCl3)
d
7.29–7.27 (m, 2H), 7.24 (d, 1H,
J = 2.2 Hz), 7.22 (d, 1H, J = 2.3 Hz), 7.06–7.02 (m, 4H), 6.95 (d, 2H,
J = 8.3 Hz), 6.91–6.88 (m, 2H), 6.04–5.93 (m, 2H), 5.18 (s, OH),
5.11–5.03 (m, 4H), 3.88 (s, 3H), 3.87 (s, 3H), 3.69 (t, 2H,
J = 6.5 Hz), 3.64 (t, 2H, J = 6.2 Hz), 3.42 (d, 2H, J = 6.7 Hz), 3.35 (d,
2H, J = 6.8 Hz), 2.76 (t, 2H, J = 7.3 Hz), 2.66 (t, 2H, J = 7.5 Hz),
1.91–1.85 (m, 4H); LRMS (ESI) m/z 321.1 (M+Na+)
3.1.13. 40-Methoxy-30,5-bis(3-methylbut-2-en-1-yl)-[1,10-
biphenyl]-2-ol (17)
3.1.17. 3-(5-Allyl-2-methoxyphenyl)-5-propylisoxazole (21a)
To a CH2Cl2 solution (0.5 mL) of 4-O-methylhonokiol (28 mg,
0.1 mmol) and 2-methyl-2-butene (1.2 mL) in sealed tube was
added 2nd generation Grubbs’ catalyst (4 mg, 4.7 lmol). After stir-
To
0.16 mmol) and allyltributyltin (103 mg, 0.31 mmol) tetrakis(tri-
phenylphosphine)palladium(0) (10 mg, 7.8 mol) was added. After
a DMF (1 mL) solution of the bromide 20a (46 mg,
l
ring for 24 h, the reaction mixture was concentrated and the resi-
due was purified by flash column chromatography on silica gel
(ethyl acetate/hexanes = 1:9) to afford the prenylated compound
17 (32 mg, 95%). 1H NMR (400 MHz, CDCl3) d 7.41 (m, 2H), 7.19
(m, 2H), 7.10 (d, 1H, J = 8.2 Hz), 7.04 (d, 1H, J = 7.8 Hz), 5.49 (m,
2H), 4.02 (s, 3H), 3.51 (d, 2H, J = 7.3 Hz), 3.45 (d, 2H J = 7.3 Hz),
1.88 (m, 12H); 13C NMR (100 MHz, CDCl3) d 157.0, 150.5, 133.8,
132.9, 132.2, 131.1, 129.9, 129.8, 128.9, 128.3, 127.8, 127.4,
123.5, 121.9, 115.3, 110.7, 55.4, 33.4, 28.4, 25.8, 25.7, 17.7; IR
(neat) 2965, 1691, 1499, 1246 cmꢀ1; LRMS (ESI) m/z 337.1 (M+H+).
stirring for 5 h at 90 °C, the reaction mixture was cooled to ambient
temperature, diluted with ethyl acetate and the organic phase was
washed with water and brine, dried over MgSO4 and concentrated
under reduced pressure. The residue was purified by flash column
chromatography (ethyl acetate/hexanes = 1:10) to afford the iso-
xzaole 21a (30 mg, 71%); 1H NMR (400 MHz, CDCl3) d 7.70 (d, 1H,
J = 2.6 Hz), 7.22 (dd, 1H, J = 2.4 and 8.2 Hz), 6.93 (d, 1H,
J = 8.8 Hz), 6.47 (s, 1H), 6.01 (m, 1H), 5.10 (m, 2H), 3.87 (s, 3H),
3.37 (d, 2H, J = 6.3 Hz), 2.78 (t, 2H, J = 7.3 Hz), 1.82(m, 2H), 1.04
(t, 3H, J = 7.3 Hz); 13C NMR (100 MHz, CDCl3) d 172.7, 159.9,
155.6, 137.4, 132.4, 130.8, 129.5, 118.2, 115.7, 111.5, 102.2, 55.6,
39.2, 28.7, 20.9, 13.7. IR (KBr) 3425, 2965, 2933, 1736, 1604,
1512, 1468, 1380, 1254 cmꢀ1; LRMS (ESI) m/z 296.0 (M+K+).
3.1.14. 5-Bromo-2-methoxybenzaldehyde oxime (19)
To
a CH2Cl2 (50 mL) solution of 5-bromo-2-anisaldehyde
(2.15 g, 10 mmol) and pyridine (5 mL), hydroxylamine HCl

2866
B. Lee et al. / Bioorg. Med. Chem. 20 (2012) 2860–2868
3.1.18. (3-(5-Allyl-2-methoxyphenyl)isoxazol-5-yl)methanol
(21b)
3.1.22. 2-Bromo-4-propylphenyl acetate (25)
To CHCl3 (13 mL) solution of 4-propylphenol (1.36 g,
a
To
0.3 mmol) and allyltributyltin (148 mg, 0.45 mmol) tetrakis-(tri-
phenylphosphine)palladium(0) (17 mg, 15 mol) was added. After
a
DMF (1 mL) solution of the isoxazole 20b (80 mg,
10 mmol) bromine (10 g, 7.8 mmol) was added at ambient temper-
ature. After stirring for 12 h, the reaction mixture was quenched
with aqueous NaHSO3. The reaction mixture was extracted with
ethyl ether, washed with water and brine, dried over MgSO4 and
concentrated under reduced pressure to afford the crude bromide
compound. To a CH2Cl2 (20 mL) solution of the crude 2-bromo-4-
phenol (ca. 2.15 g, 10 mmol) triethylamine (2.8 mL, 20 mmol)
and acetic anhydride (1.4 mL, 15 mmol) were added at 0 °C. After
stirring for 2 h at ambient temperature, the reaction mixture was
diluted with CH2Cl2 and the organic phase was washed with water
and brine, dried over MgSO4 and concentrated under reduced pres-
sure. The residue was purified by flash column chromatography
(ethyl acetate/hexanes = 1:12) to afford the acetate 25 (2.2 g,
86%); 1H NMR (400 MHz, CDCl3) d 7.40 (d, 1H, J = 2.4 Hz), 7.11
(dd, 1H, J = 1.9 and 8.3 Hz), 7.00 (d, 1H, J = 7.8 Hz), 2.55 (t, 2H,
J = 7.3 Hz), 2.32 (s, 3H), 1.65 (m, 2H), 0.94 (t, 3H, J = 7.3 Hz); 13C
NMR (100 MHz, CDCl3) d 168.5, 145.9, 142.1, 132.9, 128.3, 123.1,
115.6, 36.9, 24.2, 20.6, 13.6. IR (KBr) 3424, 2965, 1773, 1491,
1372, 1191 cmꢀ1; LRMS (ESI) m/z 278.9 (M+Na+).
l
stirring for 5 h at 90 °C, the reaction mixture was cooled to ambient
temperature, diluted with ethyl acetate and the organic phase was
washed with water and brine, dried over MgSO4 and concentrated
under reduced pressure. The residue was purified by flash column
chromatography (ethyl acetate/hexanes = 1:2) to afford the isoxaz-
ole 21b (60 mg, 82%); 1H NMR (400 MHz, CDCl3) d 7.69 (d, 1H,
J = 1.9 Hz), 7.24 (dd, 1H, J = 8.8 and 2.4 Hz), 6.94 (d, 1H,
J = 8.8 Hz), 6.75 (s, 1H), 6.00 (m, 1H), 5.10 (m, 2H), 4.81 (br s,
2H), 3.81 (s, 3H), 3.37 (d, 2H, J = 6.3 Hz); 13C NMR (100 MHz, CDCl3)
d 170.3, 160.1, 155.6, 137.3, 132.5, 131.2, 129.5, 117.6, 115.8,
111.5, 103.5, 56.6, 55.6, 39.1. IR (KBr) 3404, 2956, 2845, 1649,
1406, 1033 cmꢀ1; LRMS (ESI) m/z 268.0 (M+Na+).
3.1.19. 4-Allyl-2-(5-propylisoxazol-3-yl)phenol (22a)
To a CH2Cl2 (2 mL) solution of the isoxazole 21a (70 mg,
0.27 mmol) boron tribromide (1.0 M solution of CH2Cl2, 680 lL,
0.68 mmol) was added at ꢀ78 °C. After stirring for 1 h, the reaction
mixture was heated to ambient temperature, quenched with meth-
anol, and stirred for 30 min. The organic phase was washed with
water and brine, dried over MgSO4 and concentrated under re-
duced pressure. The residue was purified by flash column chroma-
tography (ethyl acetate/hexanes = 1:15) to afford the phenol 22a
(40 mg, 61%); 1H NMR (400 MHz, CDCl3) d 9.47 (br s, 1H), 7.27
(d, 1H, J = 1.9 Hz), 7.16 (dd, 1H, J = 8.3 and 2.4 Hz), 7.01 (d, 1H,
J = 8.3 Hz), 6.4 (s, 1H), 6.01 (m, 1H), 5.10 (m, 2H), 3.37 (d, 2H,
J = 6.8 Hz) 2.81 (t, 2H, J = 7.3 Hz) 1.84 (m, 2H), 1.05 (t, 3H,
J = 7.3 Hz); 13C NMR (100 MHz, CDCl3) d 173.0, 162.4, 154.9,
137.4, 131.7, 131.7, 130.9, 127.5, 117.3, 115.7, 113.2, 98.4, 39.1,
28.3, 20.8, 13.5; LRMS (ESI) m/z 244.1 (M+H+) and 266.1 (M+Na+).
3.1.23. 4-Propyl-2-((trimethylsilyl)ethynyl)phenyl acetate
To a triethylamine (12 mL) solution of the bromide 25 (1.26 g,
4.9 mmol) and ethynyltrimethylsilane (727 mg, 7.4 mmol), bis(tri-
phenylphosphine)palladium(II) dichloride (172 mg, 0.25 mmol)
and CuI (48 mg, 0.25 mmol) were added. After stirring for 5 h at
80 °C, the reaction mixture was cooled to ambient temperature
and concentrated under reduced pressure. The residue was puri-
fied by flash column chromatography (ethyl acetate/hex-
anes = 1:15) to afford the TMS-acetylene compound (1.09 g, 81%);
1H NMR (400 MHz, CDCl3) d 7.31 (d, 1H, J = 1.9 Hz), 7.14 (m, 1H),
7.02 (d, 1H, J = 8.3 Hz), 2.57 (dd, 2H, J = 15 and 7.8 Hz), 2.31 (s,
3H), 1.65 (m, 2H), 0.96 (dd, 3H, J = 15 and 6.8 Hz), 0.23 (s, 9H).
3.1.20. 4-Allyl-2-(5-(hydroxymethyl)isoxazol-3-yl)phenol (22b)
3.1.24. 2-Ethynyl-4-propylphenyl acetate (26)
To a CH2Cl2 (2 mL) solution of the isoxazole 21b (55 mg,
0.22 mmol) boron trichloride (1.0 M solution of CH2Cl2, 880 lL,
To an acetonitrile (5 mL) solution of the TMS-acetylene com-
pound (506 mg, 1.85 mmol) aqueous solution (1 mL) of Cs2CO3
(60 mg, 0.18 mmol) was added. After stirring for 2 h at ambient
temperature, the reaction mixture was concentrated under re-
duced pressure. The residue was purified by flash column chroma-
tography (diethyl ether/hexanes = 1:10) to afford the acetate 26
(220 mg, 59%) and the deacetylated compound (74 mg, 25%); For
the acetate, 1H NMR (400 MHz, CDCl3) d 7.41 (d, 1H, J = 1.9 Hz),
7.12 (dd, 1H, J = 4.4 and 1.9 Hz), 6.97 (d, 1H, J = 8.2 Hz), 2.57 (dd,
2H, J = 15 and 7.8 Hz), 2.34 (s, 3H), 1.63 (m, 2H), 0.96 (dd, 3H,
J = 15 and 6.8 Hz). For the deacetylated compound, 1H NMR
(400 MHz, CDCl3) d 7.18 (d, 1H, J = 2.4 Hz), 7.07 (d, 1H, J = 8.3 Hz),
6.85 (d, 1H, J = 8.3 Hz), 5.63 (s, 1H), 3.43 (s, 1H), 2.50 (t, 3H,
J = 7.3 Hz), 1.59 (m, 2H), 0.96 (t, 3H, J = 7.3 Hz).
0.88 mmol) was added at ꢀ78 °C. After stirring for 1 h, the reaction
mixture was heated to ambient temperature, quenched with meth-
anol, and stirred for 30 min. The organic phase was washed with
water and brine, dried over MgSO4 and concentrated under re-
duced pressure. The residue was purified by flash column chroma-
tography (ethyl acetate/hexanes = 1:2) to afford the phenol 22b
(35 mg, 69%); 1H NMR (400 MHz, CDCl3) d 9.32 (s, 1H), 7.27 (d,
1H, J = 1.9 Hz), 7.18 (dd, 1H, J = 8.8 and 2.4 Hz), 7.01 (d, 1H,
J = 8.3 Hz), 6.66 (s, 1H), 6.00 (m, 1H), 5.10 (m, 2H), 4.84 (s, 2H),
3.36 (d, 2H, J = 6.8 Hz), 2.37 (br s, 1H); 13C NMR (100 MHz, CDCl3)
d 170.7, 162.5, 154.8, 137.3, 132.1, 131.2, 127.7, 117.4, 115.9,
112.8, 99.6, 56.5, 39.1; LRMS (ESI) m/z 254.0 (M+Na+).
3.1.21. 4-Bromo-2-(5-propylisoxazol-3-yl)phenol (23)
3.1.25. 2-(1-Allyl-1H-1,2,3-triazol-4-yl)-4-propylphenyl acetate
(27a)
To a CH2Cl2 (2 mL) solution of the isoxazole 20a (30 mg,
0.10 mmol) boron trichloride (1.0 M solution of CH2Cl2, 300
l
L,
A DMSO (2 mL) solution of allyl bromide (145 mg, 1.2 mmol)
and sodium azide (78 mg, 1.2 mmol) was added to sealed tube.
After stirring for 12 h at ambient temperature, aqueous solution
(0.5 mL) of sodium ascorbate (25 mg, 0.1 mmol), CuSO4ꢁ5H2O
(16 mg, 0.05 mmol) and the acetylene 26 (200 mg, 1 mmol) were
added. After stirring for 1 h at ambient temperature and for 12 h
at 70 °C, the reaction mixture was cooled to ambient temperature,
quenched with cold 1 N NH4OH solution (not to generate explosive
CuN3), and extracted with EtOAc. The organic phase was washed
with brine, dried over MgSO4 and concentrated under reduced pres-
sure. The residue was purified by flash column chromatography
(ethyl acetate/hexanes = 1:2) to afford the triazole 27a (148 mg,
62%); 1H NMR (400 MHz, CDCl3) d 7.88 (d, 1H, J = 1.9 Hz), 7.73 (s,
0.30 mmol) was added at ꢀ78 °C. After stirring for 1 h, the reaction
mixture was heated to ambient temperature, quenched with meth-
anol, and stirred for 30 min. The organic phase was washed with
water and brine, dried over MgSO4 and concentrated under re-
duced pressure. The residue was purified by flash column chroma-
tography (ethyl acetate/hexanes = 1:15) to afford the phenol 23
(18 mg, 64%); 1H NMR (400 MHz, CDCl3) d 9.63 (s, 1H), 7.58 (d,
1H, J = 2.4 Hz), 7.41 (dd, 1H, J = 2.4 and 8.7 Hz), 6.97 (d, 1H,
J = 8.8 Hz), 6.37 (s, 3H), 2.82 (t, 2H, J = 7.8 Hz), 1.8 (m, 2H), 1.05
(t, 3H, J = 7.3 Hz); 13C NMR (100 MHz, CDCl3) d 173.6, 161.3,
155.7, 133.9, 130.1, 119.3, 115.3, 111.3, 98.4, 28.4, 20.8, 13.6; LRMS
(ESI) m/z 281.0 (M+H+).

B. Lee et al. / Bioorg. Med. Chem. 20 (2012) 2860–2868
2867
1H), 7.18 (dd, 1H, J = 8.2 and 2.4 Hz), 7.07 (d, 1H, J = 8.3 Hz), 6.09 (m,
1H), 5.39 (d, 1H, J = 10.2 Hz), 5.35 (d, 1H, J = 17.0 Hz), 5.03 (d, 2H,
J = 6.3 Hz), 2.64 (t, 2H, J = 7.8 Hz), 2.3 (s, 3H), 1.72 (m, 2H), 0.97 (t,
3H, J = 7.3 Hz); 13C NMR (100 MHz, CDCl3) d 169.1, 145.1, 143.5,
140.9, 131.3, 129.0, 128.4, 122.8, 122.6, 121.3, 120.1, 52.6, 37.4,
24.4, 21.2, 13.8. IR (KBr) 3417, 2962, 2935, 2873, 1767, 1733,
1497, 1374, 1192 cmꢀ1; LRMS (ESI) m/z 308.1 (M+Na+).
inhibitor celecoxib were used as positive control. Compounds were
dissolved in DMSO and were tested in at least three independent
experiments. Results were given as 50% inhibition concentration
(IC50) or means (GraphPad Software, San Diego, CA, USA).
3.2.2. Inhibitory activity on macrophage NO production and the
cell viability
The bioassay for inhibition of nitric oxide (NO) production in
lipopolysaccharide (LPS)-treated RAW264.7 macrophage cell lines
was carried out by Griess assay.15 RAW 264.7 macrophages were
purchased from American Type Culture Collection (Manassas, VA,
USA). The cells were cultured in Dulbecco’s modified Eagle’s media
supplemented with heat-inactivated 10% fetal bovine serum (FBS),
benzylpenicillin potassium (143 U/ml) and streptomycin sulfate
3.1.26. 2-(1-Allyl-1H-1,2,3-triazol-4-yl)-4-propylphenol (28a)
To
a MeOH (1 mL) solution of the triazole 27a (18 mg,
0.06 mmol) K2CO3 (20 mg, 0.15 mmol) was added. After stirring
for 2 h at 50 °C, the reaction mixture was concentrated under re-
duced pressure. The residue was purified by flash column chroma-
tography (diethyl ether/hexanes = 1:1) to afford the triazole 28a
(10 mg, 68%); 1H NMR (400 MHz, CDCl3) d 10.6 (br s, 1H), 7.84 (s,
1H), 7.20 (d, 1H, J = 2.0 Hz), 7.06 (dd, 1H, J = 2.4 and 8.3 Hz), 6.98
(d, 1H, J = 8.3 Hz), 6.12 (m, 1H), 5.44–5.36 (m, 2H), 5.06–5.04 (m,
2H), 2.54 (t, 2H, J = 7.8 Hz), 1.66–1.57 (m, 2H), 0.95 (t, 3H,
J = 7.3 Hz); 13C NMR (100 MHz, CDCl3) d 142.8, 137.1, 122.5,
119.7, 118.9, 114.4, 109.7, 107.5, 106.4, 102.4, 42.0, 26.1, 13.7,
27.3; LRMS (ESI) m/z 244.1 (M+H+).
(100 lg/ml) under 37 °C and 5% CO2 atmosphere. The cells were
stimulated with LPS (1 g/ml) for 24 h in the presence or absence
of compounds. Aliquots of the culture media were reacted with
the same volume of 0.05% sulfanilamide and 0.05% N-(1-naph-
thyl)ethylenediamine, and then absorbance values at 540 nm were
measured. Sodium nitrite used to make the standard curve. Cell
viability was examined by the propidium iodide (PI) nuclear stain-
ing method. Cells were stained with 1 lg/ml of PI and analyzed
3.1.27. 2-(1-(Cyclobutylmethyl)-1H-1,2,3-triazol-4-yl)-4-
propylphenol (28b)
with a FACSCanto flow cytometer (BD Biosciences, San Jose, CA,
USA). Cells stained with PI were considered dead cells.16
A DMSO (1 mL) solution of (bromomethyl)cyclobutane (90 mg,
0.6 mmol) and sodium azide (40 mg, 0.6 mmol) was added to sealed
tube. After stirring for 12 h at ambient temperature, aqueous solu-
tion (0.5 mL) of sodium ascorbate (13 mg, 0.05 mmol), CuSO4ꢁ5H2O
(8 mg, 0.03 mmol) and the acetylene 26 (100 mg, 0.5 mmol) were
added. After stirring for 1 h at ambient temperature and for 12 h at
70 °C, the reaction mixture was cooled to ambient temperature,
quenched with cold 1 N NH4OH solution(not to generate explosive
CuN3), and extracted with EtOAc. The organic phase was washed
with brine, dried over MgSO4 and concentrated under reduced pres-
sure. The residue was used without further purification. To a MeOH
(1 mL) solution of crude triazole compound K2CO3 (15 mg, 0.1 mmol)
was added. After stirring for 2 h at 50 °C, the reaction mixture was
concentrated under reduced pressure. The residue was purified by
flash column chromatography (diethyl ether/hexanes=1:1) to afford
the phenol 28b (90 mg, 67%); 1H NMR (400 MHz, CDCl3) d 10.6 (br s,
1H), 7.78 (s, 1H), 7.20 (d, 1H, J = 2.4 Hz), 7.05 (dd, 1H, J = 2.4 and
8.3 Hz), 6.97 (d, 1H, J = 8.3 Hz), 4.44 (d, 2H, J = 7.8 Hz), 2.93 (m, 1H),
2.55 (m, 2H), 2.17–2.10 (m, 2H), 2.04–1.81 (m, 4H), 1.66–1.56 (m,
2H), 0.95 (t, 3H, J = 7.3 Hz); 13C NMR (100 MHz, CDCl3) d 153.5,
147.6, 133.5, 129.7, 125.4, 118.5, 117.3 117.2, 55.5, 37.1, 35.4, 25.7,
24.7, 17.9, 13.7; LRMS (ESI) m/z 272.1 (M+Na+).
4. Conclusion
18 compounds based on 4-O-methylhonokiol were designed
and synthesized in order to improve metabolic stabilization and
anti-inflammatory activity. They were synthesized through the di-
rect modification of 4-O-methylhonokiol in three different ways
that are phenol direct modification of A ring, ortho-halogenation
of phenol group and olefin modification. Azole analogs such as
isoxazole and triazole were replaced with B ring in order to give
higher solubility and less reactivity of phenol in A ring through
hydrogen bond with nitrogen of the azole rings. The inhibitory ef-
fects of the newly synthesized compounds on COX-2 and PGF1 pro-
duction were evaluated with the goal of identifying new anti-
inflammatory agents. We found that aryl carbamates (10 and 11)
exhibited a potent inhibitory activity against the production of
COX-2 enzyme and the inflammatory modulator PGF1 and could
serve as new therapeutic agents for the treatment of diseases med-
iated by PGs. The structure–property relationship studies on these
novel 4-O-methylhonokiol analogs are underway and the results of
the DMPK study will be reported in due course.
Acknowledgments
3.2. Biologic assays
This work was supported in part by the Basic Science Research
Program (2010-0004496 to S.-Y.S.) and the Medical Research
Center program (2010-0029480 to J.-K.J.) through the National
Research Foundation of Korea, and by the Ministry of Land, Trans-
port and Maritime Affairs, Republic of Korea (J.-K.J.).
3.2.1. Inhibitory activity of COX-2 enzyme and PGF1 production
in macrophages
Inhibition of cyclooxygenase-2 (COX-2) enzyme was performed
by using COX inhibitor screening kit (Cat # 560131, Cayman Chem-
ical Company, Ann Arbor, MI, USA). This kit directly measures pros-
taglandin F1/2 produced by SnCl2 reduction of COX-derived PGH2
produced in the COX reaction. All procedures were performed as
indicated in the assay procedures. Briefly, the reaction buffer
(Tris–HCl buffer, pH 8.0, containing 5 mM EDTA and 2 mM phenol)
and heme were placed in test tubes. The samples and recombinant
human COX-2 were added to test tubes and pre-incubated for
10 min at 37 °C. After the substrate arachidonic acid was added,
the test tubes were incubated for 2 min at 37 °C. The concentration
of PGF2 was measured using the EIA kit.7 Inhibition of PGF1 produc-
tion in cells was determined by using RAW 264.7 cells. Cells were
treated with LPS in the presence of compounds for 24 h. The concen-
tration of PGF1 in culture media was measured using EIA kit. COX-2
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmc.2012.03.028.
These data include MOL files and InChiKeys of the most important
compounds described in this article.
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