Design and synthesis of biphenyl derivatives as mushroom tyrosinase inhibitors

Article abstract of DOI:10.1016/j.bmc.2010.07.062

Two new series of biphenyls, analogs of aglycone of natural product fortuneanoside E, were prepared using Suzuki-Miyaura cross-coupling and selective magnesium iodide demethylation/debenzylation, and their mushroom tyrosinase inhibitory activity was evaluated. Most of the 4-hydroxy-3,5- dimethoxyphenyl biphenyl compounds (series II, 20-36) were in general more active than 3,4,5-trimethoxyphenyl biphenyl compounds (series I, 1-19). Structure-activity relationships study showed that monosaccharide substituents, such as glucose, were not necessary and the presence of 4-hydroxy-3,5- dimethoxyphenyl moiety was crucial for inhibitory activity. Among the compounds synthesised, compound 21 (IC₅₀ = 0.02 mM) was found to be the most active one, which exhibited an activity that was 7 times higher than that of fortuneanoside E (IC₅₀ = 0.14 mM) and 10 times higher than that of arbutin (IC₅₀ = 0.21 mM), known as potent tyrosinase inhibitors. The inhibition kinetics analyzed by Lineweaver-Burk plots revealed that compound 21 was a competitive inhibitor (K_i = 0.015 mM).

Full text of DOI:10.1016/j.bmc.2010.07.062

Bioorganic & Medicinal Chemistry 18 (2010) 6708–6714
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Design and synthesis of biphenyl derivatives as mushroom tyrosinase inhibitors
a,c,
Kai Bao a,b, , Yi Dai a, , Zhi-Bin Zhu b, Feng-Juan Tu c, Wei-Ge Zhang b, , Xin-Sheng Yao
*
*
a Institute of Traditional Chinese Medicine & Natural Products, Jinan University, Guangzhou 510632, PR China
b School of Pharmaceutical Engineering, Shenyang Pharmaceutical University, Shenyang 110016, PR China
c School of Traditional Chinese Materia Medica, Shenyang Pharmaceutical University, Shenyang 110016, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Two new series of biphenyls, analogs of aglycone of natural product fortuneanoside E, were prepared
using Suzuki–Miyaura cross-coupling and selective magnesium iodide demethylation/debenzylation,
and their mushroom tyrosinase inhibitory activity was evaluated. Most of the 4-hydroxy-3,5-dimethoxy-
phenyl biphenyl compounds (series II, 20–36) were in general more active than 3,4,5-trimethoxyphenyl
biphenyl compounds (series I, 1–19). Structure–activity relationships study showed that monosaccharide
substituents, such as glucose, were not necessary and the presence of 4-hydroxy-3,5-dimethoxyphenyl
moiety was crucial for inhibitory activity. Among the compounds synthesised, compound 21
(IC50 = 0.02 mM) was found to be the most active one, which exhibited an activity that was 7 times higher
than that of fortuneanoside E (IC50 = 0.14 mM) and 10 times higher than that of arbutin (IC50 = 0.21 mM),
known as potent tyrosinase inhibitors. The inhibition kinetics analyzed by Lineweaver–Burk plots
revealed that compound 21 was a competitive inhibitor (Ki = 0.015 mM).
Received 17 June 2010
Revised 26 July 2010
Accepted 27 July 2010
Available online 1 August 2010
Keywords:
Fortuneanoside E
Biphenyls
Tyrosinase inhibitors
Synthesis
Ó 2010 Elsevier Ltd. All rights reserved.
1. Introduction
II: 4-hydroxy-3,5-dimethoxyphenyl series, 20–36) with different
substitution patterns. We then examined the inhibitory effects of
Tyrosinase (EC 1.14.18.1), is a multifunctional copper-contain-
ing enzyme, that is, key in melanin biosynthesis, melanisation in
animals, and browning in plants. Tyrosinase inhibitors can there-
fore be clinically useful for the treatment of some dermatological
disorders associated with melanin hyperpigmentation. In addition,
these inhibitors are also known to be useful in cosmetics for whit-
ening and depigmentation after sunburn.1,2 In the past few dec-
ades, a number of polyphenol tyrosinase inhibitors from both
natural and synthetic sources, including flavonoids, stilbenes, and
terpenoids, have been intensively investigated.3–7 To our knowl-
edge, however, only a few biphenyl inhibitors have been reported
to date.8,9
Fortuneanoside E is a new biphenyl glycoside isolated from the
fruit of Pyracantha fortuneana that has shown tyrosinase inhibi-
tory activity. We previously reported the first total synthesis of this
compound,10,11 and it was noted that a synthetic intermediate of
fortuneanoside E, compound I, was also found to show potent
activities in the tyrosinase inhibition assay (Fig. 1).
these compounds on mushroom tyrosinase to study the struc-
ture–activity relationships with these potential inhibitors.
2. Results and discussion
2.1. Chemistry
The synthetic procedures employed to obtain the target com-
pounds 1–19 and 20–36 are depicted in Scheme 1.
Generally, there are several methods for the construction of
biphenyls, including Stille coupling, the Gomberg–Bachmann reac-
tion, the Ullmann reaction, and Suzuki–Miyaura cross-coupling.
Considered the particularities of these methods and the character-
istics of the target compounds, Suzuki–Miyaura cross-coupling
OCH3
OR3
From examining the structural characteristics of fortuneanoside
E and compound I, we designed and synthesized two series of
biphenyls (series I: 3,4,5-trimethoxyphenyl series, 1–19 and series
R1O
OCH3
R2O
* Corresponding authors. Tel.: +86 24 23986422; fax: +86 24 23986393 (W.-G.Z.);
tel.: +86 20 85225849; fax: +86 20 85221559 (X.-S.Y.).
E-mail addresses: zhangweige2000@sina.com (W.-G. Zhang), yaoxinsheng@vip.-
tom.com (X.-S. Yao).
Fortuneanoside E
Compound I
R1=H, R2=Glc, R3=H
R1=Bn, R2=H, R3=CH3
Figure 1. Chemical structures of fortuneanoside E and compound I.
These authors contributed equally to this work.
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2010.07.062

K. Bao et al. / Bioorg. Med. Chem. 18 (2010) 6708–6714
6709
OCH3
OCH3
OH
R1
R4
OCH3
R1
R4
OCH3
R1
R4
R2
R3
Br
OCH3
a
b
R2
R3
R2
R3
OCH3
OCH3
R5
(HO)2B
OCH3
R5
R5
R1-R5 = H, Me, OH, Ac, etc
1-19
20-36
Scheme 1. Synthesis of biphenyls 1–19 and 20–36. Reagents and conditions: (a) K2CO3 (2.0 equiv), Pd(OAc)2 (0.01 equiv), DMF, 110 °C; (b) MgI2 (3.0 or 6.0 equiv), 80 °C.
was applied in the first step. To find the optimal conditions, a mod-
el reaction was performed using 2-(benzyloxy)-4-bromobenzalde-
hyde and 3,4,5-trimethoxy-phenylboronic acid under different
conditions. As presented in Table 1, this reaction was found to be
most efficient in the presence of K2CO3 and Pd(OAc)2 in DMF at
110 °C and using these conditions, compound 1 could be prepared
in 72% yield (Table 1, entry 9). With these optimized condition,
biphenyls 2–19 were prepared from various bromobenzenes (com-
mercially available or prepared by conventional methods) by Suzu-
ki–Miyaura cross-coupling with 3,4,5-trimethoxy-phenylboronic
acid, resulting in yields ranging from 68% to 92%.
Methyl and benzyl ethers have been widely used as stable pro-
tecting groups for hydroxyl groups in organic synthesis. A number
of methods have been reported to perform cleavage of the methyl
and benzyl groups in these ethers.12,13 In our previous report,11 we
described a highly selective demethylation/debenzylation method
that was effective in different kinds of pyrogallol trimethyl ethers
using MgI2 under solvent-free conditions. In these reactions, we
found that a variety of functional groups were tolerated under
our reaction conditions. Consequently, we applied this method in
the synthesis of biphenyls 20–36. Treatment of compounds 1–19
with MgI2 at 80 °C under solvent-free conditions gave compounds
20–36 in good to excellent yields. If necessary, the amount of MgI2
and the reaction temperature could be adjusted to improve the
reaction yield. Analytical data of the previously reported com-
pounds corresponded to the expected values and all new com-
pounds were analyzed by 1H, 13C NMR, and MS.
not necessary for good inhibition activity. Some of the tested
biphenyls, especially those containing the 4-hydroxy-3,5-dime-
Table 2
Tyrosinase inhibitory activities of compounds 1–19 and 20–36
a
Compound
R1
R2
R3
R4
R5 IC50 (mM)
OCH3
R1
OCH3
B
R2
R3
OCH3
A
R5
R4
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
H
H
H
H
H
H
H
H
F
H
F
OMe
OMe
H
OBn
OH
OBn
OH
OH
OBn
OH
F
CHO
CHO
COOH
COOH
COOEt
CONHBn
Me
H
H
H
H
H
H
H
H
F
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
>2.0
H
F
OMe
H
H
OH
OBn
H
H
H
F
F
H
H
H
H
H
H
H
H
H
Ac
CHOHCH3
H
H
H
H
H
H
H
H
Me
H
H
H
0.55
>2.0
>2.0
>2.0
>2.0
17
18
19
COOH
CONHBu
CONHBn
OH
2.2. Inhibition of tyrosinase
H
OBn
Compound I
H
0.17
The inhibition abilities of our synthetic biphenyls were initially
investigated in vitro on mushroom tyrosinase using arbutin as the
positive control (Table 2). From our preliminary investigation, we
found that monosaccharide substituents, such as glucose, were
OCH3
R1
OH
B
R2
R3
OCH3
A
R5
R4
OH
OH
OH
OH
OH
F
H
F
OMe
OH
H
Table 1
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
H
H
H
H
H
H
F
H
F
F
OMe
OMe
H
Me
H
CHO
COOH
COOEt
CONHBn
Me
H
F
F
H
H
H
H
H
H
H
H
H
H
H
F
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
0.06
0.02
0.05
0.03
0.09
0.08
0.06
0.07
0.06
0.08
0.14
0.19
0.09
0.17
0.05
0.07
0.12
0.14
Suzuki–Miyaura cross-coupling of 2-(benzyloxy)-4-bromobenzaldehyde
Entry Base
Catalyst
Solvent
Cond.
Time
(h)
Yield
(%)
1
2
3
4
5
6
7
8
9
KF
KF
KF
Pd/C
CH3OH/
H2O = 1:1
rt
3.0
3.0
3.0
5.0
12.0
3.0
5.0
5.0
5.0
—
(ultrasound)
rt
(ultrasound)
rt
(ultrasound)
Reflux
Pd(PPh3)4 CH3OH/
H2O = 1:1
Pd(PPh3)4 THF/
H2O = 4:1
—
H
H
Ac
—
K2CO3 Pd(PPh3)4 THF/
H2O = 4:1
K2CO3 Pd(PPh3)4 Dioxane/
H2O = 6:1
K2CO3 Pd(OAc)2
K2CO3 Pd(OAc)2
K2CO3 Pd(OAc)2
K2CO3 Pd(OAc)2
10
—
H
OH
H
H
H
CHOHCH3
H
H
H
H
H
H
90 °C
COOH
CONHBu
CONHBn
OGlc
THF/
H2O = 4:1
THF/
H2O = 4:1
DMF
rt
—
H
H
H
0.21
(ultrasound)
Reflux
36
H
OH
32
36
72
Fortuneanoside E
Arbutin
rt
(ultrasound)
110 °C
a
The results are from the three concurrent readings, and each SD was usually
within 2% of the mean.
DMF

6710
K. Bao et al. / Bioorg. Med. Chem. 18 (2010) 6708–6714
Table 3
thoxyphenyl moiety, were more potent inhibitors than fortuneano-
side E and arbutin. With the exception of compound I, the 3,4,5-tri-
methoxyphenyl biphenyl compounds synthesized in series I,
compounds 1–19, showed very weak inhibitory activities against
tyrosinase. However, when the methyl moiety at the C-4 position
on ring B was cleaved, the activities sharply increased. From an
examination of the structure–activity relationships of the two
types of biphenyls in this study, it appears that the inhibitory activ-
ity is mainly dependent on the C-4 hydroxyl of ring B, whereas the
presence of a hydroxy moiety on ring A does not contribute much
to the inhibitory potency.
Kinetics and inhibition constants of compounds 21, 23, 34, and 35 on the activity of
mushroom tyrosinase
Compound
Inhibition type
Inhibition constant (Ki) (mM)
21
23
34
35
Competitive
Competitive
Competitive
Competitive
0.015
0.060
0.038
0.031
3. Conclusions
In series II, the most active compound was 3,40-dihydroxy-30,50-
dimethoxybiphenyl-4-carboxylic acid 21, which exhibited an
activity that was an order of magnitude higher than that of arbutin,
7 times higher than that of fortuneanoside E and 8.5 times higher
than that of compound I. Comparing the IC50 of compound 21
(IC50 = 0.02) with those of compounds 32 and 34, it is clear that
when hydroxyl and carboxyl groups are both present on the A ring,
the potency of the resulting compound 21 increases. In the case of
the fluorine containing derivatives 25–29, there was no remarkable
difference in their activities. In addition, it was interesting to find
that the activity of compound 34 (IC50 = 0.05), having 4-carboxyl
substituents, substituents, was comparable to that of compound
21. This finding also suggests that the hydroxy moiety on ring A
did not have a great influence on the activity.
Among the tested biphenyls, compounds 21 showed the highest
inhibitory activity, and hence, we carried out the kinetic analysis of
21 for tyrosinase inhibition with respect to L-tyrosine as a sub-
strate. Lineweaver–Burk plots for the inhibition of tyrosinase by
21 were obtained with variable concentrations of 21 and the sub-
strate (Fig. 2). The intersection of these lines on the vertical axis
indicated that 21 was a competitive inhibitor of tyrosinase with re-
In conclusion, two new series of biphenyls were prepared using
Suzuki–Miyaura cross-coupling and selective magnesium iodide
demethylation, and their activity as tyrosinase inhibitors was
examined. Most of the compounds in series II possessed higher
inhibitory activity compared to arbutin, fortuneanoside E and a
synthetic intermediate that was known to be active in these assays,
which suggested that the 4-hydroxy-3,5-dimethoxyl substituent
on the B ring is crucial for tyrosinase inhibitory activity in this class
of compounds. The present study revealed that activity could also
be influenced by the type and position of substituents on the A
ring—with electron withdrawing groups being somewhat advanta-
geous—although explicit correlations would require further study.
Among the compounds synthesized, compound 21 showed the
strongest inhibitory activity. Kinetic study revealed that compound
21 acted as a competitive inhibitor of mushroom tyrosinase with Ki
value of 0.015 mM. Further investigations into the structure–activ-
ity relationships of these compounds are currently in progress.
4. Experimental section
spect to L-tyrosine as a substrate, with a Ki value of 0.015 mM.
Unless otherwise noted, all the materials were obtained from
commercially available sources and were used without purifica-
tion. Thin-layer chromatography was performed on GF254 silica
gel plates to monitor the reaction, and the plates were examined
under UV light. The purification of the products was performed
using column chromatography (60 Å, 200–300 mesh, Qingdao
Ocean Chemicals) or silica gel plates (0.25 mm layer, Qingdao
Ocean Chemicals) with the designated solvents. Melting points
were measured on a hot-stage microscope (X-4, Beijing Taike
Ltd) and are uncorrected. ESI-MS spectra were performed on a Fin-
igan LCQ Advantage MAX or Agilent 1100 series MSD TRAP mass
spectrometer. Elemental analyzes were conducted by the Analyti-
cal Centre of Jilin University, China. 1H and 13C NMR spectra were
taken in CDCl3 solution on Bruker ARX-300, Bruker AV-400, and
Bruker AV-600 spectrometers with TMS as the internal reference.
Chemical shifts were reported in ppm downfield from tetramethyl-
silane and proton–proton coupling constants (J) in Hz.
These data strongly suggested that 21 effectively inhibited the en-
zyme by binding to its active site. The inhibition kinetics of the se-
lected compounds 23, 34, and 35 were also analyzed by
Lineweaver–Burk plots and the Ki of these compounds binding
were shown in Table 3. As shown in Table 3, the Ki of compound
21 was less than that of compounds 23, 34, and 35, suggesting that
compound 21 had most potent inhibitory effect.
350
300
250
200
150
100
50
4.1. General procedure for the preparation of compounds 1–19
4.1.1. 3-(Benzyloxy)-30,40,50-trimethoxybiphenyl-4-
carbaldehyde (1)
To a solution of 2-(benzyloxy)-4-bromobenzaldehyde (0.87 g,
3.00 mmol) and 3,4,5-trimethoxyphenyl-boronic acid (0.42 g,
2.00 mmol) in DMF (20 mL), was added anhydrous K2CO3 (0.55 g,
4.00 mmol) and Pd(OAc)2 (0.01 g, 0.06 mmol). The reaction mixture
was stirred at 110 °C for 5 h, then cooled to room temperature and
poured into water, filtered and recrystallized with ethyl acetate to
afford I as a colorless solid (0.54 g, 72%). Mp 139–142 °C; 1H NMR
0
-0.5
0
0.5
1
1.5
2
2.5
(400 MHz, CDCl3, TMS):
d 10.57 (s, 1H, CHO), 7.90 (d, 1H,
-50
1/[S] (mM)-1
J = 8.0 Hz, 5-H), 7.48–7.36, 5.29 (OBn), 7.20 (dd, 1H, J = 8.0, 1.1 Hz,
6-H), 7.15 (d, 1H, J = 1.1 Hz, 2-H), 6.70 (s, 2H, 20,60-H), 3.91 (s, 6H,
30,50-CH3O), 3.90 (s, 3H, 40-CH3O); 13C NMR (100 MHz, CDCl3,
Figure 2. Lineweaver–Burk plots of mushroom tyrosinase and
(j) and with (ꢀ) 0.02 mM and (N) 0.04 mM of compound 21.
L-tyrosine without

K. Bao et al. / Bioorg. Med. Chem. 18 (2010) 6708–6714
6711
TMS): d 189.2 (CHO), 161.2 (C-3), 153.6 (C-30/C-50), 149.0 (C-1),
138.9 (C-40), 136.2, 128.8, 128.3, 127.3, 70.8 (OBn), 135.9 (C-10),
129.0 (C-5), 124.1 (C-4), 120.0 (C-6), 112.0 (C-2), 104.8 (C-20/C-60),
61.0 (40-CH3O), 56.3 (30,50-CH3O); MS (ESI, m/z): 401.4 [M+Na+],
779.1 [2M+Na+]; Anal. Calcd for C23H22O5: C, 73.00; H, 5.86. Found:
C, 72.89; H, 5.90.
TMS): d 168.1 (CONH), 157.1 (C-3), 153.7 (C-30/C-50), 146.0 (C-1),
139.2 (C-40), 138.5, 128.6, 127.7, 127.2, 44.0 (NHBn), 134.7,
128.9, 128.7, 127.8, 71.6 (OBn), 133.1 (C-10), 127.2 (C-5), 126.0
(C-4), 118.7 (C-6), 111.5 (C-2), 104.8 (C-20/C-60), 61.0 (40-CH3O),
56.4 (30,50-CH3O); MS (ESI, m/z): 484.1 [M+H+], 506.4 [M+Na+];
Anal. Calcd for C30H29NO5: C, 74.52; H, 6.04; N, 2.90. Found: C,
74.46; H, 5.98; N, 2.93.
4.1.2. 3-Hydroxy-30,40,50-trimethoxybiphenyl-4-carbaldehyde
(2)
4.1.7. 30,40,50-Trimethoxy-4-methylbiphenyl-3-ol (7)
White solid, yield 83%. Mp 149–151 °C; 1H NMR (300 MHz,
CDCl3, TMS): d 11.15 (s, 1H, CHO), 9.92 (s, 1H, OH), 7.62 (d, 1H,
J = 8.1 Hz, 5-H), 7.23 (dd, 1H, J = 8.1, 1.6 Hz, 6-H), 7.18 (d, 1H,
J = 1.6 Hz, 2-H), 6.82 (s, 2H, 20,60-H), 3.94 (s, 6H, 30,50-CH3O), 3.91
(s, 3H, 40-CH3O); 13C NMR (75 MHz, CDCl3, TMS): d 194.3 (CHO),
161.9 (C-3), 156.9 (C-1), 153.7 (C-30/C-50), 137.9 (C-40), 136.8 (C-
10), 128.4 (C-5), 124.0 (C-6), 118.9 (C-4), 115.4 (C-2), 104.5 (C-20/
C-60), 61.3 (40-CH3O), 56.5 (30,50-CH3O); MS (ESI, m/z): 289.4
[M+H+], 287.2 [MꢀHꢀ]; Anal. Calcd for C16H16O5: C, 66.66; H,
5.59. Found: C, 66.57; H, 5.62.
White solid, yield 90%. Mp 138–139 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 7.16 (d, 1H, J = 7.8 Hz, 5-H), 7.03 (dd, 1H, J = 7.8,
1.8 Hz, 6-H), 6.96 (d, 1H, J = 1.8 Hz, 2-H), 6.72 (s, 2H, 20,60-H),
5.15 (s, 1H, OH), 3.88 (s, 9H, 30,40,50-CH3O), 2.29 (s, 3H, 4-CH3);
13C NMR (100 MHz, CDCl3, TMS): d 154.1 (C-3), 153.4 (C-30/C-50),
140.5 (C-1), 137.5 (C-40), 136.9 (C-10), 131.2 (C-5), 122.9 (C-4),
119.2 (C-6), 113.6 (C-2), 104.3 (C-20/C-60), 61.0 (40-CH3O), 56.2
(30,50-CH3O), 15.4 (4-CH3); MS (ESI, m/z): 275.8 [M+H+], 571.0
[2M+Na+], 272.7 [MꢀHꢀ]; Anal. Calcd for C16H18O4: C, 70.06; H,
6.61. Found: C, 70.14; H, 6.55.
4.1.3. 3-(Benzyloxy)-30,40,50-trimethoxybiphenyl-4-carboxylic
acid (3)
4.1.8. 30,50-Difluoro-3,4,5-trimethoxybiphenyl (8)
White solid, yield 75%. Mp 72–73 °C; 1H NMR (300 MHz, CDCl3,
TMS): d 7.07 (d, 2H, J = 6.5 Hz, 20,60-H), 6.79 (m, 1H, 40-H), 6.72 (s,
2H, 2,6-H), 3.94 (s, 6H, 3,5-CH3O), 3.89 (s, 3H, 4-CH3O); 13C NMR
(75 MHz, CDCl3, TMS): d 164.8, 161.6 (C-30/C-50), 153.6 (C-3/C-5),
144.6 (C-10), 138.5 (C-4), 134.8 (C-1), 109.9 (C-20/C-60), 104.3 (C-
2/C-6), 102.6 (C-40), 61.0 (4-CH3O), 56.3 (3,5-CH3O); MS (ESI, m/
z): 281.3 [M+H+], 303.4 [M+Na+]; Anal. Calcd for C15H14F2O3: C,
64.28; H, 5.03. Found: C, 64.15; H, 4.97.
White solid, yield 76%. Mp 133–135 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 8.23 (d, 1H, J = 8.1 Hz, 5-H), 7.49–7.41, 5.39 (OBn),
7.30 (dd, 1H, J = 8.1, 1.2 Hz, 6-H), 7.22 (d, 1H, J = 1.2 Hz, 2-H),
6.70 (s, 2H, 20,60-H), 3.92 (s, 6H, 30,50-CH3O), 3.90 (s, 3H, 40-
CH3O); 13C NMR (100 MHz, CDCl3, TMS): d 165.2 (COOH), 157.5
(C-3), 153.7 (C-30/C-50), 148.3 (C-1), 138.9 (C-40), 135.2, 129.2,
127.9, 72.4 (OBn), 134.5 (C-10), 134.3 (C-5), 121.1 (C-6), 116.7 (C-
4), 112.0 (C-2), 104.7 (C-20/C-60), 61.0 (40-CH3O), 56.3 (30,50-
CH3O); MS (ESI, m/z): 811.9 [2M+Na+], 827.7 [2M+K+], 787.7
[2MꢀHꢀ]; Anal. Calcd for C23H22O6: C, 70.04; H, 5.62. Found: C,
70.16; H, 5.51.
4.1.9. 2,4-Difluoro-30,40,50-trimethoxybiphenyl (9)
White solid, yield 69%. Mp 40–42 °C; 1H NMR (300 MHz, CDCl3,
TMS): d 7.40 (m, 1H, 6-H), 6.98–6.87 (m, 2H, 3,5-H), 6.70 (s, 2H,
20,60-H), 3.90 (s, 9H, 30,40,50-CH3O); 13C NMR (75 MHz, CDCl3,
TMS): d 164.1 (C-4), 159.0 (C-2), 153.5 (C-30/C-50), 138.1 (C-40),
131.6 (C-6), 130.8 (C-10), 125.6 (C-1), 111.8 (C-5), 106.6 (C-20/C-
60), 104.7 (C-3), 61.2 (40-CH3O), 56.5 (30,50-CH3O); MS (ESI, m/z):
281.4 [M+H+], 303.3 [M+Na+], 319.3 [M+K+]; Anal. Calcd for
4.1.4. 3-Hydroxy-30,40,50-trimethoxybiphenyl-4-carboxylic acid
(4)
Light yellow solid, yield 84%. Mp 107–109 °C; 1H NMR
(300 MHz, CDCl3, TMS): d 7.86 (d, 1H, J = 8.0 Hz, 5-H), 7.13–7.10
(m, 2H, 2,6-H), 6.86 (s, 2H, 20,60-H), 3.90 (s, 6H, 30,50-CH3O), 3.78
(s, 3H, 40-CH3O); 13C NMR (75 MHz, CDCl3, TMS): d 161.6 (COOH),
156.2 (C-3), 151.7 (C-30/C-50), 145.4 (C-1), 136.4 (C-40), 133.8 (C-
10, C-5), 118.1 (C-6), 115.6 (C-4), 113.2 (C-2), 102.8 (C-20/C-60),
58.8 (40-CH3O), 54.5 (30,50-CH3O); MS (ESI, m/z): 302.9 [MꢀHꢀ],
607.1 [2MꢀHꢀ]; Anal. Calcd for C16H16O6: C, 63.15; H, 5.30. Found:
C, 63.23; H, 5.27.
C15H14F2O3: C, 64.28; H, 5.03. Found: C, 64.21; H, 5.00.
4.1.10. 30,40-Difluoro-3,4,5-trimethoxybiphenyl (10)
White solid, yield 87%. Mp 69–72 °C; 1H NMR (300 MHz, CDCl3,
TMS): d 7.38–7.31 (m, 1H, 20-H), 7.28–7.16 (m, 2H, 50,60-H), 6.70 (s,
2H, 2,6-H), 3.92 (s, 6H, 3,5-CH3O), 3.89 (s, 3H, 4-CH3O); 13C NMR
(75 MHz, CDCl3, TMS): d 163.2 (C-30), 156.4 (C-40), 153.9 (C-3/C-
5), 138.7 (C-10), 138.3 (C-4), 135.4 (C-1), 123.2 (C-60), 117.7 (C-
20), 116.2 (C-50), 104.6 (C-2/C-6), 61.2 (4-CH3O), 56.5 (3,5-CH3O);
MS (ESI, m/z): 281.4 [M+H+], 303.3 [M+Na+], 583.3 [2M+Na+]; Anal.
Calcd for C15H14F2O3: C, 64.28; H, 5.03. Found: C, 64.31; H, 4.94.
4.1.5. Ethyl 3-hydroxy-30,40,50-trimethoxybiphenyl-4-
carboxylate (5)
White solid, yield 92%. Mp 100–103 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 10.91 (s, 1H, OH), 7.89 (d, 1H, J = 8.3 Hz, 5-H),
7.18 (d, 1H, J = 1.5 Hz, 2-H), 7.08 (dd, 1H, J = 8.3, 1.5 Hz, 6-H),
6.81 (s, 2H, 20,60-H), 4.44 (q, 2H, J = 7.1 Hz, OCH2CH3), 3.94 (s, 6H,
30,50-CH3O), 3.90 (s, 3H, 40-CH3O), 1.44 (t, 3H, J = 7.1 Hz, OCH2CH3);
13C NMR (100 MHz, CDCl3, TMS): d 170.1 (COOEt), 161.8 (C-3),
153.5 (C-30/C-50), 148.4 (C-1), 138.6 (C-40), 135.5 (C-10), 130.3 (C-
5), 117.9 (C-6), 115.6 (C-4), 111.4 (C-2), 104.6 (C-20/C-60), 61.4
(OCH2CH3), 61.0 (40-CH3O), 56.3 (30,50-CH3O), 14.2 (OCH2CH3); MS
(ESI, m/z): 355.2 [M+Na+], 687.9 [2M+Na+], 331.2 [MꢀHꢀ]; Anal.
Calcd for C18H20O6: C, 65.05; H, 6.07. Found: C, 64.91; H, 6.14.
4.1.11. 2-Fluoro-3,30,40,50-tetramethoxybiphenyl (11)
White solid, yield 83%. Mp 82–85 °C; 1H NMR (400 MHz, CDCl3,
TMS): d 7.11 (m, 1H, 6-H), 7.00 (m, 1H, 5-H), 6.94 (m, 1H, 4-H), 6.76
(s, 2H, 20,60-H), 3.92 (s, 3H, 3-CH3O), 3.90 (s, 3H, 40-CH3O), 3.89 (s,
13
6H, 30,50-CH3O); C NMR (100 MHz, CDCl3, TMS): d 153.1 (C-30/C-
50), 150.7 (C-3), 148.3 (C-2), 137.9 (C-40), 131.1 (C-10), 129.8 (C-1),
123.8 (C-5), 121.8 (C-6), 112.2 (C-4), 106.5/106.4 (C-20/C-60), 60.8
(40-CH3O), 56.3 (3-CH3O), 56.1 (30,50-CH3O); MS (ESI, m/z): 315.7
[M+Na+], 607.2 [2M+Na+]; Anal. Calcd for C16H17FO4: C, 65.74; H,
5.86. Found: C, 65.58; H, 5.99.
4.1.6. N-Benzyl-3-(benzyloxy)-30,40,50-trimethoxybiphenyl-4-
carboxamide (6)
4.1.12. 1-(30,40,50,6-Tetramethoxybiphenyl-3-yl)ethanone (12)
White solid, yield 91%. Mp 114–116 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 7.02 (d, 1H, J = 8.1 Hz, 5-H), 7.96 (dd, 1H, J = 8.1,
2.3 Hz, 4-H), 7.94 (d, 1H, J = 2.3 Hz, 2-H), 6.73 (s, 2H, 20,60-H),
3.90 (s, 6H, 30,50-CH3O), 3.89 (s, 6H, 6,40-CH3O), 2.59 (s, 3H, 3-
Colorless oil, yield 90%. 1H NMR (300 MHz, CDCl3, TMS): d 8.34
(d, 1H, J = 8.3 Hz, 5-H), 7.32–7.17 (m, 12H, Ar–H), 6.73 (s, 2H, 20,60-
H), 5.21 (s, 2H, OCH2), 4.59 (d, 2H, J = 5.5 Hz, NHCH2), 3.92 (s, 6H,
30,50-CH3O), 3.90 (s, 3H, 40-CH3O); 13C NMR (75 MHz, CDCl3,

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COCH3); 13C NMR (100 MHz, CDCl3, TMS): d 196.8 (5-CO), 160.3 (C-
6), 152.9 (C-30/C-50), 137.7 (C-40), 133.0 (C-10), 131.2 (C-2), 130.7
(C-3), 130.3 (C-4), 129.8 (C-1), 110.6 (C-5), 107.0 (C-20/C-60), 60.9
(40-CH3O), 56.3 (30,50-CH3O), 55.9 (6-CH3O), 26.4 (3-COCH3); MS
(ESI, m/z): 317.8 [M+H+], 655.9 [2M+Na+]; Anal. Calcd for
3.93 (s, 6H, 30,50-CH3O), 3.90 (s, 3H, 40-CH3O); 13C NMR (100 MHz,
CDCl3, TMS): d 167.1 (CONH), 153.6 (C-30/C-50), 144.2 (C-1), 138.3
(C-40), 136.0 (C-10), 133.5 (C-4), 127.3 (C-3, C-5), 127.1 (C-2, C-6),
104.6 (C-20/C-60), 61.0 (40-CH3O), 56.3 (30,50-CH3O), 39.8, 31.8,
20.2, 13.8 (NHBu); MS (ESI, m/z): 344.2 [M+H+], 709.3 [2M+Na+];
Anal. Calcd for C20H25NO4: C, 69.95; H, 7.34; N, 4.08. Found: C,
69.87; H, 7.46; N, 4.02.
C18H20O5: C, 68.34; H, 6.37. Found: C, 68.44; H, 6.33.
4.1.13. 1-(30,40,50,6-Tetramethoxybiphenyl-3-yl)ethanol (13)
Colorless oil, yield 92%. 1H NMR (400 MHz, CDCl3, TMS): d 7.34–
7.32 (m, 2H, 2,4-H), 6.97 (d, 1H, J = 9.1 Hz, 5-H), 6.74 (s, 2H, 20,60-
H), 4.91 (q, 1H, J = 6.4 Hz, 3-CHOHCH3), 3.89 (s, 3H, 40-CH3O),
3.88 (s, 6H, 30,50-CH3O), 3.82 (s, 3H, 6-CH3O), 1.52 (d, 3H,
J = 6.4 Hz, 3-CHOHCH3); 13C NMR (100 MHz, CDCl3, TMS): d 155.8
(C-6), 152.8 (C-30/C-50), 138.2 (C-40), 134.0 (C-10), 137.3 (C-3),
130.7 (C-2), 128.1 (C-4), 125.6 (C-1), 111.4 (C-5), 107.0 (C-20/C-
60), 70.0 (3-CHOHCH3), 60.9 (40-CH3O), 56.2 (30,50-CH3O), 55.8 (6-
CH3O), 25.1 (3-CHOHCH3); MS (ESI, m/z): 319.1 [M+H+], 341.2
[M+Na+], 659.8 [2M+Na+]; Anal. Calcd for C18H22O5: C, 67.91; H,
6.97. Found: C, 67.80; H, 6.93.
4.1.19. N-Benzyl-30,40,50-trimethoxybiphenyl-4-carboxamide
(19)
White solid, yield 68%. Mp 119–121 °C; 1H NMR (300 MHz,
CDCl3, TMS):
d 7.86 (d, 2H, J = 8.2 Hz, 3,5-H), 7.61 (d, 2H,
J = 8.2 Hz, 2,6-H), 7.38–7.31(5H, Ar–H), 6.79 (s, 2H, 20,60-H), 4.69
(d, 2H, J = 5.7 Hz, NHCH2), 3.93 (s, 6H, 30,50-CH3O), 3.90 (s, 3H, 40-
CH3O); 13C NMR (75 MHz, CDCl3, TMS): d 167.3 (CONH), 153.9
(C-30/C-50), 144.8 (C-1), 138.5 (C-40), 136.2 (C-10), 133.3 (C-4),
129.1 (C-3, C-5), 128.2 (C-2, C-6), 104.8 (C-20/C-60), 61.3 (40-
CH3O), 56.6 (30,50-CH3O), 138.4, 128.0, 127.8, 127.5, 44.5 (NHBn);
MS (ESI, m/z): 378.4 [M+H+], 400.4 [M+Na+], 416.3 [M+K+], 375.8
[MꢀHꢀ]; Anal. Calcd for C23H23NO4: C, 73.19; H, 6.14; N, 3.71.
Found: C, 73.07; H, 6.03; N, 3.86.
4.1.14. 30,40,50-Trimethoxybiphenyl-3-ol (14)
White solid, yield 75%. Mp 98–101 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 7.27 (m, 1H, 5-H), 7.09 (m, 1H, 6-H), 7.03 (m, 1H,
2-H), 6.83 (dd, 1H, J = 8.1, 2.0 Hz, 4-H), 6.75 (s, 2H, 20,60-H), 3.89
(s, 3H, 40-CH3O), 3.88 (s, 6H, 30,50-CH3O); 13C NMR (100 MHz, CDCl3,
TMS): d 156.2 (C-3), 153.1 (C-30/C-50), 143.0 (C-1), 137.6 (C-40),
137.0 (C-10), 129.9 (C-5), 119.3 (C-6), 114.3 (C-2), 114.2 (C-4),
104.5 (C-20/C-60), 61.0 (40-CH3O), 56.2 (30,50-CH3O); MS (ESI, m/z):
259.7 [MꢀHꢀ], 519.6 [2MꢀHꢀ]; Anal. Calcd for C15H16O4: C,
69.22; H, 6.20. Found: C, 69.38; H, 6.05.
4.2. General procedure for the preparation of compounds 20–36
4.2.1. 3,40-Dihydroxy-30,50-dimethoxybiphenyl-4-carbaldehyde
(20)
MgI2 (0.88 g, 3.18 mmol) in dry ether (10 mL) was added to a
solution of 1 (0.2 g, 0.53 mmol) in CH2Cl2 (5 mL). The solvent was
removed under reduced pressure and the residuary solid mixture
was heated at 80 °C for 4 h. Then H2O (20 mL) and Na2S2O3 was
added and the mixture was poured into 5% hydrochloric acid
(10 mL), extracted with EtOAc (2 ꢁ 15 mL). The combined organic
phase was washed with saturated sodium bicarbonate solution
(20 mL) and saturated brine (20 mL), dried over MgSO4 and evapo-
ration in vacuo. The crude product was purified by silica gel flash
chromatography (P/E: 2/1) to give 20 as a white solid (0.12 g,
84%). Mp 60–62 °C; 1H NMR (400 MHz, CDCl3, TMS): d 11.12 (s,
1H, CHO), 9.90 (s, 1H, 3-OH), 7.58 (d, 1H, J = 8.0 Hz, 5-H), 7.20
(dd, 1H, J = 8.0, 1.5 Hz, 6-H), 7.17 (d, 1H, J = 1.5 Hz, 2-H), 6.85 (s,
2H, 20,60-H), 5.64 (s, 1H, 40-OH), 3.96 (s, 6H, 30,50-CH3O); 13C NMR
(100 MHz, CDCl3, TMS): d 195.8 (CHO), 162.0 (C-3), 150.0 (C-1),
147.5 (C-30/C-50), 134.0 (C-40), 130.8 (C-10), 121.0 (C-4), 119.4 (C-
5), 118.5 (C-6), 115.3 (C-2), 104.5 (C-20/C-60), 56.5 (30,50-CH3O);
MS (ESI, m/z): 275.8 [M+H+], 273.3 [MꢀHꢀ]; Anal. Calcd for
4.1.15. 30-(Benzyloxy)-3,4,5-trimethoxybiphenyl (15)
White solid, yield 77%. Mp 87–89 °C; 1H NMR (400 MHz, CDCl3,
TMS): d 7.47–7.14 (m, 8H, Ar–H), 6.96 (m, 1H, 40-H), 6.75 (s, 2H,
2,6-H), 5.13 (s, 2H, OBn), 3.90 (s, 6H, 3,5-CH3O), 3.89 (s, 3H,
13
4-CH3O); C NMR (100 MHz, CDCl3, TMS): d 159.1 (C-30), 153.4
(C-3/C-5), 142.9 (C-10), 137.9 (C-4), 137.0, 128.6, 128.0, 127.5,
70.2 (OBn), 136.9 (C-1), 129.7 (C-50), 119.8 (C-60), 114.2 (C-20),
113.3 (C-40), 104.6 (C-2/C-6), 60.9 (4-CH3O), 56.2 (3,5-CH3O); MS
(ESI, m/z): 373.1 [M+Na+], 723.5 [2M+Na+]; Anal. Calcd for
C22H22O4: C, 75.41; H, 6.33. Found: C, 75.56; H, 6.18.
4.1.16. 30,40,50-Trimethoxy-2-methylbiphenyl (16)
Light yellow oil, yield 86%. 1H NMR (300 MHz, CDCl3, TMS): d
7.27–7.24 (m, 4H, 3,4,5,6-H), 6.52 (s, 2H, 20,60-H), 3.90 (s, 3H, 40-
CH3O), 3.86 (s, 6H, 30,50-CH3O), 2.30 (s, 3H, 2-CH3); 13C NMR
(75 MHz, CDCl3, TMS): d 153.1 (C-30/C-50), 142.3 (C-40), 137.9 (C-
2), 137.1 (C-10), 135.7 (C-1), 130.6 (C-3), 129.8 (C-4), 127.6 (C-5),
126.0 (C-6), 106.7 (C-20/C-60), 61.2 (40-CH3O), 56.4 (30,50-CH3O),
20.8 (2-CH3); MS (ESI, m/z): 259.4 [M+H+], 281.4 [M+Na+]; Anal.
Calcd for C16H18O3: C, 74.39; H, 7.02. Found: C, 74.32; H, 7.11.
C15H14O5: C, 65.69; H, 5.15. Found: C, 65.76; H, 5.04.
4.2.2. 3,40-Dihydroxy-30,50-dimethoxybiphenyl-4-carboxylic acid
(21)
Light yellow solid, yield 86%. Mp 77–79 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 9.90 (s, 1H, 3-OH), 7.58 (d, 1H, J = 7.9 Hz, 5-H), 7.21
(dd, 1H, J = 7.9, 1.5 Hz, 6-H), 7.16 (d, 1H, J = 1.5 Hz, 2-H), 6.85 (s, 2H,
20,60-H), 5.69 (s, 1H, 40-OH), 3.96 (s, 6H, 30,50-CH3O); 13C NMR
(100 MHz, CDCl3, TMS): d 195.8 (COOH), 161.9 (C-3), 150.0 (C-30/
C-50), 147.5 (C-1), 136.0 (C-40), 134.0 (C-10), 130.7 (C-5), 119.3 (C-
6), 115.3 (C-4), 118.5 (C-2), 104.4 (C-20/C-60), 56.5 (30,50-CH3O);
MS (ESI, m/z): 290.9 [M+H+], 312.9 [M+Na+], 288.7 [MꢀHꢀ]; Anal.
Calcd for C15H14O6: C, 62.07; H, 4.86. Found: C, 61.94; H, 4.77.
4.1.17. 30,40,50-Trimethoxybiphenyl-4-carboxylic acid (17)
Light yellow solid, yield 79%. Mp 154–156 °C; 1H NMR
(400 MHz, CDCl3, TMS): d 8.14 (d, 2H, J = 8.0 Hz, 3,5-H), 7.62 (d,
2H, J = 8.0 Hz, 2,6-H), 6.80 (s, 2H, 20,60-H), 3.93 (s, 6H, 30,50-CH3O),
3.90 (s, 3H, 40-CH3O); 13C NMR (100 MHz, CDCl3, TMS): d 173.0
(COOH), 153.6 (C-30/C-50), 146.2 (C-1), 138.5 (C-40), 135.8 (C-10),
130.7 (C-4), 127.0 (C-2, C-3, C-5, C-6), 104.7 (C-20/C-60), 60.9 (40-
CH3O), 56.3 (30,50-CH3O); MS (ESI, m/z): 287.4 [MꢀHꢀ]; Anal. Calcd
for C16H16O5: C, 66.66; H, 5.59. Found: C, 66.49; H, 5.50.
4.2.3. Ethyl 3,40-dihydroxy-30,50-dimethoxybiphenyl-4-
carboxylate (22)
White solid, yield 81%. Mp 127–129 °C; 1H NMR (600 MHz,
CDCl3, TMS): d 10.91 (s, 1H, 3-OH), 7.87 (d, 1H, J = 8.4 Hz, 5-H),
7.08 (dd, 1H, J = 8.4, 1.8 Hz, 6-H), 7.16 (d, 1H, J = 1.8 Hz, 2-H),
6.84 (s, 2H, 20,60-H), 5.62 (s, 1H, 40-OH), 4.44 (q, 2H, J = 7.1 Hz,
COOCH2CH3), 3.96 (s, 6H, 30,50-CH3O), 1.44 (t, 3H, J = 7.1 Hz,
COOCH2CH3); 13C NMR (150 MHz, CDCl3, TMS): d 170.2 (COOEt),
4.1.18. N-Butyl-30,40,50-trimethoxybiphenyl-4-carboxamide (18)
White solid, yield 74%. Mp 116–118 °C; 1H NMR (400 MHz,
CDCl3, TMS):
d 7.83 (d, 2H, J = 8.5 Hz, 3,5-H), 7.60 (d, 2H,
J = 8.5 Hz, 2,6-H), 6.79 (s, 2H, 20,60-H), 3.49, 1.62, 1.43, 0.98 (NHBu),

K. Bao et al. / Bioorg. Med. Chem. 18 (2010) 6708–6714
6713
161.9 (C-3), 148.5 (C-1), 147.4 (C-30/C-50), 138.5 (C-40), 135.5 (C-10),
130.3 (C-5), 117.8 (C-6), 115.3 (C-4), 111.1 (C-2), 104.2 (C-20/C-60),
61.4 (OCH2CH3), 56.4 (30,50-CH3O), 14.2 (OCH2CH3); MS (ESI, m/z):
319.1 [M+H+], 341.1 [M+Na+], 316.9 [MꢀHꢀ]; Anal. Calcd for
150.8 (C-20), 148.4 (C-30), 147.0 (C-3/C-5), 134.8 (C-4), 130.1 (C-
10), 126.7 (C-1), 123.8 (C-50), 121.9 (C-60), 112.0 (C-40), 106.2 (C-
2/C-6), 56.4 (3,5-CH3O); MS (ESI, m/z): 301.6 [M+Na+], 277.4
[MꢀHꢀ]; Anal. Calcd for C15H15FO4: C, 64.74; H, 5.43. Found: C,
64.88; H, 5.51.
C
17H18O6: C, 64.14; H, 5.70. Found: C, 64.08; H, 5.87.
4.2.4. N-Benzyl-3,40-dihydroxy-30,50-dimethoxybiphenyl-4-
carboxamide (23)
4.2.10. 2-Fluoro-30,50-dimethoxybiphenyl-3,40-diol (29)
Colorless oil, yield 58%. 1H NMR (400 MHz, CDCl3, TMS): d 7.07
(m, 1H, 6-H), 6.99 (m, 2H, 4,5-H), 6.77 (s, 2H, 20,60-H), 3.93 (s, 6H,
30,50-CH3O); 13C NMR (100 MHz, CDCl3, TMS): d 149.5 (C-2),
147.0 (C-30/C-50), 144.2 (C-3), 134.9 (C-40), 129.7 (C-1), 126.5 (C-
10), 124.4 (C-5), 121.6 (C-6), 115.8 (C-4), 106.1 (C-20/C-ꢀ60), 56.4
(30,50-CH3O); MS (ESI, m/z): 287.5 [M+Na+], 263.3 [MꢀH ]; Anal.
Calcd for C14H13FO4: C, 63.63; H, 4.96. Found: C, 63.48; H, 4.82.
Light yellow solid, yield 79%. Mp 179–181 °C; 1H NMR
(600 MHz, CDCl3, TMS): d 12.43 (s, 1H, 3-OH), 7.39–7.36 (m, 6H,
Ar–H), 7.19 (d, 1H, J = 1.7 Hz, 2-H), 7.02 (dd, 1H, J = 8.3, 1.7 Hz, 6-
H), 6.82 (s, 2H, 20,60-H), 5.61 (s, 1H, 40-OH), 4.67 (d, 2H, J = 5.5 Hz,
NHCH2), 3.95 (s, 6H, 30,50-CH3O); 13C NMR (150 MHz, CDCl3,
TMS): d 168.1 (CONH), 162.0 (C-3), 147.3 (C-30/C-50), 144.5 (C-1),
137.5 (C-40), 135.4, 128.9, 128.8, 127.9, 43.7 (NHBn), 132.3 (C-10),
130.9 (C-5), 117.2 (C-6), 116.4 (C-2), 111.6 (C-4), 104.1 (C-20/C-
60), 56.4 (30,50-CH3O); MS (ESI, m/z): 402.9 [M+Na+], 378.7 [MꢀHꢀ];
Anal. Calcd for C22H21NO5: C, 69.64; H, 5.58; N, 3.69. Found: C,
69.74; H, 5.43; N, 3.85.
4.2.11. 1-(40-Hydroxy-30,50,6-trimethoxybiphenyl-3-yl)ethanone
(30)
White solid, yield 74%. Mp 144–145 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 7.01 (d, 1H, J = 7.0 Hz, 5-H), 7.96–7.94 (m, 2H, 2,4-
H), 6.74 (s, 2H, 20,60-H), 5.58 (s, 1H, 40-OH), 3.92 (s, 6H, 30,50-
CH3O), 3.90 (s, 3H, 6-CH3O), 2.59 (s, 3H, 3-COCH3); 13C NMR
(100 MHz, CDCl3, TMS): d 196.9 (5-CO), 160.4 (C-6), 146.8 (C-30/
C-50), 134.5 (C-40), 131.2 (C-10), 130.9 (C-2), 130.4 (C-3), 129.5 (C-
4), 128.6 (C-1), 110.6 (C-5), 106.7 (C-20/C-60), 56.5 (30,50-CH3O),
55.9 (6-CH3O), 26.4 (3-COCH3); MS (ESI, m/z): 303.7 [M+H+],
627.7 [2M+Na+], 301.7 [MꢀHꢀ]; Anal. Calcd for C17H18O5: C,
67.54; H, 6.00. Found: C, 67.59; H, 6.08.
4.2.5. 30,50-Dimethoxy-4-methylbiphenyl-3,40-diol (24)
Colorless oil, yield 91%. 1H NMR (400 MHz, CDCl3, TMS): d 7.16
(d, 1H, J = 7.8 Hz, 5-H), 7.03 (dd, 1H, J = 7.8, 1.3 Hz, 6-H), 6.97 (d,
1H, J = 1.3 Hz, 2-H), 6.77 (s, 2H, 20,60-H), 5.50 (s, 1H, 3-OH), 4.72
(s, 1H, 40-OH), 3.94 (s, 6H, 30,50-CH3O), 2.28 (s, 3H, 4-CH3); 13C
NMR (100 MHz, CDCl3, TMS): d 154.1 (C-3), 147.3 (C-30/C-50),
141.0 (C-1), 134.5 (C-40), 132.6 (C-10), 131.3 (C-5), 122.5 (C-4),
119.3 (C-6), 113.5 (C-2), 104.0 (C-20/C-60), 56.4 (30,50-CH3O), 15.4
(4-CH3); MS (ESI, m/z): 261.4 [M+H+], 259.2 [MꢀHꢀ]; Anal. Calcd
for C15H16O4: C, 69.22; H, 6.20. Found: C, 69.36; H, 6.29.
4.2.12. 50-(1-Hydroxyethyl)-20,3,5-trimethoxybiphenyl-4-ol (31)
White solid, yield 71%. Mp 103–104 °C; 1H NMR (300 MHz,
CDCl3, TMS): d 6.96 (d, 1H, J = 9.3 Hz, 30-H), 7.32 (m, 2H, 40,60-H),
6.76 (s, 2H, 2,6-H), 4.92 (q, 1H, J = 6.5 Hz, 50-CHOHCH3), 3.91 (s,
6H, 3,5-CH3O), 3.82 (s, 3H, 20-CH3O), 1.52 (d, 3H, J = 6.5 Hz, 50-
4.2.6. 30,50-Difluoro-3,5-dimethoxybiphenyl-4-ol (25)
Light yellow solid, yield 88%. Mp 60–61 °C; 1H NMR (300 MHz,
CDCl3, TMS): d 7.04 (m, 2H, 20,60-H), 6.80 (m, 1H, 40-H), 6.75 (s, 2H,
2,6-H), 5.65 (s, 1H, 4-OH), 3.95 (s, 6H, 3,5-CH3O); 13C NMR
(75 MHz, CDCl3, TMS): d 163.6 (C-30/C-50), 147.7(C-3/C-5), 145.0
(C-10), 135.6 (C-4), 135.1 (C-1), 109.9 (C-20/C-60), 104.2 (C-2/C-6),
102.3 (C-40), 56.7 (3, 5-CH3O); MS (ESI, m/z): 267.6 [M+H+], 289.6
[M+Na+], 265.1 [MꢀHꢀ]; Anal. Calcd for C14H12F2O3: C, 63.16; H,
4.54. Found: C, 63.05; H, 4.39.
13
CHOHCH3); C NMR (75 MHz, CDCl3, TMS): d 155.8 (C-20), 146.7
(C-3/C-5), 138.1 (C-4), 134.2 (C-50), 130.8 (C-1), 129.5 (C-60),
128.0 (C-40), 125.3 (C-10), 111.4 (C-30), 106.6 (C-2/C-6), 70.0 (50-
CHOHCH3), 56.4 (3,5-CH3O), 55.8 (20-CH3O), 25.1 (50-CHOHCH3);
MS (ESI, m/z): 327.9 [M+Na+], 632.2 [2M+Na+], 303.5 [MꢀHꢀ];
Anal. Calcd for C17H20O5: C, 67.09; H, 6.62. Found: C, 66.93; H, 6.53.
4.2.13. 30,50-Dimethoxybiphenyl-3,40-diol (32)
4.2.7. 20,40-Difluoro-3,5-dimethoxybiphenyl-4-ol (26)
White solid, yield 76%. Mp 121–124 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 7.29 (m, 1H, 5-H), 7.12 (m, 1H, 2-H), 7.02 (m, 1H,
6-H), 6.79 (s, 2H, 20,60-H), 6.78 (m, 1H, 4-H), 3.95 (s, 6H, 30,50-
CH3O); 13C NMR (100 MHz, CDCl3, TMS): d 156.7 (C-3), 147.3 (C-
30/C-50), 143.3 (C-1), 134.7 (C-40), 132.4 (C-10), 129.9 (C-5), 119.6
(C-6), 114.0 (C-4), 113.8 (C-2), 104.2 (C-20/C-60), 56.4 (30,50-
CH3O); MS (ESI, m/z): 269.4 [M+Na+], 245.1 [MꢀHꢀ]; Anal. Calcd
for C14H14O4: C, 68.28; H, 5.73. Found: C, 68.16; H, 5.57.
Light yellow solid, yield 80%. Mp 54–57 °C; 1H NMR (300 MHz,
CDCl3, TMS): d 7.38 (m, 1H, 60-H), 6.95–6.84 (m, 2H, 30,50-H), 6.71
(d, 2H, 2,6-H), 5.67 (s, 1H, 4-OH), 3.91 (s, 9H, 3,5-CH3O); 13C
NMR (75 MHz, CDCl3, TMS): d 162.3 (C-40), 159.9 (C-20), 147.3
(C-3/C-5), 135.0 (C-4), 131.5 (C-60), 126.4 (C-1), 125.7 (C-10),
111.7 (C-50), 106.2 (C-2/C-6), 104.8 (C-30), 56.7 (3,5-CH3O); MS
(ESI, m/z): 267.3 [M+H+], 289.3 [M+Na+], 555.3 [2M+Na+]; Anal.
Calcd for C14H12F2O3: C, 63.16; H, 4.54. Found: C, 63.01; H, 4.43.
4.2.14. 3,5-Dimethoxy-20-methylbiphenyl-4-ol (33)
4.2.8. 30,40-Difluoro-3,5-dimethoxybiphenyl-4-ol (27)
Light yellow oil, yield 90%. 1H NMR (300 MHz, CDCl3, TMS): d
7.24–7.23 (m, 4H, 30,40,50,60-H), 6.53 (s, 2H, 2,6-H), 3.88 (s, 6H,
3,5-CH3O), 2.28 (s, 3H, 20-CH3); 13C NMR (75 MHz, CDCl3, TMS): d
147.0 (C-3/C-5), 142.4 (C-4), 135.8 (C-20), 134.0 (C-1), 133.5 (C-
10), 130.6 (C-30), 130.0 (C-40), 127.5 (C-50), 126.0 (C-60), 106.4 (C-
2/C-6), 56.6 (3,5-CH3O), 20.8 (20-CH3); MS (ESI, m/z): 245.3
[M+H+], 267.3 [M+Na+], 511.4 [2M+Na+]; Anal. Calcd for
White solid, yield 85%. Mp 70–72 °C; 1H NMR (300 MHz, CDCl3,
TMS): d 7.35–7.28 (m, 1H, 20-H), 7.25–7.14 (m, 2H, 50,60-H), 6.71 (s,
2H, 2,6-H), 5.58 (s, 1H, 4-OH), 3.95 (s, 6H, 3,5-CH3O); 13C NMR
(75 MHz, CDCl3, TMS): d 150.8 (C-30), 149.8 (C-40), 147.7 (C-3/C-
5), 138.9 (C-10), 135.2 (C-4), 131.1 (C-1), 123.0 (C-60), 117.7 (C-
20), 116.0 (C-50), 104.2 (C-2/C-6), 56.7 (3,5-CH3O); MS (ESI, m/z):
267.3 [M+H+], 289.3 [M+Na+], 264.6 [MꢀHꢀ]; Anal. Calcd for
C14H12F2O3: C, 63.16; H, 4.54. Found: C, 63.06; H, 4.36.
C15H16O3: C, 73.75; H, 6.60. Found: C, 73.62; H, 6.76.
4.2.15. 40-Hydroxy-30,50-dimethoxybiphenyl-4-carboxylic acid
(34)
4.2.9. 20-Fluoro-3,30,5-trimethoxybiphenyl-4-ol (28)
White solid, yield 67%. Mp 100–102 °C; 1H NMR (400 MHz,
CDCl3, TMS): d 7.10 (m, 1H, 50-H), 6.99 (m, 1H, 60-H), 6.93 (m, 1H,
40-H), 6.78 (s, 2H, 2,6-H), 5.57 (s, 1H, 4-OH), 3.92 (s, 6H, 3,5-
CH3O), 3.93 (s, 3H, 30-CH3O); 13C NMR (100 MHz, CDCl3, TMS): d
Light yellow solid, yield 88%. Mp 228–230 °C; 1H NMR
(600 MHz, CD3SOCD3, TMS): d 8.05 (d, 2H, J = 8.2 Hz, 3,5-H), 7.63
(d, 2H, J = 8.2 Hz, 2,6-H), 6.86 (s, 2H, 20,60-H), 3.95 (s, 6H, 30,50-
CH3O); 13C NMR (150 MHz, CD3SOCD3, TMS): d 172.0 (COOH),

6714
K. Bao et al. / Bioorg. Med. Chem. 18 (2010) 6708–6714
148.2 (C-30/C-50), 144.9 (C-1), 136.2 (C-40), 131.7 (C-10), 129.7 (C-2,
C-6), 129.6 (C-4), 126.1 (C-3, C-5), 104.4 (C-20/C-60), 56.1 (30,50-
CH3O); MS (ESI, m/z): 274.9 [M+H+], 296.9 [M+Na+], 273.3 [MꢀHꢀ];
Anal. Calcd for C15H14O5: C, 65.69; H, 5.15. Found: C, 65.81; H, 5.05.
MeOH solution, a 20% MeOH solution was added to a blank solu-
tion. Before and after incubation, the amount of dopachrome pro-
duced in the reaction mixture was measured at 492 nm in the
microplate reader. The percentage of the inhibition of tyrosinase
activity was calculated by the following equation: inhibition
(%) = [(A ꢀ B) ꢀ (C ꢀ D)]/(A ꢀ B) ꢁ 100, where A is absorbance of
blank solution after incubation, B is absorbance of blank solution
before incubation, C is absorbance of sample solution after incuba-
tion, and D is absorbance of sample solution before incubation.
The inhibitory kinetics of the selected compounds 21, 23, 34,
4.2.16. N-Butyl-40-hydroxy-30,50-dimethoxybiphenyl-4-
carboxamide (35)
Light yellow solid, yield 79%. Mp 136–139 °C; 1H NMR
(300 MHz, CDCl3, TMS): d 7.86 (d, 2H, J = 8.3 Hz, 3,5-H), 7.69 (d,
2H, J = 8.3 Hz, 2,6-H), 6.93 (s, 2H, 20,60-H), 3.92 (s, 6H, 30,50-CH3O),
3.40, 1.63, 1.44, 0.99 (NHBu); 13C NMR (75 MHz, CDCl3, TMS): d
167.5 (CONH), 147.7 (C-30/C-50), 144.6 (C-1), 135.4 (C-40), 133.4
(C-10), 131.9 (C-4), 127.6 (C-3, C-5), 127.2 (C-2, C-6), 104.4 (C-20/
C-60), 56.7 (30,50-CH3O), 40.1, 32.1, 20.5, 14.1 (NHBu); MS (ESI, m/
z): 681.9 [2M+Na+], 328.5 [MꢀHꢀ]; Anal. Calcd for C19H23NO4: C,
69.28; H, 7.04; N, 4.25. Found: C, 69.12; H, 6.95; N, 4.33.
and 35 on mushroom tyrosinase for the oxidation of L-tyrosine
was determined from Lineweaver–Burk double reciprocal plots.
The velocity equation for the competitive inhibition in reciprocal
form is: 1/V = Km/Vmax(1 + [I]/Ki)1/[S]+1/Vmax
.
The inhibition
constants (Ki) of the competitive inhibitors were calculated by
the following equation: Kmapp = Km[1 + ([I]/Ki)] where Kmapp is the
apparent Km, in the presence of an inhibitor.
4.2.17. N-Benzyl-40-hydroxy-30,50-dimethoxybiphenyl-4-
carboxamide (36)
Acknowledgment
Light yellow oil, yield 86%. 1H NMR (300 MHz, CDCl3, TMS): d
7.85 (d, 2H, J = 8.5 Hz, 3,5-H), 7.60 (d, 2H, J = 8.5 Hz, 2,6-H), 7.39–
7.37 (5H, Ar–H), 6.81 (s, 2H, 20,60-H), 4.69 (d, 2H, J = 5.7 Hz, NHCH2),
3.97 (s, 6H, 30,50-CH3O); 13C NMR (75 MHz, CDCl3, TMS): d 162.6
(CONH), 155.7 (C-30/C-50), 147.8 (C-1), 138.5 (C-40), 137.7 (C-10),
132.9 (C-4), 129.1 (C-3, C-5), 127.8 (C-2, C-6), 104.5 (C-20/C-60),
56.8 (30,50-CH3O), 135.4, 128.3, 128.0, 127.2, 46.2 (NHBn); MS
(ESI, m/z): 362.1 [MꢀHꢀ]; Anal. Calcd for C22H21NO4: C, 72.71; H,
5.82; N, 3.85. Found: C, 72.64; H, 5.76; N, 3.68.
This work was supported by the National Science Foundation
for Post-doctoral Scientists of China (Grant No. 20080440774).
References and notes
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4.3. Tyrosinase assay
This assay was performed according to the procedure of Mason
et al. with slight modifications, using
Forty microliters of mushroom tyrosinase solution (100 units/
mL), 40 L of 0.1 mg/mL -tyrosine solution in phosphate-buffered
saline (PBS) solution (25 mM, pH 6.8), 80 L of phosphate-buffered
saline (PBS) solution (25 mM, pH 6.8), and 40 L of sample in 20%
L-tyrosine as a substrate.
l
L
10. Dai, Y.; Zhou, G. X.; Kurihara, H.; Ye, W. C.; Yao, X. S. J. Nat. Prod. 2006, 69, 1022.
11. Bao, K.; Zhang, L.; Zhang, W. G.; Cheng, M. S.; Yao, X. S. Org. Biomol. Chem. 2009,
7, 5084.
l
l
12. Nishioka, H.; Nagasawa, M.; Yoshida, K. Synthesis 2000, 2, 243.
13. Punna, S.; Meunier, S.; Finn, M. G. Org. Lett. 2004, 6, 2777.
MeOH solution were added to a 96-well microplate. The assay mix-
ture was incubated at 37 °C for 30 min. Instead of a sample in 20%