Synthesis and biological activity of halophenols as potent antioxidant and cytoprotective agents

Article abstract of DOI:10.1016/j.bmcl.2010.05.068

A series of new bromophenols and chlorophenols were prepared by a practical route. The in vitro antioxidative activity of the halophenols was evaluated by the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging assay, and their cytoprotective activity was also tested on hydrogen peroxide (H₂O₂)-induced injury in human umbilical vein endothelial cells (HUVEC). All halophenols tested displayed moderate to good DPPH radical-scavenging activity, and two bromophenols, 2,3′-dibromo-4,5,6′-trihydroxydiphenylmethanone (16c) and 2,3-dibromo-4,5-dihydroxydiphenylmethanone (17c) exhibited high protective activity against H₂O₂-induced injury in HUVEC with EC₅₀ values of 0.4 and 0.8 μM, respectively. The preliminary structure-activity relationships of these compounds were also investigated in order to determine the essential structures required for their bioactivities.

Full text of DOI:10.1016/j.bmcl.2010.05.068

Bioorganic & Medicinal Chemistry Letters 20 (2010) 4132–4134
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological activity of halophenols as potent antioxidant
and cytoprotective agents
Wanyi Zhao a, Xiue Feng a, Shurong Ban a, Wenhan Lin b, Qingshan Li a,b,
*
a School of Pharmaceutical Science, Shanxi Medical University, Taiyuan 030001, China
b State Key Laboratory of Natural and Biomimetic Drugs, Peking University, Beijing 100083, China
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of new bromophenols and chlorophenols were prepared by a practical route. The in vitro antiox-
idative activity of the halophenols was evaluated by the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-
scavenging assay, and their cytoprotective activity was also tested on hydrogen peroxide (H2O2)-induced
injury in human umbilical vein endothelial cells (HUVEC). All halophenols tested displayed moderate to
good DPPH radical-scavenging activity, and two bromophenols, 2,30-dibromo-4,5,60-trihydroxydiphenyl-
methanone (16c) and 2,3-dibromo-4,5-dihydroxydiphenylmethanone (17c) exhibited high protective
activity against H2O2-induced injury in HUVEC with EC50 values of 0.4 and 0.8 lM, respectively. The pre-
liminary structure–activity relationships of these compounds were also investigated in order to deter-
mine the essential structures required for their bioactivities.
Received 11 February 2010
Revised 16 May 2010
Accepted 18 May 2010
Available online 8 June 2010
Keywords:
Bromophenols
Halophenols
DPPH radical-scavenging activity
Cytoprotective activity
HUVEC
Ó 2010 Elsevier Ltd. All rights reserved.
Bromophenols frequently isolated from various marine algae,
ascidians, and sponges have attracted much research interest due
to their wide spectrum of bioactivities including antioxidative,1,2
antithrombotic,3 antimicrobial,4,5 anti-inflammatory,6 enzyme inhi-
bition,7 cytotoxic,4,8 and feeding deterrent9 activities. Two natural
bromophenols with diphenyl skeletons, bis(2,3,6-tribromo-4,5-
dihydroxyphenyl)methane (1)2,7 and (+)-3-(2,3-dibromo-4,5-dihy-
droxyphenyl)-4-bromo-5,6-dihydroxy-1,3-dihydroisobenzofuran
(2),3,8 exhibited good in vitro antioxidative and in vivo antithrom-
botic activity, respectively (Fig. 1). However, their corresponding
structure–activity relationships are still unclear.
Encouraged by these interesting pharmacological activities, we
synthesized analogs of the diphenyl bromophenols. In this paper,
we report on the synthesis and in vitro activities of a series of brom-
ophenols and chlorophenols along with their primary structure–
activity relationships.
The general procedure for the synthesis of the halophenols
started with preparation of the methoxylated diphenyl intermedi-
ates 11–14 (Scheme 1). Compounds 7–10 were obtained by treating
3–6 with dry SOCl2 and a catalytic amount of DMF, and Friedel–
Crafts acylation of 7–10 with 1,2-dimethoxybenzene catalyzed by
AlCl3 which gave 11–14,10,11 in which methoxy groups were used
as hydroxy protecting groups.
yield of 15a.12 Reduction of the benzylic ketone under standard con-
ditions then afforded 15b.13,14 Demethylation of the methoxy
groupsin 15a and 15b with BBr3 gave two bromophenols, 2,30-dibro-
mo-4,40,5-trihydroxydiphenylmethanone (15c) and 2,30-dibromo-
4,40,5-trihydroxydiphenylmethane (15d), respectively.15
To investigate the positionaleffects of hydroxylgroups in the aro-
matic ring on bioactivity, two analogs of 15c and 15d, 2,30-dibromo-
4,5,60-trihydroxydiphenylmethanone (16c) and 2,30-dibromo-
4,5,60-trihydroxydiphenylmethane (16d), were prepared in the
same manner (Scheme 3).
In order to examine the changes in bioactivity resulting from the
different numbers and positions of the bromine atoms attached to
the phenol ring, two monobrominated analogs 2-bromo-4,5-
dihydroxydiphenylmethanone (18c), 2-bromo-4,5-dihydroxydiphe
nylmethane (18d), and two dibrominated analogs 2,3-dibromo-4,5-
dihydroxydiphenylmethanone (17c), 2,3-dibromo-4,5-dihydroxydi
phenylmethane (17d) were synthesized by the same method de-
scribed in Scheme 2. The number of brominated atoms was deter-
mined by the equivalents of 13 and bromine, reaction time and
temperature (Scheme 4).
Asshownin Scheme2, thebrominationof 11with 2.2equivalents
of bromine in acetic acid at room temperature provided an excellent
* Corresponding author. Tel.: +86 351 4690322; fax: +86 351 4690114.
E-mail address: qingshanl@yahoo.com (Q. Li).
Figure 1. Bioactive bromophenols.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.05.068

W. Zhao et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4132–4134
4133
Scheme 1. Reagents and conditions: (a) SOCl2, DMF, reflux, 5 h; (b) AlCl3, CH2Cl2, rt, 2 h, 80–85%.
Scheme 2. Reagents and conditions: (a) Br2 (2.2 equiv), AcOH, rt, 4 h, 95%; (b)
LiAlH4, AlCl3, reflux, 2 h, 61%; (c) BBr3 (10 equiv), CH2Cl2, ꢀ78 °C to rt, 4 h, 72–75%.
Scheme 5. Reagents and conditions: (a) Br2 (2.2 equiv), AcOH, rt, 4 h, 91%; (b)
SO2Cl2 (2.5 equiv), CH2Cl2, reflux, 6 h, 89%; (c) LiAlH4, AlCl3, reflux, 2 h, 60–62%; (d)
BBr3 (10 equiv), CH2Cl2, ꢀ78 °C to rt, 4 h, 65–67%.
Chlorination of 14 with SO2Cl2 refluxing in CH2Cl2 provided 20a in
good yield. Two chlorophenols, 2,6-dichloro-3,30,4,40,5-penta-
hydroxydiphenylmethanone (20c) and 2,6-dichloro-3,30,4,40,5-pen-
tahydroxydiphenylmethane (20d) were prepared after reduction
and deprotection, and the bromophenols, 2,6-dibromo-3,30,4,40,5-
pentahydroxydiphenylmethanone (19c) and 2,6-dibromo-3,30,4,
40,5-pentahydroxydiphenylmethane (19d) were also obtained by
the same method.
In the above reactions, regioselective halogenation of 11, 12, 13,
and 14 gave 15a, 16a, 17a, 18a, 19a, and 20a. The substitution num-
ber of halogen atoms and the substitution position were determined
by analysis of ESI-MS, MS–MS, 1H NMR, and 13C NMR spectra.
Chlorination of 11–13 were also undertaken but gave a mixture
of mono-chlorinated and poly-chlorinated phenols. In order to ob-
tain more chlorophenols, we used o-, m-, and p-chlorobenzoyl
chlorides as starting materials, and six chlorophenols were pre-
pared as described above. The corresponding chlorophenols were
identified as 21c–23c16 and 21d–23d (Scheme 6).
Scheme 3. Reagents and conditions: (a) Br2 (2.2 equiv), AcOH, rt, 4 h, 84%; (b)
LiAlH4, AlCl3, reflux, 2 h, 52%; (c) BBr3 (10 equiv), CH2Cl2, ꢀ78 °C to rt, 4 h, 70–73%.
As shown in Table 1, some of the halophenols were subjected to
in vitro antioxidative activity testing using the DPPH radical-scav-
enging assay as previously reported.17 In addition, a number of the
intermediates with methoxy groups were also investigated in or-
der to determine their SARs. All the halophenols displayed moder-
ate to good antioxidative activity, while the intermediates showed
no activity, this demonstrated that the hydroxyl groups were
essential for the antioxidative activity of the halophenols.
All the halophenols and some of the intermediates were subse-
quently evaluated for their cytoprotective effects on H2O2-induced
injury in HUVEC.18 Some of the bromophenols and chlorophenols
exhibited varying degrees of moderate to good activity, while the
intermediates displayed no activity. Two bromophenols, 2,30-dibro-
mo-4,5,60-trihydroxydiphenylmethanone (16c) and 2,3-dibromo-
4,5-dihydroxydiphenylmethanone (17c) exhibited high cytoprotec-
Scheme 4. Reagents and conditions: (a) Br2 (2.2 equiv), AcOH, rt, 24 h, 86%; (b) Br2
(1.1 equiv), AcOH, rt, 4 h, 91%; (c) LiAlH4, AlCl3, reflux, 2 h, 50–55%; (d) BBr3
(10 equiv), CH2Cl2, –78 °C to rt, 4 h, 69–72%.
We subsequently focused on developing the SARs with respect to
different halogen atoms. Halophenols with bromine or chlorine
atoms on the same position were prepared according to Scheme 5.
tive activity with EC50 values of 0.4 and 0.8
lM, respectively, and
three chlorophenols showed moderate activity.

4134
W. Zhao et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4132–4134
Scheme 6. Reagents and conditions: (a) AlCl3, CH2Cl2, rt, 2 h, 85–90%; (b) LiAlH4, AlCl3, reflux, 2 h, 65–70%; (c) BBr3 (10 equiv), CH2Cl2, ꢀ78 °C to rt, 4 h, 75–86%.
Table 1
DPPH radical-scavenging assay. All the halophenols were found
to display moderate to good activity. The cytoprotective activity
of the halophenols on H2O2-induced injury in HUVEC was also
determined, and some of the bromophenols and chlorophenols
exhibited promising activity. Among these, two bromophenols,
2,30-dibromo-4,5,60-trihydroxydiphenylmethanone (16c) and 2,3-
dibromo-4,5-dihydroxydiphenylmethanone (17c) had high protec-
tive activity against H2O2-induced injury in HUVEC, with EC50 val-
DPPH radical-scavenging activity and cytoprotective activity against H2O2-induced
injury in HUVEC
Compound
DPPH
Cytoprotective
scavenging activity
activity
a
b
IC50
(
lM)
EC50
(lM)
13
15a–20a
23a
15b–23b
15c
16c
17c
18c
19c
20c
15d
16d
17d
18d
19d
20d
21c
22c
NA
NA
NA
NA
61.6
79.6
87.7
84.0
96.2
87.3
87.4
88.5
88.0
NA
NA
NA
NA
2.0
0.4
0.8
11.5
NA
NA
13.0
1.1
3.9
12.2
NA
NA
NA
5.0
8.9
NA
NA
8.8
NDc
18.0
ues of 0.4 and 0.8 lM, respectively. Further in vivo tests of these
compounds are now under way.
Acknowledgments
The authors thank Professor Guanhua Du, Dr. Lianhua Fang, and
Dr. Guorong He for the in vitro assay of the compounds. This work
was financially supported by the National ‘863’ Program of China
(No. 2006AA09Z446), by the State Key Laboratory of Natural and
Biomimetic Drugs, Peking University, by the Program for the Top
Young and Middle-aged Innovative Talents of Higher Learning
Institutions of Shanxi, and by the Shanxi Province Foundation for
Oversea Returnees.
163.8
84.2
102.5
112.9
109.7
102.8
130.9
222.9
202.5
169.1
NDc
23c
21d
22d
23d
BHT
Quercetin
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.05.068.
a
b
c
Values are means of three experiments; NA, not active at 100
Values are means of three experiments; NA, not active at 5
ND, not detected.
lg/mL.
lg/mL.
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It should be noted that compounds 19c, 20c, 19d, and 20d with
more hydroxyl groups than the other halophenols displayed no
activity, this showed that the cytoprotective activity of the hal-
ophenols was not directly related to the number of hydroxyl
groups. The dibrominated phenolic compounds, 17c and 17d dis-
played better activity than the monobrominated phenols, 18c
and 18d. The o-chlorinated phenolic compounds 21c and 21d
showed no activity, while some of the m- and p-chlorophenols dis-
played moderate protective effects against H2O2-induced injury in
HUVEC.
From these findings, we can conclude that hydroxyl groups on
the diphenyl backbone are essential for the in vitro antioxidative
activities of these compounds, and the presence of one or more
halogen atoms on the phenol ring is also necessary. The number
and position of the hydroxyl groups and halogen atoms may influ-
ence the potency of activity.
In summary, a number of new halophenols were synthesized
and evaluated for their in vitro antioxidative activity using the