Design, synthesis and biological evaluation of novel mansonone e derivatives prepared via CuAAC click chemistry as topoisomerase II inhibitors

Article abstract of DOI:10.1016/j.ejmech.2013.07.011

Two series of novel C-9 chloro- and bromo-substituted mansonone E derivatives with triazole moieties at the C-3 position were prepared by using copper-catalysed azide-alkyne cycloaddition click chemistry. These compounds were found as potent inhibitors of

Full text of DOI:10.1016/j.ejmech.2013.07.011

Accepted Manuscript
Design, synthesis and biological evaluation of novel mansonone E derivatives
prepared via CuAAC click chemistry as topoisomerase II inhibitors
Zhi-Hong Huang, Shi-Tian Zhuo, Chun-Yan Li, Hua-Ting Xie, Ding Li, Jia-Heng Tan,
Tian-Miao Ou, Zhi-Shu Huang, Lian-Quan Gu, Shi-Liang Huang
PII:
S0223-5234(13)00459-5
10.1016/j.ejmech.2013.07.011
EJMECH 6298
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 26 February 2013
Revised Date: 18 July 2013
Accepted Date: 24 July 2013
Please cite this article as: Z.-H. Huang, S.-T. Zhuo, C.-Y. Li, H.-T. Xie, D. Li, J.-H. Tan, T.-M. Ou, Z.-
S. Huang, L.-Q. Gu, S.-L. Huang, Design, synthesis and biological evaluation of novel mansonone E
derivatives prepared via CuAAC click chemistry as topoisomerase II inhibitors, European Journal of
Medicinal Chemistry (2013), doi: 10.1016/j.ejmech.2013.07.011.
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ACCEPTED MANUSCRIPT
Graphical abstract
Design, synthesis and biological evaluation of novel mansonone E derivatives
prepared via CuAAC click chemistry as topoisomerase II inhibitors
Zhi-Hong Huang, Shi-Tian Zhuo, Chun-Yan Li, Hua-Ting Xie, Ding Li, Jia-Heng Tan,
Tian-Miao Ou, Zhi-Shu Huang, Lian-Quan Gu, Shi-Liang Huang*
Two series of mansonone E derivates 9-Cl and 9-Br were synthesized through introducing
triazoles at position 3 through a CuAAC click chemistry approach, their cytotoxicities and
topoisomerases inhibition were evaluated.

ACCEPTED MANUSCRIPT
Title Page
Design, synthesis and biological evaluation of novel mansonone E derivatives
prepared via CuAAC click chemistry as topoisomerase II inhibitors
Zhi-Hong Huang, Shi-Tian Zhuo, Chun-Yan Li, Hua-Ting Xie, Ding Li, Jia-Heng Tan,
Tian-Miao Ou, Zhi-Shu Huang, Lian-Quan Gu, Shi-Liang Huang*
School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006, People’s
Republic of China
*To whom correspondence should be addressed. Tel.: 8620- 39943052, Fax.: 8620-39943056,
e-mail: lsshsl@mail.sysu.edu.cn.
Abstract
Two series of novel C-9 chloro- and bromo-substituted mansonone E derivatives with triazole
moieties at the C-3 position were prepared by using copper-catalysed azide-alkyne
cycloaddition click chemistry. These compounds were found as potent inhibitors of
topoisomerase II (Topo II) and topoisomerase I (Topo I). The TopoII-mediated pBR322 DNA
relaxation and cleavage assay showed that the derivatives might act as catalytic inhibitors.
Their cytotoxic activitives against A549, HL-60, K562 and HeLa cells were evaluated,
indicating that these compounds were potent antitumor agents. Their structure activity
relationships and molecular docking study revealed that the substituents of the triazole were
particularly important for cytotoxicity.
Keywords:
Mansonone E
Antitumor
Click chemistry
Topoisomerase
Cytotoxic activity
Abbreviations:
HPLC: High Performance Liquid Chromatography
HRMS: High Resolution Mass Spectrometer
MTT: methyl thiazolyl tetrazolium
NMR: nuclear magnetic resonance
Topo: topoisomerase
ME: mansonone E
MF: mansonone F
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1
. Introduction
Quinone-containing compounds have attracted a great deal of attention since the National
Cancer Institute initially identified quinone moiety as a pharmacophoric element of potent
antitumor compounds by mass screening [1]. 1,2- and 1,4-Naphthoquinones are widely
distributed in nature and have important physiological functions in animals and plants.
Several reports have shown that fused furan or pyran rings are essential elements for the
cytotoxicities of 1,2- and 1,4-naphthoquinones to cancer cell lines, and other studies have
revealed that 1,2-naphthoquinones exhibit a relatively more effective activity [2, 3]. Many
derivatives of 1,2-naphthoquinones have been used as potent antitumor drugs [4]. Some
illustrative
examples
of
antitumoral
1,2-naphthoquinones
(Figure1)
are
furano-1,2-naphthoquinone [5, 6], mansonones [3, 7, 8], β-lapachone [9-12], duinnones [13],
rhinacanthone [14, 15] and biflorin [16-18]. Among these compounds, β-lapachone has
been found to inhibit several human cancer cell lines by DNA repair enzymes [19]. Studies
have also shown that β-lapachone is a potent antitumor agent. However, few studies have
reported mansonone E (ME) and F (MF), two naturally occurring sesquiterpene o-quinones
that contain a fused pyran ring with different orientations from those of β-lapachone and
rhinacanthone.
Several studies have shown that ME, MF (Figure 1) and their analogues exihibit biological
activities, such as anti-bacterial [20], anti-MRSA [21] and anti-proliferative effects [3, 7, 8].
Structure-activity relationship (SAR) studies have also shown that 3- and 9-substituted MF
derivatives possess potential cytotoxic and topoisomerase (Topo) inhibitory activity [3, 20].
However, the exact inhibitory mechanism remains unknown. ME shows better cytotoxicity
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than MF, although the difference is only at the 2,3-position [8]. In the present study, the
mechanism of ME was investigated and novel analogues of ME were prepared to exhibit
more effective activities
Our previous study showed that MF analogues containing chlorine or bromine at C-9
exhibited better cytotoxic activity than 9-alkyl or aryl substituted derivatives [3]. Therefore,
these halogenated analogues were used to optimise the C-9 of ME. Click chemistry [22],
particularly the copper-catalyzed azide–alkyne cycloaddition (CuAAC), has been widely
applied in drug discovery and pharmaceutical sciences [23]. 1,2,3-Triazole, which has been
very useful in lead optimisation of natural products and their analogues, is formed and can
function as a bioisostere and a linker [24, 25]. In the present study, two series of ME
derivatives, including 9-chloro-3-triazole and 9-bromo-3-triazole derivatives were designed
and synthesized (Figure 2) by click chemistry approach. Their cytotoxicities against four
cancer cell lines, in vitro inhibition of human Topo I and II, molecular docking and a
preliminary analysis of SARs were performed. Our results provide insight into the mechanism
of the cytotoxicity of these types of compounds.
2
. Chemistry
Two series of ME derivatives were designed and synthesised to investigate the effect of C-9
halogenation and C-3 substitution on ME. Fourteen general alkynes were also introduced by
CuAAC click chemistry.
Compounds 5 were prepared based on a five-step procedure according to a previously
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described method with minor modifications (Scheme 1) [26]. The carbonyl group of 5 was
reduced to its corresponding product 6 by using lithium aluminum hydride as a reducing
agent. Lithium aluminum hydride produces a larger yield than sodium borohydride.
Intermediates 10 were synthesized by CuAAC click chemistry. Given those intermediates 9
are insolubility in the classical CuAAC solvent (t-BuOH/H
intermediates 10a and 10b under these conditions with CuI as a catalyst was only a moderate
0% after 24 hours. This yield significantly increased to 75% by changing the solvent system
to THF/H O with CuSO ·5H O and sodium ascorbate as catalysts.
2
O), the isolated yield of
4
2
4
2
The nitro group of 9-chloro-substituted intermediates (10a-1 to 10a-14) was reduced to
amines by using Rany Ni/H as a reducing agent. The nitro groups of 9-bromo-substituted
2
intermediates (10b-1 to 10b-14) were reduced to amines by using sodium hydrosulfite as a
reducing agent to prevent the formation of 9-debrominated by-products. Fremy’s salt was
used as an oxidant to obtain the products 12. All of the structures of compounds 12 were
1
13
confirmed by H NMR, C NMR and HRMS (ESI).
3
. Results and Discussion
3
.1 Cytotoxicity
The cell lines used in the present study were adenocarcinomic human alveolar basal epithelial
cell (A549), human promyelocytic leukaemia cell (HL-60), human chronic myelogenous
leukaemia cell (K562) and human cervical cancer cell (HeLa). Etoposide was used as a
positive control treatment. The growth inhibitory effect (IC ) of the ME derivatives was
5
0
evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT)
4

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assay described by Mosmann with minor modifications [7].
As shown in Table 1, the cytotoxicity results indicated that almost all of the compounds
showed better activity in HL-60 cell line than the three other tested cell lines. Among the
tested cells, the inhibition of the suspension cell lines (HL-60, K562) showed better activity
than the attached cell lines (A549, HeLa). This finding is consistent with that observed for the
positive control etoposide. Compounds 12a-1 to 3 and 12b-1 to 3 exhibited poor cytotoxicity
against the tested cancer cell lines. Compounds 12a-8 to 12, 12a-14, 12b-8 to 11 and 12b-14
displayed significant cytotoxicities against most of the cell lines compared with positive
control etoposide. For HL-60, these compounds exhibited IC50 < 5 µM.
3.2 SARs study
SARs study revealed that the substituents at the C-9 position and the triazole ring exhibited
various influences on the cytotoxicity against several human cancer cell lines. First, most of
the C-9 bromo substituted derivatives showed better cytotoxicity than the C-9
chloro-substituted ones (Table 1). Second, the substituents on the triazole ring performed
important functions in cytotoxicity. The aromatic substituents show better activity than
cyclopropyl, carboxylic acid ethyl ester or carboxylic acid methyl ester. The 2-pyridyl
substituents (12a-14, 12b-14) had greater activity than the phenyl substituents (12a-4, 12b-4).
Furthermore, the substituents at positions 4 on the phenyl ring play essential roles for their
cytotoxicity. Compounds 12a-9 to 11 and 12b-9 to 11, which contain alkyl substituents at
position 4 of the phenyl ring showed more effective cytotoxicities. The cytotoxicity sequence
of the aromatic-substituted derivatives was observed as follows: alkyl > halogen, -OCH > H.
3
The alkyl chain length at the positions 4 of the aromatic substituents may play an essential
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roles in their cytotoxicity, in which an increasing alkyl chain length is generally beneficial to
cytotoxicity. A propyl substituent was possibly optimal. For example, 12b-9 exhibited the
strongest cytotoxicity with IC50 = 1.49 µM for the HL-60 cells. A summary of the SARs is
illustrated in Figure 3.
3
.3 Topo inhibition assay
Topo is possibly one of the most important molecular targets of many antitumor agents
currently in use. Some quinones have already been shown to act on Topo [27-30]. Our
previous study showed that MF and its analogues are potent Topo inhibitors. Thus, the
inhibitory activities of the synthesized compounds against Topo, including Topo I and Topo II,
were evaluated.
3
.3.1 Topo I inhibition assay
Topo I is an essential nuclear enzyme involved in DNA repair, replication, transcription and
chromosome segregation during mitosis. Topo I has been identified as a target of β-lapachone
[
27, 31]. In this study, Topo I-mediated DNA relaxation assays were performed to investigate
the inhibitory activities of the novel synthesized o-quinones.
The inhibitory activity of the ME derivatives against Topo I is shown in Figure 4. Most of
these compounds exhibited weak Topo I inhibitory activity at 50 ꢀM. Their inhibitory
activities were slightly linear correlated with their cytotoxicities. For example, 12a-8, one of
the most potent anti-proliferative compounds, showed no significant Topo I inhibitory activity.
This finding may indicate that Topo I was only a minor target of their antitumor activities.
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3.3.2 Topo II inhibition assay
Several o-quinones, such as mansonone F [3], salvicine [30], β-lapachone [32], and
pyranonaphthoquinones [28] have been found to be potent Topo II inhibitors. Topo II has two
isoforms, α and β; α isoform has been recognised as an important target in cancer therapy and
drug discovery [33]. The inhibitory activity activities against Topo IIα of the synthesized
compounds were determined based on a Topo IIα relaxation assay by using human Topo IIα
(
TopoGEN) with etoposide as a positive control treatment.
The inhibitory activities of the ME derivatives against Topo IIα are shown in Figure 5. Their
inhibitory effects varied with their different substituents at 2 ꢀM. Almost all of the C-9
bromo-substituted derivatives showed better activities than the C-9 chloro-substituted
derivatives. As a representative, 12b-11 exhibited the strongest inhibitiory activity against
Topo IIα with an IC50 = 0.92 µM compared with the parent compound ME (IC50 = 18.6 µM),
which is almost 20-fold more efficacious and showed an improved efficacy in the enzymatic
assay (Figure 6). In addition, the preliminary SARs showed that the aromatic-substituted
derivatives with alkyl substituents at position 4 showed better activity than those with -F, -Cl,
3
-OCH or –H. their inhibitory activities against Topo IIα increased with increasing alkyl chain
length at position 4 of the aromatic ring. These results are similar to their anti-proliferative
activities in vitro. For example, 12a-9 to 11 and 14 as well as 12b-9 to 11 and 14 showed
Topo IIα inhibitory and anti-proliferative activities. This finding may indicate that Topo IIα
was a major target of their anti-proliferative activities. Interestingly, all of the compounds
completely inhibited the Topo IIα activity in a Topo II relaxation assay at 20 ꢀM
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concentration (data not shown). However this result is not correlated with their cytotoxic
activity, for example, esters (12a-1 to 12a-3 and 12b-1 to 12b-3) and cyclopropyl-subsitituted
compound (12a-13 and 12b-13) had no cytotoxic activities. Similar to small molecule
therapeutics, it is possible that these compounds have other biological targets, which could
involve a more complex action process in vivo. Further mechanistic studies on the Topo II
inhibition of these ME derivatives are currently underway.
3.3.3 Topo II-mediated DNA cleavage assay
Topo II inhibitors are commonly classified as poisons and catalytic inhibitors. To understand
the mode of the action of our novel compounds, Topo II-mediated pBR322 plasmid DNA
cleavage assays were performed according to the supplier’s protocol (TopoGEN) with
etoposide as a positive control. Compound 12b-9 was chosen to represent this class because it
exhibits the most effective cytotoxicity and excellently inhibits Topo II. As shown in Figure 7,
etoposide induced the formation of linear DNA; by contrast, compound 12b-9 did not induce
linear DNA formation. This finding may indicate that the compound 12b-9 was a catalytic
inhibitor rather than a poison.
3.4 Molecular docking
A molecular modelling analysis utilising the docking program called CDOCKER with
Discovery Studio 2.5 was conducted to investigate the interaction and binding mode between
our synthetic compounds and Topo IIα (PDB code: 1ZXM). We initially docked 12b-9 in
various possible binding pockets in Topo IIα. We obtained the best result in the ATP pocket.
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This finding is also consistent with the observation previously discussed, in which 12b-9
functions as a catalytic inhibitor of Topo II. We found that 12b-9 was superimposed in the
binding site with favorable interactions with the protein. Compound 12b-9 was predicted to
form several strong hydrogen bonds with the enzyme (Figure 8). In particular, two carbonyl
groups of 12b-9 formed hydrogen bonds with Lys-157 and Arg-98 residues. The 2’-N on the
triazole ring formed hydrogen bonds with Ser-149. The 3’-N on the triazole ring also formed
hydrogen bonds with both Ser-149 and Ser-148. The hydrogen bonds formed between the
nitrogen atoms of the triazole containing the residues also indicated that the triazaole
functioned as a pharmacophore rather than a linker. This result explained why the modified
ME derivatives exhibited higher Topo II inhibitiory activities than the natural product ME. In
addition, the side chain of the triazole ring of 12b-9 overlapped well with phosphate and
ribose moieties of AMPPNP in the ATP binding site. The alkyl substituent at the 4’-position of
the phenyl group in the triazole ring is predicted to fill a narrow pocket in the active site.
Favourable van der Waal’s interactions from the filling of this pocket explain why compounds
with longer alkyl substituents on the phenyl group of the triazole ring have greater potency
against Topo II. The ability of the model to explain biological and SAR further supported our
hypothesis that the compounds are ATP competitive inhibitors.
Conclusion
In summary, two series of ME derivatives including compounds with chlorine or bromine at
C-9 positions, and the CuAAC-catalysed triazole rings at the C-3 positions, were designed,
synthesized and screened for anti-proliferative activity against four cancer cell lines. MTT
9

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assay showed that HL-60 and K-562 cell lines were more sensitive to the compounds than
HeLa and A549 cell lines. Although in vitro Topo II inhibition and in vitro cytotoxicity were
not completely correlated, their aromatic substituents and alkyl chain length at position 4 of
the phenyl group are important for inhibitory potency and in vitro activity. The correlation
between in vitro inhibition and cytotoxicity may be affected by differences in permeability,
cell metabolism, and selectivity. Some of the ME derivatives exhibited significant inhibition
against Topo II at 2 ꢀM and weak inhibition of Topo I at 50 ꢀM. These results indicated that
Topo II might be the major target of this novel class of compounds. Moreover, Topo
II-mediated cleavage of pBR322 DNA assay and molecular docking analysis suggested that
our ME derivatives may function as catalytic inhibitors of Topo II, occupying its ATP binding
pocket. These results provided important information as basis of further structural
modifications of ME to develop a the rational design of more effective antitumor drugs.
4
. Experimental Section
4.1 Chemistry
All reactions were monitored by TLC. Melting points were determined by using capillary
1
tubes with a MSRS-OptiMelt Automated Melting point system and are uncorrected. H and
13
C NMR spectra were recorded using TMS as the internal standard with a Bruker BioSpin
GmbH spectrometer at 400 MHz and 100 MHz, respectively. High resolution mass spectra
HRMS) were recorded on Shimadzu LCMS-IT-TOF spectrometer. The purities of
(
synthesized compounds were confirmed by using analytical HPLC with a dual pump
Shimadzu LC-20AB system equipped with a Ultimate-QB-C18 column (4.6 × 250 mm, 5
1
0

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µm), the detection wavelength was 254 nm and the mobile phase was MeOH/H O (65:35 ~
2
90:10, v:v) with a flow rate of 0.8 mL/min.
The 9-chlorobenzo[de]chromen-3(2H)-one (5a) and 9-bromobenzo[de]chromen-3(2H)-one
5b) were synthesized according to our previously reported procedure [3].
(
General procedure for the preparation of 6a and 6b
45.7 mmol of compound 5 was dissolved in 100 mL anhydrous THF, and cooled in ice bath.
3
4.3 mmol of LiAlH was added to the solvent slowly in four portions over 30 minutes. The
4
reaction mixture was warmed to room temperature and stirred for another 1 hour. After the
reaction was completed, the reaction was terminated by addition of 1.3 mL water. After 15
minutes, 1.3 mL 10% NaOH aqueous solution was added dropwise over 5 minutes, and
further 3.9 mL water was added dropwise over 30 minutes. The reaction was filtered, and the
residue was washed with THF (4 × 30 mL). The filtrate was dried over Na SO , and
2
4
concentrated to provide crude solid product. The solid residue was subjected to flash column
chromatography (silica gel), eluting with 25% ethyl acetate in petroleum to give the desired
products.
9-chloro-2,3-dihydrobenzo[de]chromen-3-ol (6a)
1
Brown light solid, yield 78.6%. H NMR (400 MHz, CDCl
3
) δ 7.78 (t, J = 4.8 Hz, 1H), 7.48 (d,
J = 4.8 Hz, 2H), 7.43 (d, J = 8.8 Hz, 1H), 7.40 (d, J = 8.8 Hz, 1H), 5.02 (br s, 1H), 4.51 (dd, J
11.2, 4.0 Hz, 1H), 4.41 (dd, J = 11.2, 2.8 Hz, 1H), 2.28 (br s, 1H).
=
1
1

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9-bromo-2,3-dihydrobenzo[de]chromen-3-ol(6b)
1
Brown light solid, yield 74.6%. H NMR (400 MHz, CDCl
J = 8.8 Hz, 1H), 7.52 (d, J = 4.0 Hz, 2H), 7.34 (d, J = 8.8 Hz, 1H), 5.07 (br s, 1H), 4.56 (dd, J
11.2, 4.4 Hz, 1H), 4.46 (dd, J = 11.2 Hz, 2.4H), 2.11 (br s, 1H).
3
) δ 7.80 (t, J = 4.8 Hz, 1H), 7.58 (d,
=
General procedure for the preparation of 7a and 7b
1.6 mmol of the compound 6 was dissolved in 100 mL anhydrous CH
an ice bath to 0 °C. 1.5 mL PBr (15.8 mmol, 0.5 equiv.) was dissolved in 50 mL anhydrous
3
2 2
Cl , and was cooled in
3
2 2
CH Cl , and added dropwise to the reaction over 30 minutes. The reaction mixture was
warmed to room temperature and stirred for a further 2 hours. After the substrate was
completely consumed up, the reaction was neutralized to pH = 6-7 by addition of the
saturated aqueous sodium bicarbonate through a dropping funnel. The mixture was separated
2 2
and the water layer was extracted with CH Cl (3 × 30 mL). The combined extracts were
2 4
washed with brine (20 mL), and dried over Na SO , filtered and concentrated. The crude solid
was subjected to flash column chromatography (silica gel), eluting with 10% ethyl acetate in
petroleum to provide the desired compounds.
3-bromo-9-chloro-2,3-dihydrobenzo[de]chromene (7a)
1
White solid, yield 78.4%. m.p. 117.3-118.4 °C. H NMR (400 MHz, CDCl
3
) δ 7.78 (dd, J =
8
.4, 4.1 Hz, 1H), 7.50 (d, J = 8.4 Hz, 2H), 7.44 (dd, J = 8.6, 2.7 Hz, 2H), 5.60 (t, J = 2.9 Hz,
H), 4.77 (dd, J = 12.1, 3.5 Hz, 1H), 4.65 (dd, J = 12.1, 3.5 Hz, 1H).
1
1
2

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3,9-dibromo-2,3-dihydrobenzo[de]chromene(7b)
1
White solid, yield 84.1%. m.p.112.6-113.6 °C. H NMR (400 MHz, CDCl
Hz, 1H), 7.46 (dd, J = 7.8, 2.3 Hz, 2H), 7.42 (dd, J = 7.7, 3.4 Hz, 2H), 5.58 (t, J = 2.6 Hz, 1H),
.82 (dd, J = 12.1, 3.5 Hz, 1H), 4.61 (dd, J = 12.0, 3.5 Hz, 1H).
3
) δ 7.76 (d, J = 7.6
4
General procedure for the preparation of 8a and 8b
To a solution of 31.7 mmol compound 7 in 100 mL mixed solvent of acetone and water (4:1,
v:v), 2.06 g (31.7 mmol, 1.0 equiv.) sodium azide was added. The reaction was heated to
reflux until the reaction was completed. The solution was evaporated under reduced pressure,
and the residue was extracted with dichloromethane (2 × 30 mL). The combined extracts were
washed with brine (50 mL), dried over Na SO , filtered and evaporated to dryness. The solid
2
4
residue was subjected to chromatography column (silica gel), eluting with 10% ethyl acetate
in petroleum to give the desired products.
3-azido-9-chloro-2,3-dihydrobenzo[de]chromene (8a)
1
White solid, yield 91.1%. m.p. 67.3-69.9 °C. H NMR (400 MHz, CDCl ) δ 7.85 (d, J = 8.3
3
Hz, 1H), 7.53 (dd, J = 7.2, 1.2 Hz, 1H), 7.46 (d, J = 8.0 Hz, 2H), 7.42 (d, J = 7.6 Hz, 1H),
4.82 (t, J = 2.8 Hz, 1H), 4.63 (dd, J= 11.5, 3.2 Hz, 1H), 4.45 (dd, J = 11.4, 2.6 Hz, 1H).
3-azido-9-bromo-2,3-dihydrobenzo[de]chromene (8b)
1
White solid, yield 83.5%. m.p. 72.1-73.4 °C. H NMR (400 MHz, CDCl ) δ 7.86 (d, J = 8.3
3
Hz, 1H), 7.56 (t, J = 7.7 Hz, 1H), 7.51 (d, J = 6.9 Hz, 2H), 7.41 (d, J = 8.8 Hz, 1H), 4.84 (br s,
1
3

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1H), 4.64 (dd, J = 9.6, 1.2 Hz, 1H), 4.45 (dd, J = 9.6, 1.2 Hz, 1H).
General procedure for the preparation of 9a and 9b
0.4 mmol of compound 8 was dissolved in acetic anhydride (40 mL) and cooled to 0 °C in
2
an ice bath. 6.0 g of cupric nitrate (24.8 mmol, 1.2 equiv.) was added with vigorous stirring.
The reaction was warmed to room temperature, and stirring was continued until the reaction
was completed. The mixture was diluted with brash ice (300 g), stirred for another 2 hours,
and filtered to collect the residue. The residue was subjected to chromatography column
(
silica gel), eluting with 30% dichloromethane in petroleum to give the desired compounds.
3-azido-9-chloro-7-nitro-2,3-dihydrobenzo[de]chromene (9a)
1
White solid, yield 75.2%. m.p. 149.9-151.2 °C. H NMR (400 MHz, CDCl
3
) δ 8.84 (d, J = 8.9
Hz, 1H), 8.63 (s, 1H), 7.82 (dd, J = 8.9, 1.6 Hz, 1H), 7.58 (d, J = 7.0 Hz, 1H), 4.91 (t, J = 2.6
Hz, 1H), 4.73 (dd, J = 11.6, 3.1 Hz, 1H), 4.52 (dd, J = 11.6, 2.6 Hz, 1H).
3-azido-9-bromo-7-nitro-2,3-dihydrobenzo[de]chromene (9b)
1
White solid, yield 80.5%. m.p. 158.7-159.9 °C. H NMR (400 MHz, CDCl
3
) δ 8.87 (d, J = 9.4
Hz, 1H), 8.63 (s, 1H), 7.82 (dd, J = 8.7, 1.2 Hz, 1H), 7.58 (d, J = 7.0 Hz, 1H), 4.92 (br s, 1H),
.74 (dd, J = 11.6, 2.3 Hz, 1H), 4.53 (dd, J = 11.6, 1.6 Hz, 1H).
4
General procedure for the preparation of 10a-1 ~ 10a-14, 10b-1 ~ 10b-14
.6 mmol of compound 9, alkynes (0.65 mmol, 1.1 equiv), 10 mol% L-ascorbic acid (13 mg)
0
1
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and 5 mol% CuSO ·5H O (8 mg) were solubilized with THF/H O (20.0 mL, 1:1). After
4
2
2
stirring for 6 h at 50 °C, the reaction mixture was quenched with water (15 mL) and extracted
with ethyl acetate (2 × 20 mL). The combined extracts were washed with water (2 ×10 mL),
2 4
brine (10 mL), dried over Na SO , filtered and evaporated to dryness. The solid residue was
subjected to flash column chromatography (silica gel), eluting with chloroform to give the
desired products.
Ethyl1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazole-4-carboxyl-
ate (10a-1).
1
Yellow solid, yield 76.6%. m.p. 233.2-234.8 °C. H NMR (400 MHz, CDCl
3
) δ 8.90 (d, J =
8
1
2
7
1
.8 Hz, 1H, 6-CH-), 8.55 (s, 1H, 8-CH-), 7.82 (t, J = 7.2 Hz, 1H, 5-CH-), 7.65 (d, J = 7.2 Hz,
H, 4-CH-), 7.62 (s, 1H, triazole-H), 6.36 (br s, 1H, 3-CH-), 5.17 (d, J = 12.1 Hz, 1H,
-CH -), 4.79 (d, J = 12.1 Hz, 1H, 2-CH -), 4.37 (q, J = 7.2 Hz, 2H, -COOCH -), 1.35 (t, J =
2
2
2
13
3 6
.1 Hz, 3H, -CH ). C NMR (100 MHz, DMSO-d ) δ 159.99, 152.53, 138.88(2C), 130.49,
28.71, 127.42, 127.31, 126.36, 124.50, 124.37, 120.83, 113.78, 69.49, 60.56, 55.68, 14.08.
-
HRMS (ESI): calcd for C H ClN O [M-H] 387.0502, found 387.0508.
17
13
4
5
Methyl 1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazole-4-carboxy-
late (10a-2)
1
Yellow solid, yield 77.6%. m.p. 233.5-234.9 °C. H NMR (400 MHz, CDCl
3
) δ 8.93 (d, J =
8
.9 Hz, 1H, 6-CH-), 8.56 (s, 1H, 8-CH-), 7.83 (t, J = 8.9 Hz, 1H, 5-CH-), 7.67 (d, J = 7.2 Hz,
H, 4-CH-), 7.63 (s, 1H, triazole-H), 6.37 (br s, 1H, 3-CH-), 5.19 (dd, J = 12.2, 2.1 Hz, 1H,
1
1
5

ACCEPTED MANUSCRIPT
1
3
2
-CH -), 4.80 (dd, J = 12.1, 2.6 Hz , 1H, 2-CH -), 3.90 (s, 3H, -OCH ). C NMR (100 MHz,
2
2
3
DMSO-d
6
) δ 160.43, 152.50, 138.92, 138.65, 130.50, 128.77, 127.42, 127.35, 126.29, 124.52,
-
1
24.39, 120.83, 113.78, 69.50, 55.70, 51.76. HRMS (ESI): calcd for C16
73.0345, found 373.0314.
4 5
H11ClN O [M-H]
3
Dimethyl-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazole-4,5-di-
carboxylate (10a-3)
1
Yellow solid, yield 70.0%. m.p. 155.6-158.6 °C. H NMR (400 MHz, CDCl
3
) δ 8.87 (d, J =
8
1
3
1
.8 Hz, 1H, 6-CH-), 8.54 (s, 1H, 8-CH-), 7.73 (t, J = 8.0 Hz, 1H, 5-CH-), 7.35 (d, J = 7.0 Hz,
H, 4-CH-), 6.67 (br s, 1H, 3-CH-), 5.05 (dd, J = 11.8, 4.2 Hz, 1H, 2-CH -), 4.84 (dd, J = 11.7,
.1 Hz, 1H, 2-CH -), 3.98 (s, 6H, -OCH ). C NMR (100 MHz, CDCl ) δ 160.30, 159.06,
2
13
2
3
3
52.51, 140.26, 139.27, 129.75, 128.24, 126.01, 125.69, 125.67, 125.31, 125.25, 121.55,
-
115.62, 69.21, 56.50, 53.76, 52.86. HRMS (ESI): calcd for C18
H13ClN
4
O
7
[M-H] 431.0400,
found 431.0385.
1
-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-fluorophenyl)-1H-1,2,3-triazole
10a-4)
Yellow solid, yield 85.1%. m.p. 207.3-209.2 °C. H NMR (400 MHz, CDCl
(
1
3
) δ 8.88 (d, J =
.8 Hz, 1H, 6-CH- ), 8.54 (s, 1H, 8-CH-), 7.81 (t, J = 7.2 Hz, 1H, 5-CH-), 7.71–7.65 (m, 3H,
-CH- and 2’,6’-CH-), 7.33 (s, 1H, triazole-H), 7.05 (t, JH-H, H-F = 8.6 Hz, 2H, 3’,5’-CH-), 6.35
8
4
(
br s, 1H, 3-CH-), 5.17 (dd, J = 12.0, 2.5 Hz, 1H, 2-CH -), 4.82 (dd, J = 12.0, 2.9 Hz, 1H,
2
13
2
2 3
-CH -). C NMR (100 MHz, CDCl ) δ 162.78 (d, JC-F = 246 Hz), 152.00, 147.56, 139.69,
1
6

ACCEPTED MANUSCRIPT
130.21, 128.30, 127.55, 127.47 (2C), 126.18 (d, JC-F = 3.0 Hz), 125.98, 125.48, 125.33,
120.87, 118.29, 116.00, 115.80 (d, JC-F = 22.0 Hz, 2C), 69.90, 56.57. HRMS (ESI): calcd for
-
C
20
H12ClFN
4
O
3
[M-H] 409.0509, found 409.0517.
1
-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-chlorophenyl)-1H-1,2,3-triazole
10a-5)
Yellow solid, yield 86.8%. m.p. 238.6-240.9 °C. H NMR (400 MHz, CDCl
(
1
3
) δ 8.89 (d, J =
.8 Hz, 1H, 6-CH-), 8.55 (s, 1H, 8-CH-), 7.82 (t, J = 8.8 Hz, 1H, 5-CH-), 7.67–7.64 (m, 3H,
-CH- and 2’,6’-CH-), 7.41–7.30 (m, 3H, 3’,5’-CH- and triazole-H), 6.37 (br s, 1H, 3-CH-),
8
4
5
13
2 2
.18 (d, J = 12.0 Hz, 1H, 2-CH -), 4.83 (d, J = 12.0 Hz, 1H, 2-CH -). C NMR (100 MHz,
DMSO-d ) δ 152.60, 145.39, 138.86, 132.38, 130.56, 129.19, 128.79(2C), 127.45(2C),
6
126.89(2C), 126.53, 124.54, 124.28, 121.28, 120.84, 113.85, 69.85, 55.51. HRMS (ESI):
-
calcd for C20
H12Cl
2
N
4
O
3
[M-H] 425.0214, found 425.0233.
1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-phenyl-1H-1,2,3-triazole (10a-6)
1
Yellow solid, yield 82.6%. m.p. 206.7-207.9 °C. H NMR (400 MHz, CDCl ) δ 8.89 (d, J =
3
8
2
1
2
.9 Hz, 1H, 6-CH-), 8.55 (s, 1H, 8-CH-), 7.82 (t, J = 8.8 Hz, 1H, 5-CH-), 7.72 (d, J = 7.8 Hz,
H, 2’,6’-CH-), 7.66 (d, J = 7.1 Hz, 1H, 4-CH-), 7.37 (d, J = 6.8 Hz, 2H, 3’,5’-CH-), 7.36 (s,
H, triazole-H), 7.32 (t, J = 8.2 Hz, 1H, 4’-CH-), 6.36 (br s, 1H, 3-CH-), 5.17 (dd, J = 11.9,
13
2 2 6
.7 Hz, 1H, 2-CH -), 4.83 (dd, J = 12.0, 3.0 Hz, 1H, 2-CH -). C NMR (100 MHz, DMSO-d )
δ 157.90, 151.71, 144.14, 135.83, 135.55, 134.01(2C), 133.20, 132.70, 132.62, 131.98,
30.46(2C), 129.80, 129.49, 126.21, 126.13, 119.06, 75.11, 60.68. HRMS (ESI): calcd for
1
17

ACCEPTED MANUSCRIPT
-
C H ClN O [M-H] 391.0603, found 391.0591.
2
0
13
4
3
1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(p-tolyl)-1H-1,2,3-triazole (10a-7)
1
Yellow solid, yield 88.3%. m.p. 217.5-218.8 °C. H NMR (400 MHz, CDCl
3
) δ 8.88 (d, J =
8
1
2
.9 Hz, 1H, 6-CH-), 8.54 (s, 1H, 8-CH-), 7.80 (t, J = 8.9 Hz, 1H, 5-CH-), 7.64 (d, J = 6.8 Hz,
H, 4-CH-), 7.60 (d, J = 8.1 Hz, 2H, 2’,6’-CH-), 7.33 (s, 1H, triazole-H), 7.16 (d, J = 8.1 Hz,
H, 3’,5’-CH-), 6.34 (t, J = 2.8 Hz, 1H, 3-CH-), 5.16 (dd, J = 11.9, 2.9 Hz, 1H, 2-CH -), 4.82
). C NMR (100 MHz, CDCl ) δ
2
13
(
dd, J = 11.9, 3.1 Hz, 1H, 2-CH
2
-), 2.34 (s, 3H, 7’-CH
3
3
157.90, 151.78, 144.11, 142.51, 135.81, 134.54(2C), 132.78, 132.69, 132.59, 132.00,
130.38(2C), 129.80, 129.46, 126.11, 125.74, 119.06, 75.11, 60.64, 26.00. HRMS (ESI): calcd
-
for C H ClN O [M-H] 405.0760, found 405.0774.
21
15
4
3
1
-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-ethylphenyl)-1H-1,2,3-triazole
10a-8)
Yellow solid, yield 85.4%. m.p. 231.1-233.4 °C. H NMR (400 MHz, CDCl
(
1
3
) δ 8.88 (d, J =
.9 Hz, 1H, 6-CH-), 8.54 (s, 1H, 8-CH-), 7.80 (t, J = 8.8 Hz, 1H, 5-CH-), 7.65–7.62 (m, 3H,
-CH- and 2’,6’-CH-), 7.33 (s, 1H, triazole-H), 7.19 (d, J = 8.0 Hz, 2H, 3’,5’-CH-), 6.35 (br s,
H, 3-CH-), 5.16 (dd, J = 11.9, 2.4 Hz, 1H, 2-CH -), 4.82 (dd, J = 11.9, 2.6 Hz, 1H, 2-CH -),
8
4
1
2
2
2
13
.64 (q, J = 7.6 Hz, 2H, 7’-CH
) δ 152.65, 146.53, 143.58, 138.87, 130.57, 128.10(2C), 127.79, 127.44, 127.33,
26.78, 125.23(2C), 124.54, 124.21, 120.86, 120.54, 113.79, 69.85, 55.38, 27.84, 15.36.
2 3
-), 1.21 (t, J = 7.6 Hz, 3H, 8’-CH ). C NMR (100 MHz,
DMSO-d
6
1
-
4 3
HRMS (ESI): calcd for C22H17ClN O [M-H] 419.0916, found 419.0936.
1
8

ACCEPTED MANUSCRIPT
1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-propylphenyl)-1H-1,2,3-triazole
(
10a-9)
Yellow solid, yield 88.4%.m.p. 207.5-209.5 °C. H NMR (400 MHz, CDCl
1
3
) δ 8.88 (d, J =
.9 Hz, 1H, 6-CH-), 8.55 (s, 1H, 8-CH-), 7.80 (t, J = 8.8 Hz, 1H, 5-CH-), 7.66–7.62 (m, 3H,
-CH- and 2’,6’-CH-), 7.37 (s, 1H, triazole-H), 7.18 (d, J = 7.6 Hz, 2H, 3’,5’-CH-), 6.36 (br s,
H, 3-CH-), 5.17 (dd, J = 11.8, 2.0 Hz, 1H, 2-CH -), 4.84 (dd, J = 11.7, 2.2 Hz, 1H, 2-CH -),
8
4
1
2
9
1
2
2
2 2
.58 (t, J = 7.6 Hz, 2H, 7’-CH -), 1.68–1.56 (m, 2H, 8’-CH -), 0.92 (t, J = 7.3 Hz, 3H,
13
3 6
’-CH ). C NMR (100 MHz, DMSO-d ) δ 157.90, 151.80, 147.21, 144.10, 135.81,
33.93(2C), 133.06, 132.69, 132.57, 132.03, 130.41(2C), 129.80, 129.46, 126.10, 125.79,
-
119.05, 75.11, 60.63, 42.15, 29.12, 18.76. HRMS (ESI): calcd for C H ClN O [M-H]
23 19 4 3
433.1073, found 433.1091.
4-(4-butylphenyl)-1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazole
(10a-10)
1
Yellow solid, yield 77.0%. m.p. 206.1-208.2 °C. H NMR (400 MHz, CDCl ) δ 8.87 (d, J =
3
7
.8 Hz, 1H, 6-CH-), 8.53 (s, 1H, 8-CH-), 7.80 (t, J = 8.8 Hz, 1H, 5-CH-), 7.65–7.60 (m, 3H,
-CH- and 2’,6’-CH-), 7.33 (s, 1H, triazole-H), 7.17 (t, J = 7.3 Hz, 2H, 3’,5’-CH-), 6.34 (t, J
4
=
2.7 Hz, 1H, 3-CH-), 5.17 (dd, J = 11.9, 2.9 Hz, 1H, 2-CH
-CH -), 2.59 (t, J = 8.0 Hz, 2H, 7’-CH -), 1.62–1.51 (m, 2H, 8’-CH
’-CH -), 0.90 (t, J = 7.3 Hz, 3H, 10’-CH ). C NMR (100 MHz, DMSO-d ) δ 152.65,
2
-), 4.82 (dd, J = 11.9, 3.0 Hz, 1H,
2
9
1
2
2
2
-), 1.37–1.27 (m, 2H,
13
2
3
6
46.54, 142.18, 138.87, 130.56, 128.63(2C), 127.76, 127.43, 127.32, 126.78, 125.17(2C),
1
9

ACCEPTED MANUSCRIPT
124.54, 124.20, 120.85, 120.53, 113.78, 69.85, 55.37, 34.48, 32.92, 21.65, 13.69. HRMS
-
(
ESI): calcd for C24
H21ClN O
4 3
[M-H] 447.1229, found 447.1215.
1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-pentylphenyl)-1H-1,2,3-triazole
(10a-11)
1
Yellow solid, yield 82.0%. m.p. 201.3-203.2 °C. H NMR (400 MHz, CDCl ) δ 8.88 (d, J =
3
8
4
1
8
0
1
1
.9 Hz, 1H, 6-CH-), 8.54 (s, 1H, 8-CH-), 7.80 (t, J = 8.8 Hz, 1H, 5-CH-), 7.67–7.58 (m, 3H,
-CH- and 2’,6’-CH-), 7.33 (s, 1H, triazole-H), 7.17 (d, J = 6.9 Hz, 2H, 3’,5’-CH-), 6.35 (br s,
2 2
H, 3-CH-), 5.16 (d, J = 11.8 Hz, 1H, 2-CH -), 4.82 (d, J = 11.9 Hz, 1H, 2-CH -), 2.58 (t, J =
2 2 2 2
.0 Hz, 2H, 7’-CH -), 1.61–1.55 (m, 2H, 8’-CH -), 1.32–1.26 (m, 4H, 9’-CH - and 10’-CH -),
1
3
.87 (t, J = 6.1 Hz, 3H, 11’-CH ). C NMR (100 MHz, DMSO-d ) δ 152.65, 146.55, 142.20,
3
6
38.86, 130.56, 128.62(2C), 127.77, 127.44, 127.32, 126.78, 125.17(2C), 124.54, 124.20,
20.85, 120.53, 113.79, 69.85, 55.37, 34.77, 30.78, 30.42, 21.86, 13.83. HRMS (ESI): calcd
-
for C25
H23ClN
4
O
3
[M-H] 461.1386, found 461.1416.
1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-methoxyphenyl)-1H-1,2,3-triazo
le (10a-12)
1
Yellow solid, yield 87.30%. m.p. 231.9-233.8 °C. H NMR (400 MHz, CDCl
3
) δ 8.88 (d, J =
8
4
1
3
.9 Hz, 1H, 6-CH-), 8.54 (s, 1H, 8-CH-), 7.81 (t, J = 8.7 Hz, 1H, 5-CH-), 7.67–7.61 (m, 3H,
-CH- and 2’,6’-CH-), 7.28 (s, 1H, triazole-H), 6.88 (d, J = 8.3 Hz, 2H, 3’,5’-CH- ), 6.34 (br s,
H, 3-CH-), 5.15 (dd, J = 11.9, 2.9 Hz, 1H, 2-CH -), 4.82 (dd, J = 11.9, 3.1 Hz, 1H, 2-CH -),
2
2
13
.81 (s, 3H, -OCH
3
). C NMR (100 MHz, DMSO-d
6
) δ 159.03, 152.65, 146.40, 138.87,
2
0

ACCEPTED MANUSCRIPT
130.57, 127.44, 127.31, 126.82, 126.58(2C), 124.53, 124.19, 122.88, 120.85, 119.95,
-
4 4
114.16(2C), 113.78, 69.85, 55.34, 55.08. HRMS (ESI): calcd for C21H15ClN O [M-H]
421.0709, found 421.0707.
1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-cyclopropyl-1H-1,2,3-triazole
(
10a-13)
Yellow solid, yield 77.0%. m.p. 178.2-179.6 °C. H NMR (400 MHz, CDCl
1
3
) δ 8.87 (d, J =
.8 Hz, 1H, 6-CH-), 8.54 (s, 1H, 8-CH-), 7.80 (t, J = 7.2 Hz, 1H, 5-CH-), 7.59 (d, J = 5.8 Hz,
H, 4-CH-), 6.97 (s, 1H, triazole-H), 6.26 (br s, 1H, 3-CH-), 5.09 (d, J = 11.5 Hz, 1H,
-CH -), 4.79 (d, J = 10.9 Hz, 1H, 2-CH -), 1.93–1.80 (m, 1H, cyclopropyl 1’-CH-),
.98–0.87 (m, 2H, cyclopropyl 2’-CH - or 3’-CH -), 0.87–0.76 (m, 2H, cyclopropyl 3’-CH -
8
1
2
2
2
0
2
2
2
1
3
2 6
or 2’-CH -). C NMR (100 MHz, DMSO-d ) δ 153.83, 149.02, 139.06, 130.57, 129.83,
127.08, 126.98, 124.82, 124.10, 120.64, 120.32, 102.37, 69.89, 55.07, 7.54, 7.49, 6.36.
-
4 3
HRMS (ESI): calcd for C17H13ClN O [M-H] 355.0603, found 355.0604.
2
-(1-(9-chloro-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazol-4-yl)pyridine
10a-14)
Yellow solid, yield 94.2%. m.p. 214.9-216.5 °C. H NMR (400 MHz, CDCl
(
1
3
) δ 8.90 (d, J =
.9 Hz, 1H, 6-CH-), 8.55 (s, 1H, 8-CH-), 8.48 (d, J = 4.1 Hz, 1H, pyridyl-3’-H), 8.19 (d, J =
.9 Hz, 1H, pyridyl-6’-H), 7.83–7.70 (m, 3H, 5-CH-, pyridyl-5’-H and triazole-H), 7.67 (d, J
8
7
=
7.1 Hz, 1H, 4-CH-), 7.22 (t, J = 6.0 Hz, 1H, pyridyl-4’-H), 6.37 (br s, 1H, 3-CH-), 5.21 (dd,
1
3
2 2
J = 12.0, 2.5 Hz, 1H, 2-CH -), 4.82 (dd, J = 12.0, 2.8 Hz, 1H, 2-CH -). C NMR (100 MHz,
2
1

ACCEPTED MANUSCRIPT
CDCl ) δ 152.06, 149.71, 149.29, 148.77, 139.60, 137.01, 130.16, 128.35, 127.55, 126.06,
3
125.38, 125.19, 123.12, 121.11, 120.94, 120.36, 116.05, 69.91, 56.71. HRMS (ESI): calcd for
-
C
19
H12ClN
5
O
3
[M-H] 392.0556, found 392.0582.
Ehyl 1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazole-4-carboxylate
10b-1)
Yellow solid, yield 90.9%. m.p. 204.4-205.8 °C. H NMR (400 MHz, CDCl
(
1
3
) δ 8.90 (d, J =
.9 Hz, 1H, 6-CH-), 8.68 (s, 1H, 8-CH-), 7.84 (t, J = 8.8 Hz, 1H, 5-CH-), 7.65 (d, J = 7.1 Hz,
H, 4-CH-), 7.61 (s, 1H, triazole-H), 6.35 (br s, 1H, 3-CH-), 5.17 (dd, J = 12.2, 2.0 Hz, 1H,
-CH -), 4.79 (dd, J = 12.1, 2.7 Hz, 1H, 2-CH -), 4.37 (q, J = 7.2 Hz, 2H, -OCH -), 1.36 (t, J
7.1 Hz, 3H, -CH3). C NMR (100 MHz, DMSO-d ) δ 160.00, 153.77, 139.16, 138.87,
8
1
2
2
2
2
13
=
6
130.54, 129.85, 128.74, 127.23, 126.35, 124.86, 124.43, 120.75, 102.49, 69.61, 60.57, 55.73,
-
14.09. HRMS (ESI): calcd for C17
H13BrN
4
O
5
[M-H] 430.9997, found 431.0025.
Methyl 1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazole-4-carboxy-
late (10b-2)
1
Yellow solid, yield 86.7%. m.p. 216.6-218.9 °C. H NMR (400 MHz, CDCl
3
) δ 8.90 (d, J =
8
1
2
.9 Hz, 1H, 6-CH-), 8.56 (s, 1H, 8-CH-), 7.83 (t, J = 8.8 Hz, 1H, 5-CH-), 7.66 (d, J = 7.1 Hz,
H, 4-CH-), 7.62 (s, 1H, triazole-H), 6.36 (br s, 1H, 3-CH-), 5.18 (dd, J = 12.1, 1.9 Hz, 1H,
13
2 2 3
-CH -), 4.79 (dd, J = 12.1, 2.5 Hz, 1H, 2-CH -), 3.89 (s, 3H, -OCH ). C NMR (100 MHz,
DMSO-d ) δ 160.43, 153.74, 139.13, 138.64, 130.53, 129.85, 128.80, 127.27, 126.26, 124.88,
6
-
11 4 5
124.45, 120.73, 102.51, 69.62, 55.76, 51.77. HRMS (ESI): calcd for C16H N O Br [M-H]
2
2

ACCEPTED MANUSCRIPT
416.9840, found 416.9810.
Dimethyl
-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazole-4,5-dicarboxylate
10b-3)
Yellow solid, yield 77.6%. m.p. 184.5-186.3 °C. H NMR (400 MHz, CDCl ) δ 8.85 (d, J =
1
(
1
3
8.9 Hz, 1H, 6-CH-), 8.66 (s, 1H, 8-CH-), 7.75 (t, J = 8.4 Hz, 1H, 5-CH-), 7.33 (d, J = 7.1 Hz,
1
H, 4-CH-), 6.65 (t, J = 3.8 Hz, 1H, 3-CH-), 5.04 (dd, J = 11.9, 4.5 Hz, 1H, 2-CH -), 4.84 (dd,
2
13
2 3 3
J = 11.8, 3.3 Hz, 1H, 2-CH -), 3.98 (s, 3H, -OCH ) , 3.97 (s, 3H, -OCH ). C NMR (100
MHz, DMSO-d
6
) δ 160.23, 158.26, 153.92, 139.03, 130.50, 130.08, 129.82, 127.26, 126.05,
124.63, 124.57, 124.51, 121.32, 102.34, 69.67, 55.83, 53.62, 52.66. HRMS (ESI): calcd for
-
C
18 4 7
H13BrN O [M-H] 474.9895, found 474.9930.
1
-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-fluorophenyl)-1H-1,2,3-triazole
10b-4)
Yellow solid, yield 82.8%. m.p. 210.1-212.8 °C. H NMR (400 MHz, CDCl ) δ 8.87 (d, J =
(
1
3
8
.9 Hz, 1H, 6-CH-), 8.66 (s, 1H, 8-CH-), 7.83 (t , J = 7.9 Hz, 1H, 5-CH-), 7.73–7.63 (m, 3H,
-CH- and 2’,6’-CH-), 7.32 (s, 1H, triazole-H), 7.05 (t, JH-H, H-F= 8.5 Hz, 2H, 3’,5’-CH-), 6.35
4
(
br s, 1H, 3-CH-), 5.17 (dd, J = 11.9, 2.1 Hz, 1H, 2-CH
2
-), 4.82 (dd, J = 12.0, 2.6 Hz, 1H,
1
3
2
1
1
-CH -). C NMR (100 MHz, CDCl ) δ 162.81 (d, JC-F = 246 Hz), 153.24, 147.60, 139.99,
2
3
30.78, 130.26, 127.53 (d, JC-F = 8.0 Hz, 2C), 127.36, 126.19 (d, JC-F = 3.0 Hz), 126.05,
25.76, 125.50, 120.79, 118.25, 115.79 (d, JC-F = 21.0 Hz, 2C), 104.13, 70.01, 56.64. HRMS
2
3

ACCEPTED MANUSCRIPT
-
(
ESI): calcd for C H BrFN O [M-H] 453.0004, found 453.0059.
2
0
12
4
3
1
-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-chlorophenyl)-1H-1,2,3-triazole
10b-5)
Yellow solid, yield 82.1%. m.p. 221.6-233.8 °C. H NMR (400 MHz, CDCl
(
1
3
) δ 8.89 (d, J =
.5 Hz, 1H, 6-CH-), 8.68 (s, 1H, 8-CH-), 7.84 (t, J = 7.9 Hz, 1H, 5-CH-), 7.71–7.63 (m, 3H,
-CH- and 2’,6’-CH- ), 7.39–7.33 (m, 3H , triazole-H and 3’,5’-CH-), 6.36 (br s, 1H, 3-CH-),
8
4
5
1
3
2 2
.18 (d, J = 11.9 Hz, 1H, 2-CH -), 4.83 (d, J = 12.0 Hz, 1H, 2-CH -). C NMR (100 MHz,
DMSO-d
6
) δ 153.84, 145.36, 139.17, 132.38, 130.62, 129.86, 129.21, 128.81(2C), 127.39,
126.91(2C), 126.53, 124.91, 124.34, 121.32, 120.78, 102.57, 69.97, 55.57. HRMS (ESI):
-
calcd for C H BrClN O [M-H] 468.9709, found 468.9753.
2
0
12
4
3
1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-phenyl-1H-1,2,3-triazole (10b-6)
1
Yellow solid, yield 84.7%. m.p. 235.1-238.0 °C. H NMR (400 MHz, CDCl
3
) δ 8.89 (d, J =
8
2
7
1
1
1
.9 Hz, 1H, 6-CH-), 8.68 (s, 1H, 8-CH-), 7.84 (t, J = 7.2 Hz, 1H, 5-CH-), 7.73 (d, J = 7.2 Hz,
H, 2’,6’-CH-), 7.66 (d, J = 7.1 Hz, 1H, 4-CH-), 7.41–7.34 (m, 3H, triazole-H, 3’,5’-CH-),
.32 (t, J = 7.1 Hz, 1H, 4’-CH-), 6.36 (t, J = 2.7 Hz, 1H, 3-CH-), 5.18 (dd, J = 11.9, 2.8 Hz,
1
3
2 2 6
H, 2-CH -), 4.84 (dd, J = 11.9, 3.0 Hz, 1H, 2-CH -). C NMR (100 MHz, DMSO-d ) δ
53.88, 146.44, 139.12, 130.61, 130.29, 129.87, 128.75(2C), 127.94, 127.27, 126.70,
25.20(2C), 124.91, 124.28, 120.97, 120.77, 102.53, 69.96, 55.48. HRMS (ESI): calcd for
-
C H BrN O [M-H] 435.0098, found 435.0106.
2
0
13
4
3
2
4

ACCEPTED MANUSCRIPT
1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(p-tolyl)-1H-1,2,3-triazole (10b-7)
1
Yellow solid, yield 85.0%. m.p. 223.0-224.1 °C. H NMR (400 MHz, CDCl
3
) δ 8.87 (d, J =
8
4
1
7
1
7
.9 Hz, 1H, 6-CH-), 8.67 (s, 1H, 8-CH-), 7.82 (t, J = 7.9 Hz, 1H, 5-CH-), 7.67–7.56 (m, 3H, ,
-CH- and 2’,6’-CH-), 7.34 (s, 1H, triazole-H), 7.17 (d, J = 7.7 Hz, 2H, 3’,5’-CH-), 6.34 (br s,
H, 3-CH-), 5.17 (d, J = 11.9 Hz, 1H, 2-CH -), 4.83 (d, J = 11.9 Hz, 1H, 2-CH -), 2.35 (s, 3H,
2
2
13
’-CH3). C NMR (100 MHz, CDCl ) δ 153.36, 148.45, 139.84, 138.33, 130.79, 130.29,
3
29.47(2C), 127.27, 127.10, 125.91, 125.75, 125.70, 125.60(2C), 120.79, 118.24, 104.04,
-
0.01, 56.53, 21.24. HRMS (ESI): calcd for C21
H15BrN
4
O
3
[M-H] 449.0255, found 449.0279.
1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-ethylphenyl)-1H-1,2,3-triazole
(
10b-8)
Yellow solid, yield 85.3%. m.p. 248.5-249.5 °C. H NMR (400 MHz, CDCl
1
3
) δ 8.87 (d, J =
.8 Hz, 1H, 6-CH-), 8.67 (s, 1H, 8-CH-), 7.82 (t, J = 8.4 Hz, 1H, 5-CH-), 7.67–7.59 (m, 3H,
-CH- and 2’,6’-CH-), 7.32 (s, 1H, triazole-H), 7.19 (d, J = 7.9 Hz, 2H, 3’,5’-CH-), 6.34 (br s,
H, 3-CH-), 5.16 (d, J = 12.0 Hz, 1H, 2-CH -), 4.82 (d, J = 11.7 Hz, 1H, 2-CH -), 2.64 (q, J =
.6 Hz, 2H, 7’-CH -), 1.21 (t, J = 7.6 Hz, 3H, 8’-CH ). C NMR (100 MHz, DMSO-d ) δ
8
4
1
7
1
1
2
2
13
2
3
6
53.88, 146.51, 143.58, 139.13, 130.61, 129.85, 128.11(2C), 127.79, 127.25(2C), 126.76,
25.23, 124.90, 124.26, 120.78, 120.56, 102.50, 69.96, 55.44, 27.84, 15.36. HRMS (ESI):
-
calcd for C22
H17BrN
4
O
3
[M-H] 463.0411, found 463.0432.
1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-propylphenyl)-1H-1,2,3-triazole
(10b-9)
2
5

ACCEPTED MANUSCRIPT
1
Yellow solid, yield 84.3%. m.p. 191.3-192.7 °C. H NMR (400 MHz, CDCl ) δ 8.88 (d, J =
3
8
4
1
7
.9 Hz, 1H, 6-CH-), 8.68 (s, 1H, 8-CH-), 7.81 (t, J = 8.9 Hz, 1H, 5-CH-), 7.68–7.61 (m, 3H,
-CH- and 2’,6’-CH-), 7.33 (s, 1H, triazole-H), 7.18 (d, J = 8.0 Hz, 2H, 3’,5’-CH-), 6.36 (br s,
2 2
H, 3-CH-), 5.17 (d, J = 11.8 Hz, 1H, 2-CH -), 4.84 (d, J = 9.8 Hz, 1H, 2-CH -), 2.58 (t, J =
1
3
.6 Hz, 2H, 7’-CH
2 2 3
-), 1.66–1.58 (m, 2H, 8’-CH -), 0.92 (t, J = 7.3 Hz, 3H, 9’-CH ). C NMR
(
100 MHz, DMSO-d ) δ 153.88, 146.53, 141.96, 139.10, 130.59, 129.86, 128.68(2C), 127.81,
6
127.23, 126.76, 125.15(2C), 124.91, 124.26, 120.76, 120.56, 102.51, 69.96, 55.44, 36.90,
-
23.87, 13.50. HRMS (ESI): calcd for C23
H19BrN
4
O
3
[M-H] 477.0568, found 477.0581.
1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-butylphenyl)-1H-1,2,3-triazole
(
10b-10)
Yellow solid, yield 80.8%. m.p. 205.3-206.9 °C. H NMR (400 MHz, CDCl
1
3
) δ 8.87 (d, J =
.9 Hz, 1H, 6-CH-), 8.67 (s, 1H, 8-CH-), 7.82 (t, J = 8.4 Hz, 1H, 5-CH-), 7.67–7.56 (m, 3H,
-CH- and 2’,6’-CH-), 7.33 (s, 1H, triazole-H), 7.18 (d, J = 8.0 Hz, 2H, 3’,5’-CH-), 6.35 (br s,
H, 3-CH-), 5.16 (dd, J = 11.9, 2.8 Hz, 1H, 2-CH -), 4.83 (dd, J = 11.9, 3.0 Hz, 1H, 2-CH -),
8
4
1
2
0
1
1
2
2
.61 (t, J = 7.6 Hz, 2H, 7’-CH -), 1.63–1.52 (m, 2H, 8’-CH -), 1.39–1.28 (m, 2H, 9’-CH -),
2
2
2
1
3
.92 (t, J = 7.3 Hz, 3H, 10’-CH
3
). C NMR (100 MHz, DMSO-d
6
) δ 153.86, 146.52, 142.16,
39.09, 130.59, 129.85, 128.61(2C), 127.74, 127.22, 126.74, 125.16(2C), 124.89, 124.25,
20.75, 120.54, 102.50, 69.95, 55.43, 34.48, 32.91, 21.64, 13.67. HRMS (ESI): calcd for
-
C
24
H21BrN
4
O
3
[M-H] 491.0724, found 491.0762.
1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-pentylphenyl)-1H-1,2,3-triazole
2
6

ACCEPTED MANUSCRIPT
(10b-11)
1
Yellow solid, yield 81.6%. m.p. 200.8-201.9 °C. H NMR (400 MHz, CDCl
3
) δ 8.88 (d, J =
8
4
1
7
.8 Hz, 1H, 6-CH-), 8.68 (s, 1H, 8-CH-), 7.83 (t, J = 7.9 Hz, 1H, 5-CH-), 7.68–7.58 (m, 3H,
-CH- and 2’,6’-CH-), 7.34 (s, 1H, triazole-H), 7.18 (d, J = 7.7 Hz, 2H, 3’,5’-CH-), 6.35 (br s,
H, 3-CH-), 5.17 (d, J = 11.0 Hz, 1H, 2-CH -), 4.84 (d, J = 11.5 Hz, 1H, 2-CH -), 2.60 (t, J =
2
2
.5 Hz, 2H, 7’-CH -), 1.64–1.57 (m, 2H, 8’-CH -), 1.34–1.29 (m, 4H, 9’-CH - and 10’ ,
2
2
2
1
3
-
CH
2
-), 0.88 (t, J = 6.2 Hz, 3H, 11’-CH
3
). C NMR (100 MHz, DMSO-d
6
) δ 153.88, 146.53,
142.19, 139.10, 130.59, 129.86, 128.61(2C), 127.76, 127.23, 126.76, 125.16(2C), 124.91,
124.26, 120.75, 120.54, 102.51, 69.95, 55.43, 34.77, 30.78, 30.42, 21.86, 13.83. HRMS (ESI):
-
calcd for C25
H23BrN
4
O
3
[M-H] 505.0881, found 505.0917.
1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-(4-methoxyphenyl)-1H-1,2,3-triazo
le (10b-12)
1
Yellow solid, yield 86.4%. m.p. 228.0-229.1 °C. H NMR (400 MHz, CDCl
3
) δ 8.87 (d, J =
8
4
1
.9 Hz, 1H, 6-CH-), 8.67 (s, 1H, 8-CH-), 7.82 (t, J = 8.4 Hz, 1H, 5-CH-), 7.73–7.58 (m, 3H,
-CH- and 2’,6’-CH-), 7.31 (s, 1H, triazole-H), 6.90 (d, J = 8.2 Hz, 2H, 3’,5’-CH-), 6.34 (br s,
H, 3-CH-), 5.16 (d, J = 11.7 Hz, 1H, 2-CH -), 4.83 (d, J = 11.7 Hz, 1H, 2-CH -), 3.82 (s, 3H,
) δ 159.03, 153.88, 146.38, 139.10, 130.60, 129.86,
2
2
13
-OCH
3
). C NMR (100 MHz, DMSO-d
6
127.21, 126.79, 126.57(2C), 124.90, 124.24, 122.87, 120.75, 119.97, 114.15(2C), 102.50,
-
69.96, 55.40, 55.07. HRMS (ESI): calcd for C21
H15BrN
4
O
4
[M-H] 465.0204, found 465.0152.
1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-4-cyclopropyl-1H-1,2,3-triazole
27

ACCEPTED MANUSCRIPT
(10b-13)
1
Yellow solid, yield 74.3%. m.p. 176.4-177.8 °C. H NMR (400 MHz, CDCl
3
) δ 8.85 (d, J =
8
.9 Hz, 1H, 6-CH-), 8.52 (s, 1H, 8-CH-), 7.79 (t, J = 7.9 Hz, 1H, 5-CH-), 7.57 (d, J = 7.0 Hz,
H, 4-CH-), 7.01 (s, 1H, triazole-H), 6.25 (br s, 1H, 3-CH-), 5.07 (d, J = 11.7 Hz, 1H,
-CH -), 4.78 (d, J = 11.7 Hz, 1H, 2-CH -), 1.91–1.77 (m, 1H, cyclopropyl 1’-CH-),
.95–0.85 (m, 2H, cyclopropyl 2’-CH - or 3’ -CH -), 0.85–0.73 (m, 2H, cyclopropyl 3’ -CH -
1
2
2
2
0
2
2
2
13
2 6
or 2’ -CH -). C NMR (100 MHz, DMSO-d ) δ 153.83, 149.02, 139.06, 130.57, 129.83,
127.08, 126.98, 124.82, 124.10, 120.64, 120.32, 102.37, 69.89, 55.07, 7.54, 7.49, 6.36.
-
4 3
HRMS (ESI): calcd for C17H13BrN O [M-H] 399.0098, found 399.0073.
2
-(1-(9-bromo-7-nitro-2,3-dihydrobenzo[de]chromen-3-yl)-1H-1,2,3-triazol-4-yl)pyridine
10b-14)
Yellow solid, yield 94.1%. m.p. 249.4251.1 °C. H NMR (400 MHz, CDCl
(
1
3
) δ 8.88 (d, J = 8.7
Hz, 1H, 6-CH-), 8.67 (s, 1H, 8-CH-), 8.47 (d, J = 4.1 Hz, 1H, pyridyl-3’-H), 8.17 (d, J = 7.6
Hz, 1H, pyridyl-6’-H), 7.85–7.72 (m, 3H, 5-CH-, pyridyl-5’-H and triazole-H), 7.65 (d, J =
6.0 Hz, 1H, 4-CH-), 7.22 (t, J = 6.0 Hz, 1H, pyridyl-4’-H), 6.35 (br s, 1H, 3-CH-), 5.20 (d, J =
13
1
1.7 Hz, 1H, 2-CH
2
-), 4.81 (dd, J = 12.0, 1.8 Hz, 1H, 2-CH
2
-). C NMR (100 MHz,
DMSO-d
6
) δ 153.81, 149.52(2C), 147.29, 139.08, 137.17, 130.57, 129.89, 127.09, 126.85,
124.89, 124.31, 123.08, 122.70, 120.68, 119.45, 102.43, 69.76, 55.51. HRMS (ESI): calcd for
-
C
19
H
5 3
12BrN O [M-H] 436.0051, found 436.0034.
General procedure for the preparation of 12a-1~ 12a-14
2
8