Introduction of amino moiety enhances the inhibitory potency of 1-tetralone chalcone derivatives against LPS-stimulated reactive oxygen species production in RAW 264.7 macrophages

Article abstract of DOI:10.1016/j.bioorg.2019.03.055

The design and synthesis of a series of thirty-two halogenated 1-tetralone or 6-amino-1-tetralone chalcone derivatives was achieved by the Claisen-Schmidt condensation reaction and were evaluated for their inhibitory effects against ROS production in LPS-stimulated RAW 264.7 macrophages. It was observed that the introduction of amino moiety into 1-tetralone skeleton greatly increased the inhibitory potency compared to corresponding 1-tetralone chalcones. Among the synthesized compounds, compound 18 which consists of 6-amino-1-tetralone skeleton together with o-fluorobenzylidene showed the most potent ROS inhibitory effect with IC₅₀ value of 0.25 ± 0.13 μM. SAR analysis revealed that amino moiety at the 6th position of 1-tetralone chalcones have an important role for exerting the greater ROS inhibitory potency in LPS-stimulated RAW 264.7 macrophages than those exhibited by 1-tetralone chalcones alone.

Full text of DOI:10.1016/j.bioorg.2019.03.055

Accepted Manuscript
Intro duction of amino moiety enhances the inhibitory potency of 1-tetralone
chalcone derivatives against LPS-stimulated reactive oxygen species production
in RAW 264.7 macrophages
Pramila Katila, Aastha Shrestha, Aarajana Shrestha, Ritina Shrestha, Pil-Hoon
Park, Eung-Seok Lee
PII:
S0045-2068(19)30125-7
DOI:
https://doi.org/10.1016/j.bioorg.2019.03.055
YBIOO 2881
Reference:
Bioorganic Chemistry
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Revised Date:
Accepted Date:
28 January 2019
18 March 2019
19 March 2019
Please cite this article as: P. Katila, A. Shrestha, A. Shrestha, R. Shrestha, P-H. Park, E-S. Lee, Intro duction of
amino moiety enhances the inhibitory potency of 1-tetralone chalcone derivatives against LPS-stimulated reactive
oxygen species production in RAW 264.7 macrophages, Bioorganic Chemistry (2019), doi: https://doi.org/10.1016/
j.bioorg.2019.03.055
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Introduction of amino moiety enhances the inhibitory potency of 1-tetralone chalcone
derivatives against LPS-stimulated reactive oxygen species production in RAW 264.7
macrophages
Pramila Katila1, Aastha Shrestha1, Aarajana Shrestha, Ritina Shrestha, Pil-Hoon Park*, Eung-
Seok Lee*
College of Pharmacy, Yeungnam University, Gyeongsan 38541, Republic of Korea
* Corresponding authors.
Tel.: +82 53 810-2826; fax: +82 53 810-4654; e-mail: parkp@yu.ac.kr (P.-H. Park)
Tel.: +82 53 810-2827; fax: +82 53 810-4654; e-mail: eslee@yu.ac.kr (E.-S. Lee)
1 Authors have contributed equally.
1

Introduction of amino moiety enhances the inhibitory potency of 1-tetralone chalcone
derivatives against LPS-stimulated reactive oxygen species production in RAW 264.7
macrophages
Pramila Katila1, Aastha Shrestha1, Aarajana Shrestha, Ritina Shrestha, Pil-Hoon Park*, Eung-
Seok Lee*
College of Pharmacy, Yeungnam University, Gyeongsan 38541, Republic of Korea
Abstract
The design and synthesis of a series of thirty-two halogenated 1-tetralone or 6-amino-1-
tetralone chalcone derivatives was achieved by the Claisen-Schmidt condensation reaction
and were evaluated for their inhibitory effects against ROS production in LPS-stimulated
RAW 264.7 macrophages. It was observed that the introduction of amino moiety into 1-
tetralone skeleton greatly increased the inhibitory potency compared to corresponding 1-
tetralone chalcones. Among the synthesized compounds, compound 18 which consists of 6-
amino-1-tetralone skeleton together with o-fluorobenzylidene showed the most potent ROS
inhibitory effect with IC50 value of 0.25±0.13 µM. SAR analysis revealed that amino moiety
at the 6th position of 1-tetralone chalcones have an important role for exerting the greater
ROS inhibitory potency in LPS-stimulated RAW 264.7 macrophages than those exhibited by
1-tetralone chalcones alone.
Keywords: Inflammation; 1-Tetralone chalcone derivatives; 6-Amino-1-tetralone chalcone
derivatives; Inhibition of LPS-stimulated ROS production; Structure-activity relationship
study
2

1. Introduction
Inflammation is an important part of the body’s immune response to any kind of infections,
injury, pain, toxin, or stimuli. During the processes of wound healing, these responses are of
vital importance for removing infections. However if they persist longer than necessary, can
cause harmful effects, resulting in chronic inflammation. These continuous chronic
inflammatory responses that cause detrimental effects to one’s body and tissues are thought to
be mediated through reactive oxygen species (ROS) [1, 2], and has been associated with
many diseases such as rheumatoid arthritis [3], atherosclerosis [4], Alzheimer’s disease [5],
inflammatory bowel disease [6], and cancer [7]. ROS are the products of molecular oxygen,
and even while respiring we tend to break down oxygen and generate ROS. They are highly
reactive in nature and can oxidize biologically important molecules such as proteins, lipids,
including DNA as well. ROS has dual role acting both as a signaling molecule under
physiological conditions and mediator of inflammatory processes at high concentrations [8].
Our body is at constant attack from ROS, and in such instances there is a defense mechanism
system of antioxidant in our body that keeps it under control. Whenever this balance between
the ROS and defense mechanism is disrupted, oxidative stress arises which plays an
imminent role in the progression of many pathological disorders [9]. Therefore, controlling
the overproduction of ROS and balancing the oxidative stress can be considered as one of the
vital strategies in the treatment of inflammation.
Macrophages, as a part of the immune system, play an important role in defending our body
against the pathogenic stimuli via engulfing process called phagocytosis. During
phagocytosis, many inflammatory mediators like cytokines, chemokines, and nitric oxide
(NO) work together with inflammatory response assisting and ensuring a quick resolution and
3

restoration of the normal tissue architecture [10]. Lipopolysaccharide (LPS), acting as a
potent toxin, activates macrophages even at very low concentration [11]. Stimulation of the
macrophages with LPS induces rapid generation of ROS, which leads to the production of
various inflammatory mediators, including tumor necrosis factor-α, interferons and
interleukins [12]. Furthermore, the ROS may interact with NO, resulting in the formation of
reactive nitrogen species (RNS) and ultimately increasing the oxidative and nitrosative stress
responses [13]. Therefore, pharmacological intervention involving the control and balance of
ROS production in macrophages would be a promising strategy in the treatment of
inflammatory diseases.
Chalcone and its various derivatives (Fig. 1a and Fig. 1b) are naturally occurring compounds
widely found in many plants, fruits, spices, vegetables, etc. possessing valuable properties as
anti-inflammatory [14], anti-cancer [15], anti-fungal [16], and anti-oxidant [17]. The use of
chalcones as medicines dates back from very ancient times to the present day. These days
interest in the synthesis of new chalcone analogues have evolved tremendously owing to their
simple structure along with numerous pharmacological activities. The two aryl rings present
in the chalcone structure aids flexibility in the structural modifications for designing and in
the innovation of various novel pharmaceuticals. Recently, many chalcone-based compounds
are available in the market as metochalcone (choleretic drug) and sofalcone (antiulcer and
mucoprotective drug) [18]. Many studies illustrating the role of chalcones in inflammation
have been reported lately. Compounds with polymethoxychalcones showed potential
properties as anti-inflammatory agents, inflammatory cytokine expression inhibitors, and
prostaglandin production inhibitors [19]. Dimethylaminophenyl chalcone derivatives (Fig. 1c)
have been reported to be potential NO and prostaglandin E2 generation inhibitors in RAW
4

264.7 macrophage cell line [20]. Additionally, our research group has continuously explored
several modifications in chalcone skeleton and synthesized 3-benzylidenechroman-4-ones
(Fig. 1d) and 2-benzylidene-1-indanones (Fig. 1e) as rigid chalcone derivatives with
moderate to strong inhibitory activity against ROS production in RAW 264.7 macrophages
[21, 22].
Figure 1. Structures of a) chalcone, b) diphenyl substituted chalcones, c)
dimethylaminophenyl chalcone derivatives d) 3-benzylidenechroman-4-one derivatives e) 2-
benzylidene-1-indanone substituted derivatives, and f) 4-amino-3,4-dihydro-2H-napthalen-1-
one derivatives.
Halogenation has evolved as a prominent technique in the discovery of new drugs, and about
40% of drugs used in clinical trials are incorporated by addition of halogen functionalities.
Similarly, many interesting features of halogen atoms like high electronegativity, its ability to
enhance CNS penetration, improvement in metabolic and chemical stability are of great
importance in the design and optimization of halogenated drugs [23]. Recent findings
5

reported that insertion of chlorine or nitro groups in the phenyl ring of chalcone resulted in
better anti-inflammation properties as well as were selective in the inhibition of NO
generation [24]. Studies carried out on chlorinated chalcones suggested that their potent anti-
inflammatory and antioxidant activity was possible due to the presence of electron
withdrawing groups like chlorine [25]. Similarly, the chemical substitution of bromine in
methyl hydroquinone revealed the importance of bromination in anti-inflammation [26].
Rojas et al. evaluated a series of fluorinated trimethoxychalcone derivatives, and found that
the compounds inhibited the NO production and PGE2 overproduction in LPS-stimulated
RAW 264.7 macrophages [27]. 1-Tetralones, also known as 3,4-dihydro-2H-napthalen-1-one,
are important class of compounds, and several derivatives of them have shown important
biological activities such as anti-tumor [28], neurological disorders [29, 30], and
antimicrobial activity [31]. Similarly, the novel amine derivatives of 4-amino-3,4-dihydro-
2H-napthalen-1-one (Fig. 1f) studied by Barlow et al. displayed significant mast cell
stabilizing property, and it was also interesting to know that the 1-tetralone nucleus along
with aminocyclopentyl group was necessary for exerting the anti-inflammatory effects [32].
Considering these valuable biological properties of 1-tetralone along with the important
significance of amino and halogenated functionalities has led us to incorporate these
functional groups in this study as shown in Fig. 2.
6

Figure 2. Strategy design of halogenated 2-benzylidene-3,4-dihydronaphthalen-1(2H)-one
and 6-amino-2-benzylidene-3,4-dihydronaphthalen-1(2H)-one derivatives.
In this present study, we report the design and synthesis of a series of thirty-two 1-tetralone or
6-amino-1-tetralone along with halogenated functionalities on 2-benzylidene moiety at ortho,
meta, or para-position which were evaluated for their inhibitory effects against the ROS-
production in LPS-stimulated 264.7 macrophages. A Structure-activity relationship (SAR)
study was performed with respect to ortho, meta, or para- substitution along with different
halogenated functionality on the phenyl ring, and with presence or absence of amino group in
the 1-tetralone skeleton as well as with the previously synthesized other rigid skeleton of
chalcones. Malvidin is a flavonoid present abundantly in blueberries, strawberries, and
cranberries which renders them the characteristics color pigments. One of the studies
7

involving malvidin, their mixtures of malvidin-3-glucoside and malvidin-3-galactoside
considerably decreased the levels of ROS and oxidative stress in endothelial cells possessing
a potent antioxidant activity [33]. Another study on malvidin reported that it inhibits 50% of
LPS-induced ROS production in RAW 264.7 macrophages at the concentration of 9.0± 0.8
µM and used as the positive control in this study [34].
2. Results and discussion
2.1. Chemistry
At first, halogenated 1-tetralone or 6-amino-1-tetralone derivatives (1-32) were synthesized
by previously reported [35, 36] condensation reaction with 1-tetralone / 6-amino-1-tetralone
and halogenated aryl aldehyde (R1= a-p) as summarized in Scheme 1 and Scheme 2. Base-
catalyzed Claisen-Schmidt condensation reaction was used in which 50% aqueous NaOH or
KOH (0.5 mL) was added to a solution of 1-tetralone / 6-amino-1-tetralone (1.0 mmol) and
halogenated aryl aldehyde (R1= a-p) (1.2 mmol) in ethanol (4 mL), which resulted in the pure
desired chalcone derivatives in 20.0 - 96.3% yield.
The preparation of compounds 1-13, 15, and 17 have been reported in previous studies [37-43]
whereas characterization of compounds 8, 11, 12, and 15 were not found so we have included
in this study. It was interesting to know that among non-aminolated compounds 1-16,
compounds 14 and 16 were novel and not found to be reported. On the other hand, among
aminolated compounds 17-32, except compound 17, all of the compounds 18-32 are also
novel and are reported for the first time in this study.
8

Scheme 1. Schematic representation for the synthesis of halogenated 1-tetralone substituted
chalcone derivatives 1-16. Reagents and conditions: (i) 50% aq. NaOH (4.5 equiv., 0.5 mL),
EtOH (4 mL), 3-36 h, 25 ℃, 21.5 - 96.3% yield.
9

Scheme 2. Schematic representation for the synthesis of halogenated 6-amino-1-tetralone
substituted chalcone derivatives 17-32. Reagents and conditions: (i) 50% aq. KOH (4.5 equiv.,
0.5 mL), EtOH (4 mL), 3-24 h, 25 ℃, 20.0 - 52.1% yield.
The structures of the synthesized target compounds (1-32) are illustrated as shown in Fig. 3
and Fig. 4. Amongst, the first sixteen synthesized 1-tetralone chalcones, compound 1
contained 1-tetralone moiety along with phenyl moiety whereas compounds 2-4, 5-7, 8-10,
11-13, and 14-16 contained 1-tetralone moiety along with ortho-, meta-, or para-
fluorophenyl, chlorophenyl, bromophenyl, trifluoromethylphenyl, and trifluoromethoxy-
phenyl moiety, respectively (Fig. 3). Similarly, amongst, the next sixteen synthesized 6-
amino-1-tetralone chalcones, compound 17 contained 6-amino-tetralone moiety along with
phenyl moiety whereas compounds 18-20, 21-23, 24-26, 27-29, and 30-32 contained 6-
amino-tetralone moiety along with ortho-, meta-, or para-fluorophenyl chlorophenyl,
bromophenyl, trifluoromethyl-phenyl, and trifluoromethoxyphenyl moiety, respectively (Fig.
4). The SAR was determined according to the different position of fluoro, chloro, bromo,
trifluoromethyl, or trifluoromethoxy group, as well as the presence or absence of the amino
group of the 1-tetralone or 6-amino-1-tetralone chalcone derivatives.
10

Figure 3. Synthesized halogenated 1-tetralone substituted chalcone derivatives 1-16.
Figure 4. Synthesized halogenated 6-amino-1-tetralone substituted chalcone derivatives 17-
32.
11

2.2. Inhibitory activities against the ROS production stimulated by LPS in RAW 264.7
macrophages
The compounds 1-32 have been evaluated for their inhibitory activities against the ROS
production stimulated by LPS in RAW 264.7 macrophages, and are illustrated in Table 1.
Most of the prepared compounds exhibited potent ROS inhibitory activities, especially 6-
amino-1-tetralone substituted chalcone derivatives. Amongst the first sixteen series of
synthesized compounds 1-16, that had the basic skeleton of 1-tetralone together with fluorine,
chlorine, bromine, trifluoromethyl, or trifluoromethoxy substituent at the ortho, meta, or
para-position on the 2-benzylidene ring of chalcone, compounds 1, 5-10, and 12-16
significantly inhibited LPS-stimulated ROS production in RAW 264.7 macrophages (1.27 to
6.24 µM of IC50). Compared to 1-tetralone with unsubstituted 2-benzylidene moiety of
chalcone (compound 1), compounds 5, 7, 10, 12, and 16 which employed o-chloro, p-chloro,
p-bromo, m-trifluoromethyl, and p-trifluoromethoxy benzylidene moiety, respectively, greatly
increased inhibitory potency, which indicated o-chloro, p-chloro, p-bromo, m-trifluoromethyl,
and p-trifluoromethoxy substitution on 2-benzylidene moiety is important for strong
inhibition against LPS-stimulated ROS production in RAW 264.7 macrophages. Likewise,
among the next sixteen series of the synthesized compounds 17-32, that had the basic
skeleton of 6-amino-1-tetralone together with fluorine, chlorine, bromine, trifluoromethyl, or
trifluoromethoxy substituent at the ortho, meta, or para-position on the 2-benzylidene ring of
chalcone, all of the compounds 17-32 showed strong inhibitory activity, and most of the
compounds greatly increased inhibitory activity against LPS-stimulated ROS production in
RAW 264.7 macrophages (0.25 to 8.77 µM of IC50). Compared to 6-amino-1-tetralone with
unsubstituted 2-benzylidene moiety of chalcone (compound 17), compounds 18, 20, and 28
12

which employed o-fluoro, p-fluoro, and m-trifluoromethyl benzylidene moiety, respectively,
greatly increased inhibitory potency, which indicated o-fluoro, p-fluoro, and m-
trifluoromethyl substitution on 2-benzylidene moiety, is essential for strong inhibition against
LPS-stimulated ROS production in RAW 264.7 macrophages. These findings are quite
unexpected results since inhibitory effects depending on substitution of functional groups on
2- benzylidene moiety for 1-tetralone and 6-amino-1-tetralone are not concordance. Among
the thirty-two synthesized 1-tetralone / 6-amino-1-tetralone substituted chalcone compounds
(1-32), none of the compounds showed toxicity in RAW 264.7 macrophages from three
different experiments performed in triplicate.
Structure-activity relationship study (SAR) of the first sixteen compounds having 1-tetralone
as a basic skeleton revealed that the compound possessing p-bromophenyl moiety showed the
most significant inhibition (1.27±0.29 µM of IC50) whereas compound possessing fluoro and
o-trifluoromethylphenyl moiety showed the least significant inhibition (>10 µM of IC50) of
LPS-stimulated ROS production in RAW 264.7 macrophages. Comparing inhibitory activities
between different halogenated functionalities, it was observed that p-bromophenyl (1.27±0.29
µM of IC50) substituent possessed the most significant ROS inhibition followed by p-
chlorophenyl (1.55±0.64 µM of IC50), p-trifluoromethoxyphenyl (1.90±0.23 µM of IC50), m-
trifluoromethylphenyl (3.02±0.72 µM of IC50), and phenyl (5.43±1.80 µM of IC50)
substituent, respectively. SAR between ortho, meta, and para substituents of halogen
functionalities generally indicated that para substituent displayed the most significant ROS
inhibition followed by meta, and then ortho substituents.
13

Table 1. Inhibitory activities of compounds 1-32 on ROS production stimulated by LPS in RAW 264.7 macrophages and relative ROS
inhibitory potencies of 1-tetralone chalcones (1-16) and 6-amino-1-tetralone chalcones (17-32) with respect to positive control, malvidin.
Compounds
ROS Inhibition
(IC50,a µM)
Toxicity
b Relative ROS
Inhibitory Potency of
(1-16) with respect to
malvidin
Compounds
ROS Inhibition Toxicity
(IC50,a µM)
b Relative ROS
Inhibitory Potency of
(17-32) with respect
to malvidin
5.43±1.80
>10
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
1.7
<0.9
<0.9
<0.9
2.9
0.76±0.11
0.25±0.13
0.76±0.09
0.40±0.15
1.04±0.02
1.12±0.06
2.93±0.49
2.88±0.81
0.80±0.11
1.29±0.31
8.77±1.31
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
11.8
36.0
11.8
22.5
8.7
1
2
17
18
19
20
21
22
23
24
25
26
27
>10
3
>10
4
3.11±1.11
6.24±0.99
1.55±0.64
5.35±0.23
4.31±0.67
1.27±0.29
>10
5
1.4
8.0
6
5.8
3.1
7
1.7
3.1
8
2.1
11.3
7.0
9
7.1
10
11
<0.9
1.0
14

3.02±0.72
4.18±1.40
5.35±0.75
3.99±0.51
1.90±0.23
NT
NT
NT
NT
NT
3.0
2.2
1.7
2.3
4.7
0.41±0.04
4.51±1.39
3.75±1.38
0.84±0.12
1.06±0.29
NT
NT
NT
NT
NT
22.0
2.0
12
13
14
15
16
28
29
30
31
32
2.4
10.7
8.5
Malvidin
[34]
9.0±0.8
NT: Non-toxic: Cytotoxicity of the compounds was evaluated by cell viability assay (MTS assay). For this, as indicated in the experimental
methods, macrophages were treated with the compound for 24 h up to 10 µM. All the compounds used in this study did not affect cell
viability in this condition.
aEach data represents mean ± S.D. from three different experiments performed in triplicate.
bRelative ROS inhibitory potency: IC50 (µM) of Malvidin / IC50 (µM) of synthesized compounds (1-32).
15

Inhibitory effects possessing 6-amino-1-tetralone (compounds 17–32) as a basic skeleton
indicated that the compound with o-fluorophenyl moiety showed the most significant
inhibition (0.25±0.13 µM of IC50) whereas compound possessing o-trifluoromethyl moiety
showed the least significant inhibition (8.77±1.31 µM of IC50) of LPS-stimulated ROS
production in RAW 264.7 macrophages. SAR of compounds 17–32 possessing 6-amino-1-
tetralone moiety according to ortho, meta, and para substituents of halogen functionalities
generally exhibited that meta substituents showed better inhibitory activities with the
exception of fluoro functionalities. Since there is no solid relationship between different
functionalities of compounds, it is difficult to determine strict SAR.
However, SAR study revealed that the substitution of amino group at the 6th position of 1-
tetralone ring significantly enhanced the ROS inhibition indicating that the 6-amino-1-
tetralone moiety has a greater contribution to ROS inhibitory activity than the 1-tetralone
moiety as shown in Table 1. Comparing inhibitory activities between 1-tetralone and 6-
amino-1-tetralone substituted chalcone derivatives, there was significant increase in the level
of ROS inhibition by all the 6-amino-1-tetralone chalcone derivatives 17-32, except
compounds 23 and 29, than their corresponding 1-tetralone chalcone derivatives 1-16.
Furthermore, among all the halogenated functionalities, fluorine containing 1- tetralone
chalcone derivatives 2-4 did not possess ROS inhibitory activities at all, whereas 6-amino-1-
tetralone chalcone derivatives 18-20, respectively, significantly enhanced the ROS inhibition
potency even in sub-micromolar range (0.25 to 0.76 µM of IC50) as shown in Table 1.
Compounds 18 and 20 also exhibited the most potent ROS inhibition compared to all the
synthesized 1-tetralone and 6-amino-1-tetralone chalcone derivatives. Further, relative ROS
16

inhibitory potencies of 6-amino-1-tetralone chalcones (17-32) and 1-tetralone chalcones (1-
16) with respect to malvidin showed that 6-amino substituted compounds 17, 18, 19, 20, 25,
28, and 31 were 11.8, 36.0, 11.8, 22.5, 11.3, 22.0, and 10.7 times more potent, respectively,
compared to positive control, malvidin, whereas their non-amino substituted compounds 1, 2,
3, 4, 9, 12, and 15 were only 1.7, <0.9, <0.9, <0.9, 2.1, 3.0, and 2.3 times potent, respectively,
as shown in Table 1. This relative potency revealed that introduction of amino group at the 6th
position of 1-tetraone chalcones significantly enhanced the ROS inhibition in RAW 264.7
macrophages.
Figure 5. Comparison of 1-tetralone, 6-amino-1-tetralone chalcone derivatives, and
previously reported rigid structure [21, 22] for inhibition of ROS production stimulated by
LPS in RAW 264.7 macrophages.
Finally, SAR was also analyzed by comparing the various rigid chalcone derivatives reported
previously by our research group [21, 22] with 1-tetralone and 6-amino-1-tetralone
substituted chalcone derivatives. Among various rigid structures, 6-amino-1-tetralone
derivatives showed the most potent ROS inhibition whereas 4-chromanone and 1-indanone
showed moderate and weak ROS inhibition, respectively, as depicted in Fig. 5.
17

3. Conclusion
In conclusion, we have systematically designed and synthesized a series of thirty-two
halogenated 1-tetralone and 6-amino-1-tetralone chalcone derivatives using Claisen-Schmidt
condensation reaction. The synthesized compounds were evaluated for the inhibitory effect
on ROS production stimulated by LPS in RAW 264.7 macrophages. Among them,
compounds 18 possessing 6-amino-1-tetralone chalcone moiety showed the strongest
inhibitory activity of ROS production stimulated by LPS in RAW 264.7 macrophages. SAR
study revealed that chalcone derivatives having 1-tetralone skeleton is responsible for
exhibiting the potent ROS inhibition than that of previously reported 1-indanone or 4-
chromanone skeleton bearing chalcone derivatives. In addition, amino substitution at the 6th
position of 1-tetralone skeleton greatly increased inhibitory activity of ROS production
stimulated by LPS in RAW 264.7 macrophages. These results would provide new insights to
the researchers working to develop novel anti-inflammatory agents.
4. Experimental section
4.1. Chemistry section
All the commercially available starting materials and reagents were purchased from Sigma-
Aldrich Chemical Co. (Darmstadt, Germany), TCI Chemicals (Tokyo, Japan), Junsei (Tokyo,
18

Japan) and Alfa-Aesar (Heysham, England) and used without any further purification. HPLC
grade acetonitrile (ACN) and methanol were purchased from Budrick and Jackson
(Shanghai,China). Thin layer chromatography (TLC) was performed with kieselgel 60 F254
(Merck, Darmstadt, Germany). Since all the prepared compounds contain aromatic ring, they
were visualized and detected on TLC plates with UV light (short wave, long wave or both).
1
NMR spectra were recorded on a Bruker AMX 250 (250 MHz, FT) for H NMR and 63 MHz
for 13C NMR, and chemical shifts were calibrated according to TMS. Chemical shifts (δ)
were recorded in ppm and coupling constants (J) in hertz (Hz). Melting points were
determined in open capillary tubes on electrothermal 1A 9100 digital melting point apparatus
and were uncorrected.
The HPLC analysis were performed using an HPLC system consisted of a pump (LC-20AD),
an autoinjector (SIL-20-A), a UV-visible detector (SPD-20A), and communications bus
module (CBM-20A) from Shimadzu Scientific Instruments (Kyoto, Japan). A Waters
COSMOSIL 5C18-MS-II column (5μm, 4.6 x 250 mm) was used with a gradient solvent
system of 95:5 for 20 min with 95% ACN: doubly distilled water, at a flow rate of 0.5
mL/min at 254 nm UV detection. The purity of compound was described as percentage (%),
and retention time was given in minutes (min). Mass spectra were measured in positive
electrospray ionization (ESI) mode on LCMS-2020 system from Shimadzu Scientific
Instruments (Tokyo, Japan).
4.2. General method for preparation of compounds 1-32
The general synthetic method for the preparation of compounds 1-32 has been reported
previously [35, 36]. The melting points of the reported compounds and those obtained in the
19

present study are also included. The characterization of the compounds are mentioned as
follows;
4.2.1. 2-benzylidene-3,4-dihydronaphthalen-1(2H)-one (1) [29, 37, 38]
o
o
mp: 109.8–110.7 C (ref[29]: 106.0-107.3 C (ethanol)), HPLC: Retention time: 7.74 min,
purity: 98.2%.
4.2.2. 2-(2-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (2) [37, 39]
o
o
mp: 60.1–61.1 C (ref[39]: 56.0-58.0 C), HPLC: Retention time: 8.51 min, purity: 97.2%,
ESI LC/MS: m/z calcd for C17H13FO [MH] + 253.09; found 253.70.
4.2.3. 2-(3-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (3) [37, 39]
o
o
mp: 87.9–88.7 C (ref[39]: 89.0-91.0 C), HPLC: Retention time: 8.41 min, purity: 95.2%,
ESI LC/MS: m/z calcd for C17H13FO [MH] + 253.09; found 253.70.
4.2.4. 2-(4-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (4) [37, 39, 40]
o
o
mp: 113.1–114.1 C (ref[40]: 112.0-116.0 C), HPLC: Retention time: 8.50 min, purity:
95.5%, ESI LC/MS: m/z calcd for C17H13FO [MH]+ 253.09; found 253.70.
4.2.5. 2-(2-chlorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (5) [40]
o
o
mp: 72.4-73.2 C (ref[40]: 109.0-112.0 C), HPLC: Retention time: 9.19 min, purity: 95.2%,
ESI LC/MS: m/z calcd for C17H13ClO [MH] + 269.06; found 269.65.
20

4.2.6. 2-(3-chlorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (6) [40]
o
o
mp: 111.4–112.1 C (ref[40]: 106.0-108.0 C), HPLC: Retention time: 9.57 min, purity:
96.2%, ESI LC/MS: m/z calcd for C17H13ClO [MH] + 269.06; found 269.65.
4.2.7. 2-(4-chlorobenzylidene)-3,4dihydronaphthalen-1(2H)-one (7) [38, 40]
o
o
mp: 139.0–139.7 C (ref[40]: 134.0-136.0 C), HPLC: Retention time: 9.59 min, purity:
95.2%.
4.2.8. 2-(2-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (8) [41]
Yield: 86.6%, white solid; TLC (ethyl acetate/hexanes = 1:27 v/v) Rf: 0.26, mp: 77.9–78.5 oC,
o
(ref[41]: 78.0-80.0 C), HPLC: Retention time: 9.52 min, purity: 97.2%, ESI LC/MS: m/z
calcd for C17H13BrO [MH] + 313.01; found 313.55.
1H NMR (250 MHz, CDCl3) δ 8.15 (d, J = 7.8 Hz, 1H), 7.82 (s, 1H), 7.63 (d, J = 7.9 Hz, 1H),
7.47 (dd, J = 10.5, 4.4 Hz, 1H), 7.32 (dt, J = 8.3, 6.6 Hz, 3H), 7.25 – 7.15 (m, 2H), 2.93 (s,
4H).
13C NMR (63 MHz, CDCl3) δ 187.57, 143.42, 136.89, 136.34, 135.58, 133.43, 133.27,
132.93, 130.38, 129.67, 128.30, 128.27, 127.05, 126.97, 124.87, 29.01, 27.27.
4.2.9. 2-(3-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (9) [40]
o
o
mp: 110.9–111.4 C (ref[40]: 109.0-110.0 C), HPLC: Retention time: 9.99 min, purity:
98.7%, ESI LC/MS: m/z calcd for C17H13BrO [MH] + 313.01; found 313.55.
4.2.10. 2-(4-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (10) [40]
21

o
o
mp: 153.5– 154.2 C (ref[40]: 139.0-141.0 C), HPLC: Retention time: 8.84 min, purity:
97.8%, ESI LC/MS: m/z calcd for C17H13BrO [MH] + 313.01; found 313.55.
4.2.11. 2-(2-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (11) [42]
Yield: 92.0%, white solid; TLC (ethyl acetate/hexanes = 1:27 v/v) Rf: 0.21, mp: 77.4–78.4 oC,
HPLC: Retention time: 8.62 min, purity: 98.5%, ESI LC/MS: m/z calcd for C18H13F3O [MH]
+ 303.09; found 303.60.
1H NMR (250 MHz, CDCl3) δ 8.15 (dd, J = 7.8, 1.3 Hz, 1H), 7.96 (s, 1H), 7.72 (d, J = 7.7 Hz,
1H), 7.56 (t, J = 7.4 Hz, 1H), 7.52 – 7.39 (m, 2H), 7.39 – 7.27 (m, 2H), 7.23 (d, J = 7.5 Hz,
1H), 2.92 (dd, J = 9.2, 4.0 Hz, 2H), 2.87 – 2.78 (m, 2H).
13C NMR (63 MHz, CDCl3) δ 187.39, 143.50, 138.05, 134.88 (d, J = 1.9 Hz), 133.50, 133.15,
132.94, 131.47, 130.34, 129.21 (d, J = 30.2 Hz), 128.35, 128.30, 128.04, 127.07, 126.07 (q, J
= 5.3 Hz), 121.75, 29.03, 27.36.
4.2.12. 2-(3-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (12) [42, 43]
Yield: 71.4%, off-white solid; TLC (ethyl acetate/hexanes = 1:29 v/v) Rf: 0.30, mp: 106.5–
o
107.5 C, HPLC: Retention time: 8.86 min, purity: 97.6%, ESI LC/MS: m/z calcd for
C18H13F3O [MH] + 303.09; found 303.60.
1H NMR (250 MHz, DMSO) δ 7.96 (d, J = 7.8 Hz, 1H), 7.81 (d, J = 8.1 Hz, 2H), 7.78 – 7.64
(m, 3H), 7.58 (td, J = 7.4, 1.3 Hz, 1H), 7.39 (dd, J = 13.1, 7.5 Hz, 2H), 3.11 – 3.00 (m, 2H),
3.00 – 2.89 (m, 2H).
22

13C NMR (63 MHz, DMSO) δ 186.73, 143.65, 137.34, 136.51, 133.96, 133.86, 133.68,
132.83, 129.81, 129.60 (d, J = 31.7 Hz), 128.75, 127.59, 127.21, 126.39 (q, J = 3.9 Hz),
125.25 (q, J = 3.6 Hz), 122.04, 27.99, 26.73.
4.2.13. 2-(4-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (13) [38, 42, 43]
mp: 176.9–177.9 oC, HPLC: Retention time: 9.10 min, purity: 99.5%.
4.2.14. 2-(2-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (14)
Yield: 88.9%, light yellowish liquid; TLC (ethyl acetate/hexanes = 1:27 v/v) Rf: 0.28, HPLC:
Retention time: 8.47 min, purity: 95.5%, ESI LC/MS: m/z calcd C18H13F3O2 [MH] + 319.08;
found 319.60
1H NMR (250 MHz, CDCl3) δ 8.14 (dd, J = 7.8, 1.2 Hz, 1H), 7.83 (s, 1H), 7.48 (td, J = 7.4,
1.5 Hz, 1H), 7.43 – 7.27 (m, 5H), 7.23 (d, J = 7.4 Hz, 1H), 2.93 (s, 4H).
13C NMR (63 MHz, CDCl3) δ 187.36, 147.50, 143.43, 138.05, 133.46, 133.19, 130.78,
130.38, 129.76, 129.58, 128.30 (2C), 127.08, 126.54, 121.18 (d, J = 1.3 Hz), 120.49 (d, J =
258.2 Hz), 28.96, 27.55.
4.2.15. 2-(3-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (15) [43]
Yield: 91.2%, white solid; TLC (ethyl acetate/hexanes = 1:18 v/v) Rf: 0.27, mp: 150.8–151.2
oC, HPLC: Retention time: 9.06 min, purity: 97.5%, ESI LC/MS: m/z calcd C18H13F3O2 [MH]
+ 319.08; found 319.55.
1H NMR (250 MHz, CDCl3) δ 8.12 (d, J = 7.8 Hz, 1H), 7.79 (s, 1H), 7.53 – 7.39 (m, 2H),
7.35 (t, J = 7.9 Hz, 2H), 7.28 – 7.16 (m, 3H), 3.14 – 3.03 (m, 2H), 3.01 – 2.90 (m, 2H).
23

13C NMR (63 MHz, CDCl3) δ 187.50, 149.26, 143.16, 137.85, 136.85, 134.68, 133.49,
133.26, 129.85, 128.29, 128.24, 128.15, 127.13, 121.98, 120.74, 120.47 (d, J = 257.5 Hz),
28.75, 27.10.
4.2.16. 2-(4-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (16)
Yield: 40.6%, white solid; TLC (ethyl acetate/hexanes = 1:27 v/v) Rf: 0.25, mp: 136.8–137.8
oC, HPLC: Retention time: 8.58 min, purity: 98.1%, ESI LC/MS: m/z calcd C18H13F3O2 [MH]
+ 319.08; found 319.55.
1H NMR (250 MHz, CDCl3) δ 8.12 (dd, J = 7.7, 1.0 Hz, 1H), 7.81 (s, 1H), 7.52 – 7.40 (m,
3H), 7.35 (t, J = 7.5 Hz, 1H), 7.24 (d, J = 8.1 Hz, 3H), 3.08 (dd, J = 8.9, 4.3 Hz, 2H), 2.94 (t,
J = 6.1 Hz, 2H).
13C NMR (63 MHz, CDCl3) δ 187.58, 149.00, 143.12, 136.19, 134.89, 134.47, 133.42,
133.32, 131.27 (2C), 128.27, 128.21, 127.11, 120.81, 120.43 (d, J = 257.6 Hz), 28.77, 27.11.
4.2.17. 6-amino-2-benzylidene-3,4-dihydronaphthalen-1(2H)-one (17) [29]
o
mp: 175.3–176.2oC, (ref[29]: 139.0-141.0 C (ethanol)), HPLC: Retention time: 7.02 min,
purity: 98.9%.
4.2.18. 6-amino-2-(2-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (18)
Yield: 34.1%, orange solid, TLC (ethyl acetate/hexanes = 1:6v/v) Rf: 0.21, mp: 136.4–137.1
oC, HPLC: Retention time: 7.17 min, purity: 98.9%, ESI LC/MS: m/z calcd for C17H14FNO
[MH] + 268.10; found 268.70.
24

1H NMR (250 MHz, DMSO) δ 7.71 (d, J = 8.6 Hz, 1H), 7.50 (s, 1H), 7.49 – 7.37 (m, 2H),
7.27 (dt, J = 8.4, 6.3 Hz, 2H), 6.51 (dd, J = 8.6, 2.1 Hz, 1H), 6.34 (d, J = 1.9 Hz, 1H), 6.20 (s,
2H), 2.90 – 2.79 (m, 2H), 2.79 – 2.69 (m, 2H).
13C NMR (63 MHz, DMSO) δ 183.88, 162.16, 154.31, 145.99, 139.01, 130.96 (d, J = 3.0 Hz),
130.59 (d, J = 8.5 Hz), 130.40, 125.56 (d, J = 3.2 Hz), 124.54 (d, J = 3.4 Hz), 123.65 (d, J =
14.0 Hz), 121.49, 115.81 (d, J = 21.8 Hz), 112.88, 110.79, 28.64, 27.25.
4.2.19. 6-amino-2-(3-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (19)
Yield: 20.2%, dark yellow solid, TLC (ethyl acetate/hexanes = 1:6 v/v) Rf: 0.21, mp: 150.5–
o
151.5 C, HPLC: Retention time: 7.37 min, purity: 99.9%, ESI LC/MS: m/z calcd for
C17H14FNO [MH] + 268.10; found 268.70.
1H NMR (250 MHz, CDCl3) δ 7.98 (d, J = 8.5 Hz, 1H), 7.71 (s, 1H), 7.34 (dd, J = 14.0, 7.8
Hz, 1H), 7.20 – 6.94 (m, 3H), 6.58 (dd, J = 8.5, 2.0 Hz, 1H), 6.40 (s, 1H), 4.18 (s, 2H), 3.02
(t, J = 5.9 Hz, 2H), 2.81 (t, J = 6.3 Hz, 2H).
13C NMR (63 MHz, CDCl3) δ 185.99, 151.43, 145.82, 138.43 (d, J = 7.7 Hz), 137.12, 133.67
(d, J = 2.3 Hz), 130.98, 129.84 (d, J = 8.4 Hz), 125.54 (d, J = 2.9 Hz), 124.47, 116.21 (d, J =
21.6 Hz), 114.96 (d, J = 21.3 Hz), 113.54, 112.02, 99.98, 29.07, 27.13.
4.2.20. 6-amino-2-(4-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (20)
Yield: 26.9%, brown solid; TLC (ethyl acetate/hexanes = 1:6 v/v) Rf: 0.20, mp: 215.4–216.3
oC, HPLC: Retention time: 7.1 min, purity: 99.8%, ESI LC/MS: m/z calcd for C17H14FNO
[MH] + 268.10; found 268.70.
25

1H NMR (250 MHz, DMSO) δ 7.83 – 7.68 (m, J = 8.5 Hz, 4H), 7.61 – 7.48 (m, 3H), 7.26 (t,
J = 8.9 Hz, 2H), 6.75 (dd, J = 8.5, 1.9 Hz, 1H), 6.62 (s, 1H), 2.97 (t, J = 6.0 Hz, 2H), 2.78 (t,
J = 6.3 Hz, 2H).
13C NMR (63 MHz, DMSO) δ 184.68, 163.97, 145.55, 136.19 (d, J = 1.2 Hz), 133.05, 132.30,
132.24, 132.21, 132.07, 130.06, 124.43, 115.83, 115.49, 115.27, 113.97, 28.42, 26.77.
4.2.21. 6-amino-2-(2-chlorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (21)
Yield: 22.4%, orange solid; TLC (ethyl acetate/hexanes = 2:5 v/v) Rf: 0.26, mp: 60.7–61.7 oC,
HPLC: Retention time: 7.58 min, purity: 95.1%, ESI LC/MS: m/z calcd for C17H14ClNO [MH]
+ 284.07; found 284.60.
1H NMR (250 MHz, CDCl3) δ 8.00 (d, J = 8.5 Hz, 1H), 7.80 (s, 1H), 7.41 (dd, J = 7.9, 4.0 Hz,
1H), 7.30 – 7.21 (m, 3H), 6.58 (dd, J = 8.5, 2.2 Hz, 1H), 6.39 (d, J = 2.1 Hz, 1H), 4.16 (s,
2H), 2.94 – 2.84 (m, 2H), 2.85 – 2.75 (m, 2H).
13C NMR (63 MHz, CDCl3) δ 185.98, 151.40, 146.02, 137.78, 134.89, 134.71, 132.02,
130.98, 130.44, 129.65, 129.21, 126.26, 124.52, 113.55, 112.08, 29.30, 27.37.
4.2.22. 6-amino-2-(3-chlorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (22)
Yield: 24.3%, yellow solid; TLC (ethyl acetate/hexanes = 1:2v/v) Rf: 0.28, mp: 185.2–186.0
oC, HPLC: Retention time: 7.86 min, purity: 96.9%, ESI LC/MS: m/z calcd for C17H14ClNO
[MH] + 284.07; found 284.65.
1H NMR (250 MHz, CDCl3) δ 7.98 (d, J = 8.5 Hz, 1H), 7.69 (s, 1H), 7.36 (s, 1H), 7.33 – 7.22
(m, 3H), 6.58 (dd, J = 8.5, 2.3 Hz, 1H), 6.40 (d, J = 2.1 Hz, 1H), 4.16 (s, 2H), 3.07 – 2.95 (m,
2H), 2.87 – 2.74 (m, 2H).
26

13C NMR (63 MHz, CDCl3) δ 185.94, 151.41, 145.82, 138.10, 137.27, 134.25, 133.47,
131.00, 129.61, 129.39, 128.08, 127.88, 124.50, 113.56, 112.04, 29.10, 27.15.
4.2.23. 6-amino-2-(4-chlorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (23)
Yield: 24.6%, yellow solid; TLC (ethyl acetate/hexanes = 1:2 v/v) Rf: 0.29, mp: 193.4–194.3
oC, HPLC: Retention time: 7.86 min, purity: 99.5%, ESI LC/MS: m/z calcd for C17H14ClNO
[MH] + 284.07; found 284.65.
1H NMR (250 MHz, CDCl3) δ 7.97 (d, J = 8.5 Hz, 1H), 7.70 (s, 1H), 7.40 – 7.27 (m, 4H),
6.57 (dd, J = 8.5, 2.3 Hz, 1H), 6.39 (d, J = 2.2 Hz, 1H), 4.14 (s, 2H), 3.06 – 2.93 (m, 2H),
2.87 – 2.73 (m, 2H).
13C NMR (63 MHz, CDCl3) δ 185.96, 151.40, 145.74, 136.68, 134.77, 134.01, 133.73,
130.98 (3C), 128.60 (2C), 124.64, 113.57, 112.05, 29.12, 27.17.
4.2.24. 6-amino-2-(2-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (24)
Yield: 20.8%, a yellow solid; TLC (ethyl acetate/hexanes = 2:5 v/v) Rf: 0.24, mp: 121.4–
o
122.3 C, HPLC: Retention time: 8.05 min, purity: 99.7%, ESI LC/MS: m/z calcd for
C17H14BrNO [MH] + 328.02; found 328.50.
1H NMR (250 MHz, CDCl3) δ 8.00 (d, J = 8.5 Hz, 1H), 7.73 (s, 1H), 7.61 (d, J = 7.9 Hz, 1H),
7.35 – 7.22 (m, 2H), 7.21 – 7.11 (m, 1H), 6.58 (dd, J = 8.5, 2.2 Hz, 1H), 6.39 (d, J = 1.9 Hz,
1H), 4.17 (s, 2H), 2.91 – 2.83 (m, 2H), 2.83 – 2.76 (m, 2H).
13C NMR (63 MHz, CDCl3) δ 186.03, 151.40 , 146.05, 137.45, 136.72, 134.22, 132.82,
130.96, 130.46, 129.36, 126.88, 124.84, 124.48, 113.54, 112.08, 29.29, 27.28.
27

4.2.25. 6-amino-2-(3-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (25)
Yield: 20.1%, yellow solid; TLC (ethyl acetate/hexanes = 1:2 v/v) Rf: 0.24, mp: 177.2–178.2
oC, HPLC: Retention time: 8.10 min, purity: 97.7%, ESI LC/MS: m/z calcd for C17H14BrNO
[MH] + 328.02; found 328.45.
1H NMR (250 MHz, CDCl3) δ 8.00 (d, J = 8.5 Hz, 1H), 7.71 (s, 1H), 7.54 (s, 1H), 7.46 (d, J
= 7.3 Hz, 1H), 7.36 – 7.22 (m, 2H), 6.60 (dd, J = 8.5, 2.1 Hz, 1H), 6.43 (d, J = 1.7 Hz, 1H),
4.20 (s, 2H), 3.09 – 2.97 (m, J = 5.9 Hz, 2H), 2.90 – 2.78 (m, 2H).
13C NMR (63 MHz, CDCl3) δ 185.90, 151.42, 145.81, 138.40, 137.31, 133.35, 132.27,
130.99, 130.98, 129.87, 128.31, 124.47, 122.40, 113.56, 112.03, 29.09, 27.12.
4.2.26. 6-amino-2-(4-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one (26)
Yield: 28.3 %, yellow solid; TLC (ethyl acetate/hexanes = 1:2 v/v) Rf: 0.26, mp: 206.8– 207.7
oC, HPLC: Retention time: 8.20 min, purity: 99.1%, ESI LC/MS: m/z calcd for C17H14BrNO
[MH] + 328.02; found 328.45.
1H NMR (250 MHz, CDCl3) δ 7.99 (d, J = 8.5 Hz, 1H), 7.69 (s, 1H), 7.51 (dd, J = 8.3, 1.3 Hz,
2H), 7.30 – 7.24 (m, 2H), 6.58 (dd, J = 8.5, 1.8 Hz, 1H), 6.40 (s, 1H), 4.13 (s, 2H), 3.05 –
2.95 (m, J = 6.6 Hz, 2H), 2.86 – 2.75 (m, 2H).
13C NMR (63 MHz, CDCl3) δ 185.95, 151.40, 145.76, 136.82, 135.27, 133.76, 131.59 (2C),
131.25 (2C), 131.01, 124.71, 122.24, 113.62, 112.09, 29.14, 27.20.
4.2.27. 6-amino-2-(2-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (27)
28

Yield: 20.0%, a yellow solid; TLC (ethyl acetate/hexanes = 1:2v/v) Rf: 0.22 mp: 144.3–145.2
oC, HPLC: Retention time: 7.43 min, purity: 99.12%, ESI LC/MS: m/z calcd for C18H14F3NO
[MH] + 318.10; found 318.55.
1H NMR (250 MHz, CDCl3) δ 8.00 (d, J = 8.5 Hz, 1H), 7.88 (s, 1H), 7.69 (d, J = 7.7 Hz, 1H),
7.53 (t, J = 7.4 Hz, 1H), 7.40 (t, J = 7.5 Hz, 1H), 7.28 (d, J = 7.6 Hz, 1H), 6.57 (dd, J = 8.5,
2.3 Hz, 1H), 6.38 (d, J = 2.2 Hz, 1H), 4.18 (s, 2H), 2.76 (s, 4H).
13C NMR (63 MHz, CDCl3) δ 185.82, 151.51, 146.12, 138.62, 135.35, 131.51, 131.37,
130.98, 130.48, 129.14 (d, J = 30.1 Hz), 127.73, 125.95 (q, J = 5.4 Hz), 124.35, 123.96 (d, J
= 273.9 Hz), 113.55, 112.09, 29.31, 27.37.
4.2.28. 6-amino-2-(3-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (28)
Yield: 20.7%, yellow solid; TLC (ethyl acetate/hexanes = 1:3 v/v) Rf: 0.27 mp: 161.7–162.7
oC, HPLC: Retention time: 7.53 min, purity: 99.3% ESI LC/MS: m/z calcd for C18H14F3NO
[MH] + 318.10; found 318.55.
1H NMR (250 MHz, DMSO) δ 7.80 – 7.61 (m, 5H), 7.60 (s, 1H), 6.51 (dd, J = 8.5, 2.2 Hz,
1H), 6.34 (d, J = 1.9 Hz, 1H), 6.20 (s, 2H), 2.95 (t, J = 5.8 Hz, 2H), 2.74 (t, J = 6.2 Hz, 2H).
13C NMR (63 MHz, DMSO) δ 184.01, 154.30, 145.81, 138.43, 137.11, 133.57, 131.61,
130.37, 129.71, 128.83, 126.13 (d, J = 3.9 Hz), 124.72 (dd, J = 7.0, 3.0 Hz), 124.27 (d, J =
272.5 Hz), 121.54, 112.86, 110.73, 28.55, 26.86.
4.2.29. 6-amino-2-(4-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1(2H)-one (29)
29