Application of a catalyst-free Domino Mannich/Friedel-Crafts alkylation reaction for the synthesis of novel tetrahydroquinolines of potential antitumor activity

Article abstract of DOI:10.1016/j.tet.2017.12.049

A useful and efficient method to construct diversely substituted 1,2,3,4-tetrahydroquinolines in good to excellent yields has been developed through a catalyst-free Domino Mannich and intramolecular Friedel-Crafts alkylation reactions of N-arylamines with

Full text of DOI:10.1016/j.tet.2017.12.049

Accepted Manuscript
Application of a catalyst-free Domino Mannich/Friedel-Crafts alkylation reaction for
the synthesis of novel tetrahydroquinolines of potential antitumor activity
Juan-Carlos Castillo, Elizabeth Jiménez, Jaime Portilla, Braulio Insuasty, Jairo
Quiroga, Rodolfo Moreno-Fuquen, Alan R. Kennedy, Rodrigo Abonia
PII:
S0040-4020(17)31333-9
10.1016/j.tet.2017.12.049
TET 29205
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 30 November 2017
Revised Date: 22 December 2017
Accepted Date: 23 December 2017
Please cite this article as: Castillo J-C, Jiménez E, Portilla J, Insuasty B, Quiroga J, Moreno-Fuquen R,
Kennedy AR, Abonia R, Application of a catalyst-free Domino Mannich/Friedel-Crafts alkylation reaction
for the synthesis of novel tetrahydroquinolines of potential antitumor activity, Tetrahedron (2018), doi:
10.1016/j.tet.2017.12.049.
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Application of catalyst-free Domino
Mannich/Friedel-Crafts
alkylation
reaction for the synthesis of novel
tetrahydroquinolines
antitumor activity
of
potential
Juan-Carlos Castilloa,b,*, Elizabeth Jiménezc, Jaime Portillad,
Braulio Insuastya, Jairo Quirogaa, Rodolfo Moreno-Fuquene, Alan R. Kennedyf, Rodrigo
Aboniaa,*
a Research Group of Heterocyclic Compounds, Department of Chemistry, Universidad del Valle, A. A.
25360, Cali, Colombia
b Grupo de Investigación Núcleo, Departamento de Química y Bioquímica, Universidad de Boyacá,
Carrera 2ª Este No. 64-169, Tunja, Colombia
c Grupo de Investigación en Bioquímica Aplicada, Department of Chemistry, Universidad de los Andes,
Carrera 1 No. 18A-10, Bogotá, Colombia
d Bioorganic Compounds Research Group, Department of Chemistry, Universidad de los Andes, Carrera 1
No. 18A-10, Bogotá, Colombia
e Department of Chemistry, Universidad del Valle, A. A. 25360, Cali, Colombia
f WestCHEM, Departament of Pure and Applied Chemistry, University of Strathclyde, 295 Cathedral
Street, Glasgow G1 1XL, Scotland

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Application
of
catalyst-free
Domino
Leave this area blank for abstract info.
Mannich/Friedel-Crafts alkylation reaction for
the synthesis of novel tetrahydroquinolines of
potential antitumor activity
Juan-Carlos Castilloa,b,*, Elizabeth Jiménezc, Jaime Portillad,
Braulio Insuastya, Jairo Quirogaa, Rodolfo Moreno-Fuquene, Alan R. Kennedyf, Rodrigo Aboniaa,*
a Research Group of Heterocyclic Compounds, Department of Chemistry, Universidad del Valle, A. A. 25360, Cali,
Colombia
b Grupo de Investigación Núcleo, Departamento de Química y Bioquímica, Universidad de Boyacá, Carrera 2ª Este
No. 64-169, Tunja, Colombia
c Grupo de Investigación en Bioquímica Aplicada, Department of Chemistry, Universidad de los Andes, Carrera 1
No. 18A-10, Bogotá, Colombia
d Bioorganic Compounds Research Group, Department of Chemistry, Universidad de los Andes, Carrera 1 No.
18A-10, Bogotá, Colombia
e Department of Chemistry, Universidad del Valle, A. A. 25360, Cali, Colombia
f WestCHEM, Departament of Pure and Applied Chemistry, University of Strathclyde, 295 Cathedral Street,
Glasgow G1 1XL, Scotland

1
ACCEPTED MANUSCRIPT
Tetrahedron
journal homepage: www.elsevier.com
Application of a catalyst-free Domino Mannich/Friedel-Crafts alkylation reaction for
the synthesis of novel tetrahydroquinolines of potential antitumor activity
Juan-Carlos Castilloa,b, , Elizabeth Jiménezc, Jaime Portillad, Braulio Insuastya, Jairo Quirogaa, Rodolfo
Moreno-Fuquene, Alan R. Kennedyf, Rodrigo Aboniaa,
a Research Group of Heterocyclic Compounds, Department of Chemistry, Universidad del Valle, A. A. 25360, Cali, Colombia
b Grupo de Investigación Núcleo, Departamento de Química y Bioquímica, Universidad de Boyacá, Carrera 2ª Este No. 64-169, Tunja, Colombia
c Grupo de Investigación en Bioquímica Aplicada, Department of Chemistry, Universidad de los Andes, Carrera 1 No. 18A-10, Bogotá, Colombia
d Bioorganic Compounds Research Group, Department of Chemistry, Universidad de los Andes, Carrera 1 No. 18A-10, Bogotá, Colombia
e Department of Chemistry, Universidad del Valle, A. A. 25360, Cali, Colombia
f WestCHEM, Departament of Pure and Applied Chemistry, University of Strathclyde, 295 Cathedral Street, Glasgow G1 1XL, Scotland
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
A useful and efficient method to construct diversely substituted 1,2,3,4-tetrahydroquinolines in
good to excellent yields has been developed through a catalyst-free Domino Mannich and
intramolecular Friedel-Crafts alkylation reactions of N-arylamines with paraformaldehyde and
electron-rich olefins via the formation of N-aryl-N-alkylmethyleneiminium ions as the key
intermediates to afford the target products. Nine of the new compounds were evaluated in the
US National Cancer Institute (NCI), where compound 5f (R1 = 6-MeO, R2 = p-ClC6H4 and X =
pyrrolidin-2-onyl) presented a remarkable activity against 57 cancer cell lines, with the most
important GI50 values ranging from 1.46 to 8.28 µM from in vitro assays. Further studies
performed over the active compound 5f on HCT116 colon cancer cells indicated that its effect
on cell death is exerted through a cell cycle arrest (S phase) in a dose dependent manner, as well
as suppression on the cell proliferation process.
Available online
Keywords:
Tetrahydroquinolines
Multicomponent reaction
Mannich-type reaction
Anticancer activity
Cell cycle arrest
2009 Elsevier Ltd. All rights reserved
.
———
Corresponding author. Tel.: +57-8-745-0000; e-mail: juacastillo@uniboyaca.edu.co
Corresponding author. Tel.: +57-5-339-3248; fax: +57-2-339-3248; e-mail: rodrigo.abonia@correounivalle.edu.co

2
Tetrahedron
starting materials,6a-e we are reporting here an efficient metal- and
1. Introduction
ACCEPTED MANUSCRIPT
catalyst-free approach to construct novel and diversely
substituted 1,2,3,4-tetrahydroquinolines of biological interest
from N-aryl-N-benzylamines as precursors under mild reaction
Tetrahydroquinolines are important “privileged scaffolds” due
to their presence in natural occurring products and their wide
range of applications as drugs,1a-c pharmaceuticals,1d
agrochemicals,1e dye-sensitized solar cells (DSSC),1f-h and chiral
ligands in asymmetric synthesis.1i-j Particularly, the 1,2,3,4-
tetrahydroquinoline is an important moiety which is found in
numerous biologically active natural products, as well as
synthetic pharmacologically relevant agents with interesting
conditions.
Previous work
Entry 1. Involving the formation of a C4ꢀC4a new bond
Conditions
ꢀLewis and Bronsted acid catalyzed intramolecular cyclizations
ꢀPdꢀmediated intramolecular Heck reactions
ꢀIntramolecular radical cyclizations
ꢀIntramolecular iodoꢀarylations via iodonium ions
ꢀOrganocatalytic intramolecular hydroarylations
ꢀIntramolecular epoxide ringꢀopening reactions
4
4a
C4ꢀC4a
3
2
N
N
1
H
H
antiviral,2a
antibacterial,2b
antifungal,2c
antimalarial,2d
4
5
antitrypanosomal,2e antidepressant,2f and vasodilator activities.2g
Regarding the cancer chemotherapy, the 1-aroyl-1,2,3,4-
tetrahydroquinoline 1 had antiproliferative activity correlated
with the inhibition of tubulin polymerization,3a whereas the
torcetrapib 2,3b was developed to treat hypercholesterolemia and
Entry 2. Involving the formation of two new bonds in oneꢀstep via iminium ions
R2
R3
Conditions
R2CH=CHR3
Benzotriazole
methodology
"acid catalysis"
and related approaches
N
X
N
N
R1
R1
X
R1
6 (X = Bt, OMe, SPh,
7
5
SO2Ar, CN, O2CMe)
prevent
cardiovascular
disease
(Figure
1).
The
tetrahydroquinoline derivatives 3,3c were found to have moderate
to high modulating activity in multidrug resistance (MDR),
which is one of the obstacles in the chemotherapy of cancer.
Previously, we reviewed the most common synthetic routes to
construct the 1,2,3,4-tetrahydroquinoline framework 5. The first
one and more common route involves an intramolecular
formation of the new C4–C4a bond starting with materials
containing a benzene ring 4 (Entry 1, Scheme 1). Thus, numerous
synthetic strategies with this requirement have been reported,
involving Lewis and Brønsted acid-catalyzed intramolecular
Friedel-Crafts reactions,4a-c Pd-catalyzed intramolecular Heck
cyclizations of 2-iodo-N-arylamine containing allyl moieties,4d-e
and intramolecular radical cyclizations of N-allyl-N-o-
iodoacrylamides with Bu3SnH/Et3B,4f or secondary amides
containing a xanthate moiety with lauroyl peroxide.4g Other less
frequent synthetic approaches involved an intramolecular iodo-
arylation of N-protected-N-allylaniline derivatives with iodonium
ion (IPy2BF4),4h an organocatalytic asymmetric intramolecular
hydroarylation of aniline derivatives bearing an α,β-unsaturated
aldehyde moiety,4i an intramolecular epoxide ring-opening of
diphenylamine,4j and N,N′-di(2-naphthyl)-1,4-diaminobenzene
containing 2,3-epoxypropyl as N-substituent.4k
Scheme 1. More common synthetic approaches to construct the 1,2,3,4-
tetrahydroquinoline framework 5.
2. Results and discussion
This idea was based on the mechanistic hypothesis that a
catalyst-free domino Mannich/intramolecular Friedel-Crafts
alkylation sequence would be possible through the formation of
an N-aryl-N-alkylmethyleneiminium ion type 10 as the key
intermediate (Scheme 2). The present study was initiated with the
stirring of a mixture of N-benzylaniline 8a (1.0 equiv.),
polyformaldehyde (1.5 equiv.) and N-vinyl-2-pyrrolidone 9a (1.1
equiv.) in methanol at room temperature for 24 h. The reaction
was complete after this time but a mixture of products was
obtained. After performing a careful column chromatography, it
was possible to isolate the tetrahydroquinoline 5a in only 16%
yield.6b Encouraged by this result, a solvent screening revealed
that solvents significantly influence the reaction yields because of
the relative insolubility of the polyformaldehyde. Low yields
were observed when solvents such as MeOH, AcOEt, THF or
DMF were used. However, CH3CN showed the best result,
affording the desired compound 5a in 90% yield as unique
product (Scheme 2). The overall eco-compatibility of the process
is highlighted.
R2
MeO
CF3
OH
Me
MeO2C
The main spectroscopic features of the structure of compound
5a corresponded to the presence of C=O absorption band at 1682
cm-1 in the IR spectrum. Six signals corresponding to methylenic
protons between 1.92 and 4.50 ppm and a doublet of doublets
integrating for 1H at 5.43 ppm (dd, J = 5.5, 9.0 Hz) assigned to
the new aza–methynic (N–CH) proton, along with nine aromatic
N
N
F3C
MeO
MeO
N
CF3
O
R1
O
N
3: R1 = pꢀtolyl, R2 = NHMe,
CO2Et
OMe
1
2: Torcetrapib
NHBn, pyrrolidyl, etc.
1
Figure 1. Some tetrahydroquinoline derivatives with remarkable biological
activity.
protons are the most relevant signals in the H NMR spectrum.
The presence of six methylene carbon atoms between 18.4–55.3
ppm, a signal of the aza–methynic carbon atom (N–CH) at 48.2
ppm, seven aromatic CH, three quaternary Cq carbon atoms and
the (C=O) at 175.5 ppm in the 13C NMR spectrum, are in
agreement with the proposed structure for compound 5a. Finally,
a molecular ion with m/z 306 and a base peak with m/z 83, also
confirmed its structure.
Alternatively, Katritzky et al. showed that N-aryl-1H-
benzotriazole-1-methanamines 6 react with electron-rich alkenes
under acid catalysis to led tetrahydroquinolines 5 via the in situ
formation of N-arylmethyleneiminium ions 7 as the key
intermediates (Entry 2, Scheme 1).5a-d Other reagents have also
been used in the generation of the key methyleneiminium ions 7
for the synthesis of tetrahydroquinolines.5e Although the above
methods undoubtedly provide access to a wide variety of
structurally diverse tetrahydroquinolines, they had the
disadvantages of requiring high reaction temperatures, undesired
side reactions, metal-based or Lewis/Brønsted acid catalyzed
reaction conditions. Nowadays, the development of alternative
routes to construct substituted tetrahydroquinolines continue to
be an important goal in contemporary organic synthesis, where
the sequential creation of two or more C−C bonds in a single
chemical operation is significantly valuable. Thus, in
a
continuation of our earlier studies on multiple bond-forming
transformations (MBFTs) involving benzylamine derivatives as

3
Scheme 2. Selective synthesis of the tetrahydroquinoline 5a from the N-
benzylaniline 8a in CH3CN at room temperature.
ACCEPTED MANUSCRIPT
O
O
O
O
N
N
N
N
N
N
N
With the optimal reaction conditions in hand, we turned our
attention to obtain various N-arylimines from solvent-free
reactions between equimolar amounts of several anilines and
diverse aromatic aldehydes. Subsequently, the reduction of the
imines by treatment with NaBH4 in methanol at room
temperature afforded the corresponding N-aryl-N-benzylamines
8b−l in quantitative yields. The appearance in each case of an IR
absorption band attributable to the N–H functionality in the
3373–3442 cm–1 range confirmed the reduction of their
corresponding N-arylimines. The fully characterization of
compounds 8b−l is reported in the Experimental Section.
Cl
N
O
O
Cl
5c: 81%
11c: 12%
5d: 83%
11d: 8%
5a: 90%
5b: 72%
O
O
O
O
O
N
N
N
N
N
N
N
O
O
Cl
Cl
N
O
O
5g: 70%
Cl
Cl
O
11g: 14%
5e: 75%
5f: 93%
5h: 68%
O
O
O
O
N
N
N
N
N
O
N
In order to explore the scope and limits of this Domino
Mannich/Friedel-Crafts alkylation type reaction, some electron-
rich alkenes 9a−c and polyformaldehyde were subjected to
reaction with the previously synthesized amines 8 along with the
commercially available heterocyclic amines 8p−q and primary
amine 16 under the established procedure. The results are listed
in Table 1. The reaction was found to be general, and diversely
substituted tetrahydroquinolines 5a−p were synthesized. It was
found that both electron-donating and electron-withdrawing
substituents on the benzene ring of 8 were suitable for this
reaction, leading efficiently to the corresponding products 5b−i
in 68−93% yield (Table 1). The reaction appeared difficult with
quinolin-2-one based N-arylamines 8j−l, requiring a gentle
heating to obtain the desired products 5j−l in 61−83% yield
(Table 1). Satisfactorily, when N-methylaniline 8m and N-(prop-
2-yn-1-yl)aniline 8n were reacted with 9a, products 5m and 5n
were isolated in 71% and 81% yield, respectively. Additionally,
1-vinylazepan-2-one 9b and dodecyl vinyl ether 9c were used as
electron-rich olefins (Table 1). Therefore, reaction of amine 8f
with alkene 9b provided the corresponding product 5o in 89%
yield. When dodecyl vinyl ether 9c was reacted with 8o, product
5p was isolated in 72% yield. Meanwhile, we found that styrene,
2,3-dihydrofuran and 3,4-dihydro-2H-pyran were inappropriate
substrates for this catalyst-free protocol. From the results of our
earliest studies,6b it was anticipated that the γ-aminoalcohols 11
could also be formed, as shown in Scheme 2. Fortunately,
compounds 11 were isolated as minority side-products in 8−14%
yields (Table 1 and Experimental Section).
Cl
O
N
N
Cl
N
H
O
N
H
O
N
H
5i: 86%
5j: 69%[b]
5k: 61%[b]
5l: 83%[b]
O
O
N
N
N
N
O
N
N
N
O
10
O
O
O
O
O
5m: 71%
11m: 14%
Cl
5p: 72%
5n: 81%
5o: 89%
a
Reaction conditions: N-arylamine 8b−o (1.0 equiv.), alkene 9a−c (1.1
equiv.) and polyformaldehyde (1.5 equiv.) in 2.0 mL of CH3CN at room
temperature for 24 h. b CH3CN/DMSO (3:1, 2.0 mL) at 60 °C for 24 h.
On the basis of the aforementioned results and literature
precedents,5e,7 a plausible mechanistic route was proposed for the
generation of the tetrahydroquinolines, as depicted in Scheme 3.
It starts with the formation of a hemiaminal 12 from the reaction
of N-arylamine 8 with polyformaldehyde, which should be in
reversible equilibrium with the N-arylmethyleneiminium ion type
7. This species should be trapped by the alkene 9 via a Mannich-
type reaction affording a new cationic species type 10 (stabilized
by a resonant effect with the free electronic pair of the X atom)
which should undergo an intramolecular Friedel-Crafts alkylation
on the ortho position of the aromatic ring to generate the
intermediate 13. In the following step, the abstraction of the
hydrogen atom on 13 by the hydroxide anion triggers the rapid
aromatization of the intermediate 13, leading to
tetrahydroquinoline derivatives 5 and releasing water as the
unique byproduct. Otherwise, the formation of γ-aminoalcohols
11 is believed to result from an intermolecular addition of the
hydroxide ion over the N-acyliminium or oxonium cation type 10
(X = N, O) which generated in situ by a Mannich-type reaction
following a stepwise pathway instead of an asynchronous
concerted mechanism (Scheme 3).5e,7
Table 1. Domino Mannich/Friedel-Crafts alkylation reaction for the synthesis
of 1,2,3,4-tetrahydroquinolines 5.a
Scheme 3. Plausible mechanism for the formation of tetrahydroquinolines 5
and γ-aminoalcohols 11 via the iminium ion 7.

4
Tetrahedron
ACCEPTED MANUSCRIPT
The interesting results exhibited in Scheme 2 and Table 1
encouraged us to expand the application scope of this domino
reaction. Thus, cyclic amines 8p and 8q were allowed to react
with polyformaldehyde and N-vinyl-2-pyrrolidone 9a,
successfully generating the expected tetracyclic product 14 and
the julolidine derivative 15 in 91% and 89% yield, respectively
(Scheme 4). To our delight, the same reaction in the presence of
p-toluidine 16 gave the symmetric julolidine 17 in 32% yield
after 14 days of stirring. In a slightly modified protocol, the
addition of acetic acid (0.2 mL) afforded the same product 17 in
an increased 61% yield after 24 h of stirring (Scheme 4). NMR
spectra of the crude product 17 showed the formation of two
diastereomers in a 1.5:1 ratio. However, the NMR spectral data
of the individual diastereomers were insufficiently different to
allow definitive assignment of their exact configurations.
Remarkably, this methodology provided an interesting and
alternative synthetic access to the valuable julolidine scaffold,
which could be used to synthesize julolidine-based fluorescent
probes and labels with potential applications in pharmaceutical
and medicinal chemistry, as well as in material science.8
Figure 2. ORTEP drawing of the asymmetric unit for the compound 14;
ellipsoids are displayed at the 50% probability level.
3. In vitro anticancer activity
The two-stage screening process started with the selection of
nine of the obtained compounds (i.e. 5b-f, 5h-j and 5m) by the
Drug Evaluation Branch of National Cancer Institute (NCI-
USA). The selected compounds were subjected to a primary in
vitro evaluation against 57 cell lines at a single dose of 10 µM (it
is referred to as one-dose assay). The 57 cell panel is derived
from nine different cancer strains: Leukemia, Lung, Melanoma,
Colon, CNS, Ovary, Renal, Breast and Prostate cancers. The
output from the single dose screening was reported as a mean
graph available for analysis by the COMPARE program,[9] see
Supporting Information for details. Due to tetrahydroquinoline 5f
displayed the highest activity of the series at one-dose assays (see
Supporting Information), this compound was subjected to a
second screening in order to determine its cytostatic activity
against the full 57 cell panel at five concentrations at 10-fold
dilution (i.e. 100, 10, 1.0, 0.1 and 0.01 µM) (it is referred to as
five-dose assay). The test consisted of a 48 h continuous drug
exposure protocol by using sulforhodamine B (SRB) protein
assay to estimate cell growth. More details of this evaluation
method and the complementary information which is encoded by
the activity pattern over all cell lines have been published
elsewhere.[10] Table 2 shows that compound 5f displayed good
activity against 57 human tumor cell lines, and RPMI-8226
(Leukemia, GI50= 1.46 µM), SF-295 (CNS, GI50 = 1.67 µM) and
UO-31 (Renal, GI50 = 1.79 µM) are the most sensitive strains.
The cytotoxic effect associated with compound 5f was measured
as LC50 displaying values greater than 100 µM for most of the
cell lines evaluated, indicating a low toxicity of this compound
for normal human cell lines. These findings make compound 5f a
promising target to perform structural modifications in order to
improve its anticancer activity.
Scheme 4. Tricomponent synthesis of tetracyclic derivative 14 and julolidines
15 and 17.
In order to unequivocally confirm the formation of the 1,2,3,4-
tetrahydroquinoline core, single-crystals suitable for X-ray
diffraction analysis were grown for compound 14 in methanol at
room temperature. As shown in Figure 2, the proposed structure
14 was confirmed without ambiguity.
Table 2. In vitro testing results expressed as growth inhibition of cancer cell lines for compound 5f.a
Most sensitive cancer cell lines (GI50< 20 µM)
Panel
Cell line
GI50 (µM)[b]
LC50 (µM)[c]
Panel
Cell line
GI50 (µM)[b]
LC50 (µM)[c]
Leukemia
CCRF-CEM
HL-60(TB)
K-562
MOLT-4
RPMI-8226
SR
4.74
5.34
4.60
4.90
1.46
6.37
>100
>100
>100
>100
64.6
Ovarian
IGROV1
3.51
4.42
6.60
4.60
3.72
>100
>100
>100
>100
>100
OVCAR-3
OVCAR-5
OVCAR-8
NCI/ADR-RES
>100
Renal
A498
ACHN
CAKI-1
3.22
4.85
3.49
>100
>100
>100
Non-small cell lung
A549/ATCC
EKVX
4.48
3.39
>100
>100

5
HOP-62
HOP-92
4.27
2.75
2.89
4.37
4.05
4.22
4.33
>100
RXF 393
2.77
4.68
6.98
1.79
>100
>100
>100
>100
ACCEPTED MANUSCRIPT
>100
>100
>100
>100
>100
>100
SN12C
TK-10
UO-31
NCI-H226
NCI-H23
NCI-H322M
NCI-H460
NCI-H522
Prostate
Breast
PC-3
DU-145
3.00
2.66
>100
>100
Colon
COLO 205
HCC-2998
HCT-116
HCT-15
HT29
5.33
5.32
5.24
4.00
7.20
4.84
6.48
57.8
MCF-7
MDA-MB-231/ATCC
HS 578T
BT-549
T-47D
3.50
4.83
2.76
3.23
2.85
3.41
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
KM12
SW-620
MDA-MB-468
CNS
SF-268
SF-295
SF-539
SNB-19
SNB-75
U251
5.41
1.67
4.94
4.30
3.22
3.28
>100
39.7
>100
>100
>100
>100
Melanoma
LOX IMVI
MALME-3M
M14
MDA-MB-435
SK-MEL-2
SK-MEL-28
SK-MEL-5
UACC-257
UACC-62
3.54
3.99
4.42
3.77
6.00
8.28
2.05
3.07
3.25
>100
>100
60.4
78.8
>100
>100
11.1
43.3
69.8
a Data obtained from NCI’s in vitro disease-oriented human tumor cell lines screen. b GI50 was the drug concentration resulting in a 50% reduction in the net
protein increase (as measured by SRB staining) in control cells during the drug incubation. Determined at five concentration levels (100, 10, 1.0, 0.1, and 0.01
ꢁM). c LC50 is a parameter of cytotoxicity and reflects the molar concentration needed to kill 50% of the cells.
3.1 Additional biological studies
Once the in vitro screening was performed by the NCI, further
studies were carried out in order to get a better understanding of
how the active compound 5f had its effect on the tested cancer
cell lines. HCT116, a colon cancer cell line, was chosen as a
model to perform all additional studies due to the high toxicity
effect of compound 5f to this cell line (Table 2). Most of the
synthetic compounds exert their effect on tumoral cell lines by
inducing apoptosis, affecting the cell proliferation, or cell cycle
arrest. Therefore, we performed these analyses in order to assess
the effect of tetrahydroquinoline 5f on this cell line.
3.2 Annexin V/ propidium iodide binding assay
Figure 3. Evaluation of cell death by apoptosis using annexin V-FITC/PI
To determine the percentage of apoptosis induced by
staining. Dot plot profile of HCT cells untreated and treated with compound
compound 5f, the annexin V-FITC/propidium iodide staining
5f at 6µM and 10µM. HCT116 cells were cultured in the presence and
assay was carried out using colon cancer HCT116 which were
absence of compound 5f and labeled with Annexin V-FITC and propidium
treated with compound 5f with ranging concentrations from 2µM
iodide to assess the percentage of apoptotic cells on days 2 and 3 of culture.
to 10µM for 3 days. At concentrations ranging from 2µM to 5µM
Untreated cells were used as control.
no differences were observed when compared to untreated cells
(data not shown). Cells were collected and analyzed by flow
cytometry as depicted in Figure 3. With 5f−6µM and 5f−10µM
3.3 Impact of compound 5f on cell cycle
HCT116 cells displayed an increase in the early apoptotic cells
on days 2 and 3 of culture cells in a dose dependent manner.
HCT116 cells displayed a small rise in the total percentage of
early apoptotic cells from 0.469% in control (untreated cells) to
1.66% in 5f−10µM treated cells on day 2 of culture. Likewise,
HCT116 cells displayed rise in the total percentage of early
apoptotic cells from 0.20% in control (untreated cells) to 7.10%
of cells in S phase from 26.34% untreated cells (control) to
in 5f−10µM treated cells on day 3 of culture. Based on this
Many of the cytotoxic compounds have their growth inhibitory
effect by arresting the cell cycle at a specific phase. In vitro assay
was performed on cell cycle of treated and untreated HCT116
cells with compound 5f using the propidium iodide staining
method for 24 h to 72 h. Cells were collected and analyzed using
flow cytometry and the data showed an increase in the percentage
31.79% in 5f−10µM treated cells during 48 h of incubation.
results, it appears that the percentage of late apoptotic cells was
Contrast results were observed in the G0/G1 population, where
not affected by treatment with compound 5f.
the percentage decreased from 48.46% in untreated cells (control)
to 29.73% in 5f−10µM treated cells. In the same manner, an
increment in the S phase population was observed. The
percentage of increasing from 26.75% in untreated cells (control)

6
Tetrahedron
ACCEPTED MANUSCRIPT
to 36.28% in 5f−10µM treated cells for 72 h. The percentage of
not shown). Compared to untreated cells, 5f−6µM and 5f−10µM
S phase population for cells treated with 5f−6µM in 48 and 72 h
increased from 23.96% to 30.30%, respectively. These results
show that compound 5f has its effect on the cell cycle of HCT116
in a dose dependent manner (Figure 4, panel A). The ratio of
each population is presented in a bar graph (panel B). It can be
observed from the figure 4 an increasing in the population of S
phase in 5f−treated cells both 6µM and 10µM (yellow bar) and a
decreasing in the ratio in G0/G1 population for 5f−treated cells
both 6µM and 10µM (green bar). These results indicate S phase
cell cycle arrest by the compound 5f in HCT116 colon cancer
cells.
colon cancer cells HCT116 exhibited a decrease in the
proliferation profile on day 3 of culture (Figure 5, panel A).
Proliferation index is perhaps one of the most useful values to
compare sample to sample, as it only takes into account cells that
have a response within the population. In other words, the
proliferative index only accounts for cells that have undergone at
least one division. Figure 5, panel B shows a decrease in the
proliferative index for both 5f−6µM and 5f−10µM on day 3 of
culture when compared to untreated cells.
Figure 5. Impact of compound 5f on cell proliferation. Cell proliferation
analysis was assessed by flow cytometry. HCT116 cells were labeled with
CFSE, and cultured in the absence and presence of compound 5f. Panel A
shows histograms of untreated and treated HCT116 cells with compound 5f at
6µM and 10µM on day 3. Panel B shows bar graph of proliferation index for
untreated and treated HCT116 cells with compound 5f-6µM and 10µM on
day 3. Untreated cells were used as control.
4. Conclusions
Figure 4. Impact of compound 5f on cell cycle. Cell-cycle analysis by flow
cytometry in HCT116 treated with compound 5f. Panel A shows histograms
of HCT116 cells untreated and treated with compound 5f at 6µM and 10µM
and the percentage of each phase in the cell cycle on days 2 and 3 of culture.
Analysis was performed using FlowJo software with built-in model for cell
cycle analysis. Panel B shows the bar graph representing the ratio of each
population in the cell cycle of treated and untreated cells on days 2 and 3 of
culture. Untreated cells were used as control.
In summary, we have successfully developed a useful and
efficient method under mild reaction conditions to construct
novel and diversely substituted 1,2,3,4-tetrahydroquinolines 5
and julolidines 15 and 17 in good to excellent yields through a
catalyst-free Domino Mannich/intramolecular Friedel-Crafts
alkylation reaction sequence. The fact that three new bonds (one
C-N and two C-C) are formed in only one step under eco-
compatible conditions and the releasing of water as the unique
byproduct, gives this approach an outstanding bond-forming
efficiency, atom economy and an environmentally friendly
quality. The antitumor in vitro assays showed that, among the
nine tetrahydroquinolines 5 evaluated by the NCI, derivative 5f
exhibited the highest activity against 57 cancer cell lines, with
the most important GI50 values ranging from 1.46 to 8.28 µM.
Further studies performed over the active compound 5f on
HCT116 colon cancer cells indicated that its effect on cell death
is exerted through a cell cycle arrest (S phase) in a dose
3.4 Compound 5f suppresses cell proliferation in HCT116
colon cancer cells
In vitro analysis on cell proliferation was performed on treated
and untreated cells with compound 5f. HCT116 cells were
harvested and labeled with CFSE kit in order to monitor the cell
proliferation on cells treated with concentrations of compound 5f
from 2.0 to 10.0 µM. At the concentrations ranging from 1.0 to
5.0 ꢁM, compound 5f was not toxic to colon cancer cells (data

7
dependent manner, as wells as suppression on the cell
proliferation process.
m/z 230/232 [M + H]+, 252/254 [M + Na]+. The NMR data
ACCEPTED MANUSCRIPT
matches with the previously reported data.11a
5.1.1.3. (E)-N-(2-Chlorobenzylidene)-4-methylaniline. Following
the general procedure, the reaction between p-methylaniline (222
µL, 2.02 mmol) and o-chlorobenzaldehyde (229 µL, 2.03 mmol)
afforded N-arylimine as a yellow solid. Yield: 93% (430 mg).
M.p. 52–53 °C (Lit. 52–54 °C).11b FTIR (KBr): ν = 2934, 2853,
1614 (C=N), 1589, 1564 cm-1. 1H NMR (300 MHz, acetone-d6): δ
= 2.35 (s, 3H), 7.21 (d, J = 8.7 Hz, 2H), 7.25 (d, J = 8.5 Hz, 2H),
7.43–7.53 (m, 3H), 8.24 (d, J = 6.2 Hz, 1H), 8.95 (s, 1H) ppm.
13C NMR (75 MHz, acetone-d6): δ = 21.0 (CH3), 121.9 (CH),
128.3 (CH), 129.3 (CH), 130.7 (CH), 130.9 (CH), 133.3 (CH),
134.3 (Cq), 136.4 (Cq), 137.2 (Cq), 150.2 (Cq), 156.0 (CH) ppm.
MS (ESI+): m/z 252/254 [M + Na]+. The NMR data matches with
the previously reported data.11b
5. Experimental section
5.1. Chemistry
5.1.1. General methods
Melting points were determined on a Büchi melting point B-
450 apparatus and are uncorrected. IR spectra were recorded on a
Shimadzu FTIR 8400 spectrophotometer in KBr disks and films.
1H and 13C NMR spectra were recorded on a Bruker Avance 400
spectrophotometer operating at 400 and 100 MHz, respectively,
and Bruker AC 300 operating at 300 and 75 MHz respectively,
and using CDCl3, (CD3)2SO or (CD3)2CO as solvents and TMS as
internal standard. NMR spectroscopic data are reported as
follows: chemical shift, multiplicity (s = singlet, d = doublet, t =
triplet, m = multiplet), coupling constants [Hz]. Mass spectra
were run on a SHIMADZU-GCMS 2010-DI-2010 spectrometer
(equipped with a direct inlet probe) operating at 70 eV. Low
resolution mass spectra (MS) were recorded on ion trap Bruker
Esquire 6000, equipped with an electrospray source (ESI) using
methanol as solvent (Sigma Aldrich, chromasolv for HPLC).
Microanalyses were performed on an Agilent elemental analyzer
and the values are within ±0.4% of the theoretical values.
Crystallographic data for compound 14 were collected at 123 K
on an Oxford Diffraction Gemini S diffractometer, using Mo-Kα
X-ray radiation (λ = 0.71073 Ǻ) and deposited at Cambridge
Crystallographic Data Center (CCDC reference: 1556488). All
reactions were monitored by TLC with silica gel aluminum plates
(Merck 60 F254). Column chromatography was performed with
Merck 230–400 mesh silica gel. The starting anilines, aldehydes
and the electron-rich alkenes 9a−c were purchased from Sigma-
Aldrich, Fluka, and Across (analytical reagent grades) and were
used without further purification. Secondary amines 8a−l and 8p
were synthetized through a reductive amination from their
corresponding N-arylimines by following a similar procedure like
described previously.6a
5.1.1.1. General procedure for the synthesis of the N-arylimines.
A mixture of the aniline derivative (200–300 mg each one, 1.0
equiv.) and the corresponding aldehyde (1.0 equiv.) was heated
in an oil bath at 120 °C for 30–60 min under solvent-free
conditions. After complete disappearance of the starting
materials, as monitored by TLC using neutral alumina, the
mixture was allowed to cool to ambient temperature and treated
with hexane (2.0 mL). The precipitate formed corresponding to
the N-arylimine was filtered and dried in air. Particularly,
quinoline-2-one based N-arylimines were obtained by refluxing
for 3 h in methanol (5.0 mL). Then, the solid formed was filtered,
washed with cold methanol (1.0 mL) and dried to give the
desired products.
5.1.1.4.
Following the general procedure, the reaction between p-
methylaniline (208 µL, 1.89 mmol) and 3,4,5-
(E)-N-(3,4,5-Trimethoxybenzylidene)-4-methylaniline.
trimethoxybenzaldehyde (373 mg, 1.90 mmol) afforded N-
arylimine as a brown solid. Yield: 91% (490 mg). M.p. 99–100
°C (Lit. 74–76 °C).11c FTIR (KBr): ν =2931, 2834, 1622 (C=N),
1578, 1234, 1135, 1002 (C–O) cm-1. 1H NMR (300 MHz,
CDCl3): δ = 2.36 (s, 3H), 3.91 (s, 3H), 3.94 (s, 6H), 7.11–7.21
(m, 6H), 8.35 (s, 1H) ppm. 13C NMR (75 MHz, CDCl3): δ = 21.0
(CH3), 56.3 (CH3), 61.0 (CH3), 105.8 (CH), 120.8 (CH), 129.8
(CH), 131.8 (Cq), 135.8 (Cq), 141.0 (Cq), 149.3 (Cq), 153.6
(Cq), 159.1 (CH) ppm. MS (ESI+): m/z 286 [M + H]+, 308 [M +
Na]+. The NMR data matches with the previously reported
data.11c
5.1.1.5.
(E)-N-(Benzo[d][1,3]dioxol-5-ylmethylene)-4-
methoxyaniline. Following the general procedure, the reaction
between p-methoxyaniline (215 mg, 1.74 mmol) and 3,4-
methylenedioxybenzaldehyde (264 mg, 1.76 mmol) afforded N-
arylimine as a brown solid. Yield: 96% (426 mg). M.p. 113 °C
(Lit. 115–120 °C).11d FTIR (KBr): ν =2954, 2837, 1624 (C=N),
1602, 1208, 1104, 1034 (C–O) cm-1. 1H NMR (300 MHz,
CDCl3): δ = 3.82 (s, 3H), 6.02 (s, 2H), 6.87 (d, J = 7.9 Hz, 1H),
6.92 (d, J = 9.0 Hz, 2H), 7.21 (d, J = 9.0 Hz, 2H), 7.26 (dd, J =
1.4, 7.8 Hz, 1H), 7.53 (d, J = 1.3 Hz, 1H), 8.35 (s, 1H) ppm. 13C
NMR (75 MHz, CDCl3): δ = 55.6 (OCH3), 101.6 (OCH2O),
106.9 (CH), 108.3 (CH), 114.5 (CH), 122.2 (CH), 125.5 (CH),
131.4 (Cq), 144.9 (Cq), 148.5 (Cq), 150.4 (Cq), 157.6 (CH),
158.2 (Cq) ppm. MS (ESI+): m/z 256 [M + H]+, 278 [M + Na]+.
The NMR data matches with the previously reported data.11d
5.1.1.6.
(E)-N-(4-Chlorobenzylidene)-4-methoxyaniline.
Following the general procedure, the reaction between p-
methoxyaniline (254 mg, 2.06 mmol) and p-chlorobenzaldehyde
(292 mg, 2.08 mmol) afforded N-arylimine as a brown solid.
Yield: 92% (464 mg). M.p. 125 °C (Lit. 122–124 °C).11e FTIR
(KBr): ν = 2962, 2845, 1620 (C=N), 1597, 1572, 1254, 1093,
1030 (C–O) cm-1. 1H NMR (300 MHz, acetone-d6): δ = 3.82 (s,
3H), 6.97 (d, J = 8.9 Hz, 2H), 7.30 (d, J = 9.0 Hz, 2H), 7.52 (d, J
= 8.4 Hz, 2H), 7.95 (d, J = 8.4 Hz, 2H), 8.61 (s, 1H) ppm. 13C
NMR (75 MHz, acetone-d6): δ = 55.7 (OCH3), 115.2 (CH), 123.2
(CH), 129.7 (CH), 130.7 (CH), 136.6 (Cq), 137.0 (Cq), 145.2
(Cq), 157.2 (CH), 159.6 (Cq) ppm. MS (ESI+): m/z 246/248 [M
5.1.1.2 (E)-N-(4-Chlorobenzylidene)-4-methylaniline. Following
the general procedure, the reaction between p-methylaniline (222
µL, 2.02 mmol) and p-chlorobenzaldehyde (289 mg, 2.06 mmol)
afforded N-arylimine as a brown solid. Yield: 97% (449 mg).
M.p. 124–125 °C (Lit. 124–126 °C).11a FTIR (KBr): ν = 2939,
2876, 1622 (C=N), 1589, 1566 cm-1. 1H NMR (300 MHz,
CDCl3): δ = 2.38 (s, 3H), 7.15 (d, J = 8.3 Hz, 2H), 7.21 (d, J =
8.1 Hz, 2H), 7.44 (d, J = 8.4 Hz, 2H), 7.84 (d, J = 8.5 Hz, 2H),
8.43 (s, 1 H) ppm. 13C NMR (75 MHz, CDCl3): δ = 21.1 (CH3),
120.9 (CH), 129.1 (CH), 129.9 (CH), 130.0 (CH), 135.0 (Cq),
136.2 (Cq), 137.2 (Cq), 149.1 (Cq), 158.0 (CH) ppm. MS (ESI+):

8
Tetrahedron
+ H]+. The NMR data matches with the previously reported
5.1.1.11 (E)-3-(4-Methoxyphenylimino)methyl)quinolin-2(1H)-
ACCEPTED MANUSCRIPT
data.11e
one. Following the general procedure, the reaction between p-
methoxyaniline (191 mg, 1.55 mmol) and 1,2-dihydro-2-
oxoquinoline-3-carbaldehyde (270 mg, 1.56 mmol) afforded N-
arylimine as a yellow solid. Yield: 89% (384 mg). M.p. 245–246
°C. FTIR (KBr): ν = 3348 (NHCO), 2950, 2853, 1652 (C=O),
1606 (C=N), 1578, 1163, 1026 (C–O) cm-1. 1H NMR (300 MHz,
DMSO-d6): δ = 3.78 (s, 3H), 6.99 (d, J = 9.0 Hz, 2H), 7.22 (td,J
= 1.2, 7.5 Hz, 1H), 7.30 (d, J = 8.9 Hz, 2H), 7.35 (d, J = 8.2 Hz,
1H), 7.56 (td, J = 1.4, 7.6 Hz, 1H), 7.87 (d, J = 7.6 Hz, 1H), 8.63
(s, 1H), 8.80 (s, 1H), 12.11 (br s, 1H, NH) ppm. 13C NMR (75
MHz, DMSO-d6): δ = 55.2 (OCH3), 114.5 (CH), 115.1 (CH),
118.9 (Cq), 122.3 (CH), 122.4 (CH), 126.6 (Cq), 129.5 (CH),
131.6 (CH), 136.8 (CH), 139.6 (Cq), 144.1 (Cq), 152.6 (CH),
158.2 (Cq), 161.5 (Cq, C=O) ppm. MS (ESI+): m/z 279 [M +
H]+, 301 [M + Na]+.
5.1.1.7.
(E)-N-(4-Methoxybenzylidene)-4-chloroaniline.
Following the general procedure, the reaction between p-
chloroaniline (164 µL, 1.84 mmol) and p-methoxybenzaldehyde
(225 µL, 1.85 mmol) afforded N-arylimine as a brown solid (406
mg, 90%). M.p. 94 °C (Lit. 125–127 °C).11f FTIR (KBr): ν =
2962, 2840, 1620 (C=N), 1603, 1569, 1166, 1102, 1027 (C–O)
cm-1. 1H NMR (300 MHz, acetone-d6): δ = 3.88 (s, 3H), 7.06 (d, J
= 8.8 Hz, 2H), 7.23 (d, J = 8.7 Hz, 2H), 7.40 (d, J = 8.8 Hz, 2H),
7.91 (d, J = 8.8 Hz, 2H), 8.50 (s, 1H) ppm. 13C NMR (75 MHz,
acetone-d6): δ = 55.8 (OCH3), 115.0 (CH), 123.3 (CH), 129.9
(CH), 130.1 (Cq), 131.2 (Cq), 131.4 (CH), 152.1 (Cq), 161.0
(CH), 163.5 (Cq) ppm. MS (ESI+): m/z246/248 [M + H]+. The
NMR data matches with the previously reported data.11f
5.1.1.8. (E)-N-(4-Chlorobenzylidene)-4-chloroaniline. Following
the general procedure, the reaction between p-chloroaniline (192
µL, 2.15 mmol) and p-chlorobenzaldehyde (305 mg, 2.17 mmol)
afforded N-arylimine as a yellow solid. Yield: 95% (508 mg).
M.p. 112–113 °C (Lit. 112–114 °C).11f FTIR (KBr): ν = 2942,
2881, 1624 (C=N), 1587, 1563 cm-1. 1H NMR (300 MHz,
acetone-d6): δ = 7.28 (d, J = 8.8 Hz, 2H), 7.42 (d, J = 8.8 Hz,
2H), 7.54 (d, J = 8.5 Hz, 2H), 7.97 (d, J = 8.5 Hz, 2H), 8.60 (s,
1H) ppm. 13C NMR (75 MHz, acetone-d6): δ = 123.5 (CH), 129.8
(CH), 130.0 (CH), 131.1 (CH), 131.9 (Cq), 136.0 (Cq), 137.7
(Cq), 151.3 (Cq), 160.4 (CH) ppm. The NMR data matches with
the previously reported data.11f
5.1.1.12 (E)-3-(4-Chlorophenylimino)methyl)quinolin-2(1H)-one.
Following the general procedure, the reaction between p-
chloroaniline (153 µL, 1.72 mmol) and 1,2-dihydro-2-
oxoquinoline-3-carbaldehyde (301 mg, 1.74 mmol) afforded N-
arylimine as a yellow solid. Yield: 91% (441 mg). M.p. 262–263
°C. FTIR (KBr): ν = 3339 (NHCO), 2945, 2850, 1650 (C=O,
C=N), 1556 cm-1. 1H NMR (300 MHz, DMSO-d6): δ = 7.23 (td, J
= 1.0, 7.5 Hz, 1H), 7.30 (d, J = 8.8 Hz, 2H), 7.35 (d, J = 8.2 Hz,
1H), 7.47 (d, J = 8.8 Hz, 2H), 7.58 (td, J = 1.3, 7.7 Hz, 1H), 7.89
(d, J = 7.7 Hz, 1H), 8.67 (s, 1H), 8.77 (s, 1H), 12.15 (br s, 1H,
NH) ppm. 13C NMR (75 MHz, DMSO-d6): δ = 115.2 (CH), 118.8
(Cq), 122.4 (CH), 122.8 (CH), 126.2 (Cq), 129.2 (CH), 129.7
(CH), 130.5 (Cq), 132.0 (CH), 138.0 (CH), 139.8 (Cq), 150.2
(Cq), 156.0 (CH), 161.4 (Cq, C=O) ppm. MS (ESI+): m/z
305/307 [M + Na]+.
5.1.1.9. (E)-N-(4-Chlorobenzylidene)aniline. Following the
general procedure, the reaction between aniline (245 µL, 2.68
mmol) and p-chlorobenzaldehyde (376 mg, 2.67 mmol) afforded
N-arylimine as a yellow solid. Yield: 88% (505 mg). M.p. 62–64
°C (Lit. 75–77 °C).11f FTIR (KBr): ν =2954, 2873, 1622 (C=N),
1584, 1563 cm-1. 1H NMR (300 MHz, acetone-d6): δ = 7.22–7.28
(m, 3H), 7.41 (t, J = 7.6 Hz, 2H), 7.55 (d, J = 8.4 Hz, 2H), 7.98
(d, J = 8.5 Hz, 2H), 8.59 (s, 1H) ppm. 13C NMR (75 MHz,
acetone-d6): δ = 121.8 (CH), 127.0 (CH), 129.8 (CH), 130.0
(CH), 131.0 (CH), 136.3 (Cq), 137.5 (Cq), 152.6 (Cq), 159.7
(CH) ppm. MS (ESI+): m/z 216/218 [M + H]+. The NMR data
matches with the previously reported data.11f
5.1.1.13
Following the general procedure, the reaction between p-
methoxyaniline (241 mg, 1.96 mmol) and 3,4-
(E)-N-(3,4-Dimethoxybenzylidene)-4-methoxyaniline.
dimethoxybenzaldehyde (326 mg, 1.96 mmol) afforded N-
arylimine as a brown solid. Yield: 97% (517 mg). M.p. 129–130
°C (Lit. 128–130 °C).12g FTIR (KBr): ν =2932, 2838, 1622
(C=N), 1600, 1577, 1138, 1023 (C–O) cm-1. 1H NMR (300 MHz,
CDCl3): δ = 3.83 (s, 3H), 3.95 (s, 3H), 3.99 (s, 3H), 6.91–6.94
(m, 3H), 7.23 (d, J = 8.6 Hz, 2H), 7.30 (dd, J = 1.2, 8.1 Hz, 1H),
7.64 (s, 1H), 8.39 (s, 1H) ppm. 13C NMR (75 MHz, CDCl3): δ =
55.6 (OCH3), 56.1 (OCH3), 56.1 (OCH3), 109.0 (CH), 110.6
(CH), 114.5 (CH), 114.9 (Cq), 116.5 (Cq), 122.2 (CH), 124.3
(CH), 129.8 (Cq), 149.6 (Cq), 152.0 (Cq), 158.1 (CH) ppm. MS
(ESI+): m/z 272 [M + H]+, 294 [M + Na]+. The NMR data
matches with the previously reported data.12g
5.1.1.10 (E)-3-(4-Methylphenylimino)methyl)quinolin-2(1H)-one.
Following the general procedure, the reaction between p-
methylaniline (195 µL, 1.77 mmol) and 1,2-dihydro-2-
oxoquinoline-3-carbaldehyde (303 mg, 1.75 mmol) afforded N-
arylimine as a yellow solid. Yield: 71% (326 mg). M.p. 246–247
°C. FTIR (KBr): ν = 3282 (NHCO), 2943, 2857, 1659 (C=O),
1
1610 (C=N), 1597 cm-1. H NMR (400 MHz, DMSO-d6): δ =
5.1.2. General procedure for the synthesis of the N-arylamines 8.
Solid NaBH4 (2.0 equiv.) was added portion-wise with stirring,
over a period of 10 min, to a solution of N-arylimine (1.0 equiv.)
in methanol (5.0 mL). The stirring was continued at ambient
temperature for 1 h. After the reaction was complete (monitored
by TLC), the volume of the reaction mixture was reduced to 1.0
mL under reduced pressure, and water (5.0 mL) was added. The
aqueous solution was extracted with ethyl acetate (2 x 5.0 mL),
the combined organic extracts were dried with anhydrous Na2SO4
and after removal of the solvent the resulting products 8 were
used without further purification. All reactions started with 300
2.31 (s, 3H), 7.17–7.24 (m, 5H), 7.35 (d, J = 8.0 Hz, 1H), 7.56
(t,J = 7.5 Hz, 1H), 7.87 (d, J = 7.7 Hz, 1H), 8.65 (s, 1H), 8.78 (s,
1H), 12.13 (br s, 1H, NH) ppm.13C NMR (100 MHz, DMSO-d6):
δ = 20.5 (CH3), 115.1 (CH), 118.8 (Cq), 120.9 (CH), 122.3 (CH),
126.4 (Cq), 129.5 (CH), 129.7 (CH), 131.7 (CH), 135.7 (Cq),
137.2 (CH), 139.6 (Cq), 148.8 (Cq), 154.1 (CH), 161.4 (Cq,
C=O) ppm. MS (EI) m/z (%) = 262 (20) [M]+, 236 (6), 171 (13),
145 (46), 117 (26), 81 (59), 69 (100).

9
mg of the corresponding N-arylimine. The quinolin-2-one based
N-arylamines 8j–l were prepared following the same procedure
except that the solid formed was filtered, washed with water (3 x
5.0 mL), and dried to give the desired products.
143.8 (Cq), 147.3 (Cq), 148.7 (Cq), 152.6 (Cq) ppm. MS
ACCEPTED MANUSCRIPT
(ESI+): m/z 258 [M + H]+, 280 [M + Na]+. The NMR data
matches with the previously reported data.12c
5.1.2.5. N-(4-Chlorobenzyl)-4-methoxyaniline 8f. Following the
general procedure, the reaction between the corresponding N-
arylimine (289 mg, 1.18 mmol) and NaBH4 (89 mg, 2.35 mmol)
afforded compound 8f as a light brown solid. Yield: 88% (256
mg). M.p. 75–76 °C (Lit. 75–76 °C).12d FTIR (KBr): ν = 3442
(N–H), 2962, 2839, 1597, 1094, 1030 (C–O) cm-1. 1H NMR (300
MHz, acetone-d6): δ = 3.66 (s, 3H), 4.29 (s, 2H), 5.12 (br s, 1H,
NH), 6.58 (d, J = 9.0 Hz, 2H), 6.71 (d, J = 9.0 Hz, 2H), 7.32 (d, J
= 8.6 Hz, 2H), 7.40 (d, J = 8.6 Hz, 2H) ppm. 13C NMR (75 MHz,
acetone-d6): δ = 48.2 (CH2), 55.7 (OCH3), 114.6 (CH), 115.4
(CH), 129.1 (CH), 129.8 (CH), 130.7 (Cq), 140.6 (Cq), 143.6
(Cq), 152.7 (Cq) ppm. MS (ESI+): m/z 248/250 [M + H]+. The
NMR data matches with the previously reported data.12d
5.1.2.1. N-(4-Chlorobenzyl)-4-methylaniline 8b. Following the
general procedure, the reaction between the corresponding N-
arylimine (311 mg, 1.36 mmol) and NaBH4 (103 mg, 2.72 mmol)
afforded compound 8b as a brown solid. Yield: 68% (214 mg).
M.p. 47 °C (Lit. 47–48 °C).12a FTIR (KBr): ν = 3379 (N–H),
2945, 2851, 1615 cm-1. 1H NMR (300 MHz, acetone-d6): δ = 2.15
(s, 3H), 4.32 (d, J = 6.0 Hz, 2H), 5.30 (br s, 1H, NH), 6.54 (d, J =
8.3 Hz, 2H), 6.89 (d, J = 8.2 Hz, 2H), 7.32 (d, J = 8.6 Hz, 2H),
7.40 (d, J = 8.6 Hz, 2H) ppm. 13C NMR (75 MHz, acetone-d6): δ
= 20.4 (CH3), 47.7 (CH2), 113.6 (CH), 126.2 (Cq), 129.1 (CH),
129.7 (CH), 130.2 (CH), 132.6 (Cq), 140.6 (Cq), 147.2 (Cq)
ppm. MS (ESI+): m/z 232/234 [M + H]+. The NMR data matches
with the previously reported data.12a
5.1.2.6. N-(4-Methoxybenzyl)-4-chloroaniline 8g. Following the
general procedure, the reaction between the corresponding N-
arylimine (313 mg, 1.28 mmol) and NaBH4 (97 mg, 2.56 mmol)
afforded compound 8g as a yellow solid. Yield: 89% (281 mg).
M.p. 85–86 °C (Lit. 84–85 °C).12e FTIR (KBr): ν = 3409 (N–H),
2932, 2835, 1594, 1173, 1110, 1028 (C–O) cm-1. 1H NMR (300
MHz, CDCl3): δ = 3.82 (s, 3H), 4.06 (br s, 1H, NH), 4.23 (s, 2H),
6.56 (d, J = 8.8 Hz, 2H), 6.90 (d, J = 8.6 Hz, 2H), 7.13 (d, J = 8.9
Hz, 2H), 7.28 (d, J = 8.8 Hz, 2H) ppm. 13C NMR (75 MHz,
CDCl3): δ = 47.9 (CH2), 55.4 (OCH3), 114.0 (CH), 114.2 (CH),
122.2 (Cq), 128.8 (CH), 129.1 (CH), 130.9 (Cq), 146.7 (Cq),
159.0 (Cq) ppm. MS (ESI+): m/z 248/250 [M + H]+, 270/272 [M
+ Na]+. The NMR data matches with the previously reported
data.12e
5.1.2.2. N-(2-Chlorobenzyl)-4-methylaniline 8c. Following the
general procedure, the reaction between the corresponding N-
arylimine (297 mg, 1.30 mmol) and NaBH4 (98 mg, 2.59 mmol)
afforded compound 8c as a yellow solid. Yield: 93% (279 mg).
M.p. 44–45 °C (Lit. 45 °C).12b FTIR (KBr): ν = 3412 (N–H),
2936, 2865, 1601 cm-1. 1H NMR (300 MHz, CDCl3): δ = 2.28 (s,
3H), 4.05 (br s, 1H, NH), 4.45 (s, 2H), 6.59 (d, J = 8.5 Hz, 2H),
7.02 (d, J = 8.1 Hz, 2H), 7.22–7.28 (m, 2H), 7.40–7.47 (m, 2H)
ppm. 13C NMR (75 MHz, CDCl3): δ = 20.5 (CH3), 46.3 (CH2),
113.2 (CH), 127.0 (CH), 127.1 (Cq), 128.4 (CH), 129.1 (CH),
129.6 (CH), 129.8 (CH), 133.3 (Cq), 136.9 (Cq), 145.5 (Cq)
ppm. MS (ESI+): m/z 232/234 [M + H]+, 254/256 [M + Na]+. The
NMR data matches with the previously reported data.12b
5.1.2.3.
N-(3,4,5-Trimethoxybenzyl)-4-methylaniline
8d.
5.1.2.7. N-(4-Chlorobenzyl)-4-chloroaniline 8h. Following the
general procedure, the reaction between the corresponding N-
arylimine (334 mg, 1.34 mmol) and NaBH4 (102 mg, 2.70 mmol)
afforded compound 8h as a brown solid. Yield: 79% (267 mg).
M.p. 70 °C (Lit. 70–71 °C).12a FTIR (KBr): ν = 3402 (N–H),
2945, 2844, 1597 cm-1. 1H NMR (300 MHz, acetone-d6): δ = 4.36
(d, J = 5.2 Hz, 2H), 5.73 (br s, 1H, NH), 6.63 (d, J = 8.9 Hz, 2H),
7.06 (d, J = 8.9 Hz, 2H), 7.34 (d, J = 8.7 Hz, 2H), 7.40 (d, J = 8.7
Hz, 2H) ppm. 13C NMR (75 MHz, acetone-d6): δ = 47.3 (CH2),
114.7 (CH), 121.4 (Cq), 129.2 (CH), 129.5 (CH), 129.7 (CH),
132.8 (Cq), 139.8 (Cq), 148.3 (Cq) ppm. The NMR data matches
with the previously reported data.12a
Following the general procedure, the reaction between the
corresponding N-arylimine (303 mg, 1.06 mmol) and NaBH4 (81
mg, 2.14 mmol) afforded compound 8d as a brown solid. Yield:
87% (265 mg). M.p. 94–95 °C (Lit. 94–96 °C).12c FTIR (KBr): ν
= 3365 (N–H), 2946, 2845, 1603, 1124, 1006 (C–O) cm-1.1H
NMR (300 MHz, acetone-d6): δ = 2.16 (s, 3H), 3.69 (s, 3H), 3.77
(s, 6H), 4.23 (s, 2H), 5.15 (br s, 1H, NH), 6.58 (d, J = 8.1 Hz,
2H), 6.71 (s, 2H), 6.90 (d, J = 8.0 Hz, 2H) ppm. 13C NMR (75
MHz, acetone-d6): δ = 20.4 (CH3), 48.8 (CH2), 56.3 (OCH3), 60.4
(OCH3), 105.5 (CH), 113.7 (CH), 126.0 (Cq), 130.2 (CH), 137.0
(Cq), 138.0 (Cq), 147.6 (Cq), 154.4 (Cq) ppm. MS (ESI+): m/z
288 [M + H]+, 310 [M + Na]+, 326 [M + K]+. The NMR data
matches with the previously reported data.12c
5.1.2.8. N-(4-Chlorobenzyl)aniline 8i. Following the general
procedure,the reaction between the corresponding N-arylimine
(301 mg, 1.40 mmol) and NaBH4 (109 mg, 2.88 mmol) afforded
compound 8i as a yellow solid. Yield: 86% (269 mg). M.p. 46–
47 °C (Lit. 50–51 °C).12f FTIR (KBr): ν = 3418 (N–H), 2920,
2848, 1603 cm-1. 1H NMR (400 MHz, CDCl3): δ =3.97 (br s, 1H,
NH), 4.25 (s, 2H), 6.56 (d, J = 7.6 Hz, 2H), 6.68 (t, J = 7.3 Hz,
1H), 7.13 (t, J = 7.7 Hz, 2H), 7.24 (d, J = 8.2 Hz, 2H), 7.26 (d, J
= 8.3 Hz, 2H) ppm. 13C NMR (100 MHz, CDCl3): δ = 47.7
(CH2), 113.0 (CH), 117.9 (CH), 128.7 (CH), 128.8 (CH), 129.4
(CH), 132.9 (Cq), 138.1 (Cq), 147.9 (Cq) ppm. MS (ESI+):
m/z218/220 [M + H]+. The NMR data matches with the
previously reported data.12f
5.1.2.4. N-((Benzo[d][1,3]dioxol-5-yl)methyl)-4-methoxyaniline
8e. Following the general procedure, the reaction between the
corresponding N-arylimine (319 mg, 1.25 mmol) and NaBH4 (94
mg, 2.48 mmol) afforded compound 8e as a brown solid. Yield:
81% (322 mg). M.p. 82–83 °C (Lit. 76–78 °C).12c FTIR (KBr): ν
= 3389 (N–H), 2948, 2834, 1606, 1124, 1038 (C–O) cm-1. H
1
NMR (300 MHz, acetone-d6): δ = 3.66 (s, 3H), 4.20 (d, J = 5.0
Hz, 2H), 4.98 (br s, 1H, NH), 5.94 (s, 2H), 6.60 (d, J = 9.0 Hz,
2H), 6.71 (d, J = 9.0 Hz, 2H), 6.77 (d, J = 7.9 Hz, 1H), 6.86 (d, J
= 8.0 Hz, 1H), 6.90 (s, 1H) ppm. 13C NMR (75 MHz, acetone-
d6): δ = 48.7 (CH2), 55.7 (OCH3), 101.8 (OCH2O), 108.5 (CH),
108.7 (CH), 114.6 (CH), 115.4 (CH), 121.2 (CH), 135.5 (Cq),

10
Tetrahedron
ACCEPTED MANUSCRIPT
5.1.2.9. 3-((4-Methylphenylamino)methyl)quinolin-2(1H)-one
6.84 (t, J = 7.4 Hz, 1H), 7.27 (t, J = 7.9 Hz, 2H) ppm. 13C NMR
8j. Following the general procedure, the reaction between the
corresponding N-arylimine (296 mg, 1.13 mmol) and NaBH4 (85
mg, 2.25 mmol) afforded compound 8j as a yellow solid. Yield:
88% (263 mg). M.p. 250–251 °C. FTIR (KBr): ν = 3373 (N–H,
(100 MHz, CDCl3): δ = 33.7 (CH2), 71.3 (Cq), 81.1 (CH), 113.6
(CH), 118.7 (CH), 129.3 (CH), 146.9 (Cq) ppm. MS (ESI+): m/z
132 [M + H]+. The NMR data matches with the previously
reported data.12g
NHCO), 2966, 2854, 1652 (C=O), 1572 cm-1. H NMR (300
1
MHz, DMSO-d6): δ = 2.11 (s, 3H), 4.12 (d, J = 5.8 Hz, 2H), 5.87
(t, J = 6.0 Hz, 1H, NH), 6.49 (d, J = 8.3 Hz, 2H), 6.87 (d, J = 8.2
Hz, 2H), 7.12 (t, J = 7.5 Hz, 1H), 7.31 (d, J = 8.1 Hz, 1H), 7.43
(t, J = 7.6 Hz, 1H), 7.56 (d, J = 7.5 Hz, 1H), 7.71 (s, 1H), 11.85
(br s, 1H, NHCO) ppm. 13C NMR (75 MHz, DMSO-d6): δ = 20.0
(CH3), 42.2 (CH2), 112.3 (CH), 114.8 (CH), 119.1 (Cq), 121.7
(CH), 124.3 (Cq), 127.4 (CH), 129.3 (CH), 129.4 (CH), 131.1
(Cq), 134.7 (CH), 137.7 (Cq), 146.1 (Cq), 161.7 (Cq, C=O) ppm.
MS (ESI+): m/z 265 [M + H]+, 287 [M + Na]+.
5.1.2.13.
N-(3,4-Dimethoxybenzyl)-4-methoxyaniline
8o.
Following the general procedure, the reaction between the
corresponding N-arylimine (287 mg, 1.06 mmol) and NaBH4 (83
mg, 2.19 mmol) afforded compound 8o as a yellow solid. Yield:
86% (249 mg). M.p. 114 °C (Lit. 114–116 °C).12h FTIR (KBr): ν
= 3376 (N–H), 2953, 2839, 1601, 1119, 1042 (C–O) cm-1. H
1
NMR (400 MHz, CDCl3): δ = 3.36 (br s, 1H, NH), 3.74 (s, 3H),
3.86 (s, 3H), 3.87 (s, 3H), 4.21 (s, 2H), 6.61 (d, J = 8.8 Hz, 2H),
6.78 (d, J = 8.8 Hz, 2H), 6.83 (d, J = 8.5 Hz, 1H), 6.90 (d, J = 2.3
Hz, 1H), 6.92 (s, 1H) ppm. 13C NMR (100 MHz, CDCl3): δ =
49.3 (CH2), 55.9 (OCH3), 55.9 (OCH3), 56.0 (OCH3), 110.9
(CH), 111.3 (CH), 114.2 (CH), 115.0 (CH), 119.7 (CH), 132.3
(Cq), 142.6 (Cq), 148.2 (Cq), 149.2 (Cq), 152.3 (Cq) ppm. MS
(ESI+): m/z 274 [M + H]+, 296 [M + Na]+. The NMR data
matches with the previously reported data.12h
5.1.2.10. 3-((4-Methoxyphenylamino)methyl)quinolin-2(1H)-one
8k. Following the general procedure, the reaction between the
corresponding N-arylimine (280 mg, 1.00 mmol) and NaBH4 (76
mg, 2.01 mmol) afforded compound 8k as a yellow solid. Yield:
92% (255 mg). M.p. 246–248 °C. FTIR (KBr): ν = 3391 (N–H,
NHCO), 2937, 2846, 1653 (C=O), 1572, 1122, 1038 (C–O) cm-1.
1H NMR (300 MHz, DMSO-d6): δ = 3.61 (s, 3H), 4.09 (d, J = 5.8
Hz, 2H), 5.69 (t, J = 6.1 Hz, 1H, NH), 6.54 (d, J = 8.9 Hz, 2H),
6.70 (d, J = 8.9 Hz, 2H), 7.12 (td, J = 1.0, 7.4 Hz, 1H), 7.31 (d, J
= 8.1 Hz, 1H), 7.43 (td, J = 1.1, 7.7 Hz, 1H), 7.56 (d, J = 7.7 Hz,
1H), 7.73 (s, 1H), 11.85 (br s, 1H, NHCO) ppm. 13C NMR (75
MHz, DMSO-d6): δ = 42.7 (CH2), 55.2 (OCH3), 113.3 (CH),
114.6 (CH), 114.8 (CH), 119.1 (Cq), 121.7 (CH), 127.4 (CH),
129.4 (CH), 131.2 (Cq), 134.7 (CH), 137.7 (Cq), 142.6 (Cq),
150.8 (Cq), 161.7 (Cq, C=O) ppm. MS (ESI+): m/z 281 [M +
H]+, 303 [M + Na]+.
5.1.3. General procedure for the synthesis of 1,2,3,4-
tetrahydroquinolines 5a–p, tetracyclic product 14 and julolidines
15 and 17. A mixture of secondary amine 8 (100–200 mg each
one, 1.0 equiv.), polyformaldehyde (1.5 equiv.) and the electron-
rich alkene 9 (1.1 equiv.) in acetonitrile (2.0 mL) was stirred at
room temperature for 24 h until the starting secondary amine 8
was not detected by TLC (revealed with an ethanolic solution of
vanillin-sulfuric acid or iodine). After the solvent was removed
under reduced pressure, the oily material obtained was purified
by column chromatography on silica gel using a mixture of
CHCl3:MeOH (30:1, 20:1 or 15:1) as eluent. The
tetrahydroquinolines 5j–l were obtained using a mixture of
CH3CN/DMSO (3:1, 2.0 mL) at 60 °C for 24 h. Then, the solid
formed was filtered, washed with cold CH3CN (1.0 mL) and
dried to give the desired product. The julolidine 17 was obtained
by using 0.2 mL of acetic acid and 2.2 equiv of alkene 9a.
5.1.2.11. 3-(((4-Chlorophenylamino)methyl)quinolin-2(1H)-one
8l. Following the general procedure, the reaction between the
corresponding N-arylimine (315 mg, 1.11 mmol) and NaBH4 (85
mg, 2.25 mmol) afforded compound 8l as a yellow solid. Yield:
72% (227 mg). M.p. 240–241 °C. FTIR (KBr): ν = 3395 (N–H,
1
NHCO), 2937, 2851, 1652 (C=O), 1600, 1572 cm-1. H NMR
(400 MHz, DMSO-d6): δ = 4.13 (d, J = 6.0 Hz, 2H), 6.29 (t, J =
6.0 Hz, 1H, NH), 6.58 (d, J = 8.8 Hz, 2H), 7.06 (d, J = 8.8 Hz,
2H), 7.12 (t, J = 7.3 Hz, 1H), 7.31 (d, J = 8.3 Hz, 1H), 7.44 (t, J =
7.3 Hz, 1H), 7.57 (d, J = 7.8 Hz, 1H), 7.70 (s, 1H), 11.87 (br s,
1H, NHCO) ppm. 13C NMR (100 MHz, DMSO-d6): δ = 42.0
(CH2), 113.7 (CH), 115.0 (CH), 119.1 (Cq), 119.3 (Cq), 122.0
(CH), 127.6 (CH), 128.7 (CH), 129.7 (CH), 130.5 (Cq), 135.0
(CH), 137.8 (Cq), 147.4 (Cq), 161.8 (Cq, C=O) ppm. MS (ESI+):
m/z 285/287 [M + H]+.
5.1.3.1.
(±)-1-(1-Benzyl-1,2,3,4-tetrahydroquinolin-4-
yl)pyrrolidin-2-one 5a. Following the general procedure, the
reaction between N-arylamine 8a (119 mg, 0.65 mmol),
polyformaldehyde (29 mg, 0.96 mmol) and N-vinyl-2-
pyrrolidinone 9a (77 µL, 0.72 mmol) afforded compound 5a as a
yellow oil. Yield: 90% (179 mg) (Lit. Yellow oil).6b FTIR (film):
ν = 2928, 2877, 1682 (C=O), 1601 (C=C) cm-1. H NMR (400
1
MHz, CDCl3): δ = 1.92–2.23 (m, 4H), 2.45–2.52 (m, 2H), 3.16–
3.36 (m, 3H), 3.46–3.53 (m, 1H), 4.44 (d, J = 16.8 Hz, 1H), 4.50
(d, J = 16.8 Hz, 1H), 5.43 (dd, J = 5.5, 9.0 Hz, 1H, CH–N), 6.57
(d, J = 8.5 Hz, 1H), 6.62 (t, J = 7.3 Hz, 1H), 6.88 (d, J = 7.5 Hz,
1H), 7.03 (t, J = 7.5 Hz, 1H), 7.22–7.26 (m, 3H), 7.31 (t, J = 7.5
Hz, 2H) ppm. 13C NMR (100 MHz, CDCl3): δ = 18.4 (CH2), 26.8
(CH2), 31.5 (CH2), 44.0 (CH2), 47.7 (CH2), 48.2 (CH–N), 55.3
(CH2), 111.9 (CH), 116.4 (CH), 119.2 (Cq), 126.7 (CH), 127.0
(CH), 127.6 (CH), 128.7 (CH), 128.7 (CH), 138.4 (Cq), 146.3
(Cq), 175.5 (Cq, C=O) ppm. MS (EI) m/z (%) = 306 (21) [M]+,
215 (2), 130 (4), 85 (66), 83 (100). The NMR data matches with
the previously reported data.6b
5.1.2.12. Synthesis of N-(prop-2-ynyl)aniline 8n. A mixture of
aniline (1187 µL, 13.0 mmol), propargyl bromide solution 80 wt.
% in toluene (305 µL, 2.74 mmol) and ethanol (5.0 mL) was
stirred at room temperature for 24 h. After the excess of solvent
was removed under reduced pressure, the oily material obtained
was purified by column chromatography on silica gel using a
mixture of hexane:EtOAc (3:1) as eluent to give the product 8n
as a yellow oil. Yield: 78% (280 mg) (Lit. Yellow oil).12g FTIR
(film): ν = 3403 (N–H), 3288, 3052, 2921, 2864, 2125, 1603 cm-
1. 1H NMR (400 MHz, CDCl3): δ = 2.26 (t, J = 2.0 Hz, 1H), 3.91
(br s, 1H, NH), 3.98 (d, J = 2.0 Hz, 2H), 6.74 (d, J = 7.5 Hz, 2H),

11
5.1.3.2.
(±)-1-(1-(4-Chlorobenzyl)-6-methyl-1,2,3,4-
procedure, the reaction between N-arylamine 8d (103 mg,
ACCEPTED MANUSCRIPT
tetrahydroquinolin-4-yl)pyrrolidin-2-one 5b. Following the
general procedure, the reaction between N-arylamine 8b (120
mg, 0.52 mmol), polyformaldehyde (24 mg, 0.80 mmol) and N-
vinyl-2-pyrrolidinone 9a (62 µL, 0.58 mmol) afforded compound
5b as a brown solid. Yield: 72% (132 mg). M.p. 122–123 °C.
0.36 mmol), polyformaldehyde (16 mg, 0.53 mmol) and N-vinyl-
2-pyrrolidinone 9a (43 µL, 0.40 mmol) afforded firstly the
tetrahydroquinoline 5d (122 mg, 83%) and then γ-aminoalcohol
11d (12 mg, 8%). Yellow solid. M.p. 102–103 °C. FTIR (KBr): ν
1
= 2946, 2831, 1677 (C=O), 1590, 1125, 1002 (C–O) cm-1. H
FTIR (KBr): ν = 2933, 2858, 1675 (C=O), 1615 (C=C) cm-1. H
NMR (400 MHz, CDCl3): δ = 1.96–2.07 (m, 3H), 2.11–2.19 (m,
1H), 2.19 (s, 3H), 2.47–2.53 (m, 2H), 3.14–3.30 (m, 3H), 3.40
(ddd, J = 3.3, 8.9, 12.1 Hz, 1H), 3.80 (s, 6H), 3.82 (s, 3H), 4.32
(d, J = 16.3 Hz, 1H), 4.39 (d, J = 16.6 Hz, 1H), 5.40 (dd, J = 5.5,
8.8 Hz, 1H), 6.48 (s, 2H), 6.54 (d, J = 8.3 Hz, 1H), 6.70 (s, 1H),
6.88 (dd, J = 1.6, 8.4 Hz, 1H) ppm. 13C NMR (100 MHz, CDCl3):
δ = 18.4 (CH2), 20.4 (CH3), 27.1 (CH2), 31.6 (CH2), 44.0 (CH2),
47.7 (CH2), 48.2 (CH–N), 56.0 (CH2), 56.2 (OCH3), 60.9
(OCH3), 103.6 (CH), 112.4 (CH), 119.6 (Cq), 126.0 (Cq), 128.3
(CH), 129.3 (CH), 134.6 (Cq), 137.0 (Cq), 144.4 (Cq), 153.6
(Cq), 175.5 (Cq, C=O) ppm. MS (ESI+): m/z 411 [M + H]+.
Anal. Calcd for C24H30N2O4: C, 70.22; H, 7.37; N, 6.82. Found:
C, 70.46; H, 7.58; N, 6.59.
1
NMR (400 MHz, CDCl3): δ = 1.98–2.07 (m, 3H), 2.12–2.18 (m,
1H), 2.18 (s, 3H), 2.48–2.53 (m, 2H), 3.17–3.30 (m, 3H), 3.43
(ddd, J = 3.5, 9.1, 12.1 Hz, 1H), 4.36 (d, J = 16.8 Hz, 1H), 4.42
(d, J = 16.8 Hz, 1H), 5.40 (dd, J = 5.5, 8.8 Hz, 1H, CH–N), 6.43
(d, J = 8.5 Hz, 1H), 6.70 (s, 1H), 6.85 (dd, J = 2.0, 8.0 Hz, 1H),
7.18 (d, J = 8.5 Hz, 2H), 7.28 (d, J = 8.5 Hz, 2H) ppm. 13C NMR
(100 MHz, CDCl3): δ = 18.4 (CH2), 20.4 (CH3), 27.1 (CH2), 31.6
(CH2), 44.0 (CH2), 47.9 (CH2), 48.1 (CH–N), 55.1 (CH2), 112.1
(CH), 119.5 (Cq), 126.0 (Cq), 128.1 (CH), 128.3 (CH), 128.8
(CH), 129.3 (CH), 132.7 (Cq), 137.3 (Cq), 144.0 (Cq), 175.5
(Cq, C=O) ppm. MS (ESI+): m/z 377/379 [M + Na]+. Anal. Calcd
for C21H23ClN2O: C, 71.07; H, 6.53; N, 7.89. Found: C, 71.28; H,
6.66; N, 7.71.
5.1.3.6. (±)-1-(3-(N-(3,4,5-Trimethoxybenzyl)-N-p-tolylamino)-1-
hydroxypropyl)pyrrolidin-2-one 11d. Yellow solid. M.p. 119–
120 °C. FTIR (KBr): ν = 3477 (O–H), 2935, 2835, 1681 (C=O),
1592, 1233, 1126, 1007 (C–O) cm-1. 1H NMR (400 MHz,
CDCl3): δ = 1.80–1.88 (m, 1H), 1.92–2.05 (m, 3H), 2.24 (s, 3H),
2.33–2.39 (m, 2H), 3.23–3.30 (m, 1H), 3.35–3.43 (m, 1H), 3.48–
3.55 (m, 2H), 3.78 (s, 6H), 3.82 (s, 3H), 4.39 (s, 2H), 5.54 (dd, J
= 4.5, 8.6 Hz, 1H), 6.44 (s, 2H), 6.70 (d, J = 8.5 Hz, 2H), 7.02 (d,
J = 8.5 Hz, 2H) ppm, OH is absent. 13C NMR (100 MHz,
CDCl3): δ = 18.2 (CH2), 20.3 (CH3), 30.9 (CH2), 31.8 (CH2), 42.1
(CH2), 47.7 (CH2), 56.1 (CH2), 56.2 (OCH3), 60.9 (OCH3), 74.2
(NCH–O), 103.8 (CH), 114.1 (CH), 126.8 (Cq), 129.9 (CH),
134.9 (Cq), 137.0 (Cq), 146.5 (Cq), 153.5 (Cq), 176.1 (Cq, C=O)
ppm. MS (EI) m/z (%) = 343 (68) [M-85]+, 287 (50), 181 (100),
148 (45), 134 (33).
5.1.3.3.
(±)-1-(1-(2-Chlorobenzyl)-6-methyl-1,2,3,4-
tetrahydroquinolin-4-yl)pyrrolidin-2-one 5c. Following the
general procedure, the reaction between N-arylamine 8c (99 mg,
0.43 mmol), polyformaldehyde (19 mg, 0.63 mmol) and N-vinyl-
2-pyrrolidinone 9a (51 µL, 0.48 mmol) afforded firstly the
tetrahydroquinoline 5c (124 mg, 81%) and then γ-aminoalcohol
11c (19 mg, 12%). Brown solid. M.p. 109–110 °C. FTIR (KBr):
ν = 2963, 2820, 1680 (C=O), 1616 (C=C) cm-1. H NMR (400
1
MHz, CDCl3): δ = 1.99–2.11 (m, 3H), 2.18 (s, 3H), 2.19–2.25
(m, 1H), 2.47–2.53 (m, 2H), 3.18–3.35 (m, 3H), 3.47–3.54 (m,
1H), 4.47 (s, 2H), 5.42 (dd, J = 5.4, 8.6 Hz, 1H, CH–N), 6.29 (d,
J = 8.3 Hz, 1H), 6.71 (s, 1H), 6.82 (d, J = 8.3 Hz, 1H), 7.16–7.22
(m, 3H), 7.36–7.38 (m, 1H) ppm. 13C NMR (100 MHz, CDCl3): δ
= 18.3 (CH2), 20.3 (CH3), 27.0 (CH2), 31.5 (CH2), 44.0 (CH2),
47.9 (CH2), 48.1 (CH–N), 53.7 (CH2), 111.8 (CH), 119.1 (Cq),
125.7 (Cq), 126.9 (CH), 127.6 (CH), 128.0 (CH), 128.0 (CH),
129.2 (CH), 129.5 (CH), 132.9 (Cq), 135.4 (Cq), 143.7 (Cq),
175.4 (Cq, C=O) ppm. MS (ESI+): m/z 355/357 [M + H]+,
377/379 [M + Na]+. Anal. Calcd for C21H23ClN2O: C, 71.07; H,
6.53; N, 7.89. Found: C, 71.15; H, 6.41; N, 7.99.
5.1.3.7
(±)-1-(1-((Benzo[d][1,3]dioxol-5-yl)methyl)-1,2,3,4-
5e.
tetrahydro-6-methoxyquinolin-4-yl)pyrrolidin-2-one
Following the general procedure, the reaction between N-
arylamine 8e (139 mg, 0.54 mmol), polyformaldehyde (25 mg,
0.83 mmol) and N-vinyl-2-pyrrolidinone 9a (66 µL, 0.62 mmol)
afforded compound 5e as a brown solid. Yield: 75% (154 mg).
M.p. 130–132 °C. FTIR (KBr): ν = 2942, 2837, 1677 (C=O),
1236, 1035 (C–O) cm-1. 1H NMR (400 MHz, CDCl3): δ = 1.95–
2.06 (m, 3H), 2.09–2.18 (m, 1H), 2.48 (td, J = 2.4, 8.1 Hz, 2H),
3.16–3.28 (m, 3H), 3.38 (ddd, J = 3.4, 9.6, 11.8 Hz, 1H), 3.69 (s,
3H), 4.26 (d, J = 16.1 Hz, 1H), 4.33 (d, J = 16.3 Hz, 1H), 5.41
(dd, J = 5.8, 9.0 Hz, 1H, CH–N), 5.92 (s, 2H), 6.50 (d, J = 3.1
Hz, 1H), 6.54 (d, J = 9.0 Hz, 1H), 6.66 (dd, J = 2.9, 9.0 Hz, 1H),
6.70–6.77 (m, 3H) ppm. 13C NMR (100 MHz, CDCl3): δ = 18.4
(CH2), 27.0 (CH2), 31.5 (CH2), 43.7 (CH2), 47.8 (CH2), 48.3
(CH–N), 55.9 (CH2), 56.0 (OCH3), 101.0 (OCH2O), 107.4 (CH),
108.4 (CH), 113.4 (CH), 113.5 (CH), 114.2 (CH), 119.9 (CH),
121.1 (Cq), 132.7 (Cq), 141.0 (Cq), 146.6 (Cq), 148.0 (Cq),
151.5 (Cq), 175.5 (Cq, C=O) ppm. MS (ESI+): m/z 381 [M +
H]+, 403 [M + Na]+. Anal. Calcd for C22H24N2O4: C, 69.46; H,
6.36; N, 7.36. Found: C, 69.68; H, 6.12; N, 7.45.
5.1.3.4.
(±)-(3-((2-Chlorobenzyl)(p-tolyl)amino)-1-
hydroxypropyl)pyrrolidin-2-one 11c. Brown solid. M.p. 138–139
°C. FTIR (KBr): ν = 3269 (O–H), 2955, 2831, 1662 (C=O), 1616
(C=C), 1194, 1090, 1043 (C–O) cm-1. 1H NMR (400 MHz,
CDCl3): δ = 1.85–2.14 (m, 4H), 2.23 (s, 3H), 2.30–2.45 (m, 2H),
3.33 (ddd, J = 5.8, 8.0, 9.6 Hz, 1H), 3.45 (ddd, J = 5.3, 8.9, 14.9
Hz, 1H), 3.54–3.65 (m, 2H), 4.30 (br s, 1H), 4.52 (d, J = 18.1 Hz,
1H), 4.56 (d, J = 18.1 Hz, 1H), 5.55 (dd, J = 4.3, 8.8 Hz, 1H,
NCH–O), 6.59 (d, J = 8.5 Hz, 2H), 7.00 (d, J = 8.7 Hz, 2H),
7.13–7.19 (m, 3H), 7.37 (d, J = 7.4 Hz, 1H) ppm. 13C NMR (100
MHz, CDCl3): δ = 18.2 (CH2), 20.3 (CH3), 31.3 (CH2), 31.8
(CH2), 42.1 (CH2), 48.1 (CH2), 53.0 (CH2), 73.8 (NCH–O), 112.7
(CH), 126.1 (Cq), 126.9 (CH), 128.1 (CH), 128.1 (CH), 129.6
(CH), 130.0 (CH), 132.9 (Cq), 135.9 (Cq), 145.8 (Cq), 176.2
(Cq, C=O) ppm. MS (ESI+): m/z 395/397 [M + Na]+.
5.1.3.5. (±)-1-(1-(3,4,5-Trimethoxybenzyl)-1,2,3,4-tetrahydro-6-
5.1.3.8.
(±)-1-(1-(4-Chlorobenzyl)-1,2,3,4-tetrahydro-6-
methylquinolin-4-yl)pyrrolidin-2-one 5d. Following the general
methoxyquinolin-4-yl)pyrrolidin-2-one 5f. Following the general

12
Tetrahedron
ACCEPTED MANUSCRIPT
procedure, the reaction between N-arylamine 8f (128 mg, 0.52
vinyl-2-pyrrolidinone 9a (63 µL, 0.59 mmol) afforded compound
mmol), polyformaldehyde (23 mg, 0.76 mmol) and N-vinyl-2-
pyrrolidinone 9a (62 µL, 0.58 mmol) afforded compound 5f as a
brown solid. Yield: 93% (179 mg). M.p. 134–136 °C. FTIR
(KBr): ν = 2943, 2827, 1679 (C=O), 1575 (C=C), 1257, 1066,
1036 (C–O) cm-1. 1H NMR (400 MHz, CDCl3): δ = 1.95–2.07 (m,
3H), 2.10–2.18 (m, 1H), 2.48 (td, J = 2.4, 8.1 Hz, 2H), 3.15–3.29
(m, 3H), 3.39 (ddd, J = 3.4, 9.6, 11.6 Hz, 1H), 3.68 (s, 3H), 4.32
(d, J = 16.6 Hz, 1H), 4.37 (d, J = 16.8 Hz, 1H), 5.41 (dd, J = 5.6,
9.2 Hz, 1H, CH–N), 6.45 (d, J = 9.0 Hz, 1H), 6.50 (d, J = 2.8 Hz,
1H), 6.64 (dd, J = 3.0, 8.8 Hz, 1H), 7.18 (d, J = 8.3 Hz, 2H), 7.27
(d, J = 8.3 Hz, 2H) ppm. 13C NMR (100 MHz, CDCl3): δ = 18.4
(CH2), 27.0 (CH2), 31.5 (CH2), 43.7 (CH2), 48.1 (CH2), 48.2
(CH–N), 55.6 (CH2), 55.8 (OCH3), 113.3 (CH), 113.5 (CH),
114.2 (CH), 121.1 (Cq), 128.2 (CH), 128.8 (CH), 132.6 (Cq),
137.4 (Cq), 140.8 (Cq), 151.6 (Cq), 175.5 (Cq, C=O) ppm. MS
(ESI+): m/z 371/373 [M + H]+. Anal. Calcd for C21H23ClN2O2: C,
68.01; H, 6.25; N, 7.55. Found: C, 67.81; H, 6.13; N, 7.31.
5h as a brown solid. Yield: 68% (132 mg). M.p. 129–130 °C.
FTIR (KBr): ν = 2933, 2822, 1676 (C=O), 1598(C=C) cm-1. 1H
NMR (400 MHz, CDCl3): δ = 1.99–2.07 (m, 3H), 2.11–2.20 (m,
1H), 2.42–2.57 (m, 2H), 3.16–3.33 (m, 3H), 3.48 (ddd, J = 3.5,
9.9, 11.9 Hz, 1H), 4.37 (d, J = 17.1 Hz, 1H), 4.42 (d, J = 17.1 Hz,
1H), 5.38 (dd, J = 5.4, 9.8 Hz, 1H, CH–N), 6.39 (d, J = 8.7 Hz,
1H), 6.82 (dd, J = 1.0, 2.4 Hz, 1H), 6.95 (dd, J = 2.5, 8.8 Hz,
1H), 7.14 (d, J = 8.5 Hz, 2H), 7.27 (d, J = 8.5 Hz, 2H) ppm. 13C
NMR (100 MHz, CDCl3): δ = 18.3 (CH2), 26.5 (CH2), 31.3
(CH2), 43.6 (CH2), 47.9 (CH2), 47.9 (CH–N), 54.9 (CH2), 113.1
(CH), 121.2 (Cq), 121.5 (Cq), 127.0 (CH), 127.9 (CH), 128.5
(CH), 128.9 (CH), 132.9 (Cq), 136.4 (Cq), 144.6 (Cq), 175.6
(Cq, C=O). MS (EI) m/z (%) = 378/376/374 (1/8/12) [M]+,
251/249 (19/55), 166/164 (20/57), 127/125 (34/100). Anal. Calcd
for C20H20Cl2N2O: C, 64.01; H, 5.37; N, 7.46. Found: C, 64.19;
H, 5.62; N, 7.22.
5.1.3.12. (±)-1-(1-(4-Chlorobenzyl)-1,2,3,4-tetrahydroquinolin-4-
yl)pyrrolidin-2-one 5i. Following the general procedure, the
reaction between N-arylamine 8i (102 mg, 0.47 mmol),
polyformaldehyde (21 mg, 0.70 mmol) and N-vinyl-2-
pyrrolidinone 9a (56 µL, 0.52 mmol) afforded compound 5i as a
brown solid. Yield: 86% (137 mg). M.p. 117 °C. FTIR (KBr): ν
5.1.3.9.
(±)-1-(1-(4-Methoxybenzyl)-6-chloro-1,2,3,4-
tetrahydroquinolin-4-yl)pyrrolidin-2-one 5g. Following the
general procedure ,the reaction between N-arylamine 8g (119 mg,
0.48 mmol), polyformaldehyde (23 mg, 0.76 mmol) and N-vinyl-
2-pyrrolidinone 9a (57 µL, 0.53 mmol) afforded firstly the
tetrahydroquinoline 5g (124 mg, 70%) and then γ-aminoalcohol
11g (26 mg, 14%). Brown solid. M.p. 124–126 °C. FTIR (KBr):
ν = 2943, 2835, 1678 (C=O), 1168, 1124, 1089, 1041 (C–O) cm-
1. 1H NMR (400 MHz, CDCl3): δ = 1.98–2.15 (m, 4H), 2.46–2.52
(m, 2H), 3.16–3.33 (m, 3H), 3.45 (ddd, J = 4.0, 9.4, 12.4 Hz,
1H), 3.76 (s, 3H), 4.35 (d, J = 17.6 Hz, 1H), 4.39 (d, J = 17.8 Hz,
1H), 5.36 (dd, J = 5.3, 9.4 Hz, 1H, CH–N), 6.48 (d, J = 8.8 Hz,
1H), 6.80 (d, J = 2.3 Hz, 1H), 6.84 (d, J = 8.5 Hz, 2H), 6.95 (dd,
J = 2.6, 8.8 Hz, 1H), 7.12 (d, J = 8.5 Hz, 2H) ppm. 13C NMR
(100 MHz, CDCl3): δ = 18.3 (CH2), 26.5 (CH2), 31.4 (CH2), 43.6
(CH2), 47.6 (CH2), 48.0 (CH–N), 54.7 (CH2), 55.3 (OCH3), 113.2
(CH), 114.2 (CH), 121.0 (Cq), 121.1 (Cq), 127.0 (CH), 127.8
(CH), 128.4 (CH), 129.7 (Cq), 144.9 (Cq), 158.8 (Cq), 175.5
(Cq, C=O) ppm. MS (ESI+): m/z 371/373 [M + H]+, 393/395 [M
+ Na]+. Anal. Calcd for C21H23ClN2O2: C, 68.01; H, 6.25; N,
7.55. Found: C, 68.24; H, 6.38; N, 7.73.
1
= 2936, 2847, 1681 (C=O), 1602(C=C) cm-1. H NMR (400
MHz, CDCl3): δ = 2.00–2.08 (m, 3H), 2.14–2.23 (m, 1H), 2.50
(td, J = 2.4, 8.1 Hz, 2H), 3.15–3.34 (m, 3H), 3.48 (td, J = 3.5,
10.6 Hz, 1H), 4.40 (d, J = 17.0 Hz, 1H), 4.45 (d, J = 16.8 Hz,
1H), 5.43 (dd, J = 5.3, 9.2 Hz, 1H, CH–N), 6.51 (d, J = 8.3 Hz,
1H), 6.64 (t, J = 7.4 Hz, 1H), 6.89 (d, J = 7.5 Hz, 1H), 7.03 (td, J
= 1.4, 7.8 Hz, 1H), 7.18 (d, J = 8.5 Hz, 2H), 7.28 (d, J = 8.5 Hz,
2H) ppm. 13C NMR (100 MHz, CDCl3): δ = 18.4 (CH2), 26.8
(CH2), 31.6 (CH2), 43.9 (CH2), 47.9 (CH2), 48.2 (CH–N), 54.9
(CH2), 111.9 (CH), 116.8 (CH), 119.5 (Cq), 127.7 (CH), 128.1
(CH), 128.7 (CH), 128.9 (CH), 132.8 (Cq), 137.0 (Cq), 146.1
(Cq), 175.6 (Cq, C=O) ppm. MS (EI) m/z (%) = 342/340 (5/15)
[M]+, 215 (73), 130 (100), 127/125 (27/83). Anal. Calcd for
C20H21ClN2O: C, 70.48; H, 6.21; N, 8.22. Found: C, 70.33; H,
6.39; N, 7.99.
5.1.3.13.
(±)-3-((3,4-Dihydro-6-methyl-4-(2-oxopyrrolidin-1-
5.1.3.10.
(±)-1-(3-(N-(4-Methoxybenzyl)-N-(4-
11g.
yl)quinolin-1(2H)-yl)methyl)quinolin-2(1H)-one 5j. Following
the general procedure, the reaction between N-arylamine 8j (82
mg, 0.31 mmol), polyformaldehyde (14 mg, 0.47 mmol) and N-
vinyl-2-pyrrolidinone 9a (41 µL, 0.38 mmol) in CH3CN/DMSO
(3:1, 2.0 mL) at 60 °C for 24 h afforded compound 5j as a white
solid. Yield: 69% (83 mg). M.p. 243–244 °C. FTIR (KBr): ν =
3441 (N–H), 2950, 2862, 1668 (NC=O, NHC=O), 1615, 1577cm-
1. 1H NMR (400 MHz, CDCl3): δ = 1.99–2.16 (m, 3H), 2.20 (s,
3H), 2.22–2.30 (m, 1H), 2.49–2.58 (m, 2H), 3.21–3.36 (m, 2H),
3.41 (ddd, J = 4.2, 7.0, 11.6 Hz, 1H), 3.58 (ddd, J = 3.1, 8.8, 12.0
Hz, 1H), 4.45 (d, J = 18.6 Hz, 1H), 4.51 (d, J = 18.6 Hz, 1H),
5.46 (dd, J = 5.3, 8.4 Hz, 1H, CH–N), 6.44 (d, J = 8.5 Hz, 1H),
6.74 (s, 1H), 6.86 (d, J = 8.3 Hz, 1H), 7.19 (t, J = 7.5 Hz, 1H),
7.40 (d, J = 8.3 Hz, 1H), 7.45–7.50 (m, 2H), 7.61 (s, 1H), 11.87
(br s, 1H, NH) ppm. 13C NMR (100 MHz, CDCl3): δ = 18.5
(CH2), 20.4 (CH3), 27.3 (CH2), 31.6 (CH2), 44.2 (CH2), 48.2
(CH–N), 48.3 (CH2), 51.8 (CH2), 112.2 (CH), 115.7 (CH), 119.3
(Cq), 120.1 (Cq), 122.7 (CH), 126.0 (Cq), 127.8 (CH), 128.3
(CH), 129.0 (Cq), 129.5 (CH), 130.0 (CH), 135.7 (CH), 137.5
chlorophenyl)amino)-1-hydroxypropyl)pyrrolidin-2-one
Yellow oil. FTIR (film): ν = 3267 (O–H), 2943, 2840, 1681
(C=O), 1170, 1118, 1093, 1033 (C–O) cm-1. 1H NMR (400 MHz,
CDCl3): δ = 1.78–1.86 (m, 1H), 1.93–2.06 (m, 3H), 2.31–2.39
(m, 2H), 3.28 (ddd, J = 5.8, 8.1, 9.5 Hz, 1H), 3.41 (ddd, J = 6.3,
8.7, 14.9 Hz, 1H), 3.50–3.58 (m, 2H), 3.77 (s, 3H), 4.03 (br s,
1H), 4.43 (s, 2H), 5.51 (dd, J = 4.3, 8.8 Hz, 1H, NCH–O), 6.63
(d, J = 9.0 Hz, 2H), 6.83 (d, J = 8.5 Hz, 2H), 7.07–7.12 (m, 4H)
ppm. 13C NMR (100 MHz, CDCl3): δ = 18.2 (CH2), 31.1 (CH2),
31.8 (CH2), 42.1 (CH2), 47.7 (CH2), 54.4 (CH2), 55.4 (OCH3),
73.7 (NCH–O), 114.1 (CH), 114.2 (CH), 121.6 (Cq), 127.9 (CH),
129.1 (CH), 130.1 (Cq), 147.0 (Cq), 158.8 (Cq), 176.3 (Cq, C=O)
ppm.
5.1.3.11.
(±)-1-(1-(4-Chlorobenzyl)-6-chloro-1,2,3,4-
tetrahydroquinolin-4-yl)pyrrolidin-2-one 5h. Following the
general procedure, the reaction between N-arylamine 8h (131
mg, 0.52 mmol), polyformaldehyde (23 mg, 0.76 mmol) and N-

13
(Cq), 144.0 (Cq), 163.7 (Cq, C=O), 175.6 (Cq, C=O) ppm. MS
(EI) m/z (%) = 387 (7) [M]+, 229 (29), 144 (100), 130 (16). Anal.
Calcd for C24H25N3O2: C, 74.39; H, 6.50; N, 10.84. Found: C,
74.46; H, 6.69; N, 10.63.
11g (34 mg, 14%). Yellow oil. FTIR (film): ν = 2947, 2851,
ACCEPTED MANUSCRIPT
1678 (C=O), 1243, 1086, 1026 (C–O) cm-1. 1H NMR (400 MHz,
CDCl3): δ = 1.86–2.05 (m, 4H), 2.47 (t, J = 8.0 Hz, 2H), 2.86 (s,
3H), 3.06 (ddd, J = 5.0, 8.2, 9.6 Hz, 1H), 3.13–3.30 (m, 3H), 3.86
(s, 3H), 5.39 (dd, J = 6.8, 8.8 Hz, 1H, CH–N), 6.61 (d, J = 7.8
Hz, 1H), 6.73 (d, J = 7.5 Hz, 1H), 6.88 (t, J = 7.9 Hz, 1H) ppm.
13C NMR (100 MHz, CDCl3): δ = 18.3 (CH2), 22.0 (CH2), 31.5
(CH2), 42.0 (CH3), 43.4 (CH2), 48.1 (CH–N), 50.6 (CH2), 55.5
(OCH3), 110.0 (CH), 119.9 (CH), 121.9 (CH), 127.2 (Cq), 139.0
(Cq), 152.3 (Cq), 175.6 (Cq, C=O) ppm. MS (ESI+): m/z261 [M
+ H]+, 283 [M + Na]+. Anal. Calcd for C15H20N2O2: C, 69.20; H,
7.74; N, 10.76. Found: C, 69.13; H, 7.51; N, 10.58.
5.1.3.14. (±)-3-((3,4-Dihydro-6-methoxy-4-(2-oxopyrrolidin-1-
yl)quinolin-1(2H)-yl)methyl)quinolin-2(1H)-one 5k. Following
the general procedure, the reaction between N-arylamine 8k (95
mg, 0.34 mmol), polyformaldehyde (15 mg, 0.50 mmol) and N-
vinyl-2-pyrrolidinone 9a (45 µL, 0.42 mmol) in CH3CN/DMSO
(3:1, 2.0 mL) at 60 °C for 24 h afforded compound 5k as a brown
solid. Yield: 61% (84 mg). M.p. 229–230 °C. FTIR (KBr): ν =
3440 (N–H), 2954, 2846, 1671 (NC=O, NHC=O), 1615, 1202,
1061, 1032 (C–O) cm-1. 1H NMR (400 MHz, CDCl3): δ = 2.00–
2.16 (m, 3H), 2.21–2.30 (m, 1H), 2.50–2.55 (m, 2H), 3.22–3.41
(m, 3H), 3.57 (ddd, J = 2.9, 9.0, 10.4 Hz, 1H), 3.70 (s, 3H), 4.43
(d, J = 19.6 Hz, 1H), 4.48 (d, J = 19.6 Hz, 1H), 5.49 (dd, J = 5.3,
6.8 Hz, 1H, CH–N), 6.49 (d, J = 9.0 Hz, 1H), 6.55 (d, J = 2.5 Hz,
1H), 6.66 (dd, J = 3.0, 9.0 Hz, 1H), 7.20 (td, J = 1.2, 7.5 Hz, 1H),
7.40 (d, J = 7.8 Hz, 1H), 7.45–7.52 (m, 2H), 7.63 (s, 1H), 11.76
(br s, 1H, NH) ppm. 13C NMR (100 MHz, CDCl3): δ = 18.5
(CH2), 27.2 (CH2), 31.6 (CH2), 44.0 (CH2), 48.4 (CH–N), 48.5
(CH2), 52.3 (CH2), 56.0 (OCH3), 113.4 (CH), 113.6 (CH), 114.4
(CH), 115.7 (CH), 120.1 (Cq), 120.8 (Cq), 122.8 (CH), 127.8
(CH), 129.2 (Cq), 130.0 (CH), 135.8 (CH), 137.5 (Cq), 140.7
(Cq), 151.6 (Cq), 163.7 (Cq, C=O), 175.6 (Cq, C=O) ppm. MS
(EI) m/z (%) = 403 (18) [M]+, 313 (6), 245 (38), 160 (100), 130
(18). Anal. Calcd for C24H25N3O3: C, 71.44; H, 6.25; N, 10.41.
Found: C, 71.31; H, 6.41; N, 10.22.
5.1.3.17.
(±)-1-(3-(N-(2-Methoxyphenyl)-N-methylamino)-1-
hydroxypropyl)pyrrolidin-2-one 11m. Yellow oil (Lit. Yellow
oil).6e FTIR (film): ν = 3325 (O–H), 2952, 2849, 1674 (C=O),
1238, 1079, 1027 (C–O) cm-1. 1H NMR (400 MHz, CDCl3): δ =
1.61–1.68 (m, 1H), 1.90–2.01 (m, 3H), 2.34 (t, J = 8.2 Hz, 2H),
2.71 (s, 3H), 3.07 (ddd, J = 5.5, 7.0, 12.8 Hz, 1H), 3.16 (ddd, J =
6.5, 7.6, 13.6 Hz, 1H), 3.26–3.37 (m, 1H), 3.56 (ddd, J = 5.8, 5.8,
12.2 Hz, 1H), 3.83 (s, 3H), 5.34 (br s, 1H, OH), 5.62 (dd, J = 4.0,
8.8 Hz, 1H, NCH–O), 6.83–6.90 (m, 2H), 6.97–7.02 (m, 2H)
ppm. 13C NMR (100 MHz, CDCl3): δ = 18.1 (CH2), 30.5 (CH2),
31.8 (CH2), 40.6 (CH3), 41.8 (CH2), 52.6 (CH2), 55.3 (OCH3),
75.2 (NCH–O), 111.3 (CH), 120.1 (CH), 120.8 (CH), 123.7
(CH), 141.1 (Cq), 153.0 (Cq), 175.4 (Cq, C=O) ppm. MS (ESI+):
m/z279 [M + H]+, 301 [M + Na]+. The NMR data matches with
the previously reported data.6e
5.1.3.18.
(±)-1-(1,2,3,4-Tetrahydro-1-(prop-2-ynyl)quinolin-4-
5.1.3.15.
(±)-3-((6-Chloro-3,4-dihydro-4-(2-oxopyrrolidin-1-
yl)pyrrolidin-2-one 5n. Following the general procedure, the
reaction between N-arylamine 8n (122 mg, 0.93 mmol),
polyformaldehyde (45 mg, 1.50 mmol) and N-vinyl-2-
pyrrolidinone 9a (120 µL, 1.12 mmol) afforded compound 5n as
a red solid. Yield: 81% (191 mg). M.p. 88 °C (Lit. 88–90 °C).13
1H NMR (400 MHz, CDCl3): δ = 1.88–2.19 (m, 5H), 2.47 (t, J =
8.0 Hz, 2H), 3.09 (ddd, J = 5.3, 8.0, 9.8 Hz, 1H), 3.21–3.30 (m,
2H), 3.40 (ddd, J = 3.3, 8.8, 11.8 Hz, 1H), 3.97 (dd, J = 2.3, 18.2
Hz, 1H), 4.05 (dd, J = 2.3, 18.0 Hz, 1H), 5.39 (dd, J = 6.1, 8.4
Hz, 1H, CH–N), 6.72 (t, J = 7.5 Hz, 1H), 6.76 (d, J = 8.0 Hz,
1H), 6.90 (d, J = 7.5 Hz, 1H), 7.15 (t, J = 7.3 Hz, 1H) ppm. 13C
NMR (100 MHz, CDCl3): δ = 18.4 (CH2), 26.8 (CH2), 31.5
(CH2), 40.9 (CH2), 43.8 (CH2), 47.4 (CH2), 47.8 (CH–N), 72.1
(CH), 79.1 (Cq), 112.9 (CH), 118.2 (CH), 121.1 (Cq), 128.0
(CH), 128.5 (CH), 145.4 (Cq), 175.4 (Cq, C=O) ppm. The NMR
data matches with the previously reported data.13
yl)quinolin-1(2H)-yl)methyl)quinolin-2(1H)-one 5l. Following
the general procedure, the reaction between N-arylamine 8l (116
mg, 0.41 mmol), polyformaldehyde (20 mg, 0.67 mmol) and N-
vinyl-2-pyrrolidinone 9a (53 µL, 0.50 mmol) in CH3CN/DMSO
(3:1, 2.0 mL) at 60 °C for 24 h afforded compound 5l as a white
solid. Yield: 83% (139 mg). M.p. 278–280 °C. FTIR (KBr): ν =
3447 (N–H), 2950, 2857, 1657 (NC=O, NHC=O), 1605 cm-1. 1H
NMR (400 MHz, CDCl3): δ = 2.02–2.13 (m, 3H), 2.21–2.30 (m,
1H), 2.47–2.60 (m, 2H), 3.22–3.36 (m, 2H), 3.43 (ddd, J = 4.3,
5.8, 12.0 Hz, 1H), 3.63 (ddd, J = 2.9, 9.5, 12.1 Hz, 1H), 4.45 (d, J
= 19.1 Hz, 1H), 4.52 (d, J = 19.1 Hz, 1H), 5.46 (dd, J = 5.1, 9.6
Hz, 1H, CH–N), 6.45 (d, J = 8.8 Hz, 1H), 6.86 (d, J = 1.8 Hz,
1H), 6.98 (dd, J = 2.3, 6.7 Hz, 1H), 7.21 (t, J = 7.5 Hz, 1H), 7.39
(d, J = 8.0 Hz, 1H), 7.47–7.51 (m, 2H), 7.54 (s, 1H), 11.91 (br s,
1H, NH) ppm. 13C NMR (100 MHz, CDCl3): δ = 18.4 (CH2), 26.7
(CH2), 31.4 (CH2), 43.9 (CH2), 48.2 (CH–N), 48.3 (CH2), 51.7
(CH2), 113.3 (CH), 115.7 (CH), 119.9 (Cq), 121.1 (Cq), 121.6
(Cq), 122.9 (CH), 127.1 (CH), 127.8 (CH), 128.3 (Cq), 128.7
(CH), 130.2 (CH), 135.8 (CH), 137.6 (Cq), 144.6 (Cq), 163.7
(Cq, C=O), 175.7 (Cq, C=O) ppm. Anal. Calcd for
C23H22ClN3O2: C, 67.73; H, 5.44; N, 10.30. Found: C, 67.56; H,
5.26; N, 10.45.
5.1.3.19.
(±)-1-(1-(4-Chlorobenzyl)-1,2,3,4-tetrahydro-6-
methoxyquinolin-4-yl)azepan-2-one 5o. Following the general
procedure, the reaction between N-arylamine 8f (118 mg, 0.48
mmol), polyformaldehyde (22 mg, 0.73 mmol) and N-
vinylcaprolactam 9b (78 µL, 0.58 mmol) afforded compound 5o
as a yellow oil. Yield: 89% (170 mg). FTIR (film): ν = 2932,
2856, 1639 (C=O), 1191, 1088, 1059 (C–O) cm-1. 1H NMR (400
MHz, CDCl3): δ = 1.41–1.48 (m, 1H), 1.59–1.85 (m, 5H), 2.05–
2.10 (m, 2H), 2.62–2.72 (m, 2H), 3.08–3.27 (m, 3H), 3.42 (ddd, J
= 4.6, 6.3, 11.5 Hz, 1H), 3.71 (s, 3H), 4.32 (d, J = 16.6 Hz, 1H),
4.40 (d, J = 16.6 Hz, 1H), 5.94 (t, J = 8.2 Hz, 1H, CH–N), 6.48
(d, J = 8.8 Hz, 1H), 6.55 (d, J = 2.3 Hz, 1H), 6.65 (dd, J = 2.8,
8.8 Hz, 1H), 7.22 (d, J = 8.0 Hz, 2H), 7.30 (d, J = 8.0 Hz, 2H)
5.1.3.16. (±)-1-(1,2,3,4-Tetrahydro-8-methoxy-1-methylquinolin-
4-yl)pyrrolidin-2-one 5m. Following the general procedure, the
reaction between N-arylamine 8m (121 mg, 0.88 mmol),
polyformaldehyde (41 mg, 1.36 mmol) and N-vinyl-2-
pyrrolidinone 9a (108 µL, 1.01 mmol) afforded firstly the
tetrahydroquinoline 5m (162 mg, 71%) and then γ-aminoalcohol

14
Tetrahedron
ACCEPTED MANUSCRIPT
ppm. 13C NMR (100 MHz, CDCl ): δ = 23.7 (CH ), 27.3
monochromator. ωscans, (λ = 0.71073 Å), multiscan absorption
3
2
(CH2), 29.6 (CH2), 30.1 (CH2), 37.7 (CH2), 45.2 (CH2), 48.6
(CH2), 51.5 (CH–N), 55.8 (CH2), 55.9 (OCH3), 113.4 (CH),
113.9 (CH), 114.0 (CH), 122.4 (Cq), 128.3 (CH), 128.8 (CH),
132.7 (Cq), 137.5 (Cq), 141.5 (Cq), 151.6 (Cq), 176.6 (Cq, C=O)
ppm. MS (EI) m/z (%) = 400/398 (7/21) [M]+, 160 (54), 127/125
(21/65), 96 (91), 43 (100). Anal. Calcd for C23H27ClN2O2: C,
69.25; H, 6.82; N, 7.02. Found: C, 69.34; H, 7.01; N, 7.34.
correction (CrysAlis PRO)[14]. Tmin, Tmax 0.839, 1.000. No. of
measured, independent and observed [I > 2σ(I)] reflections 7880,
3705, 2854, Rint = 0.025, θ values (°): θmax = 27.50, θmin = 3.08;
Range h = -11→11, k = -10→10, l = -31→24, Refinement on F2:
R[F2> 2σ(F2)] = 0.0458, wR(F2) = 0.0963, S =1.039. No. of
reflections 2854, No. of parameters 217, No. of restraints 0.
Weighting scheme: w = 1/σ2(Fo2) + (0.0407P)2 + 0.4964P where
P = (Fo2 + 2Fc2)/3. (∆/σ) < 0.002, ꢂρmax, ꢂρmin (e Å-3) 0.241, -
0.232.
5.1.3.20.
(±)-1-(3,4-Dimethoxybenzyl)-4-(dodecyloxy)-1,2,3,4-
tetrahydro-6-methoxyquinoline 5p. Following the general
procedure, the reaction between N-arylamine 8o (120 mg, 0.44
mmol), polyformaldehyde (20 mg, 0.67 mmol) and dodecyl vinyl
ether 9c (140 µL, 0.54 mmol) afforded compound 5p as a yellow
oil. Yield: 72% (158 mg). FTIR (film): ν = 2946, 2854, 1261,
1132, 1089, 1033 (C–O) cm-1. 1H NMR (400 MHz, CDCl3): δ =
0.88 (t, J = 6.6 Hz, 3H), 1.25–1.35 (m, 13H), 1.52–1.64 (m, 5H),
1.92–2.00 (m, 1H), 2.09–2.15 (m, 1H), 3.17 (ddd, J = 4.4, 4.5,
11.3 Hz, 1H), 3.41–3.60 (m, 3H), 3.63 (t, J = 6.4 Hz, 2H), 3.73
(s, 3H), 3.82 (s, 3H), 3.85 (s, 3H), 4.33 (t, J = 3.6 Hz, 1H, CH–
O), 4.35 (d, J = 16.3 Hz, 1H), 4.41 (d, J = 16.5 Hz, 1H), 6.53 (d,
J = 9.0 Hz, 1H), 6.70 (dd, J = 2.8, 8.4 Hz, 1H), 6.78–6.84 (m,
4H) ppm. 13C NMR (100 MHz, CDCl3): δ = 14.2 (CH3), 22.8
(CH2), 25.8 (CH2), 26.4 (CH2), 27.6 (CH2), 29.4 (CH2), 29.5
(CH2), 29.6 (CH2), 30.2 (CH2), 32.0 (CH2), 32.9 (CH2), 45.0
(CH2), 55.5 (CH2), 55.9 (OCH3), 56.0 (OCH3), 56.0 (OCH3), 63.1
(CH2), 68.2 (CH2), 73.5 (CH–O), 110.0 (CH), 111.4 (CH), 112.9
(CH), 115.3 (CH), 116.0 (CH), 118.7 (CH), 122.7 (Cq), 131.7
(Cq), 140.3 (Cq), 148.0 (Cq), 149.3 (Cq), 150.6 (Cq) ppm. MS
(EI) m/z (%) = 497 (21) [M]+, 311 (7), 160 (17), 151 (100). Anal.
Calcd for C31H47NO4: C, 74.81; H, 9.52; N, 2.81. Found: C,
74.70; H, 9.36; N, 2.68.
5.1.3.22. (±)-1-(2-oxopyrrolidin-1-yl)julolidine1 5. Following the
general
procedure,
the
reaction
between
1,2,3,4-
tetrahydroquinoline 8q (114 µL, 0.91 mmol), polyformaldehyde
(41 mg, 1.36 mmol) and N-vinyl-2-pyrrolidinone 9a (108 µL,
1.01 mmol) afforded compound 15 as a red oil. Yield: 89% (207
mg) (Lit. Red oil).5f FTIR (film): ν = 2932, 2862, 1682 (C=O),
1
1598 cm-1. H NMR (400 MHz, CDCl3): δ = 1.95–2.15 (m, 6H),
2.47 (td, J = 1.2, 8.6 Hz, 2H), 2.68–2.82 (m, 2H), 3.05–3.17 (m,
4H), 3.20–3.27 (m, 2H), 5.39 (dd, J = 6.3, 9.0 Hz, 1H, CH–N),
6.53 (t, J = 7.4 Hz, 1H), 6.69 (d, J = 7.3 Hz, 1H), 6.84 (d, J = 7.3
Hz, 1H) ppm. 13C NMR (100 MHz, CDCl3): δ = 18.3 (CH2), 22.0
(CH2), 26.5 (CH2), 27.5 (CH2), 31.6 (CH2), 43.7 (CH2), 48.0
(CH–N), 48.4 (CH2), 50.2 (CH2), 116.4 (CH), 118.9 (Cq), 122.4
(Cq), 125.7 (CH), 128.5 (CH), 143.9 (Cq), 175.4 (Cq, C=O) ppm.
The NMR data matches with the previously reported data.5f
5.1.3.23.
(±)-1,7-Di(2-oxopyrrolidin-1-yl)julolidine
17.
Following the general procedure, the reaction between p-
toluidine 16 (150 mg, 1.40 mmol), polyformaldehyde (105 mg,
3.50 mmol), N-vinyl-2-pyrrolidinone 9a (329 µL, 3.08 mmol)
and acetic acid (0.2 mL, 3.50 mmol) afforded compound 17 as a
yellow oil in a 1.5:1 mixture of diastereomers after silica gel
purification. Yield: 61% (301 mg). FTIR (film): ν = 2936, 2865,
5.1.3.21.
(±)-1-(7,8,12,13-Tetrahydro-6H-
14.
1
benzo[6,7]azepino[3,2,1-ij]quinolin-8-yl)pyrrolidin-2-one
1687 (C=O), 1601 cm-1. H NMR (400 MHz, CDCl3): Majority
Following the general procedure, the reaction between 10,11-
dihydro-5H-dibenzo[b,f]azepine 8p (133 mg, 0.68 mmol),
polyformaldehyde (31 mg, 1.03 mmol) and N-vinyl-2-
pyrrolidinone 9a (80 µL, 0.75 mmol) afforded compound 14 as a
white solid. Yield: 91% (197 mg). M.p. 158–159 °C. FTIR
isomer δ = 1.90–2.09 (m, 8H), 2.11 (s, 3H), 2.43–2.48 (m, 4H),
2.99–3.27 (m, 8H), 5.28 (t, J = 6.5 Hz, 2H, CH–N), 6.57 (s, 2H)
ppm; Minority isomer δ = 1.90–2.09 (m, 8H), 2.10 (s, 3H), 2.43–
2.48 (m, 4H), 2.99–3.27 (m, 8H), 5.37 (dd, J = 6.8, 10.0 Hz, 2H,
CH–N), 6.53 (s, 2H) ppm. 13C NMR (100 MHz, CDCl3):
Majority isomer δ = 18.3 (CH2), 20.5 (CH3), 27.2 (CH2), 31.5
(CH2), 44.3 (CH2), 47.3 (CH–N), 48.1 (CH2), 119.6 (Cq), 126.3
(Cq), 128.2 (CH), 142.9 (Cq), 175.3 (Cq, C=O) ppm; Minority
isomer δ = 18.2 (CH2), 20.5 (CH3), 26.2 (CH2), 31.5 (CH2), 43.1
(CH2), 47.9 (CH–N), 49.0 (CH2), 120.2 (Cq), 126.7 (Cq), 127.4
(CH), 142.3 (Cq), 175.6 (Cq, C=O) ppm. MS (ESI+): m/z 354 [M
+ H]+, 376 [M + Na]+, 392 [M + K]+. Anal. Calcd for
C21H27N3O2: C, 71.36; H, 7.70; N, 11.89. Found: C, 71.14; H,
7.58; N, 11.77.
1
(KBr): ν = 2960, 2875, 1686 (C=O), 1585 cm-1. H NMR (400
MHz, CDCl3): δ = 1.88–2.06 (m, 2H), 2.21–2.30 (m, 2H), 2.49 (t,
J = 8.0 Hz, 2H), 2.98–3.07 (m, 3H), 3.19–3.27 (m, 3H), 3.74–
3.86 (m, 2H), 5.58 (t, J = 8.3 Hz, 1H, CH–N), 6.80 (t, J = 7.5 Hz,
1H), 6.88 (d, J = 7.0 Hz, 1H), 6.92–6.98 (m, 2H), 7.10 (d, J = 7.3
Hz, 1H), 7.13–7.19 (m, 2H) ppm. 13C NMR (100 MHz, CDCl3):
δ = 18.2 (CH2), 26.2 (CH2), 31.5 (CH2), 32.7 (CH2), 34.0 (CH2),
43.2 (CH2), 48.5 (CH–N), 49.1 (CH2), 119.2 (CH), 121.0 (CH),
122.6 (CH), 125.7 (Cq), 126.2 (CH), 126.8 (CH), 129.5 (CH),
129.8 (CH), 131.8 (Cq), 135.2 (Cq), 145.1 (Cq), 148.4 (Cq),
175.5 (Cq, C=O) ppm. Anal. Calcd for C21H22N2O: C, 79.21; H,
6.96; N, 8.80. Found: C, 79.38; H, 7.10; N, 9.01. Crystal data for
14 were deposited at CCDC with reference CCDC 1556488:
Chemical formula C21H22N2O, Mr 318.41, Monoclinic, P21/c, 123
K, cell dimensions a, b, c, (Å) 8.4754(3), 7.8605(3), 24.5290(10);
α, β, γ (º) 90, 95.483(4), 90. V (Å3) 1626.67(11), Z = 4, F(000) =
680, Dx (Mg m-3) = 1.300, Mo Kα, ꢁ (mm-1) = 0.080, white
prism crystal, size (mm) = 0.35 x 0.32 x 0.12. Data collection:
CrysAlis PRO.14 Oxford Diffraction Gemini S Diffractometer.
5.2. Biological studies
5.2.1. Cells cycle analysis
HCT116 cells were seeded (1 x 105) in six-well plates and
allow to grow overnight. The medium was replaced with
complete medium containing compound 5f at 2-10 µM for 3
days. After 24, 48 and 72 h of treatment control and treated cells
were harvested by trypsinization, washed with PBS, fixed with
70% ethanol in water for 12–24 h at 4 °C, and stained with
Radiation
source:
fine-focus
sealed
tube
Graphite

15
Propidium Iodide staining solution according to manufaturer’s
instruction (BD, PharmigenTM, PI, 50 µg/mL in PBS pH 7.4).
About 100.000 cells were recorded and analyzed by flow
cytometry (C6 Acuri, BD, USA). The distribution of cells
through cell cycle phases was performed using the analysis
platform FlowJo, LLC.
Malinauskas, T.; Stumbraite, J.; Gaidelis, V.; Jankauskas, V.
ACCEPTED MANUSCRIPT
Monatsh. Chem. 2006, 137, 1401–1409; (g) Agbo, S. I.; Hallas,
G.; Towns, A. D. Dyes Pigm. 2000, 47, 33–43; (h) Hallas, G.;
Zhai, K. Y. Dyes Pigm. 1996, 32, 187–192; (i) Pullmann, T.;
Engendahl, B.; Zhang, Z.; Hölscher, M.; Zanotti-Gerosa, A.;
Dyke, A.; Franció, G.; Leitner, W. Chem.—Eur. J. 2010, 16,
7517–7526; (j) Liu, W.-B.; He, H.; Dai, L.-X.; You, S.-L.
Synthesis. 2009, 12, 2076–2082.
5.2.2. Apoptosis assay
2. (a) Su, D.-S.; Lim, J.J.; Tinney, E.; Wan, B.-L.; Young, M.B.;
Anderson, K.D.; Rudd, D.; Munshi, V.; Bahnck, C.; Felock, P.J.;
Lu, M.; Lai, M.-T.; Touch, S.; Moyer, G.; DiStefano, D.J.; Flynn,
J.A.; Liang, Y.; Sanchez, R.; Rasad, S.; Yan, Y.; Perlow-
Poehnelt, R.; Torrent, M.; Miller, M.; Vacca, J.P.; Williams,
T.M.; Anthony, N.J. Bioorg. Med. Chem. Lett. 2009, 19, 5119–
5123; (b) Jarvest, R.L.; Armstrong, S.A.; Berge, J.M.; Brown, P.;
Elder, J.S.; Brown, M.J.; Copley, R.C.B.; Forrest, A.K.;
Hamprecht, D.W.; O’Hanlon, P.J.; Rittenhouse, S.; Witty, D.R.
Bioorg. Med. Chem. Lett. 2004, 14, 3937–3941; (c) Suvire, F.D.;
Sortino, M.; Kouznetzov, V.V.; Vargas, L.Y.; Zacchino, S.A.;
Cruz, U.M.; Enriz, R.D. Bioorg. Med. Chem. 2006, 14, 1851–
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Chem. 2010, 45, 3245–3264; (e) Pagliero, R.J.; Lusvarghi, S.;
Pierini, A.B.; Brun, R.; Mazzieri, M.R. Bioorg. Med. Chem.
2010, 18, 142–150; (f) Oshiro, Y.; Sakurai, Y.; Sato, S.;
Kurahashi, N.; Tanaka, T.; Kikuchi, T.; Tottori, K.; Uwahodo,
Y.; Miwa, T.; Nishi, T. J. Med. Chem. 2000, 43, 177–189; (g)
Gouault, N.; Martin-Chouly, C.A.E.; Lugnier, C.; Cupif, J.-F.;
Tonnelier, A.; Feger, F.; Lagente, V.; David, M. J. Pharm.
Pharmacol. 2004, 56, 1029–1037.
3. (a) Liou, J.-P.; Wu, Z.-Y.; Kuo, C.-C.; Chang, C.-Y.; Lu, P.-
Y.; Chen, C.-M.; Hsieh, H.-P.; Chang, J.-Y. J. Med. Chem. 2008,
51, 4351–4355; (b) Liu, H.; Dagousset, G.; Masson, G.;
Retailleau, P.; Zhu, J. J. Am. Chem. Soc. 2009, 131, 4598–4599;
(c) Hiessböck, R.; Wolf, C.; Richter, E.; Hitzler, M.; Chiba, P.;
Kratzel, M.; Ecker, G. J. Med. Chem. 1999, 42, 1921–1926.
4. (a) Ishikawa, T.; Manabe, S.; Aikawa, T.; Kudo, T.; Saito, S.
Org. Lett. 2004, 6, 2361–2364; (b) Kouznetsov, V.V.; Zubkov,
F.I.; Nikitina, E.V.; Duarte, L.D.A. Tetrahedron Lett. 2004, 45,
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Duarte, L.D.A. Eur. J. Org. Chem. 2004, 24, 5064–5074; (d)
Lautens, M.; Tayama, E.; Herse, C. J. Am. Chem. Soc. 2005, 127,
72–73; (e) Di Fabio, R.; Alvaro, G.; Bertani, B.; Donati, D.;
Giacobbe, S.; Marchioro, C.; Palma, C.; Lynn, S.M. J. Org.
Chem. 2002, 67, 7319–7328; (f) Petit, M.; Geib, S.J.; Curran,
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Barvainiene, B.; Stanisauskaite, A.; Getautis, B. Chem.
Heterocycl. Compd. 2006, 42, 123–124; (k) Degutyte, R.;
Stumbraite, J.; Getautis, V. ARKIVOC. 2009, vii, 26–32.
HCT116 cells were seeded (1 x 105) in six-well plates and
allowed to grow overnight. The medium was then replaced with
complete medium containing compound 5f at 2-10µM. After 24,
48, and 72 h of treatment, control and treated cells were
harvested by trypsinization, washed with PBS and stained with
FITC Annexin V solution (BD, PharmigenTM) and Propidium
Iodide staining solution (BD, PharmigenTM, PI, 50 µg/mL in PBS
pH 7.4) according to manufacturer’s instruction. About 100,000
cells were recorded and analyzed by flow cytometry (C6 Acuri,
BD, USA).
5.2.3. Cell proliferation assay
HCT116 cells (1 x 105) were seeded in six-well plates and
allowed to grow overnight. The medium was then replaced with
complete medium containing compound 5f at 2-10µM. After 24,
48, and 72 h of treatment, control, and treated cells, were
harvested by trypsinization, washed with PBS and stained with
10µM of Carboxyflurescein Diacetate Succidimyl Ester (CFSE in
PBS) according to manufacturer’s instruction. A small aliquot of
cells was analyzed immediately after staining by flow cytometry
as intact cell (no fixation). Data was analyzed by using the
software analysis FlowJo LLC. Proliferative index is the total
number of divisions divided by the number of cells that went into
division.
Acknowledgements
The authors wish to credit the Developmental Therapeutics
Program (DTP) of the National Cancer Institute of United States
for performing the screening of the selected compounds. Authors
thank COLCIENCIAS, Universidad del Valle-Project No CI-
7812, Chemistry Department and the Faculty of Science of the
Universidad de los Andes for financial support, and Universidad
de Boyacá. E.J. also acknowledge Dr. Weihong Tan (Department
of Chemistry, University of Florida).
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http:
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