N-substituted noscapine derivatives as new antiprotozoal agents: Synthesis, antiparasitic activity and molecular docking study

Article abstract of DOI:10.1016/j.bioorg.2019.103116

Novel N-substituted noscapine derivatives were synthesized by a three-component Strecker reaction of cyclic ether of N-nornoscapine with varied aldehydes, in the presence of cyanide ion. Moreover, the corresponding amides were synthesized by the oxidation of cyanide moieties in good yields. The in vitro antiprotozoal activity of the products was also investigated. Interestingly, some analogues did put on display promising antiparasitic activity against Trypanosoma brucei rhodesiense with IC₅₀ values between 2.5 and 10.0 μM and selectivity index (SI) ranged from 0.8 to 13.2. Eight compounds exhibited activity against Plasmodium falciparum K1 strain with IC₅₀ ranging 1.7–6.4 μM, and SI values between 2.8 and 10.5 against L6 rat myoblast cell lines. Molecular docking was carried out on trypanothione reductase (TbTR, PDB ID: 2WOW) and UDP-galactose 4′ epimerase (TbUDPGE PDB: 1GY8) as targets for studying the envisaged mechanism of action. Compounds 6j₂ and 6b₂ displayed excellent docking scores with ?8.59 and ?8.86 kcal/mol for TbTR and TbUDPGE, respectively.

Full text of DOI:10.1016/j.bioorg.2019.103116

Accepted Manuscript
N-Substituted noscapine derivatives as new antiprotozoal agents: synthesis, an-
tiparasitic activity and molecular docking study
Kosar Babanezhad Harikandei, Peyman Salehi, Samad Nejad Ebrahimi,
Morteza Bararjanian, Marcel Kaiser, Hamid Reza Khavasi, Ahmed Al-Harrasi
PII:
S0045-2068(19)30058-6
https://doi.org/10.1016/j.bioorg.2019.103116
103116
DOI:
Article Number:
Reference:
YBIOO 103116
Bioorganic Chemistry
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Revised Date:
Accepted Date:
12 January 2019
3 July 2019
10 July 2019
Please cite this article as: K. Babanezhad Harikandei, P. Salehi, S. Nejad Ebrahimi, M. Bararjanian, M. Kaiser, H.
Reza Khavasi, A. Al-Harrasi, N-Substituted noscapine derivatives as new antiprotozoal agents: synthesis,
antiparasitic activity and molecular docking study, Bioorganic Chemistry (2019), doi: https://doi.org/10.1016/
j.bioorg.2019.103116
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N-Substituted noscapine derivatives as new antiprotozoal agents: synthesis,
antiparasitic activity and molecular docking study
Kosar Babanezhad Harikandeia, Peyman Salehia*, Samad Nejad Ebrahimia, Morteza Bararjanian a,
Marcel Kaiser b,c, Hamid Reza Khavasid, Ahmed Al-Harrasie
a Department of Phytochemistry, Medicinal Plants and Drugs Research Institute, Shahid Beheshti
University, G. C., Evin, 1983963113 Tehran, Iran
b Swiss Tropical and Public Health Institute, Basel, Switzerland
c University of Basel, Basel, Switzerland
d Department of Chemistry, Shahid Beheshti University, G. C., Evin, 1983963113 Tehran, Iran
e National and Medical Sciences Research Center, University of Nizwa, P.O. Box 33, Birkat Al-Mauz,
Nizwa 611, Oman
*Corresponding author
Lead contact: Prof. Peyman Salehi, Tel.: +98 21 29904049, fax: +98 21 22431783, E-mail: p-
salehi@sbu.ac.ir. Department of Phytochemistry, Medicinal Plants and Drugs Research Institute,
Shahid Beheshti University, G. C., Evin, 1983963113 Tehran, Iran.
1

Abstract
Novel N-substituted noscapine derivatives were synthesized by a three-component Strecker reaction
of cyclic ether of N-nornoscapine with varied aldehydes, in the presence of cyanide ion. Moreover,
the corresponding amides were synthesized by the oxidation of cyanide moieties in good yields. The
in vitro antiprotozoal activity of the products was also investigated. Interestingly, some analogues did
put on display promising antiparasitic activity against Trypanosoma brucei rhodesiense with IC50
values between 2.5-10.0 µM and selectivity index (SI) ranged from 0.8 to 13.2. Eight compounds
exhibited activity against Plasmodium falciparum K1 strain with IC50 ranging 1.7-6.4 µM, and SI
values between 2.8 to 10.5 against L6 rat myoblast cell lines. Molecular docking was carried out on
trypanothione reductase (TbTR, PDB ID: 2WOW) and UDP-galactose 4' epimerase (TbUDPGE
PDB: 1GY8) as targets for studying the envisaged mechanism of action. Compounds 6j2 and 6b2
displayed excellent docking scores with -8.59 and -8.86 kcal/mol for TbTR and TbUDPGE,
respectively.
Keywords: Noscapine; Strecker reaction; Trypanosoma brucei; Plasmodium falciparum; Molecular
docking; Induced fit docking.
2

1. Introduction
Currently, millions of people worldwide suffer from protozoal diseases such as human African
trypanosomiasis, malaria, Chagas disease, and leishmaniasis caused by single-celled parasites. These
illnesses occur in developing regions of Asia, Africa and Latin America [1]. Human African
Trypanosomiasis (HAT) or sleeping sickness is caused by one of the two subspecies of Trypanosoma
brucei (Trypanosoma brucei gambiense and T. brucei rhodesiense) transmitted via tsetse fly [2,3]. T.
b. rhodesiense infection is found in East Africa and ignites an acute infection whereas T. b.
gambiense is endemic in the West and central Africa and present a chronic type of infection. HTA is
deadly if not treated in a timely manner [4,5]. Malaria is the first responsible for the deaths caused by
parasitic diseases by which over 219 million infections and 435000 deaths were reported in 2017
[6,7]. Plasmodium falciparum is the most widespread one among six Plasmodium species (infecting
the man) and nearly amongst all malaria-related deaths reasons [7,8]. Leishmaniasis is the second
widespread parasitic disease, that is caused by Leishmania species [10,11]. WHO (World Health
Organization) reports appraised that there are 12 million cases infected with Leishmania species
worldwide, and over 2 million people get the infection each year [12,13]. WHO report in 2014
revealed that 72% of Iran’s population live in the endemic areas of Leishmania [14]. Trypanosoma
cruzi is responsible for Chagas diseases or American trypanosomiasis, which is found in tropical and
subtropical countries in Latin America [15,16]. Due to the lack of vaccination against parasitic
infections, low efficacy, significant side effects, alongside with the high costs of current treatments,
plus emerging resistance to them, development of new chemotherapeutic agents are indispensable
[17,18]. Natural products and their semisynthetic derivatives serve an essential role in discovering
and expansion of new antiprotozoal drugs namely artemisinin and artesunate [19–21]
Noscapine 1 is a phthalideisoquinoline alkaloid possessing the second highest abundance in opium
which has been isolated from Papaver somniferum. It is an oral nontoxic cough suppressant
prescribed since 1960 [22]. In the last decade, noscapine has attracted the attention of many research
groups for its anticancer capabilities[23–25].
Opium was in use as an antimalarial drug for many years in the late 19th century [26]. In the last
decades, there have been several reports regarding the investigation of alkaloids as antiprotozoal
agents. [18,27,28]. As an instance, quinine and its derivatives as a class of quinoline alkaloids have
an deciding role in the development of new antimalarial medicines [29]. Having considered such
3

reports, we decide to use noscapine (an isoquinoline alkaloid) as the starting material for the
synthesis of new antiparasitic agents.
In the present paper, we report the synthesis of two series of novel N-substituted cyclic ether
derivatives of N-nornoscapine and their antiprotozoal activities against T. b. rhodesiense, T. cruzi, L.
donovani, and P. falciparum. Furthermore, the cytotoxicity of the products was investigated against
L6 cells (rat skeletal myoblasts) to determine their selectivity index. The probable mechanism of
action against T. b. rhodesiense was studied by molecular docking of the most active compounds in
the binding site of two proteins, trypanothione reductase (TbTR) and uridine diphosphate galactose 4ʹ
epimerase (TbUDPGE) by the Glide module (Schrodinger suite).
2. Results and discussion
2.1. Chemistry
To furnish the desired N-substituted cyclic ether N-nornoscapine analogues, at first commercially
available noscapine 1 was converted to N-nornoscapine 2 by N-demethylation, and then the lactone
ring of compound 2 was reduced to the corresponding cyclic ether 3 (Scheme 1) [30,31]. Secondly,
the α-aminonitrile derivatives were prepared employing the three-component Strecker reaction in the
presence of 3 as the amine source, different aldehydes 4a-4o (Fig. 1) and potassium cyanide [32]. A
series of aldehydes including aliphatic (4m and 4o), aromatic with electron withdrawing (4e, 4d, and
4f) plus electron donating groups (4c, 4g, 4i, 4j, 4n, and 4l), as well as a heterocyclic model
compound (4h) were used.
4

O
O
O
O
O
O
H
H
H
H
H
H
N
O
NH
NH
O
1. H2O2, CH3CN
NaBH4, BF3.Et2O
THF
O
O
O
2. FeSO4·7H2O
CH3OH
O
O
O
O
O
O
O
O
O
2
1
3
O
O
H
H
NH
H
O
N
R
O
CH3COOH
O
+ RCHO + KCN
O
CN
O
O
H
O
O
O
O
5a-5o
3
4a-4o
O
O
O
NH2
O
H
H
H
N
O
N
O
R
H2O2, K2CO3
O
CH3OH, DMSO
O
R
O
CN
H
O
O
O
O
6a-6l
5a-5l
Scheme 1. Synthesis of N-substituted N-nornoscapine derivatives with reduced lactone ring.
H
O
H
O
H
O
H
O
H
O
H
O
N
H
O
H
O
Cl
O
Cl
O
O
O
Cl
O
F
O
4a
4b
4c
4d
4e
4f
4g
4h
H
O
H
H
O
H
O
H
O
H
OH
O
H
O
O
4i
4j
4k
4l
4m
4n
4o
Fig. 1. Series of aromatic and aliphatic aldehydes used in the Strecker reaction.
All of the synthesized compounds 5a-5o were obtained as a mixture of two diastereomers whose ratio
1
was analyzed by H-NMR. Some derivatives were purified by crystallization, from which a single
diastereomer was obtained (Table 1). As can be seen, the majority of reactions progressed smoothly
with high diastereoselectivity, except for two ortho-chloro substituted model compounds.
5

Throughout the next step, the synthesis of amide derivatives 6a-6l was considered in the presence of
various reagents (KOH, NaOH [33], K2CO3 [34], TiCl4 [35], HCl [36], and H2SO4 [37]) along with
different solvents (MeOH, DMF, DMSO, THF, EtOH, and hexane). The best results were obtained
when the α-aminonitrile derivatives 5a-5l were treated with hydrogen peroxide and potassium
carbonate [34]. The two diastereomers of derivatives were separated and purified taking advantage of
preparative TLC (Table 1).
1
13
Structures of all synthetic compounds were elucidated by the use of H-NMR, C-NMR, FTIR, and
HRMS.
Table 1
Synthesis of compounds 5 and 6.
Compound 5
Compound 6
Aldehyde
dra
Product Time (h) Yield%
Product Time (h) Yield%
b
4a
4b
4c
4d
4e
4f
5a1
5b
5c
1.0
0.5
5.0
1.0
1.0
1.0
6.0
0.5
0.5
8.0
24.0
8.0
2.0
2.0
2.0
71
69
79
50
95
80
65
85
75
75
65
60
75
71
65
-
6a
6b
6c
5.0
12.0
24.0
24.0
8.0
65
75
77
55
90
66
58
65
44
65
54
50
88:12
84:16
66:34
61:39
81:19
91:9
5d
5e
6d
6e
5f
6f
24.0
12.0
8.0
4g
4h
4i
5g
5h
5i1
5j
6g
6h
6i1
6j
83:17
b
-
48.0
8.0
4j
84:16
84:16
86:14
80:20
4k
4l
5k
5l
6k1
6l1
2.0
24.0
4m
4n
4o
5m
5n1
5o
b
-
85:15
adr: Diastereomeric ratio (calculated by 1H-NMR), bMajor diastereomer precipitated spontaneously.
The absolute configuration of chiral center generated in the Strecker reaction of compound 5f1 was
assigned by single-crystal X-ray diffraction. The single crystal of 5f1 was prepared by dissolving it in
chloroform followed by slow evaporation of the solvent at room temperature. The ORTEP view (Fig.
2) showed that the configuration of the chiral center at C-1″ was R. Thus, 5f1 as the major
6

diastereomer possesses 3S,5'R,1″R configuration (The X-ray analysis details are reported in
supplementary data).
A
B
6"
3"
1'
9'
8'
F
5"
O
7'
7"
2"
2'
H
N6'
4"
(R)
1"
O
4'
5' (R)
3'
OMe
CN
3 (S)
4
8"
O 2
H
5
1
6
7
MeO
OMe
Fig. 2. A) ORTEP view and B) 2D structure of compound 5f1.
2.2. Antiparasitic activity
All synthesized compounds went through an investigation for growth inhibition against four
parasites; namely, Trypanosoma brucei rhodesiense strain STIB900, T. cruzi strain Tulahuen C4,
Leishmania donovani axenic amastigotes strain MHOM-ET-67/L82 and Plasmodium falciparum
strain NF54 at two concentrations of 2 and 10 µg/mL. Melarsoprol, benznidazole, miltefosine, and
artemisinin were used as positive controls respectively for T. b. rhodesiense, T. cruzi, L. donovani,
and P. falciparum (The results are reported in Table S2 in supplementary data). Twenty-eight
derivatives exhibited a high percentage of growth inhibition (higher than 50%) for T. b. rhodesiense
protozoa. Only 6g2 showed activity against T. cruzi with 60% growth inhibition at 2 µg/mL. The
most potent synthetic compound against L. donovani was 6b1 with 100% growth inhibition at 2
µg/mL.
The N-α-aminonitrile derivatives 5a1 and 5m and the amide derivatives 6b1, 6b2,6d2, 6e1, 6e2, 6g2,
6i1, and 6k1 presented higher percentage of growth inhibition than other analogues for P. falciparum.
It should be mentioned that N-nornoscapine 2 displayed poor antiprotozoal activities. Derivatives
with the growth inhibition higher than 50% for two parasites T. b. rhodesiense and P. falciparum at 2
µg/mL were opted for determination of the IC50 values. Also, the cytotoxic activity of these
analogues was evaluated against the rat myoblast L6 cells. Selectivity indices (SIs) were determined
employing the IC50 values of cytotoxicity and antiprotozoal activity. The results are presented in
Table 2. Fortunately, most of the examined products showed weak cytotoxicity.
7

Table 2
In vitro antiprotozoal and cytotoxic activities and selectivity index of selected derivatives.
T. b. rhodesiense
IC50 a(µM)
-
P. falciparum
IC50 (µM)
14.9 ± 0.0
L6 cells
IC50 (µM)
11.2 ± 5.2
91.8 ± 8.7
20.7 ±7.9
16.8 ± 2.2
29.5 ± 2.6
62.3 ± 29.7
50.1 ± 11.4
13.9 ± 1.8
20.2 ± 6.3
70.6 ± 5.2
40.5 ± 9.3
93.7 ± 3.3
31.6 ± 7.7
24.2 ± 11.3
25.6 ± 10.2
32.2 ± 2.5
25.5 ± 3.5
10.7 ± 0.3
17.2 ± 4.8
24.9 ± 2.6
85.5 ± 2.8
18.5 ± 6.7
20.4 ± 7.8
17.9 ± 9.1
13.4 ± 1.9
39.0 ± 2.2
17.1 ± 4.9
76.3 ± 6.1
7.8 ± 3.2
Compound
SIb
-
SI
0.8
-
-
-
-
-
-
-
-
-
-
-
-
-
1.4
-
-
5a1
5b1
5c
5d
5e
5f1
5f2
5g
5h
5i1
5j
5k
5l
5l1
5m
5n1
5o
6a2
6b1
6b2
6c1
6c2
6d2
6e1
6e2
6f1
6f2
6g1
6g2
6h2
6i1
6j1
6k1
6l1
17.6 ± 7.1
8.9 ± 1.8
9.8 ± 1.0
23.6 ± 0.1
10.2 ± 0.9
27.7 ± 5.7
8.8 ± 1.1
10.6 ± 1.3
29.6 ± 2.0
13.5 ± 4.6
13.0 ± 3.4
18.5 ± 8.5
11.6 ± 0.6
-
2.5 ± 1.2
12.6 ± 0.6
33.1 ± 5.9
15.3 ± 6.6
19.7 ± 5.4
6.5 ± 2.6
29.7 ± 6.4
-
5.2
2.3
1.7
1.2
6.1
1.8
1.6
1.9
2.3
3.0
7.2
1.7
2.1
-
12.9
2.0
0.3
1.1
1.3
13.2
0.6
-
-
-
-
-
-
-
-
-
-
-
-
-
-
19.0 ± 0.4
-
-
-
-
4.1 ± 0.8
3.5 ± 0.0
4.2
7.1
-
-
-
-
4.7 ± 1.1
6.4 ± 1.8
2.4 ± 0.5
4.3
2.8
5.5
-
-
-
4.6
-
6.3
-
10.5
-
-
-
-
-
-
26.5 ± 3.1
32.6 ± 4.1
25.6 ± 5.2
10.0 ± 0.6
53.4 ± 14.2
-
9.2 ± 1.1
45.2 ± 19.3
42.9 ± 16.1
0.0075c
1.5
0.5
3.0
0.8
0.2
-
3.2
0.8
0.7
-
-
-
-
1.7 ± 0.2
-
11.5 ± 0.5
37.4 ± 1.7
29.3 ± 8.3
36.9 ± 2.0
31.2 ± 3.4
0.012e
5.9 ± 0.6
-
3.5 ± 0.1
-
Positive control
0.012d
aIC50: 50% inhibitory concentration, bSelectivity Index (IC50 value of cytotoxicity/ IC50 value of antiprotozoal
activity), cMelarsoprol, dChloroquine, ePodophyllotoxin used as positive controls.
Among twenty-eight tested compounds against T. b. rhodesiense, seven compounds showed good
activities with IC50 values in the range of 2.5-10.0 µM with the selectivity index in the range of 0.8-
13.2. The most active compounds were 5n1 (IC50 = 2.5 µM, SI = 12.9) and 6c1 (IC50 = 6.5 µM, SI =
13.2) with nitrile and amide function, respectively. Derivatives with hydroxyl, dimethoxy, and
benzyloxy moieties on the phenyl ring (5n1, 5c, 5g, 6c1, 6g2, and 6j1) were the most potent analogues.
Moreover, the monohalogenated α-aminonitrile derivatives (5d, 5f1) displayed better activities
8

compared
to
other
compounds.
In
general,
the
nitrile
selectivity than the amide bearing compounds.
All derivatives were tested against P. falciparum. From which the amide derivatives 6b1, 6b2, 6d1,
6e1, 6e2, 6g2, 6i1, and 6k1, displayed high activity with IC50 values between 1.7 – 6.4 µM and
selectivity index in the range of 2.8 to 10.5. Compound 6g2 showed the lowest IC50 value equal to 1.7
µM and SI= 4.6. The highest selectivity index (SI=10.5) was observed for compound 6k1 (IC50 = 3.5
µM). In general amide derivatives were more active than their α-aminonitrile parents for P.
falciparum.
2.3. Docking studies
For more investigation on the mode of interaction between the synthetic compounds and T. b.
rhodesiense and the possible mechanism of action, molecular docking studies of all derivatives was
performed in the flavin adenine dinucleotide (FAD) binding site of trypanothione reductase (TbTRR)
(PDB ID: 2WOW) [38] and nicotinamide adenine dinucleotide (NAD) binding site of UDP-galactose
4ʹ epimerase (TbUDPGE) (PDB:1GY8) using Glide application in Maestro 10.2 platform
(Schrödinger, LLC )[39]. The TbTR has a vital and unique role in maintaining redox balance in these
parasites and is an attractive target for drug discovery studies [38]. The in silico investigation of
antitrypanosomal natural compounds by Setzer et al. has shown that TbUDPGE is an appropriate
target for investigation of the mechanism of action in indole alkaloids [39].
The docking scores of the compounds with the best IC50 values are listed in Table 3 (details were
reported in Table S3 in supplementary data). As shown in results, all derivatives illustrated great
affinity toward the binding site of the two target proteins chiefly TbUDPGE by high docking scores
(-5.29 to -8.86 kcal/mol). Compound 6j2 showed the most significant docking score (-8.59 kcal/mol)
for the ligand-TbTR protein complex (Fig. 3A). This ligand showed two hydrogen bonding
interactions between MET333 and THR335 and the ligand amide side chain. Also, the interaction
between 4-(benzyloxy) phenyl moiety with hydrophobic pocket include ILE438, ALA 363, PRO 435,
PHE367, and ALA 364, was revealed.
Then induced fit docking (IFD) was used to study the insight of the interaction between the ligands
5j2, 6j2, and 6k2 and protein in flexible mode (Fig. 3B). The IFD scores -8.01, -9.64, and -8.00
kcal/mol were obtained for these ligands, respectively. The hydrogen bond interactions in ARG287,
9

CYS52, and THR335 were observed, and Pi-Pi staking in PHE198 was revealed in the 6j2-TbTR
binding site complex. The phenyl ring was involved in the Pi-cation interaction with LYS60. The
ligand 6j2 indicated hydrophobic interactions with CYS57, PHE367, ALA365, ALA363, VAL337,
LEU334, PRO336, and CYS52 whose residues are present in the FAD binding site. Furthermore, a
polar pocket containing GLY56, SER178, ASN179, THR51, and ARG287 interacted with
dioxolo[4,5-g]isoquinoline moiety. All the mentioned interactions, especially the hydrophobic ones,
were likely to be responsible for the IFD score improving (-9.64 kcal/mol).
Table 3
Docking scores (kcal/mol) of the synthesized and reference compounds at FAD
and NAD cavity for TbTR and TbUDPGE, respectively.
Compounda
Docking score (kcal/mol)
TbTR
TbUDPGE
-7.06
-6.59
-8.24
-6.91
-6.94
-7.68
-6.8
5a1
5a2
5b2
5c1
5c2
5d1
5d2
5f2
-5.94
-5.91
-6.71
-6.54
-5.91
-5.61
-6.41
-6.08
-6.06
-5.25
-7.15
-6.72
-6.37
-7.66
-7.18
-6.18
-7.02
-6.65
-6.02
-7.18
-7.52
-7.1
-6.75
-6.96
-7.15
-6.81
-6.8
5g1
5g2
5h1
5h2
5j1
-8.03
-6.69
-8.74
-6.91
-6.81
-6.91
-8.08
-8.86
-7.6
5j2
5k2
5l2
5n1
5n2
6b1
6b2
6c2
6g2
6j1
-6.79
-7.82
-8.65
-5.29
-17.34
-7.44
-8.59
-7.94
-13.82
6j2
6k2
FAD
NAD
aDiastereomers of derivatives were docked separately.
10

Fig. 3. The 2D and 3D docking of A) the complex of 6j2 with the binding site of TbTR; B) IFD of the
complex 6j2 with the binding site of TbTR; C) the complex 6b2 with the binding site of TbUDPGE.
The docking studies of the synthesized compounds in the FAD-binding site of TbTR exhibited that in
most cases the conversion of nitrile group to amide (i.e., 5b vs. 6b, 5c vs. 6c1, and 5j vs. 6j) and
addition of hydroxyl group (5n1) or heteroatom in aromatic ring (5h) caused additional hydrogen
11

bonding and greater docking score. Insertion of groups with a bulky side chain at C-7' position (5b2,
5j2, 5k2, 6b2, 6j2, and 6k2) afforded more hydrophobic interactions with the binding site. Comparison
between two diastereomers of the synthesized derivatives docking scores did not show a notable
variation. The docking scores of the ligands-TbTR complex were in good agreement with IC50 values.
Compound 6b2 showed the highest docking score (-8.86 kcal/mol) with the TbUDPGE enzyme (Fig
3C). This ligand revealed no hydrogen bond, Pi-Pi or Pi-cation interaction with the protein residues.
The primary interaction was among the 3-phenoxyphenyl group of the ligand 6b2 and hydrophobic
residues containing TYR173, ALA100, LEU102, MET98, ILE12, and ALA9. Interestingly these
residues also play an essential role in the binding of NAD as the native ligand by hydrophobic
interaction. Moreover, polar interactions of the benzofuran group with HID221, THR220, SER219,
THR37, HID215, HID43, and Ser14 and hydrophobic interactions of isoquinoline moiety with
PHE101, TYR114, and AlA103 were all revealed in the ligand-protein complex.
Molecular docking of the ligands with NAD binding site in TbUDPGE demonstrated that bulky
groups in the C-7' position of the ligands (5b2, 5j2, 5k2, 6b1, 6b2, and 6k2) and their orientation in the
binding site had a vital role in the formation of hydrophobic interactions.
Finally, for the evaluation of drug-likeness and molecular properties of all the active compounds, the
consistency with Lipinski’s rules of five was investigated through ADMET prediction by Qikprop
4.4. This rule recommends some criteria for molecular weight (MW<500), H acceptor bonds
(HA<10), H donor bonds (HD<5), and octanol/water partition coefficient (QPlogPo/w<5). The orally
active compounds mostly adhere to three of the four Lipinski’s rules. The results proved that the
synthesized derivatives were compatible with the Lipinski’s rules (Table 4).
12

Table 4
Calculated theoretical parameters of Lipinski's rules for the active compounds.
Compound mol_MW donorHB accptHB QPlogPo/w Rule Of Five(<4)
500.55
592.647
560.602
534.995
518.54
0
0
0
0
0
0
0
0
0
0
0
0
1
0
2
2
2
2
2
2
2
2
2
2
2
2
2
2
8.95
9.45
10.45
8.95
8.95
8.95
10.45
10.45
8.95
9.7
3.331
4.372
3.013
3.786
3.497
3.118
3.099
2.501
4.56
1
1
1
1
1
1
1
1
1
1
1
0
1
0
1
1
1
1
1
1
1
1
1
1
1
1
1
1
5a1
5b1
5c
5d
5f1
5f2
5g
5h
5i1
5j
518.54
560.602
501.538
542.63
606.674
514.577
466.533
516.549
452.506
518.565
610.662
610.662
610.662
553.01
587.455
587.455
536.556
536.556
519.553
560.646
624.689
606.674
532.592
4.95
8.95
8.95
9.7
3.374
2.621
2.805
2.507
2.539
3.493
4.099
3.493
2.938
3.283
3.103
2.579
2.715
1.649
2.995
3.613
3.57
5l
5m
5n1
5o
8.95
9.95
10.45
10.45
10.45
9.95
9.95
9.95
9.95
9.95
11.45
9.95
10.7
9.95
9.95
6a1
6a2
6b1
6b2
6d2
6e1
6e2
6f1
6f2
6h2
6i1
6j1
6k1
6l1
2.941
3. Conclusion
In this paper, the synthesis procedure of two novel classes of noscapine derivatives was described.
The antiparasitic activities of new noscapinoids were evaluated and for the first time antipasitic
activity from noscapinoids were reported. Most derivatives displayed promising activity against T. b.
rhodesiense and P. falciparum. Compound 5n1 showed high potency and selectivity for T. b.
rhodesiense with IC50= 2.5μM and SI=13. Compound 6g2 was identified as the most active against P.
falciparum with IC50= 1.7μM but had toxicity. Molecular docking studies for the most active
compounds against T. b. rhodesiense were performed on two proteins TbTR and TbUDPGE, and the
results were in good agreement with the experimental data showing good affinity of the synthesized
compounds to the binding site of the target receptors. As we assumed, the noscapine derivatives
13

showed plausible antiparasitic activity but we need more manipulation for obtaining better
selectivities. Therefore, these novel semi-synthetic analogues could be explored as the new lead
compounds for protozoal diseases, and deserve more investigation for in vivo studies.
4. Experimental
4.1. General
Noscapine was purchased from Faran Shimi Pharmaceutical Co. Miltefosine, artemisinin,
chloroquine, and podophyllotoxin were purchased from Sigma, and benznidazole and melarsoprol
were received from WHO (with purity >95% according to the suppliers). Other chemicals and
solvents were provided from Merck, Sigma-Aldrich, and Kimia Exir chemical companies without
further purification. Pre-coated silica gel F254 sheets for TLC from Merck Co. were used for reaction
monitoring. Products were purified by preparative TLC plates coated with silica gel 60 PF254
containing gypsum. Melting points were measured using a Branstead/Electrothermal 9200 apparatus
1
and were uncorrected. HNMR and 13CNMR spectra were recorded in CDCl3 and DMSO-d6 as
solvents on Bruker Avance III spectrometers at 600, 500 and 300 MHz and TMS was employed as
internal standard. Chemical shifts were reported by ppm. HRESIMS data were acquired on a Bruker
micro TOF ESIMS in positive mode. FT-IR spectra were recorded on a Bruker Tensor 27
spectrometer.
4.1.2. N-Nornoscapine; (3S)-6,7-Dimethoxy-3-((5R)-4-methoxy-5,6,7,8 tetrahydro[1,3]dioxolo[4,5-
g] isoquinolin-5-yl)-2-benzofuran-1(3H)-one (2)
N-Nornoscapine was synthesized by the literature procedure with some modification [30]. 10.0g
noscapine (24.0 mmol) was added to a flask containing 200 mL H2O2 30% and 200 mL CH3CN. The
reaction mixture was stirred overnight at room temperature. When the reaction completed and
noscapine-N-oxide formed, the mixture was cooled to 0 °C and then solid MnO2 was slowly added to
remove excess amount of H2O2. The mixture was filtered and concentrated in vacuo. The reaction
mixture was acidified to pH 1with 1M HCl and extracted with CHCl3 (3×300). The organic layer was
removed under the reduced pressure and the noscapine-N-oxide·as hydrochloride salt was isolated.
13.34 g of FeSO4.7H2O (48.0 mmol) was added to a solution of noscapine-N-oxide·HCl in 750 mL
CH3OH at -8 °C and stirred. Having passed 8 hours, the TLC showed that the noscapine-N-oxide was
consumed to the full, thereafter the solvent was evaporated under reduced pressure. The residue was
14

dissolved in CHCl3 and then washed with a saturated solution of EDTA in 1M HCl. The organic layer
was washed with 1M NaOH and dried over Na2SO4. The crude product was recrystalized from
dichloromethane and hexane to give pure N-nornoscapine.
4.1.3.
(5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinoline (3)
Reduction of N-nornoscapine lactone moiety was performed according to previous reports [23,22].
1.0 g N-nornoscapine (2.5 mmol) was dissolved in 12 mL BF3.Et2O and then added dropwise to a
solution of 0.19 g NaBH4 (5.0 mmol) in 20 mL dry THF at -5°C. After 1h, the reaction mixture was
warmed to room temperature and stirred overnight. To quench the reaction, 10 mL of HCl (10%) was
added dropwise in an ice bath. After stirring for 1h, the reaction was extracted with CHCl3 (3×50)
and the organic layer was separated and dried over anhydrous Na2SO4. Concentration under reduced
pressure gave the free base of 3 which was purified by preparative TLC.
4.1.4. General Procedure for the Synthesis of N-α-Aminonitrile Derivatives
To a solution of compound 3 (1.0 mmol, 385.4 mg) in 5 mL acetic acid was added aldehyde (4) (1.1
mmol). After stirring for 30 min at room temperature, 2.0 mmol KCN was added to the reaction
mixture. After complete consumption of compound 3 approved by TLC, water was added to the
reaction mixture and extracted with CHCl3 (3×30 mL). The organic layer was separated and dried
with Na2SO4 and solvent was evaporated in vacuo. The crude product was purified by preparative
TLC or recrystallization by methanol and chloroform.
4.1.4.1.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(phenyl)acetonitrile (5a)
Aldehyde substrate: Benzaldehyde (4a), reaction time: 1h. The major diastereomer was crystalized
from methanol (5a1: major diastereomer). Yield: 71%; white crystal; m.p. 158 °C; IR (KBr): 1042,
1275, 1488, 1620, 2845, 2949 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.29 – 7.43 (m, 5H), 6.70
(d, J = 8.2 Hz, 1H), 6.36 (s, 1H), 6.02 (s, 1H), 5.98 (s, 2H), 5.88 (d, J = 8.2 Hz, 1H), 5.63 (s, 1H),
5.20 (dd, J = 12.5, 2.9 Hz, 1H), 5.10 (d, J = 12.5 Hz, 1H), 4.89 (d, J = 2.9 Hz, 1H), 4.12 (s, 3H), 3.88
(s, 3H), 3.87 (s, 3H), 2.56 (dt, J = 10.0, 3.7 Hz, 1H), 2.47 (td, J = 10.0, 3.7 Hz, 1H), 2.38 – 2.21 (m,
2H) ppm;13C NMR (75 MHz, chloroform-d) δ 151.2, 148.3, 142.6, 140.5, 134.8, 134.5, 133.0, 132.8,
15

132.4, 128.7, 128.4, 126.8, 117.7, 117.6, 117.5, 112.0, 102.2, 100.9, 88.1, 71.6, 63.1, 59.9, 59.5, 56.1,
43.6, 29.9 ppm; HRMS (ESI): [M+H]+ C29H29N2O6 calcd 501.1981, found 501.2043.
4.1.4.2. ((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(3-phenoxyphenyl) acetonitrile (5b)
Aldehyde substrate: 3-Phenoxybenzaldehyde (4b), reaction time: 0.5 h. The mixture of two
diastereomers (5b) (88:12) were purified by preparative TLC, yield: 69%.
1
5b:Off white powder; m.p. 139 °C; H NMR (300 MHz, chloroform-d) δ 7.44 – 6.86 (m, 18H,
mixture of two diastereomers), 6.67 (d, J = 8.2 Hz, 1H, minor diastereomer), 6.64 (d, J = 8.2 Hz, 1H,
major diastereomer), 6.36 (s, 1H, major diastereomer), 6.32 (s, 1H, minor diastereomer), 6.05 – 6.01
(m, 2H, minor diastereomer), 6.00 (s, 1H, major diastereomer), 5.97 (s, 2H, major diastereomer),
5.91 (brs, 2H, minor diastereomer), 5.85 (d, J = 8.2 Hz, 1H, major diastereomer), 5.60 (s, 1H, major
diastereomer), 5.41 (s, 1H, minor diastereomer), 5.33 (m, 4H, minor diastereomer), 5.18 (dd, J =
12.4, 2.9 Hz, 1H, major diastereomer), 5.08 (d, J = 12.4 Hz, 1H, major diastereomer), 4.87 (d, J = 3.0
Hz, 1H, major diastereomer), 4.74 (d, J = 3.8 Hz, 1H, minor diastereomer), 4.11 (s, 3H, major
diastereomer), 3.88 (s, 3H, minor diastereomer), 3.87 (s, 3H, minor diastereomer), 3.84 (s, 3H, minor
diastereomer), 3.83 (s, 3H, major diastereomer), 3.81 (s, 3H, major diastereomer), 2.64 – 2.19 (m,
8H, mixture of two diastereomers); HRMS (ESI): [M+H]+ C35H33N2O7 calcd 593.6560, found
593.2278.
5b1:Off white powder; m.p. 137 °C; IR (KBr): 1033, 1083, 1270, 1488, 1620, 2925 cm-1; 1H NMR
(600 MHz, chloroform-d) δ 7.36-7.33 (m, 2H), 7.27 (t, J = 7.9 Hz, 1H), 7.15 (d, J= 7.9Hz, 1H), 7.13
(t, J = 7.4 Hz, 1H), 7.09 (s, 1H), 6.97 (d, J = 8.0 Hz, 2H), 6.90 (d, J = 8.0 Hz, 1H), 6.64 (d, J = 8.2
Hz, 1H), 6.36 (s, 1H), 5.99 (s, 1H), 5.98 (s, 2H), 5.85 (d, J = 8.2 Hz, 1H), 5.61 (br s, 1H), 5.17 (dd, J
= 12.4, 2.6 Hz, 1H), 5.07 (d, J = 12.3 Hz, 1H), 4.86 (d, J = 2.7 Hz, 1H), 4.11 (s, 3H), 3.83 (s, 3H),
3.81 (s, 3H), 2.61 (dt, J = 10.0, 3.4 Hz, 1H), 2.50 (td, J = 10.4, 2.5 Hz, 1H), 2.32 (d, J = 15.2 Hz,
1H), 2.31-2.28 (m. 1H) ppm; 13C NMR (151 MHz, chloroform-d) δ 157.6, 156.7, 151.2, 148.3, 142.7,
140.5, 136.9, 134.5, 132.8, 132.7, 132.3, 129.9, 129.7, 123.4, 121.6, 118.7, 118.4, 117.7, 117.7,
117.4, 117.2, 112.1, 102.1, 100.8, 88.0, 71.6, 62.7, 60.6, 59.9, 59.5, 56.1, 43.7, 29.8 ppm; HRMS
(ESI): [M+H]+ C35H33N2O7 calcd 593.6560, found 593.2278.
4.1.4.3.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(3,4-dimethoxyphenyl)acetonitrile (5c)
16

Aldehyde substrate: 3, 4-Dimethoxybenzaldehyde (4c), reaction time: 5h. The mixture of two
diastereomers (84:16) were purified by preparative TLC. Yield: 79 %, white powder; m.p. 83 °C; IR
(KBr): 1030, 1082, 1269, 1619, 2932 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.07 (d, J = 8.4 Hz,
1H, minor diastereomer), 7.00 (d, J = 7.8 Hz, 1H, major diastereomer), 6.90 – 6.76 (m, 4H, mixture
of two diastereomers), 6.69 (d, J = 8.2 Hz, 2H, mixture of two diastereomers), 6.36 (s, 1H, major
diastereomer), 6.32 (s, 1H, minor diastereomer), 6.09 (d, J = 6.6 Hz, 1H, minor diastereomer), 5.98
(s, 4H, mixture of two diastereomers), 5.95 (s, 2H, mixture of two diastereomers), 5.91 (d, J = 7.9
Hz, 1H, major diastereomer), 5.64 (s, 1H, major diastereomer), 5.35 (s, 1H, minor diastereomer),
5.25 (dd, J = 12.6, 2.9 Hz, 2H, mixture of two diastereomers), 5.12 (d, J = 12.5 Hz, 2H, mixture of
two diastereomers), 4.87 (d, J = 2.8 Hz, 1H, major diastereomer), 4.77 (d, J = 3.5 Hz, 1H, minor
diastereomer), 4.12 (s, 3H, major diastereomer), 4.02 (s, 3H, minor diastereomer), 4.00 (s, 3H, minor
diastereomer), 3.98 (s, 3H, minor diastereomer), 3.90 (s, 6H, minor diastereomer), 3.88 (s, 3H, major
diastereomer), 3.85 (s, 3H, major diastereomer), 3.83 (s, 3H, major diastereomer), 3.73 (s, 3H, major
diastereomer), 2.70 – 2.27 (m, 8H, mixture of two diastereomer) ppm; 13C NMR (126 MHz,
chloroform-d) δ 151.1, 148.3, 142.6, 140.6, 133.0, 133.0, 132.4, 127.1, 119.2, 117.5, 117.5, 112.2,
112.1, 110.8, 110.7, 109.6, 109.5, 102.1, 100.8, 88.2, 88.1, 71.3, 62.5, 60.5, 59.8, 59.5, 56.1, 55.9,
55.6, 43.4, 30.0, 28.0 ppm; HRMS (ESI): [M+H]+ C31H33N2O8 calcd 561.6110, found 561.2258.
4.1.4.4.
(2-Chlorophenyl)((5R)-5-((1S)-4,5-dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-
5,6,7,8-tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)acetonitrile (5d)
Aldehyde substrate: 2-Chlorobenzaldehyde (4d), reaction time: 1h. The mixture of two diastereomers
(66:34) were purified by preparative TLC. Yield: 50 %, white powder; m.p. 92 °C; IR (KBr): 1037,
1
1269,1483, 1621, 2930 cm-1; H NMR (300 MHz, chloroform-d) δ 7.73 – 7.64 (m, 1H, major
diastereomer), 7.52 – 7.29 (m, 6H, mixture of two diastereomer), 7.26 – 7.14 (m, 1H, major
diastereomer), 6.68 (d, J = 8.2 Hz, 1H, minor diastereomer), 6.63 (d, J = 8.3 Hz, 1H, major
diastereomer), 6.39 (s, 1H, minor diastereomer), 6.36 (s, 1H, major diastereomer), 6.11 (d, J = 8.1
Hz, 1H, major diastereomer), 6.01 (d, J = 8.2 Hz, 1H, minor diastereomer), 5.95 (s, 2H, minor
diastereomer), 5.92 (s, 2H, major diastereomer), 5.80 (s, 1H, minor diastereomer), 5.55 (s, 1H, minor
diastereomer), 5.47 (s, 1H, major diastereomer), 5.39 (dd, J = 12.4, 2.9 Hz, 1H, minor diastereomer),
5.27 (s, 1H, major diastereomer), 5.15 (dd, J = 12.4, 2.8 Hz, 1H, major diastereomer), 4.95 (d, J =
12.0 Hz, 1H, major diastereomer), 4.92 (d, J = 12.0 Hz, 1H, minor diastereomer), 4.76 – 4.64 (m, 2H,
17

mixture of two diastereomers), 3.99 (s, 3H, minor diastereomer), 3.91 (s, 3H, major diastereomer),
3.84 (s, 3H, minor diastereomer), 3.83 (s, 6H, major diastereomer), 3.67 (s, 3H, minor diastereomer),
3.02 – 2.28 (m, 8H, mixture of two diastereomers) ppm; 13C NMR (126 MHz, chloroform-d) δ 151.1,
148.3, 148.2, 142.6, 140.6, 134.4, 134.3, 133.5, 133.0, 132.7, 132.5, 132.3, 131.6, 130.7, 130.2,
130.2, 129.8, 129.8, 129.3, 126.9, 126.9, 118.0, 117.5, 117.3, 117.3, 116.6, 111.9, 111.7, 103.0,
102.5, 102.3, 100.7, 100.6, 87.3, 86.3, 71.7, 71.4, 71.3, 61.0, 59.9, 59.7, 59.3, 59.2, 58.6, 57.6, 57.5,
56.2, 45.1, 44.4, 28.5, 26.9 ppm; HRMS (ESI): [M+H]+ C29H28ClN2O6 calcd 535.1636, found
535.2175.
4.1.4.5.
(2,4-Dichlorophenyl)((5R)-5-((1S)-4,5-dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-
methoxy-5,6,7,8-tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)acetonitrile (5e)
Aldehyde substrate: 2,4-Dichlorobenzaldehyde (4e), reaction time: 1h. The mixture of two
diastereomers (61:39) were purified by preparative TLC. Yield: 95 %, white powder; m.p. 144 °C; IR
(KBr): 1037, 1083, 1269,1483, 1621, 2930 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.59 (d, J =
8.3 Hz, 1H, minor diastereomer), 7.46 (dd, J = 6.9, 1.9 Hz, 1H, major diastereomer), 7.33 (dd, J =
8.3, 2.1 Hz, 1H, minor diastereomer), 7.26 – 7.17 (m, 3H, mixture of two diastereomer), 6.68 (d, J =
8.2 Hz, 1H, major diastereomer), 6.62 (d, J = 8.2 Hz, 1H, minor diastereomer), 6.39 (s, 1H, major
diastereomer), 6.36 (s, 1H, minor diastereomer), 6.09 (d, J = 8.2 Hz, 1H, minor diastereomer), 5.99
(d, J = 8.0 Hz, 1H, major diastereomer), 5.97 (d, J = 1.5 Hz, 1H, major diastereomer), 5.95 (d, J = 1.4
Hz, 1H, major diastereomer), 5.94 (d, J = 1.4 Hz, 1H, minor diastereomer), 5.93 (d, J = 1.5 Hz, 1H,
minor diastereomer), 5.74 (s, 1H, major diastereomer), 5.55 (s, 1H, major diastereomer), 5.40 (s, 1H,
minor diastereomer), 5.30 (s, 1H, minor diastereomer), 5.19 (dd, J = 12.1, 2.7 Hz, 1H, minor
diastereomer), 5.02 – 4.88 (m, 2H, mixture of two diastereomers), 4.71 (dd, J = 8.1, 2.9 Hz, 1H,
major diastereomer), 4.65 (s, 1H, major diastereomer), 4.64 (s, 1H, minor diastereomer), 4.01 (s, 3H,
major diastereomer), 3.95 (s, 3H, minor diastereomer), 3.85 (s, 3H, major diastereomr), 3.84 (s, 6H,
minor diastereomer), 3.70 (s, 3H, major diastereomer), 2.98 – 2.36 (m, 8H, mixture of two
13
diastereomers) ppm; C NMR (126 MHz, chloroform-d) δ 151.1, 148.4, 148.3, 140.5, 135.5, 135.1,
135.1, 134.3, 134.1, 133.2, 132.8, 132.7, 132.5, 131.3, 131.2, 131.0, 130.5, 130.4, 130.2, 130.1,
130.0, 127.3, 127.2, 117.6, 117.4, 117.3, 116.2, 112.1, 111.7, 102.5, 102.3, 100.8, 100.7, 87.3, 86.0,
71.4, 71.3, 60.94, 59.9, 59.7, 59.3, 59.3, 58.2, 57.4, 57.3, 56.2, 56.2, 45.2, 44.5, 28.4, 26.8 ppm;
HRMS (ESI): [M+H]+ C29H27Cl2N2O6 calcd 569.1246, found 569.1238.
18

4.1.4.6.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(4-fluorophenyl)acetonitrile (5f)
Aldehyde substrate: 4-Fluorobenzaldehyde (4f), reaction time: 1h. The two diastereomers (81:19)
were separated and purified by preparative TLC.
5f1 : Yield: 65%, white powder; m.p. 166 °C; IR (KBr): 1042, 1086, 1275, 1489, 1620, 2898,3056
cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.43 – 7.29 (m, 2H), 7.12 – 6.81 (m, 2H), 6.70 (d, J = 8.2
Hz, 1H), 6.36 (s, 1H), 5.98 (s, 2H), 5.96 (s, 1H), 5.88 (d, J = 8.2 Hz, 1H), 5.63 (s, 1H), 5.15 (dd, J =
12.6, 2.8 Hz, 1H), 5.09 (d, J = 12.0 Hz, 1H), 4.86 (d, J = 2.9 Hz, 1H), 4.12 (s, 3H), 3.87 (s, 3H), 3.86
(s, 3H), 2.56 – 2.41 (m, 2H), 2.39 – 2.22 (m, 2H) ppm; 13C NMR (126 MHz, chloroform-d) δ 163.6,
161.6, 151.3, 148.4, 140.5, 134.6, 133.0, 132.8, 132.3, 130.5, 128.6, 128.5, 117.5, 117.3, 115.7,
115.5, 112.2, 102.1, 100.8, 88.1, 71.5, 62.3, 60.6, 59.9, 59.5, 56.2, 43.5, 29.8 ppm; HRMS (ESI):
[M+H]+ C29H28FN2O6 calcd 519.1931, found 519.1944
5f2 : Yield: 15%, white powder; m.p. 127 °C; IR (KBr): 1042, 1086, 1275, 1489, 1620, 2898,3056
cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.43 – 7.29 (m, 4H), 6.69 (d, J = 8.1 Hz, 1H), 6.35 (s,
1H), 6.00 (s, 1H), 5.96 (s, 2H), 5.87 (d, J = 8.2 Hz, 1H), 5.62 (s, 1H), 5.18 (dd, J = 12.6, 3.2 Hz, 1H),
5.08 (d, J = 12.6 Hz, 1H), 4.88 (d, J = 3.2 Hz, 1H), 4.11 (s, 3H), 3.87 (s, 3H), 3.85 (s, 3H), 2.60 –
13
2.41 (m, 2H), 2.38 – 2.23 (m, 2H).ppm; C NMR (126 MHz, chloroform-d) 163.5, 161.5, 151.3,
148.4, 140.5, 134.6, 133.0, 132.8, 132.3, 130.5, 128.6, 128.6, 117.6, 117.3, 115.7, 115.6, 112.2,
102.2, 100.8, 88.1, 71.5, 62.3, 60.7, 60.0, 59. 6, 56.2, 43.5, 29.9 ppm; HRMS (ESI): [M+H]+
C29H28FN2O6 calcd 519.1931, found 519.2072.
4.1.4.7.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(2,4-dimethoxyphenyl)acetonitrile (5g)
Aldehyde substrate: 2, 4-Dimethoxybenzaldehyde (4g), reaction time: 6h. The mixture of two
diastereomers (91:9) were purified by preparative TLC. Yield: 65 %, yellow oil; IR (KBr): 1030,
1082, 1269, 1619, 2932 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.08 – 6.95 (m, 4H, mixture of
two diastereomers), 6.86 (d, J = 8.2 Hz, 2H, mixture of two diastereomers), 6.78 (s, 1H, minor
diastereomer), 6.77 (s, 1H, major diastereomer), 6.68 (d, J = 8.3 Hz, 1H, major diastereomer ), 6.34
(s, 1H, major diastereomer), 6.30 (s, 1H, minor diastereomer), 6.07 (d, J = 8.1 Hz, 1H, minor
diastereomer), 5.95 (s, 2H, major diastereomer), 5.89 (s, 2H, minor diastereomer), 5.87 (s, 1H, minor
diastereomer), 5.62 (s, 1H, major diastereomer), 5.47 (s, 1H, major diastereomer), 5.30 (s, 1H, minor
diastereomer), 5.22 (dd, J = 12.5, 2.8 Hz, 2H, mixture of two diastereomers), 5.09 (d, J = 12.5 Hz,
19

1H, major diastereomer), 5.04 (d, J = 12.2 Hz, 1H, minor diastereomer), 4.84 (d, J = 2.8 Hz, 1H,
major diastereomer), 4.74 (d, J = 4.0 Hz, 1H, minor diastereomer), 4.10 (s, 6H, mixture of two
diastereomers), 3.84 (s, 6H, mixture of two diastereomers), 3.83 (s, 6H, mixture of two
diastereomers), 3.81 (s, 6H, mixture of two diastereomers), 3.71 (s, 6H, mixture of two
diastereomers), 2.58 – 2.25 (m, 8H, mixture of two diastereomers) ppm; 13C NMR (126 MHz,
chloroform-d) δ 151.2, 148.2, 142.5, 140.3, 134.4, 133.4, 132.4, 131.8, 130.7, 129.0, 117.6, 117.4,
116.87, 114.9, 111.7, 105.7, 104.6, 102.1, 100.8, 99.0, 97.9, 87.5, 71.5, 60.5, 59.9, 59.7, 59.5, 56.2,
55.7, 55.6, 55.5, 48.8, 46.9, 29.6 ppm; HRMS (ESI): [M+H]+ C31H33N2O8 calcd 561.6110, found
561.2693.
4.1.4.8.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(pyridin-3-yl)acetonitrile (5h)
Aldehyde substrate: 3-Pyridinecarbaldehyde (4h), reaction time: 0.5 h. The mixture of diastereomers
(83:17) were purified by preparative TLC. Yield: 85%; off white powder; m.p. 87 °C; IR (KBr):
1032, 1083, 1271, 1486, 1621, 2939 cm-1; 1H NMR (300 MHz, chloroform-d) δ 8.77 (d, J = 2.4 Hz,
1H, minor diastereomer), 8.61 (d, J = 7.2 Hz, 1H, minor diastereomer), 8.63 – 8.53 (m, 2H, major
diastereomer), 7.85 (d, J = 8.0 Hz, 1H, minor diastereomer), 7.67 (dd, J = 7.9, 2.3 Hz, 1H, major
diastereomer), 7.35 (dd, J = 8.0, 4.9 Hz, 1H, minor diastereomer), 7.28 - 7.25 (m, 1H, major
diastereomer), 6.69 (d, J = 8.2 Hz, 1H, major diastereomer), 6.62 (d, J = 8.2 Hz, 1H, minor
diastereomer), 6.36 (s, 1H, major diastereomer), 6.32 (s, 1H, minor diastereomer), 6.07 (s, 1H, major
diastereomer), 5.98 (s, 2H, major diastereomer), 5.95 (s, 1H, minor diastereomer), 5.92 (s, 2H, minor
diastereomer), 5.88 (d, J = 8.2 Hz, 2H, mixture of two diastereomers), 5.80 (s, 1H, minor
diastereomer), 5.64 (s, 1H, major diastereomer), 5.51 (s, 1H, minor diastereomer), 5.38 (d, J = 11.7
Hz, 1H, minor diastereomer), 5.19 – 5.02 (m, 2H, major diastereomer), 4.88 (d, J = 2.9 Hz, 1H, major
diastereomer), 4.69 (dd, J = 8.9, 3.8 Hz, 1H, minor diastereomer), 4.12 (s, 3H, major diastereomer),
3.94 (s, 3H, minor diastereomer), 3.87 (s, 3H, major diastereomer), 3.86 (s, 3H, major diastereomer),
3.74 (s, 6H, minor diastereomer), 2.72 – 2.29 (m, 8H, mixture of two diastereomers) ppm; 13C NMR
(151 MHz, chloroform-d) δ 151.4, 150.0, 149.7, 149.3, 148.5, 148.4, 142.7, 140.4, 135.7, 134.7,
134.6, 132.9, 132.7, 132.5, 132.1, 130.9, 130.7, 123.4, 117.9, 117.5, 117.4, 117.4, 116.4, 112.3,
111.9, 102.9, 102.3, 102.1, 100.9, 100.7, 88.0, 86.8, 71.6, 71.4, 63.6, 60.9, 60.6, 60.1, 60.0, 59.5,
20

59.2, 58.2, 58.2, 56.2, 56.1, 43.7, 29.8, 27.8 ppm; HRMS (ESI): [M+H]+ C28H28N3O6 calcd 502.1978,
found 502.1974.
4.1.4.9.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(4-(propan-2-yl)phenyl)acetonitrile (5i)
Aldehyde substrate: 4-(Propan-2-yl)benzaldehyde (4i), reaction time: 1h. The major diastereomer
was crystalized from methanol and hexane (5i1). Yield: 75%; white crystal; mp : 154 °C; IR (KBr):
1033, 1083, 1272, 1486, 1620, 2926 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.31 (d, J = 8.1 Hz,
2H), 7.18 (d, J = 8.2 Hz, 2H), 6.69 (d, J = 8.2 Hz, 1H), 6.36 (s, 1H), 5.98 (s, 2H), 5.96 (s, 1H), 5.88
(d, J = 8.2 Hz, 1H), 5.62 (br s, 1H), 5.24 (dd, J = 12.5, 2.7 Hz, 1H), 5.11 (d, J = 12.3 Hz, 1H), 4.88
(d, J = 2.8 Hz, 1H), 4.12 (s, 3H), 3.88 (s, 6H), 2.90 (hept, J = 6.9 Hz, 1H), 2.57 (dt, J = 6.7, 3.3 Hz,
1H), 2.45 (td, J = 10.4, 3.3 Hz, 1H), 2.37 – 2.19 (m, 2H), 1.24 (d, J = 6.9 Hz, 6H) ppm; 13C NMR
(126 MHz, chloroform-d) δ 151.3, 149.2, 148.3, 142.7, 140.5, 134.6, 133.1, 132.9, 132.5, 132.05,
126.9, 126.7, 117.9, 117.6, 117.5, 112.1, 102.2, 100.8, 88.1, 71.6, 62.9, 60.8, 59.9, 59.5, 56.2, 43.60,
33.7, 29.9, 23.9, 23.8 ppm; HRMS (ESI): [M+H]+ C32H35N2O6 calcd 543.2495, found 543.2497.
4.1.4.10.
(4-(Benzyloxy)phenyl)((5R)-5-((1S)-4,5-dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-
methoxy-5,6,7,8-tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)acetonitrile (5j)
Aldehyde substrate: 4-Benzyloxybenzaldehyde (4j), reaction time: 8h. The mixture of two
diastereomers (84:16) were purified by preparative TLC. Yield: 75 %, off white powder; m.p. 98 °C;
IR (KBr): 1033, 1083, 1270, 1488, 1620, 2925 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.87 (d, J
= 8.4 Hz, 2H, minor diastereomer), 7.48 – 7.28 (m, 10H, mixture of two diastereomers), 7.10 (d, J =
8.4 Hz, 2H, minor diastereomer), 6.99 (d, J = 8.4 Hz, 2H, minor diastereomer), 6.92 (d, J = 8.3 Hz,
2H, major diastereomer), 6.69 (d, J = 8.3 Hz, 1H, major diastereomer), 6.63 (d, J = 8.1 Hz, 1H, minor
diastereomer), 6.36 (s, 1H, major diastereomer), 6.32 (s, 1H, minor diastereomer), 6.05 (d, J = 8.4
Hz, 1H, minor diastereomer), 5.98 (s, 2H, major diastereomer), 5.94 (s, 2H, minor diastereomer),
5.92 (s, 1H, major diastereomer), 5.88 (d, J = 8.5 Hz, 1H, major diastereomer), 5.84 (s, 1H, minor
diastereomer), 5.81 (s, 1H, minor diastereomer), 5.62 (s, 1H, major diastereomer), 5.46 – 5.31 (m,
4H, minor diastereomer), 5.19 (d, J = 12.34, 2.7 Hz, 1H, major diastereomer), 5.10 (d, J = 12.4 Hz,
1H, major diastereomer), 5.06 (s, 2H, major diastereomer), 4.86 (d, J = 2.9 Hz, 1H, major
diastereomer), 4.75 (d, J = 3.0 Hz, 1H, minor diastereomer), 4.11 (s, 3H, major diastereomer), 4.02
(s, 3H, minor diastereomer), 3.87 (s, 6H, mixture of two diastereomers), 3.84 (s, 6H, mixture of two
21

diastereomers), 3.10 – 2.95 (m, 1H, minor diastereomer), 2.61 – 2.22 (m, 7H, mixture of two
13
diastereomers) ppm; C NMR (126 MHz, chloroform-d) δ 158.8, 151.2, 148.3, 134.6, 133.1, 132.9,
132.4, 128.6, 128.2, 128.0, 127.4, 127.4, 126.9, 117.9, 117.6, 117.6, 114.9, 112.1, 102.1, 100.8, 88.1,
71.5, 70.0, 62.4, 60.7, 59.9, 59.5, 56.2, 43.4, 29.8 ppm; HRMS (ESI): [M+H]+ C36H35N2O7 calcd
607.2744, found 607.2747.
4.1.4.11.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(9H-fluoren-2-yl)acetonitrile (5k)
Aldehyde substrate: 9H-fluoren-2-carbaldehyde (4k), reaction time: 24h. The mixture of two
diastereomers (5k) (84:16) were purified by preparative TLC, yield: 65 %.
5k: Off white powder; m.p. 120 °C; IR (KBr): 1042, 1086, 1275, 1488, 1620, 2898 cm-1; 1H NMR
(300 MHz, chloroform-d) δ 7.80 (d, J = 7.2 Hz, 2H, minor diastereomer), 7.78 (d, J = 7.2 Hz, 2H,
major diastereomer), 7.77 – 7.67 (m, 4H, mixture of two diastereomers), 7.57 (d, J = 6.9 Hz, 2H,
mixture of two diastereomers), 7.56 (s, 2H, mixture of two diastereomer), 7.47 – 7.28 (m, 6H,
mixture of two diastereomers), 6.73 (d, J = 8.2 Hz, 1H, major diastereomer), 6.67 (d, J = 8.2 Hz, 1H,
minor diastereomer), 6.37 (s, 1H, major diastereomer), 6.32 (s, 1H, minor diastereomer), 6.09 (s, 1H,
minor diastereomer), 6.07 (s, 1H, major diastereomer), 5.99 (s, 2H, major diastereomer), 5.93 (s, 2H,
minor diastereomer), 5.89 (d, J = 8.1 Hz, 2H, mixture of two diastereomer), 5.67 (s, 1H, major
diastereomer), 5.60 (s, 1H, minor diastereomer), 5.35 (dd, J = 12.4, 2.6 Hz, 1H, minor diastereomer),
5.26 (dd, J = 12.4, 2.9 Hz, 1H, major diastereomer), 5.13 (d, J = 12.4 Hz, 1H, major diastereomer),
5.11 (d, 12.4 Hz, 1H, minor diastereomer), 4.92 (d, J = 2.8 Hz, 1H, major diastereomer), 4.83 (d, J =
2.6 Hz, 1H, minor diastereomer), 4.13 (s, 3H, major diastereomer), 4.11 (s, 3H, minor diastereomer),
3.91 (s, 6H, mixture of two diastereomers), 3.89 (s, 6H, mixture of two diastereomers), 3.88 (s, 2H,
major diastereomer), 3.84 (s, 2H, minor diastereomer), 2.64-2.60 (m, 2H, mixture of two
diastereomers), 2.58 – 2.46 (m, 2H, mixture of two diastereomers), 2.40 – 2.30 (m, 4H, mixture of
two diastereomers) ppm; 13C NMR (151 MHz, chloroform-d) δ 151.3, 148.4,143.8, 143.4, 142.7,
142.2, 140.9, 140.6, 134.5, 132.9, 132.2, 129.7, 128.4, 127.1, 127.0, 126.8, 125.7, 125.3, 125.1,
123.8, 121.1, 120.1, 120.0, 119.9, 117.5, 112.2, 102.2, 100.8, 88.0, 71.6, 62.9, 60.7, 59.9, 59.5, 56.2,
43.6, 36.8, 29.6 ppm; HRMS (ESI): [M+H]+ C36H33N2O6 calcd 589.2339, found 589.2320.
5k1 (major diastereomer): White powder; m.p. 104 °C; IR (KBr): 1042, 1086, 1275, 1488, 1620,
2898 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.78 (d, J = 7.4 Hz, 1H), 7.72 (d, J = 8.0 Hz, 1H),
7.57 (d, J = 6.5 Hz, 1H), 7.55 (s, 1H), 7.45- 7.31 (m, 3H), 6.74 (d, J = 8.2 Hz, 1H), 6.37 (s, 1H), 6.08
22

(s, 1H), 5.99 (s, 2H), 5.94 (d, J = 8.3 Hz, 1H), 5.67 (s, 1H), 5.27 (dd, J = 12.6, 2.8 Hz, 1H), 5.14 (d, J
= 12.4 Hz, 1H), 4.92 (d, J = 2.8 Hz, 1H), 4.14 (s, 3H), 3.91 (s, 3H), 3.90 (s, 3H), 3.88 (s, 2H), 2.68 –
13
2.45 (m, 2H), 2.42 – 2.30 (m, 2H) ppm; C NMR (75 MHz, chloroform-d) δ 151.3, 148.3, 143.8,
143.4, 142.7, 142.1, 140.9, 140.6, 134.6, 133.2, 133.0, 132.4, 127.0, 126.8, 125.5, 125.1, 123.7,
120.0, 119.9, 117.9, 117.7, 117.5, 112.1, 102.2, 100.8, 88.2, 71.6, 63.1, 60.8, 59.9, 59.5, 56.2, 43.6,
36.8, 29.9 ppm; HRMS (ESI): [M+H]+ C36H33N2O6 calcd 589.2339, found 589.2320.
4.1.4.12.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(4-methylphenyl)acetonitrile (5l)
Aldehyde substrate: 4-Methylbenzaldehyde (4l), reaction time: 8h. The mixture of two diastereomers
(5l) (86:14) were purified by preparative TLC, yield: 60 %.
5l: Off white powder; m.p. 74 °C; IR (KBr): 1033, 1083, 1272, 1486, 1620, 2926 cm-1; 1H NMR (600
MHz, chloroform-d) δ 7.25 (d, J = 7.9 Hz, 4H, mixture of two diastereomer), 7.01 (d, J = 7.8 Hz, 4H,
mixture of two diastereomers), 6.67 (d, J = 8.2 Hz, 1H, major diastereomer), 6.65 (d, J = 8.2 Hz, 1H,
minor diastereomer), 6.31 (s, 2H, mixture of two diastereomers), 6.03 (s, 2H, minor diastereomer),
5.92 (s, 2H, major diastereomer), 5.90 (d, J = 7.8 Hz, 2H, mixture of two diastereomers), 5.83 (s, 1H,
major diastereomer), 5.78 (s, 1H, minor diastereomer), 5.63 (s, 1H, major diastereomer), 5.56 (s, 1H,
minor diastereomer), 5.10 (d, J = 12.0, Hz, 2H, mixture of two diastereomers), 5.04 (d, J = 12.4 Hz,
2H, mixture of two diastereomers), 4.81 (d, J = 2.1 Hz, 1H, major diastereomer), 4.80 (d, J = 2.0 Hz,
1H, minor diastereomer), 4.00 (s, 3H, major diastereomer), 3.84 (s, 3H, minor diastereomer), 3.83 (s,
6H, mixture of two diastereomers), 3.82 (s, 6H, mixture of two diastereomers), 2.70-2.61 (m, 2H,
mixture of two diastereomers), 2.58 – 2.46 (m, 2H, mixture of two diastereomers), 2.40 – 2.32 (m,
13
4H, mixture of two diastereomers), 2.30 (s, 6H, mixture of two diastereomers) ppm; C NMR (126
MHz, chloroform-d) δ 151.2, 148.3, 140.5, 138.2, 134.5, 133.1, 132.9, 132.5, 131.7, 129.3, 129.3,
128.3, 126.8, 117.9, 117.6, 117.5, 112.1, 111.9, 102.3, 102.1, 101.6, 100.8, 88.1, 71.6, 70.7, 62.8,
60.8, 60.7, 60.0, 59.9, 59.5, 56.2, 43.8, 43.5, 29.9, 29.6, 28.1, 21.0 ppm; HRMS (ESI): [M+H]+
C30H31N2O6 calcd 515.2182, found 515.2192.
5l1: Off white powder; m.p. 91 °C; IR (KBr): 1033, 1083, 1272, 1486, 1620, 2926 cm-1; 1H NMR
(600 MHz, chloroform-d) δ 7.30 (d, J = 7.6 Hz, 2H), 7.15 (d, J = 7.8 Hz, 2H), 6.740 (d, J = 8.2 Hz,
1H), 6.37 (s, 1H), 6.00 - 5.94 (m, 3H), 5.84 (s, 1H), 5.70 (s, 1H), 5.11 (d, J = 12.6, 1H), 5.07 (d, J =
12.1 Hz, 1H), 4.85 (s, 1H), 4.02 (s, 3H), 3.87 (s, 3H), 3.86 (s, 3H), 2.76 – 2.73 (m, 1H), 2.64 – 2.60
23

(m, 1H), 2.42 – 2.37 (m, 2H), 2.35 (s, 3H) ppm; 13C NMR (126 MHz, chloroform-d) δ 151.2, 148.3,
138.2, 134.6, 133.1, 132.9, 132.5, 131.7, 129.4, 129.3, 126.8, 117.5, 112.1, 102.1, 100.8, 88.1, 71.6,
62.8, 60.8, 59.9, 59.5, 56.2, 43.5, 29.9, 21.0 ppm; HRMS (ESI): [M+H]+ C30H31N2O6 calcd 515.2182,
found 515.2192.
4.1.4.13.
2-((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)-3-methylbutanenitrile (5m)
Aldehyde substrate: 2-Methylpropanal (4m), reaction time: 2h. The mixture of two diastereomers
(80:20) were purified by preparative TLC. Yield: 75 %, off white powder; m.p. 68 °C; IR (KBr):
1035, 1083, 1270, 1487, 1622, 2962 cm-1; 1H NMR (300 MHz, chloroform-d) δ 6.68 (d, J = 8.2 Hz,
1H, minor diastereomer) 6.64 (d, J = 8.2 Hz, 1H, major diastereomer), 6.35 (s, 2H, mixture of two
diastereomers), 6.03 (d, J = 8.2 Hz, 1H, minor diastereomer), 5.94 (s, 4H, mixture of two
diastereomers), 5.93 (d, J = 8.2 Hz, 1H, major diastereomer), 5.47 (s, 1H, minor diastereomer), 5.43
(s, 1H, major diastereomer), 5.23 (dd, J = 12.2, 2.7 Hz, 2H, mixture of two diastereomers ), 5.12 (d, J
= 12.5 Hz, 1H, minor diastereomer), 5.05 (d, J = 12.5 Hz, 1H, major diastereomer), 4.60 (d, J = 5.2
Hz, 1H, major diastereomer), 4.55 (d, J = 4.3 Hz, 1H, minor diastereomer), 4.05 (s, 3H, major
diastereomer), 3.94 (d, J = 5.7 Hz, 2H, mixture of two diastereomers), 3.90 (s, 3H, minor
diastereomer), 3.86 (s, 6H, minor diastereomer), 3.82 (s, 6H, major diastereomer), 2.87 – 2.69 (m,
2H, mixture of two diastereomers), 2.66 – 2.55 (m, 1H, minor diastereomer), 2.49 – 2.19 (m, 6H,
mixture of two diastereomers), 2.07 – 1.86 (m, 1H, major diastereomer), 1.12 (d, J = 6.6 Hz, 6H,
mixture of two diastereomers), 0.87 (d, J = 6.6 Hz, 6H, mixture of two diastereomers) ppm; 13C
NMR (151 MHz, chloroform-d) δ 151.2, 148.3, 142.6, 140.5, 134.6, 133.1, 132.8, 132.0, 117.5,
112.0, 102.1, 100.8, 87.5, 71.6, 71.1, 66.2, 61.4, 60.0, 59.9, 59.5, 56.3, 56.2, 43.1, 29.1, 20.3, 18.7
ppm; HRMS (ESI): [M+H]+ C26H31N2O6 calcd 467.2182, found 467.2180.
4.1.4.14.
((5R)-5-((1S)-4,5-dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(2-hydroxyphenyl)acetonitrile (5n)
Aldehyde substrate: 2-Hydroxybenzaldehyde (4n), reaction time: 2h. The major diastereomer was
crystalized from methanol (5n1). Yield: 71%; orange crystal; mp : 186 °C; IR (KBr): 1081, 1276,
1488, 1618, 2912, 3269 cm-1; 1H NMR (300 MHz, chloroform-d) δ 9.22 (s, 1H), 7.46 (d, J = 7.7 Hz,
1H), 7.23 (d, J = 7.0 Hz, 1H), 6.89 (td, J = 7.5, 1.2 Hz, 1H), 6.76 (d, J = 8.0 Hz, 1H), 6.68 (d, J = 8.3
Hz, 1H), 6.40 (s, 1H), 6.18 (s, 1H), 5.99 (s, 2H), 5.82 (d, J = 8.2 Hz, 1H), 5.71 (s, 1H), 5.41 (dd, J =
24

12.3, 2.7 Hz, 1H), 5.06 (d, J = 12.3 Hz, 1H), 4.95 (d, J = 3.3 Hz, 1H), 4.13 (s, 3H), 3.93 (s, 3H), 3.85
(s, 3H), 2.73 (dt, J = 10.4, 3.6 Hz, 1H), 2.66 – 2.38 (m, 3H) ppm; 13C NMR (75 MHz, chloroform-d)
δ 156.2, 151.8, 148.8, 143.1, 140.5, 134.7, 133.9, 131.6, 130.8, 130.3, 127.9, 119.7, 117.4, 116.8,
116.5, 116.1, 115.7, 112.4, 102.1, 101.0, 86.5, 71.0, 62.1, 61.2, 59.9, 59.6, 56.1, 44.2, 29.0 ppm;
HRMS (ESI): [M+H]+ C29H29N2O7 calcd 517.1975, found 517.1987.
4.1.4.15.
2-((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)butanenitrile (5o)
Aldehyde substrate: Propanal (4o), reaction time: 2h. The mixture of two diastereomers (85:15) were
purified by preparative TLC. Yield: 65 %, yellow powder; m.p. 84 °C; IR (KBr): 1035, 1083, 1270,
1
1487, 1622, 2962 cm-1; H NMR (300 MHz, chloroform-d) δ 6.65 (d, J = 8.2 Hz, 1H, minor
diastereomer), 6.61 (d, J = 8.1 Hz, 1H, major diastereomer),6.34 (s, 1H, minor diastereomer), 6.32 (s,
1H, major diastereomer), 5.99 (d, J = 8.4 Hz, 1H, minor diastereomer), 5.94 (s, 2H, major
diastereomer), 5.92 (s, 2H, minor diastereomer), 5.83 (d, J = 8.2 Hz, 1H, major diastereomer), 5.38
(s, 1H, major diastereomer), 5.35 (s, 1H, minor diastereomer), 5.19 (d, J = 12.2 Hz, 2H, mixture of
two diastereomer), 5.09 (d, J = 12.1 Hz, 2H, mixture of two diastereomer), 4.63 (d, J = 4.1 Hz, 1H,
minor diastereomer), 4.56 (d, J = 3.9 Hz, 1H, major diastereomer), 4.50 (dd, J = 9.8, 4.1 Hz, 1H,
minor diastereomer), 4.25 (dd, J = 9.8, 6.0 Hz, 1H, major diastereomer), 4.05 (s, 3H, major
diastereomer), 3.99 (s, 3H, minor diastereomer), 3.86 (s, 6H, minor diastereomer), 3.84 (s, 3H, major
diastereomer), 3.82 (s, 3H, major diastereomer), 2.87 – 2.75 (m, 2H, mixture of two diastereomers),
2.69 – 2.49 (m, 2H, mixture of two diastereomers), 2.48 – 2.30 (m, 2H, mixture of two
diastereomers), 2.05 (td, J = 15.0, 13.8, 4.1 Hz, 2H, mixture of two diastereomers), 1.94 – 1.76 (m,
2H, mixture of two diastereomers), 1.75 – 1.61 (m, 2H, mixture of two diastereomers), 1.06 (t, J =
7.4 Hz, 6H, mixture of two diastereomers) ppm; 13C NMR (126 MHz, chloroform-d) δ 151.2, 148.1,
140.4, 134.5, 133.3, 132.7, 132.3, 119.1, 118.0, 117.54, 117.3, 111.6, 102.4, 102.0, 100.8, 100.6,
87.6, 87.30, 8.7, 71.6, 71.5, 61.5, 61.4, 59.9, 59.5, 59.0, 58.4, 56.2, 56.1, 44.5, 43.1, 42.6, 41.6, 29.7,
29.5, 28.1, 25.8, 23.5, 10.8 ppm; HRMS (ESI): [M+H]+ C25H29N2O6 calcd 453.2026, found
453.2038.
4.1.5. General Procedure for the Synthesis of Amide Derivatives
H2O2 30% (2mL) was added dropwise to a mixture of N-α-aminonitrile derivative (1.0 mmol) and
K2CO3 (5.0 mmol) in 5 mL CH3OH and 500 µL DMSO. The reaction was stirred at room
25

temperature for the specified time. The reaction was monitored by TLC and after completion, the
solvent was evaporated and water (5 mL) was added. The mixture was extracted with CHCl3 (3×30).
Organic layer was dried (Na2SO4) and concentrated under reduced pressure. Finally, the two
diastereomers were purified by preparative TLC.
4.1.5.1.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(phenyl)acetamide (6a)
Yield: 65 %, reaction time: 5h
6a1 (major diastereomer): White powder; m.p. 78 °C; IR (KBr): 1036, 1078, 1267, 1484, 1684, 2931,
3436 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.91 (s, 1H), 7.19 (s, 5H), 6.68 (d, J = 8.2 Hz, 1H),
6.27 (s, 1H), 6.09 (d, J = 8.1 Hz, 1H), 5.93 (s, 1H), 5.88 (s, 1H), 5.84 (s, 1H), 5.47 (s, 1H), 5.13 (s,
2H), 4.29 (s, 1H), 4.27 (d, J = 5.5 Hz, 1H), 3.84 (s, 3H), 3.84 (s, 3H), 3.50 (s, 3H), 2.92 – 2.75 (m,
3H), 2.35 – 2.20 (m, 1H) ppm; 13C NMR (126 MHz, chloroform-d) δ 175.8, 153.3, 149.3, 147.9,
140.1, 136.8, 134.1, 129.7, 129.0, 128.2, 127.9, 118.0, 117.2, 112.2, 112.1, 102.7, 100.4, 85.5, 72.3,
71.4, 63.7, 60.0, 58.9, 56.2, 44.0, 23.6 ppm; HRMS (ESI): [M+H]+ C29H31N2O7 calcd 519.2131,
found 519.2191.
6a2 (minor diastereomer): White powder; m.p. 82 °C; IR (KBr): 1033, 1075, 1267, 1485, 1684, 2928,
3432 cm-1; 1H NMR (500 MHz, chloroform-d) δ 7.94 (s, 1H), 7.19 (s, 5H), 6.67 (d, J = 8.1 Hz, 1H),
6.26 (s, 1H), 6.08 (d, J = 8.2 Hz, 1H), 5.97 (s, 1H), 5.87 (s, 1H), 5.84 (s, 1H), 5.47 (s, 1H), 5.12 (s,
2H), 4.28 (s, 1H), 4.27 (s, 1H), 3.83 (s, 6H), 3.49 (s, 3H), 2.94 – 2.74 (m, 3H), 2.27 (m, 1H) ppm; 13C
NMR (126 MHz, chloroform-d) δ 175.6, 149.9, 140.8, 136.6, 129.9, 128.8, 128.4, 128.1, 117.2,
112.3, 112.2, 102.6, 100.6, 84.9, 71.9, 71.8, 60.0, 59.7, 59.3, 56.2, 42.6, 40.4, 23.6 ppm; HRMS
(ESI): [M+H]+ C29H31N2O7 calcd 519.2131, found 519.2191.
4.1.5.2.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(3-phenoxyphenyl)acetamide (6b)
Yield: 75 %, reaction time: 12h
6b1 (major diastereomer): Off white powder; m.p. 54 °C; IR (KBr): 1038, 1080, 1263, 1485, 1699,
2926, 3237 cm-1; 1H NMR (300 MHz, chloroform-d) δ 8.09 (s, 1H), 7.39 – 7.23 (m, 2H), 7.16 (t, J =
7.8 Hz, 1H), 7.08 (t, J = 7.4 Hz, 1H), 7.02 – 6.77 (m, 5H), 6.65 (d, J = 8.2 Hz, 1H), 6.25 (s, 1H), 6.06
(s, 1H), 6.05 (d, J = 8.2 Hz, 1H), 5.87 (s, 1H), 5.86 (s, 1H), 5.50 (s, 1H), 5.15 (t, J = 14.2 Hz, 2H),
26

4.37 (d, J = 4.7 Hz, 1H), 4.26 (s, 1H), 3.83 (s, 3H), 3.82 (s, 3H), 3.64 (s, 3H), 2.91 – 2.67 (m, 3H),
13
2.33 – 2.11 (m, 1H) ppm; C NMR (126 MHz, chloroform-d) δ 175.3, 156.8, 151.2, 147.9, 139.0,
133.9, 133.7, 131.8, 129.7, 129.6, 129.3, 123.9, 123.1, 119.8, 118.7, 118.2, 117.8, 117.2, 112.2,
102.7, 100.4, 85.5, 72.4, 71.4, 60.0, 58.9, 56.3, 56.2, 44.4, 42.6, 23.7 ppm; HRMS (ESI): [M+H]+
C35H35N2O8 calcd 611.2393, found 611.2427.
6b2 (minor diastereomer): Off white powder; m.p. 76 °C; IR (KBr): 1038, 1080, 1263, 1486, 1698,
2928, 3417 cm-1; 1H NMR (300 MHz, chloroform-d) δ 8.22 (d, J = 3.7 Hz, 1H), 7.34 (t, J = 7.9 Hz,
2H), 7.27 (t, J = 7.8 Hz, 1H), 7.14 (s, 1H), 7.11 (d, J = 7.2 Hz, 2H), 7.00 (d, J = 7.8 Hz, 2H), 6.92 (d,
J = 8.1 Hz, 1H), 6.62 (d, J = 8.2 Hz, 1H), 6.29 (s, 1H), 5.95 (d, J = 1.5 Hz, 1H), 5.94 (d, J = 1.5 Hz,
1H), 5.88 (d, J = 8.1 Hz, 1H), 5.77 (s, 1H), 5.53 (d, J = 3.7 Hz, 1H), 5.41 (dd, J = 12.2, 2.4 Hz, 1H),
5.24 (d, J = 12.2 Hz, 1H), 4.56 (d, J = 4.9 Hz, 1H), 4.27 (s, 1H), 4.02 (s, 3H), 3.87 (s, 3H), 3.81 (s,
3H), 2.51 – 2.32 (m, 2H), 2.24 – 2.08 (m, 1H), 2.00 – 1.82 (m, 1H) ppm; 13C NMR (126 MHz,
chloroform-d) δ 175.1, 157.3, 156.9, 148.3, 143.3, 140.8, 138.7, 134.0, 133.3, 129.8, 129.7, 125.2,
123.8, 123.3, 118.9, 118.9, 118.3, 117.2, 112.3, 102.6, 101.6, 100.6, 84.8, 77.2, 77.0, 76.7, 71.8, 71.5,
60.0, 59.6, 59.3, 56.2, 40.4, 23.6 ppm; HRMS (ESI): [M+H]+ C35H35N2O8 calcd 611.2393, found
611.2444.
4.1.5.3.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(3,4-dimethoxyphenyl)acetamide (6c)
Yield: 77 %, reaction time: 24h
6c1 (major diastereomer): Yellow powder; m.p. 114 °C; IR (KBr): 1032, 1079, 1264, 1478, 1683,
2944, 3446 cm-1; 1H NMR (300 MHz, chloroform-d) δ 7.84 (s, 1H), 6.83 – 6.59 (m, 4H), 6.32 (s,
1H), 6.30 (s, 1H), 6.14 (d, J = 8.1 Hz, 1H), 5.87 (s, 1H), 5.85 (s, 1H), 5.48 (s, 1H), 5.16 (t, J = 12.9
Hz, 2H), 4.25 (d, J = 5.3 Hz, 1H), 4.21 (s, 1H), 3.84 (s, 3H), 3.83 (s, 6H), 3.69 (s, 3H), 3.55 (s, 3H),
13
2.95 – 2.73 (m, 3H), 2.48 – 2.15 (m, 1H) ppm; C NMR (126 MHz, chloroform-d) δ 175.8, 151.2,
148.6, 147.9, 143.0, 140.3, 134.2, 133.8, 131.9, 129.6, 129.4, 121.4, 118.2, 117.1, 112.2, 112.0,
110.9, 102.7, 100.4, 85.5, 71.8, 71.3, 59.9, 58.8, 56.2, 56.1, 55.9, 55.5, 43.7, 23.4 ppm; HRMS (ESI):
[M+H]+ C31H35N2O9 calcd 579.2343, found 579.2389.
6c2 (minor diastereomer): Yellow powder; m.p. 85 °C; IR (KBr): 1032, 1079, 1264, 1478, 1683,
1
2944, 3446 cm-1; H NMR (300 MHz, chloroform-d) δ 8.27 (s, 1H), 6.96 (dd, J = 8.2, 1.9 Hz, 1H),
27

6.91 (d, J = 1.9 Hz, 1H), 6.82 (d, J = 8.2 Hz, 1H), 6.63 (d, J = 8.2 Hz, 1H), 6.30 (s, 1H), 5.96 (s, 1H),
5.94 (s, 1H), 5.89 (d, J = 8.2 Hz, 1H), 5.79 (s, 1H), 5.62 (s, 1H), 5.48 (dd, J = 12.0, 2.9 Hz, 1H), 5.28
(d, J = 12.1 Hz, 1H), 4.58 (d, J = 5.0 Hz, 1H), 4.24 (s, 1H), 4.02 (s, 3H), 3.88 (s, 6H), 3.87 (s, 6H),
13
3.82 (s, 3H), 2.47 – 2.32 (m, 2H), 2.23 – 2.05 (m, 1H), 1.97 – 1.86 (m, 1H) ppm; C NMR (126
MHz, chloroform-d) δ 175.7, 159.9, 141.7, 140.8, 131.0, 130.0, 129.2, 129.1, 121.8, 121.5, 117.3,
117.2, 116.3, 112.3, 111.9, 111.1, 102.6, 100.6, 90.5, 85.0, 71.8, 71.4, 60.0, 59.6, 59.3, 56.2, 55.8,
42.6, 40.4, 23.8 ppm; HRMS (ESI): [M+H]+ C31H35N2O9 calcd 579.2343, found 579.2384.
4.1.5.4.
(2-Chlorophenyl)((5R)-5-((1S)-4,5-dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-
5,6,7,8-tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)acetamide (6d)
Yield: 55 %, reaction time: 24h
6d1 (major diastereomer): Off white powder; m.p. 66 °C; IR (KBr): 1034, 1081, 1267, 1482, 1689,
2902, 3413 cm-1; 1H NMR (300 MHz, chloroform-d) δ 8.13 (s, 1H), 7.35 – 7.29 (m, 1H), 7.27 – 7.21
(m, 1H), 7.16 – 7.02 (m, 2H), 6.65 (d, J = 8.2 Hz, 1H), 6.17 (s, 2H), 6.00 (d, J = 8.2 Hz, 1H), 5.87 (d,
J = 1.5 Hz, 1H), 5.84 (d, J = 1.5 Hz, 1H), 5.51 (s, 1H), 5.17 (dd, J = 12.4, 4.9 Hz, 1H), 5.11 (d, J =
12.2 Hz, 1H), 5.03 (s, 1H), 4.46 (d, J = 4.8 Hz, 1H), 3.83 (s, 3H), 3.82 (s, 3H), 3.72 (s, 3H), 2.93 –
13
2.61 (m, 3H), 2.30 – 2.09 (m, 1H) ppm; C NMR (126 MHz, chloroform-d) δ 174.8, 151.2, 147.9,
139.8, 135.4, 135.0, 133.7, 133.5, 131.8, 130.3, 130.1, 129.4, 128.9, 126.4, 117.8, 117.3, 112.1,
102.6, 100.4, 85.5, 71.4, 70.6, 68.1, 60.0, 58.9, 56.4, 56.2, 44.5, 24.3 ppm; HRMS (ESI): [M+H]+
C29H30ClN2O7 calcd 553.1742, found 553.1792.
6d2 (minor diastereomer): Off white powder; m.p. 67 °C; IR (KBr): 1034, 1081, 1267, 1482, 1689,
1
2942, 3413 cm-1; H NMR (300 MHz, chloroform-d) δ 8.38 (s, 1H), 7.50 (dd, J = 7.4, 2.1 Hz, 1H),
7.36 (dd, J = 7.4, 1.9 Hz, 1H), 7.22 (td, J = 7.4, 1.9 Hz, 2H), 6.62 (d, J = 8.3 Hz, 1H), 6.29 (s, 1H),
5.95 (d, J = 1.5 Hz, 1H), 5.94 (d, J = 1.5 Hz, 1H), 5.88 (d, J = 8.1 Hz, 1H), 5.80 (s, 1H), 5.76 (s, 1H),
5.50 (dd, J = 12.1, 2.9 Hz, 1H), 5.28 (d, J = 12.1 Hz, 1H), 4.99 (s, 1H), 4.70 (d, J = 5.0 Hz, 1H), 4.04
(s, 3H), 3.88 (s, 3H), 3.81 (s, 3H), 2.45 (ddd, J = 16.7, 11.2, 5.7 Hz, 1H), 2.36 – 2.03 (m, 2H), 1.92
13
(d, J = 16.6 Hz, 1H) ppm; C NMR (126 MHz, chloroform-d) δ 175.3, 151.3, 148.3, 140.7, 135.0,
134.6, 134.0, 133.7, 133.2, 132.1, 130.1, 130.1, 129.0, 126.8, 117.3, 116.1, 112.3, 102.7, 102.6,
100.6, 84.9, 71.9, 71.5, 60.1, 59.8, 59.3, 56.1, 40.9, 24.2 ppm; HRMS (ESI): [M+H]+ C29H30ClN2O7
calcd 553.1742, found 553.1773.
28

4.1.5.5.
(2,4-Dichlorophenyl)((5R)-5-((1S)-4,5-dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-
methoxy-5,6,7,8-tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)acetamide (6e)
Yield: 90 %, reaction time: 8h
6e1 (major diastereomer): Off white powder; m.p. 76 °C; IR (KBr): 1034, 1081, 1267, 1482, 1689,
2902, 3413 cm-1; 1H NMR (300 MHz, chloroform-d) δ 8.14 (s, 1H), 7.29 – 7.18 (m, 2H), 7.03 (dd, J
= 8.4, 2.2 Hz, 1H), 6.64 (d, J = 8.3 Hz, 1H), 6.50 (s, 1H), 6.16 (s, 1H), 5.97 (d, J = 8.1 Hz, 1H), 5.86
(s, 2H), 5.48 (s, 1H), 5.17 (dd, J = 12.4, 2.6 Hz, 1H), 5.11 (d, J = 12.5 Hz, 1H), 4.97 (s, 1H), 4.44 (d,
J = 4.8 Hz, 1H), 3.82 (s, 3H), 3.82 (s, 3H), 3.79 (s, 3H), 2.95 – 2.51 (m, 3H), 2.24 – 2.05 (m, 1H)
13
ppm; C NMR (126 MHz, chloroform-d) δ 174.8, 151.3, 148.1, 142.9, 139.8, 134.3, 133.9, 133.5,
133.4, 131.1, 130.0, 129.0, 126.7, 117.4, 117.2, 112.2, 102.6, 100.4, 85.3, 71.4, 60.0, 59.0, 56.4, 56.2,
44.6, 42.6, 24.3 ppm; HRMS (ESI): [M+H]+ C29H29Cl2N2O7 calcd 587.1352, found 587.1373.
6e2 (minor diastereomer): Off white powder; m.p. 79 °C; IR (KBr): 1034, 1081, 1267, 1482, 1689,
1
2942, 3413 cm-1; H NMR (300 MHz, chloroform-d) δ 8.33 (s, 1H), 7.41 (d, J = 11.4 Hz, 1H), 7.28
(s, 1H), 7.23 (dd, J = 8.4, 2.2 Hz, 1H), 6.62 (d, J = 8.2 Hz, 1H), 6.29 (s, 1H), 5.95 (s, 1H), 5.94 (s,
1H), 5.92 (s, 1H), 5.87 (d, J = 8.2 Hz, 1H), 5.78 (s, 1H), 5.48 (dd, J = 12.2, 2.8 Hz, 1H), 5.26 (d, J =
12.2 Hz, 1H), 4.98 (s, 1H), 4.70 (d, J = 4.8 Hz, 1H), 4.04 (s, 3H), 3.88 (s, 3H), 3.81 (s, 3H), 2.40
(ddd, J = 16.5, 10.9, 5.8 Hz, 1H), 2.31 – 2.08 (m, 2H), 1.94 (dt, J = 16.8, 3.8 Hz, 1H) ppm; 13C NMR
(125 MHz, chloroform-d) δ 172.3, 151.4, 148.3, 143.0, 140.7, 135.7, 134.1, 134.0, 133.3, 133.0,
132.2, 131.1, 129.9, 129.8, 127.2, 117.3, 116.0, 112.3, 102.5, 100.6, 85.0, 71.8, 60.0, 59.8, 59.3, 56.1,
41.1, 24.5 ppm; HRMS (ESI): [M+H]+ C29H29Cl2N2O7 calcd 587.1352, found 587.1363.
4.1.5.6.
((5R)-5-((1S)-4,5-Dimethoxy-1,3-dihydro-2-benzofuran-1-yl)-4-methoxy-5,6,7,8-
tetrahydro[1,3]dioxolo[4,5-g]isoquinolin-6-yl)(4-fluorophenyl)acetamide (6f)
Yield: 66 %, reaction time: 24h
6f1 (major diastereomer): Yellow powder; m.p. 64 °C; IR (KBr): 1035, 1080, 1268, 1488, 1679,
2927, 3430 cm-1; 1H NMR (300 MHz, chloroform-d) δ 8.00 (s, 1H), 7.21 – 7.09 (m, 2H), 6.93 – 6.85
(m, 2H), 6.67 (d, J = 8.2 Hz, 1H), 6.25 (s, 1H), 6.05 (d, J = 8.2 Hz, 1H), 5.97 (d, J = 4.4 Hz, 1H),
5.88 (d, J = 1.5 Hz, 1H), 5.86 (d, J = 1.4 Hz, 1H), 5.48 (s, 1H), 5.15 (m, 2H), 4.27 (s, 1H), 4.25 (d, J
= 5.2 Hz, 1H), 3.85 (s, 3H), 3.83 (s, 3H), 3.60 (s, 3H), 2.88 – 2.71 (m, 3H), 2.33 – 2.18 (m, 1H) ppm;
13C NMR (126 MHz, chloroform-d) δ 176.1, 163.3, 161.3, 151.2, 148.0, 139.9, 133.8, 133.7, 132.8,
29