Synthesis and cytotoxicity of Baylis-Hillman template derived betulinic acid-triazole conjugates

Article abstract of DOI:10.1016/j.tet.2016.11.056

Several alkynes and azides were prepared starting from betulinic acid and Baylis-Hillman reaction-derived allylic alcohols. These alkynes and azides were then coupled under click cycloaddition conditions to obtain functionalized betulinic acid-triazole conjugates. Similarly, pyrazinyl- and indolylbetulinic acid-triazoles were also prepared employing cycloaddition chemistry. All the synthetic compounds were tested for their cytotoxicity against murine breast cancer (4T1) and human pancreatic cancer (MIA PaCa-2) cell lines. Based on these in vitro assays, two series of compounds have been identified as lead compounds for further development.

Full text of DOI:10.1016/j.tet.2016.11.056

Accepted Manuscript
Synthesis and cytotoxicity of Baylis-Hillman template derived betulinic acid-triazole
conjugates
Pathi Suman, Amardeep Patel, Lucas Solano, Gayathri Jampana, Zachary S.
Gardner, Crystal M. Holt, Subash C. Jonnalagadda
PII:
S0040-4020(16)31220-0
10.1016/j.tet.2016.11.056
TET 28270
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 19 September 2016
Revised Date: 18 November 2016
Accepted Date: 21 November 2016
Please cite this article as: Suman P, Patel A, Solano L, Jampana G, Gardner ZS, Holt CM,
Jonnalagadda SC, Synthesis and cytotoxicity of Baylis-Hillman template derived betulinic acid-triazole
conjugates, Tetrahedron (2016), doi: 10.1016/j.tet.2016.11.056.
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to
our customers we are providing this early version of the manuscript. The manuscript will undergo
copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please
note that during the production process errors may be discovered which could affect the content, and all
legal disclaimers that apply to the journal pertain.

ACCEPTED MANUSCRIPT
Graphical Abstract
To create your abstract, type over the instructions in the template box below.
Fonts or abstract dimensions should not be changed or altered.
Synthesis and Cytotoxicity of Baylis-Hillman
Template Derived Betulinic Acid-Triazole
Conjugates
Leave this area blank for abstract info.
a
a
b
a
b
a
Pathi Suman, Amardeep Patel, Lucas Solano, Gayathri Jampana, Zachary S. Gardner, Crystal M. Holt, and
a,c *
Subash C. Jonnalagadda.
a
b
Department of Chemistry and Biochemistry, Rowan University, Glassboro, NJ 08028, USA, Department of Chemistry and Biochemistry, University of
c
Minnesota, Duluth, MN 55812, USA, Department of Biomedical and Translational Sciences, Rowan University, Glassboro, NJ 08028, USA.

1
ACCEPTED MANUSCRIPT
Tetrahedron
journal homepage: www.elsevier.com
Synthesis and Cytotoxicity of Baylis-Hillman Template Derived Betulinic Acid-
†
Triazole Conjugates
a
a
b
a
b§
Pathi Suman, Amardeep Patel, Lucas Solano, Gayathri Jampana, Zachary S. Gardner, Crystal M.
∗
a,c
a§
Holt, and Subash C. Jonnalagadda.
a
b
Department of Chemistry and Biochemistry, Rowan University, Glassboro, NJ 08028, USA, Department of Chemistry and Biochemistry, University of
c
Minnesota, Duluth, MN 55812, USA, Department of Biomedical and Translational Sciences, Rowan University, Glassboro, NJ 08028, USA.
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
Several alkynes and azides were prepared starting from betulinic acid and Baylis-Hillman
reaction-derived allylic alcohols. These alkynes and azides were then coupled under click
cycloaddition conditions to obtain functionalized betulinic acid-triazole conjugates. Similarly,
pyrazinyl- and indolylbetulinic acid-triazoles were also prepared employing cycloaddition
chemistry. All the synthetic compounds were tested for their cytotoxicity against murine breast
cancer (4T1) and human pancreatic cancer (MIA PaCa-2) cell lines. Based on these in vitro
assays, two series of compounds have been identified as lead compounds for further
development.
Available online
Keywords:
Betulin, Betulinic Acid
Baylis-Hillman Reaction,
Click Reaction,
Triazole,
2
016 Elsevier Ltd. All rights reserved.
Cycloaddition.
—
——
∗
Corresponding author. Tel.: +1-856-256-5452; fax: +1- 856-256-4478; e-mail: jonnalagadda@rowan.edu
Undergraduate Research Participants
§
†
Dedicated to Professor Deevi Basavaiah on the occasion of his retirement.

2
Tetrahedron
ACCEPTED MANUSCRIPT
Introduction
parent betulinic acid for their development as anti-cancer agents.
Additionally, the BH template offers multiple avenues for
chemical manipulations, which should further assist us in
understanding a detailed structure-activity relationship profile for
these analogs, once active molecules have been identified.
Pentacyclic triterpenoids such as betulin 1 and betulinic acid 2
1
(Figure 1) are readily found in >200 types of trees. Betulin is
abundantly available in nature and can be readily extracted from
the bark of birch trees. Various parts of the birch tree including
the leaves, bark, and stem have been used in herbal and folk
medicine as diuretics as well as for reducing the effects of
arthritis and rheumatism. Betulin and related analogs show
diverse medicinal activities ranging from anti-cancer, anti-
Results and Discussion
2
We initiated our synthesis with the preparation of BH reaction
derived allylic azide and alkyne motifs (Scheme 1). The reaction
of benzaldehyde with methyl acrylate in the presence of DABCO
yielded the allylic alcohol 8. The three allylic azides 9a-c were
then obtained from the alcohol 8. Reaction of 8 with HBr and
H SO followed by nucleophilic displacement of the resulting
3
microbial, anti-HIV, and anti-inflammatory properties. While the
detailed mechanism of action is not entirely clear, it is generally
understood that betulinic acid affects the mitochondrial pathways4
and increases the caspase-3 activity thereby leading to apoptosis.
Betulinic acid is also known to be selective toxic to a variety of
cancer cells while the normal cells are typically resistant to this
molecule. Recently, betulinic acid was found to exhibit in vivo
protective effect against dexamethasone induced thymocyte
apoptosis in mice. Several recent reports also indicated excellent
in vivo activity for betulin via the inhibition of NF-κB pathway.
2
4
allylic bromide with sodium azide resulted in the formation of the
azide 9a. Hydrolysis of 9a with NaOH and the coupling of the
resulting acid with N,N,N’-trimethylethylenediamine in the
presence of TBTU and Hünig’s base furnished the azide 9b.
Azide 9c was also obtained from 9a upon alkaline hydrolysis and
coupling with N-methylpiperazine (Scheme 1).
5
6
7
In addition to their medicinal effects, these natural products have8
The two β-azidoethylamide derivatives 9d-e were prepared
with utmost ease starting from the BH alcohol 8. Acetylation of 8
with acetic anhydride followed by the treatment of the acetate 10
with N,N,N’-trimethylethylenediamine and potassium carbonate
in DMF furnished the allylic amine 11. Alkaline hydrolysis of 11
and the coupling of the resulting acid with 2-azidoethylamine in
the presence of HOBt and EDCI produced the azide 9d. The
azide 9e was obtained from the BH acetate 10 via the
displacement with N-methylpiperazine followed by hydrolysis
and coupling with 2-azidoethylamine. Azide 9f was obtained via
the reaction of cinnamic acid 16 with 2-azidoethylamine
also found other applications such as their use as anti-feedants,
9
10
11
bio-based coatings, solder pastes, and bio-hybrid polymers.
Multicomponent coupling reactions offer ready access to
complex and diversely functionalized libraries of compounds and
they are valuable tools in drug design and discovery. Baylis-
Hillman (BH) reaction involves the coupling of aldehydes or
aldimines with activated olefins (acrylates, acrylonitriles,
vinylsulfones, etc.) to furnish the allylic alcohols or amines in a
2
0
12
one-step transformation. Passerini reaction is an isocyanide
based three component coupling reaction used for the preparation
13
(Scheme 1).
of α-acyloxyamides. Click chemistry involves the cycloaddition
14
of azides and alkynes for the preparation of 1,2,3-triazoles. We
have been working on the development of novel small
The alkyne precursors 13a-c were derived via HOBt, EDCI
mediated coupling of propargyl amine with the cinnamic acids
1
5
16
molecules as medicinal agents utilizing Baylis-Hillman and
1
2, 15, and 16 respectively (Scheme 1).
1
7
Passerini reactions. Previously, we had reported the synthesis
of chalcone (3), α-acetoxyamide (4), and amine (5) derivatives of
betulin utilizing aldol condensation, Passerini reaction, and
3e,18
reductive amination respectively (Figure 1).
Owing to the impressive therapeutic effects as well as the
abundant availability of these natural products, we undertook a
project involving the development of triazole derivatives of
betulinic acid as anti-cancer agents using BH reaction and Click
chemistry as the key steps in our synthesis. The synthesis of two
precursors (alkyne and azide) required for Click chemistry was
envisioned from betulinic acid and BH templates. While there
have been multiple reports on click coupling using
betulin/betulinic acid and related analogs, so far, limited success
has been achieved in terms of identifying potent lead molecule
19
for further development. We hypothesized that molecular
hybridization of modestly cytotoxic betulinic acid with the BH
reaction-derived and pharmacologically active 1,4-acceptor unit
should lead to the compounds with higher potency than the
R'
H
H
H
HN
H
R
Ph
N
R
H
X
H
H
O
R
H
O
O
H
H
H
H
HO
HO
HO
O
HO
5
4
H
H
3
H
H
X = CH OH, 1
2
X = COOH, 2
Figure 1: Functionalized Betulin/Betulinic Acid Analogs

3
ACCEPTED MANUSCRIPT
O
O
O
7
OH
O
O
a
b, c
d, e
O
O
O
O
8
N
+
N
N
6
9a
N3
9b
9c
N3
O
f
O
OAc O
d, e
N
g
O
14
N
10
N3
g
N
h
O
N
O
O
O
N
O
N
N3
OH
O
OH
N
h
i
H
1
5
N
N
12
1
1
j
9d
N
N
N
N
N
i
O
O
N
N3
N
H
N
N
j
H
H
N
9
e
13b
13a
N
N
O
O
O
N3
j
N
i
N
OH
H
H
9f
13c
16
Reaction Conditions: (a) DABCO, 25 °C, 14d, 78%; (b) HBr, H SO , 0-25 °C, 3h, 74%; (c) NaN , acetone:H O (4:1), 25 °C, 3h, 85%;
2
4
3
2
i
(
d) NaOH, MeOH:THF (1:9), 25 °C, 78%; (e) R NH, Pr NEt, TBTU, DMF, 0-25 °C, 12-16h, 72-74%; (f) Ac O, NEt , DMAP, CH Cl ,
2
2
2
3
2
2
2
5 °C, 2h, 91%; (g) R NH, K CO , DMF, 25 °C, 13-15h, 77-81%; (h) NaOH, MeOH:THF (1:9), 0-25 °C, 8-10h, 71-74%; (i) 2-
2 2 3
i i
Azidoethylamine, Pr NEt, HOBt, EDCI, CH Cl , 0-25 °C, 12-16h, 72-74%; (j) Propargyl amine, Pr NEt, HOBt, EDCI, CH Cl , 0-25
2
2
2
2
2
2
°
C, 12-16h, 71-76%.
Scheme 1: Preparation of azides 9a-f and alkynes 13a-c
Having prepared the azides 9a-f and alkynes 13a-c from BH
template, we focused our efforts on the functionalization of
betulin for the preparation of click reaction counterparts.
Oxidation of betulin 1 with Jones reagent yielded betulonic acid
1
7. Reaction of 17 with propargyl amine under peptide coupling
conditions using TBTU and Hünig’s base resulted in N-propargyl
betulonamide 18 (Scheme 2). The azide motifs 9a-f (Scheme 1)
were then utilized for click coupling with 18. Our initial efforts
towards click coupling of methyl α-azidomethylcinnamate 9a
with 18 employing copper iodide were not fruitful and the
product triazole 19a was obtained in trace quantities even after
performing the reaction in various solvents or heating the
reaction. However, cycloaddition was realized upon using
sodium ascorbate and copper sulphate in t-butyl alcohol/water as
the solvent medium and the corresponding triazole derivative 19a
21
was obtained in 89% yield (Entry 1, Table 1). Reaction of 19a
with sodium borohydride resulted in the reduction of the C3
ketone to yield the corresponding alcohol 20a. In an effort
towards increasing the hydrophilicity, alcohol 20a was further
converted to the succinic acid hemiester 21a upon refluxing with
succinic anhydride in toluene. Under identical conditions,
reaction of the remaining azides 9b-f with N-propargyl
betulonamide 18 led to the triazoles 19b-f in high yield (Entries
2
-6, Table 1). Treatment of 19b-f with NaBH afforded 20b-f,
4
which upon succinylation furnished the hemiesters 21b-f
Scheme 2). All the compounds 19-21 were characterized using
NMR and mass spectrometric analyses.
(

4
Tetrahedron
ACCEPTED MANUSCRIPT
H
H
H
H
OH
OH
b
N
a
c
H
H
H
O
O
H
H
H
1
17
18
HO
O
O
H
H
H
O
N
N
N
N
N
N
H
H
H
N R
HO
H
N R
H
N R
H
N
N
N
H
H
H
d
e
O
O
O
H
H
H
2
1a-f
O
O
O
1
9a-f
HO
20a-f
H
H
H
i
Reaction Conditions: (a) Jones reagent, acetone, 0-25 °C, 4h, 75%; (b) Propargyl amine, Pr NEt, TBTU, DMF, 0-25 °C, 14h, 73%; (c)
2
t
9
a-f, CuSO , sodium ascorbate, BuOH:H O (1:1), 25 °C, 12-15h, 85-90%; (d) NaBH , CH OH, 0-25 °C, 2-3h, 84-90%; (e) Succinic
4
2
4
3
anhydride, DMAP, toluene, 80 °C, 12-15h, 80-87%.
Scheme 2: Coupling of N-propargyl betulonamide with azido cinnamates/cinnamamides
Table 1: Preparation of betulin conjugates 19-21
Melting
Melting
Point
( C)
Melting
Point
( C)
Yield
%)
Yield
(%)
Yield
(%)
#
1
R
Compound
Point
Compound
Compound
(
o
o
o
(
C)
O
O
N
N
19a
89
88
90
124-126
98-101
20a
86
85
88
123-125
118-121
134-136
21a
87
80
83
122-125
130-132
142-145
O
O
O
2
3
19b
19c
20b
20c
21b
21c
N
N
134-137
N
H
4
19d
85
108-110
20d
90
119-122
21d
82
114-116
N
O
N
N
N
H
5
6
19e
19f
87
91
118-120
139-141
20e
20f
89
90
130-132
185-187
21e
21f
84
81
132-134
141-144
N
O
N
H
We were able to synthesize pyrazinylbetulinic acid derivative
2, via the cycloaddition of betulonic acid 17 with
ethylenediamine upon refluxing with sulphur and morpholine.
In an analogous protocol, betulonic acid 17 was converted to
2
the indolylbetulinic acid 25 upon refluxing with phenyl hydrazine
22
22d,23
in acetic acid under Fischer-Indole synthesis conditions.
The
Amide coupling of 22 with propargyl amine was accomplished
using TBTU and Hünig’s base to generate 23, which was then
subjected to cycloaddition with azides 9a-f under click reaction
reaction of 25 with propargyl amine yielded the N-propargyl
indolylbetulinamide 26. Click coupling of 26 with methyl α-
azidomethylcinnamate 9a and N-2-azidoethylcinnamamide 9f
yielded triazoles 27a and 27f respectively (Scheme 3).
conditions (CuSO and sodium ascorbate). The resulting triazoles
4
2
4a-f were purified by silica gel column chromatography and
characterized via spectroscopic techniques (Scheme 3).

5
ACCEPTED MANUSCRIPT
N
N
H
H
H
N R
H
H
N
OH
c
N
H
H
H
N
N
N
N
N
N
O
O
O
a
H
H
d
H
17
2
4a-f
23
22
H
H
H
b
N
N
H
H
H
N R
H
H
N
OH
N
H
H
H
O
O
O
H
H
H
c
d
N
H
N
N
H
27a, 27f
26
25
H
H
H
H
O
N
O
R =
O
R =
R =
R =
O
R =
O
R =
O
N
N
H
N
O
N
N
H
H
N
N
N
N
C
N
4a: 82%, 127-129o
24f: 88%, 150-152o
27f: 81%, 167-169o
C
C
2
2
C
4b: 84%, 131-133o
24c: 84%, 137-140o
C
2
C
7a: 80%, 133-136o
24d: 85%, 112-114o
24e: 87%, 132-135o
C
C
Reaction Conditions: (a) Ethylenediamine, Sulfur, Morpholine, reflux, 24h, 71%; (b) Phenylhydrazine, CH COOH, reflux, 4h, 45%; (c)
3
i
t
Propargyl amine, Pr NEt, TBTU, DMF, 0-25 °C, 14-20h, 71-80%; (d) 9a-f, CuSO , sodium ascorbate, BuOH:H O (1:1), 25 °C, 12-
2
4
2
1
5h, 80-88%.
Scheme 3: Coupling of fused heterocyclic N-propargyl betulinamides with azido cinnamates/cinnamamides
H
H
H
H
H
H
N
N
N
N3
O
N
R
N
R
H
H
H
N N
b
N N
a
O
O
O
17
H
H
c
H
3
0a-c
O
28
29a-c
HO
H
H
H
R =
O
O
R =
R =
O
N
N
H
H
N
H
N
N
N
N
2
3
9a: 79%, 108-111o
0a: 86%, 116-118o
C
C
29b: 86%, 115-117o
30b: 89%, 129-131o
C
29c: 81%, 155-158o
30c: 91%, 163-166o
C
C
C
i
Reaction Conditions: (a) 2-Azidoethylamine, Pr NEt, TBTU, DMF, 0-25 °C, 20h, 74%; (b) 13a-c, CuSO , sodium ascorbate,
2
4
t
BuOH:H O (1:1), 25 °C, 12-15h, 79-86%; (c) NaBH , CH OH, 0-25 °C, 2-3h, 86-91%.
2
4
3
Scheme 4: Coupling of N-azidoethyl betulinamide with N-alkynylcinnamamides
The triazoles shown in Schemes 2-3 were obtained upon click
coupling of betulin-derived alkynes 18, 23, and 26 with Baylis-
Hillman motif-derived azides 9a-f. In order to understand the
complete SAR profile of our template, we decided to switch the
coupling partners for the click reaction. Accordingly, we chose
the BH reaction-derived alkynes (13a-c, Scheme 1), for coupling
with N-(2-azidoethyl) betulonamide 28. The amide 28 was
obtained upon coupling betulonic acid 17 with 2-azidoethylamine
in the presence of TBTU and N,N-diisopropylethylamine. It was
noticed that an excess of the amine was required for this coupling
as the use of molar equivalent of 2-azidoethylamine furnished the
unreacted intermediate benzotriazolyl ester of betulonic acid and
led to lower yields of the amide 28. Coupling of 28 with N-
propargyl α-(dialkylaminomethyl)cinnamamides 13a-b and N-
propargyl cinnamamide 13c under click conditions proceeded
smoothly to provide triazoles 29a-c. Reduction of 29a-c with
sodium borohydride yielded the betulinic acid based triazole
derivatives 30a-c in 86-91% yield (Scheme 4).
All the synthesized betulinic acid-triazole conjugates as well
as some of the intermediates were evaluated for their general
cytotoxicity against murine breast cancer (4T1) and pancreatic
cancer (MIA PaCa-2) cell lines. The cells were purchased from
ATCC and the assays were performed under standard conditions
by seeding the cells in 96 well plates and the cell viability was
determined using MTT assay. All the candidate compounds were
tested at 50 µM, 12.5 µM, and 1.5 µM concentrations. While
betulinic acid showed moderate activity at 12.5 µM (~60% cell
death for 4T1 and ~40% cell death for MIA PaCa-2), majority of
the betulinic acid-triazole conjugates (eg. 20b-e, 24d-e, and 30a-
c, etc.) that were tested showed promising activity at this
concentration (>90% cell death for 4T1 and ~80-90% cell death

6
Tetrahedron
for MIA PaCa-2). The compounds showing
A
m
C
od
C
era
E
te
P
t
TE
o g
o
D
od MAin
NU
ter
est
S
in
C
g
R
toIP
note that the conversion of C -alcohol to the
T
3
cytotoxicity were then selected for determining the IC values
succinic acid hemi ester 21a-f (Entries 13-18) did not improve
the biological activity. Compounds 19a-f and 21a-f showed
comparable activity to that of the parent molecule betulinic acid
5
0
(Table 2).
(Entry 36) in both 4T1 and MiaPaCa-2 cells, however, the C3-
Table 2: In vitro cytotoxicity of the BH-template derived
alcohol containing analogs 20b-e (Entries 8-11) showed two-
fold (4T1) and ~20-fold (MiaPaCa-2) increase in biological
activity. While most of the pyrazine and indole containing
analogs did not show significant activity, analogs 24d-e (Entries
betulinic acid-triazole derivatives
4
T1
MIA PaCa-2
IC50 (µM)*
NT
#
Compound
IC (µM)*
5
0
2
3-24),
which
contain
the
allylic
amine
moiety
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
19a
19b
19c
19d
19e
19f
20a
20b
20c
20d
20e
20f
21a
21b
21c
21d
21e
21f
23
NT
(trimethylethylenediamine and piperazine respectively) showed
6.93 ± 1.09
7.15 ± 0.18
5.67 ± 0.10
5.17 ± 0.60
NT
9.87 ± 0.79
14.99 ± 2.12
8.11 ± 1.54
5.98 ± 0.99
NT
2-6 fold increase in potency. It was encouraging to note that the
betulinic acid derivatives 30a-c (Entries 33-35), which were
obtained upon switching coupling partners for the click reaction,
showed comparable or better cytotoxicity when compared to their
structurally analogous counterparts 20d-f (Entries 10-12).
Based on these findings, we recognize the importance of C3-
alcohol unit (eg. compounds 20 & 30), and the N,N,N’-
trimethylethylenediamine motif (eg. compounds 20b, 20d, 24d,
30a, etc.) for enhancing the biological activity of these
molecules. It should also be noted that none of the intermediates
tested showed any toxicity, which leads us to believe that the BH
motif is also required for the activity. Efforts are currently
underway to identify the mechanism of action for the lead
derivatives as well as for the identification of potent candidate for
further development.
39.75 ± 9.45
3.97 ± 0.73
4.37 ± 0.43
2.38 ± 0.45
2.62 ± 0.24
27.26 ± 12.2
5.26 ± 0.65
7.48 ± 0.59
6.13 ± 0.81
4.13 ± 0.22
7.05 ± 0.53
8.61 ± 2.27
39.33 ± 7.0
NT
NT
2.44 ± 0.36
8.69 ± 0.33
1.36 ± 0.21
1.64 ± 0.20
32.50 ± 7.92
47.58 ± 5.51
21.73 ± 5.12
14.44 ± 3.44
9.66 ± 0.51
24.28 ± 3.24
NT
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
Conclusion
In conclusion, we have prepared betulinic acid-triazole
derivatives utilizing Baylis-Hillman reaction and click chemistry
as the key protocols in our synthesis. We have also prepared
pyrazinyl and indolyl betulinic acid derivatives employing the
26.73 ± 3.70
NT
24a
24b
24c
24d
24e
24f
26
above protocol.
These compounds were tested for their
5.14 ± 0.80
17.28 ± 5.00
2.88 ± 0.06
2.88 ± 0.04
13.47 ± 2.10
NT
8.26 ± 0.91
15.21 ± 1.91
3.87 ± 0.56
4.36 ± 0.44
33.21 ± 2.80
NT
biological efficacy against murine breast cancer cell line (4T1)
and human pancreatic cancer cell line (MIA PaCa-2). Based on
these in vitro assays, we have been able to identify two series of
betulin derivatives for further SAR and pre-clinical studies. The
ready availability of betulin from natural resources as well as the
great chemical diversity afforded by the Baylis-Hillman template
imparts significance to this class of compounds for potential
development as anti-cancer agents.
27a
27f
28
NT
NT
NT
NT
NT
NT
Experimental Section
29a
29b
29c
30a
30b
5.39 ± 0.40
4.74 ± 0.93
NT
5.02 ± 0.47
8.10 ± 2.42
NT
General Methods: All operations were carried out under an inert
atmosphere of nitrogen. Glassware for all reactions was oven
dried at 125 °C and cooled under nitrogen prior to use. Liquid
reagents and solvents were introduced by oven-dried syringes or
1.49 ± 0.06
5.38 ± 0.22
0.81 ± 0.03
1.34 ± 0.19
3.86 ± 0.39
3.56 ± 0.38
cannulas through septa sealed flasks under
a nitrogen
atmosphere. THF was distilled from sodium benzophenone ketyl.
All other solvents and reagents were purchased and used without
30c
Betulinic
acid
1
13
3
6
6.29 ± 0.96
25.63 ± 3.79
further purification. The H and C NMR spectra were plotted on
a Varian-400 spectrometer fitted with a Quad probe.
*
*
IC50 values reported as average ± SEM
General amide-coupling procedure A: To a stirred solution of
the appropriate acid (1.0 mmol) in dimethylformamide (10.0
mL), was added N,N-diisopropylethylamine (2.0 mmol) followed
by TBTU (1.1 mmol) at 0 °C and stirred for 30 min. The
appropriate amine (1.0 mmol) was then added in one portion and
stirred overnight at room temperature. Upon completion (as
indicated by TLC), the reaction mixture was quenched by the
Minimum of three independent experiments.
NT = Not toxic
Based on the detailed IC50 analysis, it was revealed that the C3
ketone containing betulin analogs (19a-f, Entries 1-6, Table 2)
showed lower biological activity than the corresponding C3-
alcohol based triazoles 20a-f (Entries 7-12). Further, it was
addition of saturated
NaHCO3
and extracted with
dichloromethane (2 x 10.0 mL). The combined extracts were
washed with cold water (10.0 mL) and brine (10.0 mL). The

7
organic layer was dried over anhydrous Na SO4
A
,
C
co
C
nc
E
en
P
tr
T
ate
E
d
D
in MAPr
N
ep
methylaminomethyl)-3-phenyl
Trimethylethylenediamine (521 mg, 5.1 mmol) and K
mg, 6.40 mmol) were added to a stirred solution of 10 (1.0 g,
.27 mmol ) in N,N-dimethylformamide (10.0 mL) at room
U
ar
S
at
C
io
R
n
I
o
P
f
T
Methyl (E)-2-(N-(2-dimethylaminoethyl)-N-
acrylate, 11: N,N,N’-
CO (883
2
vacuo, and purified by column chromatography (silica gel,
hexanes:ethyl acetate) to obtain pure amides. This procedure was
utilized for the preparation of amides 9b-c, 23, 26, and 28.
2
3
4
General
diisopropylethylamine (2.0 mmol), HOBt (1.1 mmol), and EDCI
1.1 mmol) were added at 0 °C to a stirred solution of the
amide-coupling
procedure
B:
N,N-
temperature and the reaction mixture was stirred overnight. Upon
completion (TLC), the reaction was quenched with cold water
and extracted with ethyl acetate (2 x 20.0 mL). The combined
organic layers were washed thoroughly with cold water (2 x 10.0
mL), brine (2 x 10.0 mL) and dried over anhydrous Na SO . The
(
appropriate acid (1.0 mmol) in dichloromethane (10.0 mL) and
the reaction was stirred for 30 min. The appropriate amine (1.0
mmol) was added in one portion and the reaction was stirred
overnight at room temperature. After completion of the reaction
as indicated by TLC, the reaction mixture was quenched by the
addition of saturated NaHCO3 solution and worked up with
dichloromethane (2 x 10.0 mL). The combined extracts were
washed with brine (10.0 mL), dried over anhydrous Na SO ,
2
4
ethyl acetate was concentrated in vacuo and purified by column
chromatography (silica gel, hexanes: ethyl acetate, 1:4) to obtain
1
1
1 as brown liquid (912 mg, 77%). H NMR (400 MHz, CDCl ):
3
δ (ppm) 7.80 (s, 1H), 7.57 – 7.61 (m, 2H), 7.29 – 7.41 (m, 3H),
.82 (s, 3H), 3.39 (s, 2H), 2.48 – 2.54 (m, 2H), 2.37 – 2.44 (m,
H), 2.21 (s, 6H), 2.18 (s, 3H); C NMR (101 MHz, CDCl ): δ
ppm) 168.9, 142.6, 135.3, 130.4, 130.2, 128.7, 128.3, 57.2, 55.5,
3
2
13
2
4
3
concentrated in vacuo, and purified by column chromatography
silica gel, hexanes:ethyl acetate) to obtain pure amides in good
yields. This procedure was utilized for the preparation of amides
d-f and 13a-c.
(
5
(
3.1, 51.9, 45.8, 41.9; ESIMS: m/z calculated for C H N O
1
6
24
2
2
+
(M+H) 277.19, found 277.38.
9
Preparation
azidomethyl-3-phenyl acrylamide, 9b: The reaction of (E)-2-
azidomethyl)-3-phenylacrylic acid (1.0 g, 4.9 mmol) with
of
N-(2-N,N-dimethylaminoethyl)
(E)-2-
Preparation of (E)-2-(N-(2-dimethylaminoethyl)-N-methyl
aminomethyl)-3-phenyl acrylic acid, 12: aq. NaOH (2.9 mL,
2.5 M, 7.3 mmol) was added to a solution of 11 (1.0 g, 3.6 mmol)
in THF:MeOH (9:1, 20.0 mL) at 0 °C and the reaction was stirred
overnight at room temperature. Upon completion (TLC), the
solution was acidified to pH 6 with 1N HCl. The solution was
concentrated in vacuo and the resulting slurry was dissolved in
isopropyl alcohol to effect the precipitation of sodium chloride.
The reaction was then filtered and the filtrate was concentrated in
(
N,N,N’-trimethylethylenediamine (602 mg, 5.9 mmol) as per the
general amide coupling procedure A yielded 1.02 g (72%) of 9b
1
as a pale brown liquid. H NMR (400 MHz, CDCl ): δ (ppm)
3
7
2
2
1
.31 – 7.44 (m, 3H), 7.26 – 7.31 (m, 2H), 6.78 (s, 1H), 4.30 (s,
H), 3.61 (t, J = 7.1 Hz, 2H), 3.15 (s, 3H), 2.55 (t, J = 7.1 Hz,
13
H), 2.28 (s, 6H); C NMR (101 MHz, CDCl ): δ (ppm) 169.5
3
34.4, 133.5, 131.6, 128.8, 128.6, 128.4, 49.4, 45.7; ESIMS: m/z
vacuo to yield 671 mg (71%) of 12 as pale cream-colored semi-
+
1
calculated for C H N O (M+H) 288.18, found 288.38.
solid. H NMR (400 MHz, CDCl
3
): δ (ppm) 7.96 (s, 1H), 7.33 –
15
21
5
7
3
.42 (m, 3H), 7.26 – 7.31 (m, 2H), 3.81 (s, 2H), 3.45 (m, 2H),
.19 (m, 2H), 2.89 (s, 6H), 2.43 (s, 3H); C NMR (101 MHz,
Preparation of (E)-2-azidomethyl-1-(4-methylpiperazin-1-yl)-
-phenylprop-2-en-1-one, 9c: The reaction of (E)-2-
azidomethyl)-3-phenylacrylic acid (1.0 g, 4.92 mmol) with N-
13
3
(
DMSO-d ): δ (ppm) 169.5, 142.9, 134.9, 130.4, 129.5, 129.1,
6
5
2.9, 52.7, 51.3, 42.6, 41.4; ESIMS: m/z calculated for
methylpiperazine (590 mg, 5.90 mmol) as per the general amide
+
C H N O (M+H) 263.17, found 263.37.
15
22
2
2
coupling procedure A furnished 1.04 g (74%) of 9c as a pale
1
brown liquid. H NMR (400 MHz, CDCl ): δ (ppm) 7.24 – 7.41
3
Preparation of N-propargyl (E)-2-(N-(2-dimethylamino
ethyl)-N-methylaminomethyl)-3-phenyl acrylamide, 13a: The
reaction of 12 (430 mg, 1.6 mmol) with propargyl amine (99 mg,
(
2
m, 5H), 6.69 (s, 1H), 4.26 (s, 2H), 3.61 – 3.80 (m, 4H), 2.37 –
13
.50 (m, 4H), 2.29 (s, 3H); C NMR (101 MHz, CDCl ): δ
3
(
4
ppm) 169.9, 134.3, 134.0, 131.2, 129.0, 128.9, 128.8, 55.0, 49.5,
1
.8 mmol) as per the general amide-coupling procedure B
+
6.2; ESIMS: m/z calculated for C H N O (M+H) 286.16,
1
15
19
5
yielded 364 mg (72%) of 13a as a pale orange liquid. H NMR
400 MHz, CDCl ): δ (ppm) 10.08 (s, 1H), 7.98 (s, 1H), 7.21 –
found 286.30.
(
3
7
2
.39 (m, 5H), 4.11 (dd, J = 2.5, 5.3 Hz, 2H), 3.38 (s, 2H), 2.36 –
.50 (m, 4H), 2.24 (s, 6H), 2.17 (t, J = 2.6 Hz, 1H), 2.14 (s, 3H);
Preparation of N-2-azidoethyl (E)-2-(N-(2-dimethylamino
ethyl)-N-methylaminomethyl)-3-phenyl acrylamide, 9d: The
reaction of 12 (500 mg, 1.90 mmol) with 2-azidoethylamine (180
mg, 2.09 mmol) as per the general amide-coupling procedure B
1
3
C NMR (101 MHz, CDCl ): δ (ppm) 167.9, 140.3, 136.1,
3
1
4
3
30.9, 129.2, 128.4, 127.9, 80.9, 70.5, 56.8, 54.6, 52.8, 45.5,
+
1
2.1, 29.1; ESIMS: m/z calculated for C H N O (M+H)
yielded 460 mg (74%) of 9d as a pale orange liquid. H NMR
18 25
3
00.20, found 300.41.
(
400 MHz, CDCl ): δ (ppm) 10.10 (s, 1H), 7.96 (s, 1H), 7.21 –
3
7
2
.41 (m, 5H), 3.49 – 3.53 (m, 4H), 3.39 (s, 2H), 2.43 – 2.51 (m,
13
Preparation of Methyl (E)-2-(N-methylpiperazin-1-ylmethyl)-
-phenyl acrylate, 14: Procedure similar to that of 11. The
reaction of 10 (1.0 g, 4.3 mmol) with N-methylpiperazine (512
mg, 5.1 mmol) and K CO (883 mg, 6.4 mmol) provided 943 mg
H), 2.34 – 2.42 (m, 2H), 2.24 (s, 6H), 2.14 (s, 3H); C NMR
3
(101 MHz, CDCl ): δ (ppm) 168.6, 140.1, 136.1, 131.0, 129.2,
3
1
28.4, 128.0, 56.8, 54.6, 53.1, 50.8, 45.6, 42.0, 39.7; ESIMS: m/z
+
calculated for C H N O (M+H) 331.22, found 331.35.
2
3
17
26
6
1
(
81%) of 14 as a pale cream liquid. H NMR (400 MHz, CDCl ):
3
Preparation of N-(2-azidoethyl) (E)-2-(N-methylpiperazin-1-
ylmethyl)-3-phenyl acrylamide, 9e: The reaction of 15 (250
mg, 0.96 mmol) with 2-azidoethylamine (90 mg, 1.05 mmol) as
δ (ppm) 7.84 (s, 1H), 7.63 – 7.68 (m, 2H), 7.32 – 7.41 (m, 3H),
1
3
3
.81 (s, 3H), 3.35 (s, 2H), 2.31 – 2.63 (m, 8H), 2.26 (s, 3H);
C
NMR (101 MHz, CDCl ): δ (ppm) 169.3, 143.6, 135.6, 130.7,
3
per the general amide-coupling procedure B yielded 226 mg,
1
29.9, 129.1, 128.6, 55.5, 53.4, 52.8, 52.3, 46.3; ESIMS: m/z
1
+
(72%) of 9e as a pale orange liquid. H NMR (400 MHz, CDCl ):
3
calculated for C H N O (M+H) 275.17, found 275.10.
16
22
2
2
δ (ppm) 10.04 (s, 1H), 7.92 (s, 1H), 7.18 – 7.36 (m, 5H), 3.50 –
3
8
1
4
3
.56 (m, 2H), 3.45 – 3.49 (m, 2H), 3.39 (s, 2H), 2.33 – 2.68 (m,
Preparation
of
(E)-2-(N-methylpiperazin-1-ylmethyl)-3-
13
H), 2.25 (s, 3H); C NMR (101 MHz, CDCl ): δ (ppm) 168.6,
3
phenyl acrylic acid, 15: Procedure similar to that of 12. The
reaction of 14 (1.0 g, 3.6 mmol) and aq. NaOH (2.9 mL, 2.5 M,
40.7, 135.7, 129.8, 129.2, 128.5, 128.2, 55.3, 54.9, 52.5, 51.2,
+
6.2, 38.9; ESIMS: m/z calculated for C H N O (M+H)
17
24
6
7.3 mmol) yielded 710 mg (74%) of 15 as a pale cream-colored
1
29.20, found 329.33.
solid. H NMR (400 MHz, CDCl ): δ (ppm) 8.02 (s, 1H), 7.38 –
3

8
Tetrahedron
7
.42 (m, 3H), 7.30 – 7.32 (m, 2H), 3.66 (s, 2H), 2.95 – 3.10 (m,
Hz, 1H), 3.63 (m, 4H), 3.12 (dt, J = 4.3, 11.2 Hz, 1H), 2.34 –
ACCEPTED MANUSCRIPT
13
8
1
H), 2.69 (s, 3H). C NMR (101 MHz, CDCl ): δ (ppm) 169.5,
42.9, 134.6, 129.5, 128.8, 128.3, 127.9, 53.2, 52.9, 49.4, 43.4;
2.52 (m, 3H), 2.17 – 2.30 (m, 4H), 2.25 (s, 3H), 0.77 – 1.94 (m,
3
21H), 1.67 (s, 3H), 1.05 (s, 3H), 1.01 (s, 3H), 0.94 (s, 3H), 0.87
(s, 3H), 0.81 (s, 3H); C NMR (101 MHz, CDCl ): δ (ppm)
+
13
ESIMS: m/z calculated for C H N O (M ) 260.15, found
15
20
2
2
3
2
60.80.
218.4, 176.5, 169.0, 150.9, 145.3, 134.8, 134.1, 129.9, 129.1,
1
29.0, 128.9, 123.7, 109.6, 55.8, 55.2, 54.9, 50.3, 50.1, 48.8,
Preparation of N-propargyl (E)-2-(N-methylpiperazin-1-
ylmethyl)-3-phenyl acrylamide, 13b: The reaction of 15 (210
mg, 0.8 mmol) with propargyl amine (48 mg, 0.9 mmol) as per
47.5, 46.9, 46.2, 42.7, 40.9, 39.8, 38.4, 37.9, 37.1, 34.9, 34.3,
33.8, 33.7, 31.0, 29.6, 26.8, 25.8, 21.6, 21.2, 19.8, 19.7, 16.2,
+
15.8, 14.8; ESIMS: m/z 777.65 [100%, (M+H) ], HRMS-ESI:
+
the general amide-coupling procedure B yielded 176 mg (71%)
calculated for C H N O [M+Na] 799.5245, found 799.5239.
48
68
6
3
1
of 13b as a pale orange liquid. H NMR (400 MHz, CDCl ): δ
3
(
ppm) 10.08 (s, 1H), 7.90 (s, 1H), 7.27 – 7.31 (m, 2H), 7.22 –
.26 (m, 1H), 7.15 – 7.19 (m, 2H), 4.07 (dd, J = 2.6, 4.8 Hz, 2H),
Preparation of triazole 19d: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 18 with
azide 9d. Yield: 85%; cream color solid; mp 108 – 110 °C; H
7
3
=
1
.35 – 3.38 (m, 2H), 2.25 – 2.60 (m, 8H), 2.21 (s, 3H), 2.18 (t, J
13
2.6 Hz, 1H); C NMR (101 MHz, CDCl ): δ (ppm) 167.9,
NMR (400 MHz, CDCl ): δ (ppm) 9.89 (brs, 1H), 7.94 (s, 1H),
3
3
1
5
2
40.8, 135.6, 129.4, 129.2, 128.5, 128.2, 80.4, 71.4, 55.3, 54.8,
7.59 (s, 1H), 7.22 – 7.40 (m, 5H), 6.37 (brs, 1H), 4.73 (s, 1H),
4.56 – 4.60 (m, 3H), 4.51 (dd, J = 5.4, 15.0 Hz, 1H), 4.44 (dd, J =
5.5, 15.0 Hz, 1H), 3.65 – 3.82 (m, 2H), 3.34 (s, 2H), 3.12 (dt, J =
+
2.4, 46.2, 29.4; ESIMS: m/z calculated for C H N O (M+H)
18
23
3
98.19, found 298.40.
4
.2, 11.1 Hz, 1H), 2.34 – 2.52 (m, 7H), 2.19 (brs, 3H), 2.07 (s,
Preparation of triazole 19a: To a stirred solution of alkyne 18
450 mg, 0.9 mmol) and azide 9a (198 mg, 0.9 mmol) in a
6H), 0.82 – 1.97 (m, 21H), 1.67 (s, 3H), 1.05 (s, 3H), 1.01 (s,
3H), 0.94 (s, 3H), 0.90 (s, 3H), 0.84 (s, 3H); C NMR (101
13
(
mixture of t-butanol/water (1:1, 8.0 mL), was added CuSO (23
MHz, CDCl ): δ (ppm) 218.1, 176.4, 168.5, 150.8, 145.0, 140.1,
4
3
mg, 0.1 mmol) and sodium ascorbate (36 mg, 0.2 mmol). The
reaction mixture was stirred overnight at room temperature. Upon
completion (TLC), the reaction was concentrated in vacuo and
diluted with water to effect precipitation. The resulting solid was
filtered, washed with water, and further purified via column
chromatography (silica gel, methanol:dichloromethane, 2:3) to
135.6, 130.5, 128.9, 128.3, 127.9, 122.9, 109.4, 55.9, 55.5, 54.9,
53.8, 52.7, 50.0, 49.9, 49.2, 47.3, 46.7, 44.9, 42.4, 41.6, 40.6,
39.9, 39.6, 38.2, 37.7, 36.9, 34.7, 34.1, 33.6, 33.4, 30.8, 29.6,
29.3, 26.6, 25.6, 21.4, 21.0, 19.7, 19.4, 15.9, 15.6, 14.5; ESIMS:
+
m/z 822.65 [100%, (M+H) ], HRMS-ESI: calculated for
+
C H N O [M+Na] 844.5824, found 844.5831.
50
75
7
3
obtain 528 mg (89%) of 1,2,3-triazole 19a as a white solid. Mp
1
1
7
(
24 – 126 °C, H NMR (400 MHz, CDCl ): δ (ppm) 8.07 (s, 1H),
Preparation of triazole 19e: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 18 with
azide 9e. Yield: 87%; cream color solid; mp 118 – 120 °C; H
3
.74 (s, 1H), 7.62 (d, J = 7.4 Hz, 2H), 7.39 – 7.46 (m, 3H), 6.36
t, J = 5.2 Hz, 1H), 5.35 (s, 2H), 4.72 (s, 1H), 4.58 (s, 1H), 4.40 –
1
4
2
0
.56 (m, 2H), 3.84 (s, 3H), 3.11 (dt, J = 4.2, 11.0 Hz, 1H), 2.33 –
NMR (400 MHz, CDCl ): δ (ppm) 9.87 (s, 1H), 7.92 (s, 1H),
3
.49 (m, 3H), 0.70 – 2.04 (m, 21H), 1.66 (s, 3H), 1.03 (s, 3H),
7.61 (s, 1H), 7.22 – 7.37 (m, 3H), 7.20 – 7.25 (m, 2H), 6.23 –
6.29 (m, 1H), 4.73 (s, 1H), 4.59 (s, 1H), 4.38 – 4.55 (m, 4H),
3.79 – 3.92 (m, 2H), 3.34 (s, 2H), 3.08 – 3.15 (m, 1H), 2.29 –
2.52 (m, 11H), 2.25 (s, 3H), 0.76 – 1.92 (m, 21H), 1.75 (s, 3H),
1.04 (s, 3H), 1.00 (s, 3H), 0.94 (s, 3H), 0.90 (s, 3H), 0.81 (s, 3H);
13
.95 (s, 3H), 0.93 (s, 3H), 0.83 (s, 3H), 0.75 (s, 3H); C NMR
(101 MHz, CDCl ): δ (ppm) 218.4, 176.5, 167.2, 151.1, 146.2,
3
1
5
3
1
45.2, 133.7, 130.2, 129.9, 129.2, 125.1, 123.4, 109.6, 55.8, 55.1,
2.8, 50.2, 50.1, 47.5, 47.1, 46.9, 42.6, 40.8, 39.8, 38.4, 37.9,
7.0, 34.8, 34.3, 33.7, 33.6, 31.0, 29.5, 26.8, 25.8, 21.6, 21.2,
13
C NMR (101 MHz, CDCl ): δ (ppm) 218.3, 176.7, 168.9,
3
9.8, 19.7, 16.1, 15.8, 14.7; ESIMS: m/z calculated for
150.9, 145.5, 140.9, 135.4, 129.5, 129.2, 128.5, 128.3, 123.2,
109.7, 55.7, 55.2, 54.9, 54.9, 52.3, 50.2, 50.1, 49.9, 47.5, 46.9,
+
C H N O (M+H) 709.47, found 709.55.
44
60
4
4
4
5.9, 42.7, 40.9, 39.8, 39.4, 38.4, 38.0, 37.1, 35.0, 34.3, 33.8,
Preparation of triazole 19b: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 18 with
azide 9b. Yield: 88%; cream color solid; mp 98 – 101 °C; H
33.7, 31.0, 29.6, 26.8, 25.8, 21.6, 21.2, 19.8, 19.7, 16.2, 15.8,
+
14.8; ESIMS: m/z 820.70 [100%, (M+H) ], HRMS-ESI:
1
+
calculated for C H N O [M+H] 820.5848, found 820.5877.
50
73
7
3
NMR (400 MHz, CDCl ): δ (ppm) 7.62 (s, 1H), 7.33 – 7.46 (m,
3
5
4
4
4
6
3
H), 6.92 (s, 1H), 6.25 (t, J = 5.3 Hz, 1H), 5.31 – 5.42 (m, 2H),
.73 (s, 1H), 4.55 – 4.61 (m, 1H), 4.50 (dd, J = 5.5, 15.0 Hz, 1H),
.44 (dd, J = 5.5, 15.2 Hz, 1H), 3.42 – 3.50 (m, 2H), 3.12 (dt, J =
.3, 11.2 Hz, 1H), 3.00 (brs, 3H), 2.33 – 2.52 (m, 5H), 2.24 (brs,
H), 0.78 – 2.01 (m, 21H), 1.67 (s, 3H), 1.05 (s, 3H), 0.99 (s,
Preparation of triazole 19f: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 18 with
azide 9f. Yield: 91%; white solid; mp 139 – 141 °C; H NMR
1
(400 MHz, CDCl ): δ (ppm) 7.62 (d, J = 15.6 Hz, 1H), 7.60 (s,
3
1H), 7.46 – 7.51 (m, 2H), 7.32 – 7.36 (m, 3H), 6.26 – 6.42 (m,
3H), 4.70 (s, 1H), 4.57 (s, 1H), 4.46 – 4.54 (m, 3H), 4.40 (dd, J =
5.7, 15.0 Hz, 1H), 3.85 – 3.94 (m, 2H), 3.12 (dt, J = 4.4, 11.1 Hz,
1H), 2.32 – 2.51 (m, 3H), 0.75 – 1.95 (m, 21H), 1.64 (s, 3H),
1.02 (s, 3H), 0.98 (s, 3H), 0.92 (s, 3H), 0.88 (s, 3H), 0.80 (s, 3H);
13
H), 0.94 (s, 3H), 0.87 (s, 3H), 0.82 (s, 3H); C NMR (101
MHz, CDCl ): δ (ppm) 218.0, 176.2, 150.8, 145.0, 134.8, 134.0,
3
1
4
3
1
30.2, 128.8 (2C), 128.7, 123.2, 109.4, 55.5, 54.9, 50.0, 49.9,
8.5, 47.2, 46.6, 45.6, 42.4, 40.6, 39.6, 38.2, 37.7, 36.8, 34.7,
4.1, 33.6, 33.4, 30.8, 29.6, 29.3, 26.6, 25.6, 21.4, 20.9, 19.6,
13
C NMR (101 MHz, CDCl ): δ (ppm) 218.5, 176.9, 166.7,
3
+
9.4, 15.9, 15.6, 14.5; ESIMS: m/z 779.70 [100%, (M+H) ],
150.8, 145.7, 141.7, 134.8, 130.1, 129.0, 128.0, 123.6, 120.3,
109.8, 55.8, 55.1, 50.2, 50.1, 49.9, 47.5, 46.9, 42.7, 40.8, 39.8,
38.4, 37.9, 37.1, 35.0, 34.4, 33.8, 33.6, 31.0, 29.6, 26.8, 25.8,
+
HRMS-ESI: calculated for C H N O [M+H] 779.5582, found
7
48
70
6
3
79.5578.
2
1.6, 21.2, 19.8, 19.6, 16.2, 15.9, 14.7; ESIMS: m/z 708.55
+
+
Preparation of triazole 19c: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 18 with
azide 9c. Yield: 90%; cream color solid; mp 134 – 137 °C; H
[100%, (M+H) ], HRMS-ESI: calculated for C H N O [M+H]
44 61 5 3
708.4847, found 708.4833.
1
NMR (400 MHz, CDCl ): δ (ppm) 7.61 (s, 1H), 7.34 – 7.43 (m,
Preparation of Alcohol 20a: To a stirred solution of the
appropriate ketone 19a (180 mg, 0.2 mmol) in methanol at 0 °C,
3
5
H), 6.87 (s, 1H), 6.25 (m, 1H), 5.35 (s, 2H), 4.73 (s, 1H), 4.59
(s, 1H), 4.52 (dd, J = 4.0, 16.0 Hz, 1H), 4.42 (dd, J = 4.0, 16.0
was added NaBH (14 g, 0.4 mmol), and stirred for 2 h at room
4

9
+
temperature. Upon completion of reaction (as monitored by
ESI: calculated for C H N O [M+Na] 846.5980, found
846.5983.
ACCEPTED MANUSCRIPT
50 77 7 3
TLC), the reaction mixture was concentrated in vacuo, diluted
with water and extracted with ethyl acetate (2 x 10.0 mL). The
combined organic extracts were washed with brine (10.0 mL),
dried over anhydrous Na SO , concentrated under vacuum and
Preparation of 20e: Procedure similar to that of 20a. Yield:
89%; cream color solid; mp 130 – 132 °C; H NMR (400 MHz,
1
2
4
purified via column chromatography (silica gel, hexane:ethyl
CDCl ): δ (ppm) 9.86 (t, J = 4.0 Hz, 1H), 7.92 (s, 1H), 7.58 (s,
3
acetate, 1:2) to obtain 155 mg (86%) of pure alcohol 20a as a
1H), 7.20 – 7.39 (m, 5H), 6.26 (t, J = 5.7 Hz, 1H), 4.73 (s, 1H),
4.58 (s, 1H), 4.46 – 4.56 (m, 3H), 4.40 (dd, J = 5.6, 15.0 Hz, 1H),
3.79 – 3.91 (m, 2H), 3.34 (s, 2H), 3.08 – 3.18 (m, 2H), 2.21 –
2.48 (m, 10H), 2.25 (s, 3H), 0.64 – 1.92 (m, 25H), 1.67 (s, 3H),
0.94 (s, 3H), 0.93 (s, 3H), 0.80 (s, 3H), 0.76 (s, 3H), 0.74 (s, 3H);
1
white solid. Mp 123 – 125 °C; H NMR (400 MHz, CDCl ): δ
3
(
(
ppm) 8.06 (s, 1H), 7.72 (s, 1H), 7.57 – 7.64 (m, 2H), 7.39 – 7.46
m, 3H), 6.49 (m, 1H), 5.33 (s, 2H), 4.70 (s, 1H), 4.56 (s, 1H),
4
3
0
0
.50 (dd, J = 5.7, 15.0 Hz, 1H), 4.41 (dd, J = 5.5, 15.0 Hz, 1H),
.82 (s, 3H), 3.03 – 3.22 (m, 2H), 2.34 (dt, J = 3.5, 12.7 Hz, 1H),
.59 – 2.02 (m, 24H), 1.64 (s, 3H), 0.91 (s, 3H), 0.90 (s, 3H),
13
C NMR (101 MHz, CDCl ): δ (ppm) 176.5, 168.7, 150.8,
3
145.3, 140.8, 135.2, 129.3, 128.9, 128.3, 128.0, 123.1, 109.4,
78.8, 55.6, 55.3, 54.8, 54.6, 52.1, 50.5, 50.1, 49.7, 46.8, 45.7,
42.4, 40.7, 39.2, 38.8, 38.7, 38.2, 37.8, 37.2, 34.7, 34.3, 33.5,
30.8, 29.4, 27.9, 27.4, 25.6, 20.9, 19.4, 18.2, 16.2, 15.8, 15.4,
13
.72 (s, 3H), 0.70 (s, 3H), 0.69 (s, 3H); C NMR (101 MHz,
CDCl ): δ (ppm) 176.5, 167.2, 151.2, 146.2, 145.2, 133.8, 130.2,
3
1
5
3
1
7
29.9, 129.2, 125.1, 123.3, 109.6, 79.1, 55.8, 55.5, 52.8, 50.8,
0.3, 47.1, 47.0, 42.6, 40.9, 39.0, 38.9, 38.4, 37.9, 37.4, 34.8,
4.5, 33.7, 31.1, 29.6, 28.2, 27.6, 25.8, 21.1, 19.7, 18.5, 16.3,
+
14.6; ESIMS: m/z 822.70 [100%, (M+H) ], HRMS-ESI:
+
calculated for C H N O [M+Na] 844.5824, found 844.5852.
50
75
7
3
+
6.0, 15.6, 14.8; ESIMS: m/z calculated for C H N O (M+H)
44
62
4
4
11.48, found 711.70.
Preparation of 20f: Procedure similar to that of 20a. Yield:
1
9
0%; gray color solid; mp 185 – 187 °C; H NMR (400 MHz,
Preparation of 20b: Procedure similar to that of 20a. Yield:
CDCl ): δ (ppm) 7.63 (d, J = 15.6 Hz, 1H), 7.60 (s, 1H), 7.45 –
3
1
8
5%; cream color solid; mp 118 – 121 °C; H NMR (400 MHz,
7.53 (m, 2H), 7.33 – 7.60 (m, 3H), 6.38 (d, J = 15.6 Hz, 1H),
6.24 – 6.35 (m, 2H), 4.71 (s, 1H), 4.57 (s, 1H), 4.38 – 4.54 (m,
4H), 3.86 – 3.93 (m, 2H), 3.06 – 3.17 (m, 2H), 2.26 – 2.39 (m,
1H), 0.62 – 1.94 (m, 24H), 1.65 (s, 3H), 0.92 (s, 6H), 0.78 (s,
CDCl ): δ (ppm) 7.60 (s, 1H), 7.33 – 7.43 (m, 5H), 6.99 (s, 1H),
3
6
.25 – 6.34 (m, 1H), 5.35 (s, 2H), 4.72 (s, 1H), 4.58 (s, 1H), 4.49
(
(
(
0
1
1
4
2
dd, J = 5.6, 15.1 Hz, 1H), 4.42 (dd, J = 5.6, 15.1 Hz, 1H), 3.61
brs, 2H), 3.05 – 3.18 (m, 5H), 2.29 – 2.77 (m, 10H), 0.63 – 1.94
m, 23H), 1.66 (s, 3H), 0.94 (s, 3H), 0.93 (s, 3H), 0.77 (s, 3H),
13
3H), 0.75 (s, 3H), 0.73 (s, 3H); C NMR (101 MHz, CDCl ): δ
3
(ppm) 176.7, 166.4, 150.7, 145.5, 141.6, 134.6, 129.8, 128.8,
127.8, 123.4, 120.0, 109.4, 78.9, 55.6, 55.3, 50.5, 50.1, 49.7,
46.7, 42.4, 40.7, 39.5, 38.8, 38.7, 38.2, 37.8, 37.1, 34.7, 34.3,
33.4, 30.8, 29.4, 27.9, 27.4, 25.6, 20.9, 19.4, 18.2, 16.1, 15.8,
13
.76 (s, 3H), 0.73 (s, 3H); C NMR (101 MHz, CDCl ): δ (ppm)
3
76.4, 150.8, 135.1, 133.9, 129.8, 128.84, 128.80, 128.78, 123.3,
09.4, 78.9, 55.6, 55.3, 50.5, 50.1, 48.6, 46.8, 44.9, 44.8, 42.4,
0.7, 38.8, 38.7, 38.2, 37.7, 37.1, 34.8, 34.3, 33.5, 30.8, 29.4,
+
15.4, 14.6; ESIMS: m/z calculated for C H N O (M+H)
44
63
5
3
7.9, 27.4, 25.6, 20.9, 19.4, 18.3, 16.1, 15.8, 15.4, 14.6; ESIMS:
710.50, found 710.65.
+
m/z 781.70 [100%, (M+H) ], HRMS-ESI: calculated for
C H N O [M+Na] 803.5558, found 803.5558.
+
Preparation of succinic acid hemiester 21a: A stirred solution
of alcohol 20a (110 mg, 0.1 mmol), DMAP (19 mg, 0.1 mmol),
and succinic anhydride (31 mg, 0.3 mmol) in toluene (4.0 mL)
was refluxed overnight. Upon completion (TLC), the reaction
mixture was concentrated in vacuo, diluted with water, and
extracted with ethyl acetate (2 x 10.0 mL). The combined
extracts were washed with brine (10.0 mL), dried over anhydrous
Na SO , concentrated under reduced pressure, and purified using
48
72
6
3
Preparation of 20c: Procedure similar to that of 20a. Yield:
1
8
8%; white solid; mp 134 – 136 °C; H NMR (400 MHz,
CDCl ): δ (ppm) 7.60 (s, 1H), 7.34 – 7.42 (m, 5H), 6.86 (s, 1H),
3
6
4
3
3
3
.24 (t, J = 5.6 Hz, 1H), 5.34 (s, 2H), 4.73 (s, 1H), 4.58 (s, 1H),
.52 (dd, J = 5.6, 15.1 Hz, 1H), 4.42 (dd, J = 5.6, 15.1 Hz, 1H),
.66 (m, 4H), 3.06 – 3.19 (m, 2H), 2.17 – 2.35 (m, 5H), 2.26 (s,
H), 0.63 – 1.93 (m, 24H), 1.67 (s, 3H), 0.94 (s, 3H), 0.93 (s,
2
4
column chromatography (silica gel, hexanes:ethyl acetate, 1:3) to
13
H), 0.76 (s, 6H), 0.73 (s, 3H); C NMR (101 MHz, CDCl ): δ
obtain 109 mg (87%) of pure succinic acid hemi ester 21a as a
3
1
(ppm) 176.3, 168.8, 150.8, 145.1, 134.6, 133.8, 129.6, 128.8,
white solid. Mp 122 – 125 °C; H NMR (400 MHz, CDCl ): δ
3
1
4
3
1
28.8, 128.7, 123.5, 109.3, 78.8, 55.7, 55.3, 54.7, 50.5, 50.1,
8.5, 46.7, 45.8, 42.4, 40.7, 38.8, 38.7, 38.2, 37.7, 37.1, 34.7,
4.3, 33.5, 30.8, 29.4, 27.9, 27.4, 25.6, 20.9, 19.5, 18.3, 16.1,
(ppm) 8.07 (s, 1H), 7.73 (s, 1H), 7.58 – 7.63 (m, 2H), 7.41 – 7.47
(m, 3H), 6.71 (t, J = 5.9 Hz, 1H), 5.34 (s, 2H), 4.71 (s, 1H), 4.57
(s, 1H), 4.43 – 4.50 (m, 3H), 3.84 (s, 3H), 3.10 (dt, J = 4.3, 11.1
Hz, 1H), 2.57 – 2.69 (m, 4H), 2.32 (dt, J = 3.6, 12.3 Hz, 1H),
0.65 – 1.98 (m, 23H), 1.66 (s, 3H), 0.92 (s, 3H), 0.80 (s, 3H),
+
5.8, 15.4, 14.6; ESIMS: m/z 779.70 [100%, (M+H) ], HRMS-
+
ESI: calculated for C H N O [M+Na] 801.5402, found
8
48
70
6
3
13
01.5443.
0.78 (s, 3H), 0.75 (s, 3H), 0.68 (s, 3H); C NMR (101 MHz,
CDCl ): δ (ppm) 176.6, 176.3, 171.9, 166.9, 150.9, 146.1, 144.9,
3
Preparation of 20d: Procedure similar to that of 20a. Yield:
133.5, 130.0, 129.7, 128.9, 124.7, 123.4, 109.3, 81.4, 55.6, 55.5,
52.6, 50.4, 50.1, 46.9, 46.8, 42.4, 40.6, 38.4, 38.1, 37.8, 37.7,
37.0, 34.2, 34.0, 33.3, 30.8, 29.4, 29.3, 29.1, 27.8, 25.5, 23.6,
20.9, 19.4, 18.1, 16.5, 16.1, 15.7, 14.6; ESIMS: m/z 811.65
1
9
0%; cream color solid; mp 119 – 122 °C; H NMR (400 MHz,
CDCl ): δ (ppm) 9.95 (s, 1H), 7.96 (s, 1H), 7.58 (s, 1H), 7.24 –
3
7
4
3
2
1
0
1
1
4
3
1
.41 (m, 5H), 6.33 (s, 1H), 4.72 (s, 1H), 4.55 – 4.64 (m, 3H),
.51 (dd, J = 5.5, 15.0 Hz, 1H), 4.43 (dd, J = 5.5, 15.0 Hz, 1H),
.69 – 3.83 (m, 2H), 3.33 (s, 2H), 3.06 – 3.20 (m, 2H), 2.29 –
.42 (m, 4H), 2.07 (brs, 3H), 2.07 (s, 6H), 0.63 – 1.95 (m, 25H),
.67 (s, 3H), 0.94 (s, 3H), 0.93 (s, 3H), 0.80 (s, 3H), 0.79 (s, 3H),
+
+
[100%, (M+H) ], HRMS-ESI: calculated for C H N O [M+H]
48
66
4
7
811.5004, found 811.5038.
Preparation of succinic acid hemiester 21b: Procedure similar
13
.74 (s, 3H); C NMR (101 MHz, CDCl ): δ (ppm) 176.4, 168.5,
to that of 21a. Yield: 80%; cream color solid; mp 130 – 132 °C;
3
1
50.8, 144.9, 140.1, 135.6, 130.5, 129.0, 128.2, 127.9, 122.9,
09.4, 78.9, 56.1, 55.6, 55.3, 54.0, 52.5, 50.6, 50.1, 49.2, 46.8,
5.0, 42.4, 41.7, 40.7, 39.9, 38.8, 38.7, 38.2, 37.7, 37.2, 34.7,
4.3, 33.5, 30.9, 29.4, 27.9, 27.4, 25.6, 20.9, 19.4, 18.2, 16.1,
H NMR (400 MHz, CDCl ): δ (ppm) 7.60 (s, 1H), 7.33 – 7.43
3
(m, 5H), 6.99 (s, 1H), 6.29 (m, 1H), 5.33 (s, 2H), 4.72 (s, 1H),
4.58 (s, 1H), 4.39 – 4.50 (m, 3H), 3.55 – 3.70 (m, 2H), 3.07 –
3.18 (m, 1H), 3.05 (brs, 3H), 2.70 (m, 1H), 2.29 – 2.77 (m, 8H),
2.24 – 2.40 (m, 4H), 0.63 – 1.94 (m, 23H), 1.66 (s, 3H), 0.92 (s,
+
5.8, 15.4, 14.6; ESIMS: m/z 824.75 [100%, (M+H) ], HRMS-

1
0
Tetrahedron
13
3
H), 0.80 (s, 6H), 0.76 (s, 3H), 0.75 (s, 3H); C NMR (101
134.5, 129.9, 128.8, 127.9, 123.6, 119.9, 109.5, 81.4, 55.6, 55.4,
50.4, 50.0, 49.8, 46.8, 42.4, 40.7, 39.5, 38.3, 38.1, 37.8, 37.7,
37.0, 34.3, 34.2, 33.3, 30.8, 29.6, 29.3, 27.8, 25.5, 23.6, 20.9,
19.4, 18.1, 16.5, 16.2, 15.7, 14.6; ESIMS: m/z 810.75 [100%,
ACCEPTED MANUSCRIPT
MHz, CDCl ): δ (ppm) 176.5, 176.3, 172.6, 150.8, 145.4, 135.1,
3
1
5
3
2
33.8, 129.8, 129.7, 128.9, 128.7, 113.9, 109.5, 80.9, 55.6, 55.4,
0.4, 50.1, 48.6, 46.8, 43.8, 42.4, 40.7, 38.3, 38.2, 37.8, 37.7,
7.0, 34.6, 34.2, 33.4, 30.8, 30.1, 30.0, 29.7, 29.4, 27.9, 25.5,
+
+
(M+H) ], HRMS-ESI: calculated for C H N O [M+H]
832.4984, found 832.5026.
48
67
5
6
3.6, 20.9, 19.4, 18.1, 16.5, 16.2, 15.8, 14.6; ESIMS: m/z
+
calculated for C H N O (M+H) 881.59, found 881.58.
52
76
6
6
Preparation of N-Propargyl pyrazinylbetulinamide 23: The
title compound was prepared by the reaction of compound 22
(450 mg, 0.9 mmol) and propargyl amine (60 mg, 1.1 mmol) as
per the general amide-coupling procedure A to yield 387 mg
Preparation of succinic acid hemiester 21c: Procedure similar
to that of 21a. Yield: 83%; cream color solid; mp 142 – 145 °C;
1
H NMR (400 MHz, CDCl ): δ (ppm) 7.61 (s, 1H), 7.36 – 7.44
3
1
(m, 5H), 6.88 (s, 1H), 6.58 (t, J = 5.7 Hz, 1H), 5.34 (s, 2H), 4.72
(80%) of 23 as a white solid. Mp 122 – 125 °C; H NMR (400
(s, 1H), 4.58 (s, 1H), 4.43 – 4.50 (m, 3H), 3.73 (brs, 4H) 3.12 (dt,
MHz, CDCl ): δ (ppm) 8.38 (s, 1H), 8.25 (s, 1H), 5.77 – 5.89 (m,
3
J = 5.4, 11.0 Hz, 1H), 2.57 – 2.65 (m, 4H), 2.26 – 2.53 (m, 5H),
1H), 4.74 (s, 1H), 4.60 (s, 1H), 3.93 – 4.10 (m, 2H), 3.11 – 3.16
(m, 1H), 3.01 (d, J = 16.6 Hz, 1H), 2.36 – 2.56 (m, 2H), 2.16 –
2.20 (m, 1H), 0.73 – 1.96 (m, 19H), 1.68 (s, 3H), 1.27 (s, 3H),
2
0
.36 (s, 3H), 0.72 – 1.96 (m, 23H), 1.67 (s, 3H), 0.93 (s, 3H),
.82 (s, 3H), 0.81 (s, 3H), 0.77 (s, 3H), 0.74 (s, 3H); C NMR
13
13
(
101 MHz, CDCl ): δ (ppm) 176.6, 175.8, 172.3, 168.9, 150.8,
1.25 (s, 3H), 1.00 (s, 6H), 0.78 (s, 3H); C NMR (101 MHz,
3
1
8
3
2
45.4, 135.1, 133.7, 129.1, 128.9, 128.9, 128.7, 123.5, 109.4,
1.1, 55.6, 55.4, 53.8, 50.4, 50.1, 48.6, 46.8, 44.7, 42.4, 40.7,
8.4, 38.1, 37.8, 37.7, 37.0, 34.5, 34.2, 33.4, 30.8, 29.8, 29.6,
9.6, 29.4, 27.9, 25.5, 23.6, 20.9, 19.4, 18.1, 16.5, 16.2, 15.8,
CDCl ): δ (ppm) 176.2, 159.8, 151.0, 150.8, 142.4, 141.6, 109.7,
3
80.5, 71.2, 55.8, 53.2, 50.3, 49.0, 48.8, 46.8, 42.7, 40.8, 39.7,
38.3, 37.9, 36.9, 33.7, 33.6, 31.7, 31.0, 29.6, 29.2, 25.8, 24.2,
21.7, 20.3, 19.8, 16.3, 15.9, 14.8; ESIMS: m/z calculated for
+
+
1
4.6; ESIMS: m/z calculated for C H N O (M+H) 879.57,
C H N O (M+H) 528.81, found 528.60.
52
74
6
6
35 49
3
found 879.65.
Preparation of triazole 24a: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 23 with
azide 9a. Yield: 82%; white solid; mp 127 – 129 °C; H NMR
Preparation of succinic acid hemiester 21d: Procedure similar
to that of 21a. Yield: 82%; white solid; mp 114 – 116 °C; H
1
1
NMR (400 MHz, CDCl ): δ (ppm) 9.68 (s, 1H), 7.91 (s, 1H),
(400 MHz, CDCl ): δ 8.32 (s, 1H), 8.20 (s, 1H), 7.98 (s, 1H),
3
3
7
=
.60 (s, 1H), 7.27 – 7.39 (m, 3H), 7.20 – 7.25 (m, 2H), 6.66 (t, J
5.5 Hz, 1H), 4.72 (s, 1H), 4.58 (s, 1H), 4.40 – 4.57 (m, 5H),
7.68 (s, 1H), 7.49 – 7.60 (m, 2H), 7.45 – 7.29 (m, 3H), 6.30 (brs,
1H), 5.29 (s, 2H), 4.68 (s, 1H), 4.55 (s, 1H), 4.37 – 4.49 (m, 2H),
3.78 (s, 3H), 3.07 (dt, J = 4.3, 11.2 Hz, 1H), 2.93 (d, J = 16.4 Hz,
1H), 2.30 – 2.43 (m, 2H), 0.61 – 1.93 (m, 19H), 1.62 (s, 3H),
3
2
0
0
1
1
5
3
2
.73 – 3.85 (m, 2H), 3.34 (s, 2H), 3.12 (dt, J = 4.2, 10.8 Hz, 1H),
.40 – 2.68 (m, 8H), 2.34 (m, 1H), 2.28 (s, 6H), 2.05 (s, 3H),
.72 – 1.98 (m, 23H), 1.66 (s, 3H), 0.92 (s, 3H), 0.81 (s, 9H),
1.20 (s, 3H), 1.15 (s, 3H), 0.91 (s, 3H), 0.73 (s, 3H), 0.66 (s, 3H);
13
13
.77 (s, 3H); C NMR (101 MHz, CDCl ): δ (ppm) 176.5 (2C),
C NMR (101 MHz, CDCl ): δ 176.5, 167.2, 159.8, 151.1,
3
3
72.7, 168.5, 150.8, 145.3, 140.3, 135.5, 130.3, 129.0, 128.3,
27.9, 123.0, 109.4, 80.9, 55.6, 55.4, 55.2, 53.9, 53.5, 53.1, 50.6,
0.5, 50.1, 49.4, 46.7, 44.1, 42.4, 41.4, 40.7, 39.6, 38.4, 38.2,
7.8, 37.7, 37.1, 34.6, 34.2, 33.4, 30.9, 29.4, 27.9, 25.5, 23.7,
150.9, 146.2, 142.5, 141.7 (2C), 133.7 (2C), 130.2, 129.9, 129.2,
125.0, 109.7, 55.9, 53.2, 52.8, 50.3, 49.0, 48.9, 47.1, 46.9, 42.7,
40.8, 39.6, 38.4, 38.0, 36.9, 34.9, 33.7, 33.5, 31.7, 31.1, 29.6,
25.8, 24.2, 21.6, 20.3, 19.8, 16.3, 15.6, 14.8; ESIMS: m/z
+
0.9, 19.4, 18.1, 16.5, 16.2, 15.8, 14.6; ESIMS: m/z calculated
calculated for C H N O (M+H) 745.48, found 745.45.
46
60
6
3
+
for C H N O (M+H) 924.63, found 924.70.
54
81
7
6
Preparation of triazole 24b: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 23 with
azide 9b. Yield: 84%; white solid; mp 131 – 133 °C; H NMR
Preparation of succinic acid hemiester 21e: Procedure similar
1
to that of 21a. Yield: 84%; cream color solid; mp 132 – 134 °C;
1
H NMR (400 MHz, CDCl ): δ 9.64 (m, 1H), 7.94 (s, 1H), 7.64
(400 MHz, CDCl ): δ 8.39 (d, J = 2.4 Hz, 1H), 8.26 (d, J = 2.4
3
3
(
s, 1H), 7.28 – 7.40 (m, 3H), 7.19 – 7.23 (m, 2H), 6.52 (t, J = 5.0
Hz, 1H), 7.63 (s, 1H), 7.29 – 7.48 (m, 5H), 6.95 (brs, 1H), 6.23 –
6.30 (m, 1H), 5.31 – 5.39 (m, 2H), 4.76 (s, 1H), 4.61 (s, 1H),
4.51 (dd, J = 5.4, 15.1 Hz, 1H), 4.44 (dd, J = 5.6, 15.2 Hz, 1H),
3.39 – 3.54 (m, 2H), 3.13 (dt, J = 4.2, 10.9 Hz, 1H), 2.92 – 3.05
(m, 4H), 2.33 – 2.51 (m, 4H), 2.12 – 2.31 (brs, 6H), 0.73 – 1.97
(m, 19H), 1.69 (s, 3H), 1.28 (s, 3H), 1.25 (s, 3H), 0.99 (s, 3H),
Hz, 1H), 4.72 (s, 1H), 4.59 (s, 1H), 4.38 – 4.51 (m, 5H), 3.88 –
3
2
1
.92 (m, 2H), 3.38 (s, 2H), 3.11 (dt, J = 4.4, 11.0 Hz, 1H), 2.40 –
.76 (m, 10H), 2.44 (s, 3H), 2.33 (m, 3H), 0.71 – 1.95 (m, 23H),
.67 (s, 3H), 0.93 (s, 3H), 0.81 (s, 6H), 0.79 (s, 3H), 0.75 (s, 3H);
13
C NMR (101 MHz, CDCl ): δ 176.7 (2C), 172.6, 168.5, 150.7,
3
13
1
8
4
3
1
9
45.7, 141.5, 134.9, 128.9, 128.7, 128.5, 128.3, 123.5, 109.5,
0.9, 55.6, 55.4, 54.2, 53.3, 50.6, 50.4, 50.1, 50.0, 46.8, 44.0,
2.4, 40.7, 39.0, 38.3, 38.2, 37.8, 37.7, 37.1, 34.6, 34.3, 33.4,
0.8, 29.7, 29.4, 27.9, 25.5, 23.6, 20.9, 19.4, 18.1, 16.5, 16.2,
0.86 (s, 3H), 0.75 (s, 3H); C NMR (101 MHz, CDCl ): δ 176.2,
3
159.6, 150.8, 150.7, 145.0, 142.2, 141.4 (2C), 134.8, 134.0,
130.2, 128.8, 128.7, 123.2, 109.4, 55.6, 53.0, 50.0, 48.8, 48.6,
48.6, 46.6, 45.6, 42.4, 40.6, 39.4, 38.2, 37.7, 36.7, 34.7, 33.4,
33.3, 31.4, 30.9, 29.4, 25.6, 24.0, 21.4, 20.0, 19.5, 16.0, 15.4,
+
5.8, 14.6; ESIMS: m/z calculated for C H N O (M+H)
54
79
7
6
+
22.62, found 922.75.
14.6; ESIMS: m/z calculated for C H N O (M+H) 815.57,
50
70
8
2
found 815.30.
Preparation of succinic acid hemiester 21f: Procedure similar
to that of 21a. Yield: 81%; yellow solid; mp 141 – 144 °C; H
1
Preparation of triazole 24c: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 23 with
azide 9c. Yield: 84%; white solid; mp 137 – 140 °C; H NMR
NMR (400 MHz, CDCl ): δ (ppm) 7.64 (d, J = 15.6 Hz, 1H),
3
1
7
6
4
2
2
0
.63 (s, 1H), 7.46 – 7.52 (m, 2H), 7.34 – 7.38 (m, 3H), 6.60 –
.69 (m, 1H), 6.40 (d, J = 15.5 Hz, 1H), 6.26 – 6.34 (m, 1H),
.70 (s, 1H), 4.57 (s, 1H), 4.39 – 4.54 (m, 5H), 3.80 – 3.97 (m,
H), 3.09 (dt, J = 4.1, 11.0 Hz, 1H), 2.59 – 2.70 (m, 4H), 2.22 –
.33 (m, 1H), 0.66 – 1.91 (m, 23H), 1.65 (s, 3H), 0.91 (s, 3H),
(400 MHz, CDCl ): δ (ppm) 8.33 (d, J = 2.4 Hz, 1H), 8.20 (d, J =
3
2.4 Hz, 1H), 7.55 (s, 1H), 7.29 – 7.41 (m, 5H), 6.80 (s, 1H), 6.20
- 6.30 (m, 1H), 5.35 (s, 2H), 4.70 (s, 1H), 4.56 (s, 1H), 4.46 (dd,
J = 5.5, 15.0 Hz, 1H), 4.36 (dd, J = 5.5, 15.0 Hz, 1H), 3.60 (brs,
4H), 3.12 (dt, J = 4.3, 11.2 Hz, 1H), 3.01 (d, J = 16.6 Hz, 1H),
2.05 – 2.39 (m, 9H), 0.64 – 1.89 (m, 19H), 1.63 (s, 3H), 1.22 (s,
13
.80 (s, 3H), 0.79 (s, 6H), 0.71 (s, 3H); C NMR (101 MHz,
CDCl ): δ (ppm) 176.9 (2C), 172.2, 166.6, 150.7, 145.3, 141.8,
3

1
1
1
3
1
3
H), 1.19 (s, 3H), 0.93 (s, 3H), 0.78 (s, 3H), 0.69 (s, 3H);
C
of 26 as a yellow solid. Mp 251 – 254 °C; H NMR (400
ACCEPTED MANUSCRIPT
NMR (101 MHz, CDCl ): δ (ppm) 176.6, 169.0, 159.8, 151.0,
MHz, DMSO-d ): δ (ppm) 10.64 (s, 1H), 7.98 (s, 1H), 7.26 (d, J
3
6
1
1
4
3
50.9, 145.3, 142.5, 141.7 (2C), 134.8, 134.1, 129.9, 129.0,
28.9, 123.7, 109.7, 55.8, 54.9, 53.2, 50.3, 49.0, 48.9, 48.8, 46.9,
6.2, 42.7, 40.8, 39.6, 38.4, 37.9, 36.9, 34.9, 33.7, 33.5, 31.7,
= 7.7 Hz, 1H), 7.20 (d, J = 7.8 Hz, 1H), 6.81 – 6.98 (m, 2H), 4.68
(s, 1H), 4.55 (s, 1H), 3.65 – 3.92 (m, 2H), 2.99 – 3.12 (m, 1H),
2.73 (d, J = 15.2 Hz, 1H), 2.50 – 2.63 (m, 1H), 2.10 – 2.18 (m,
1H), 2.03 (d, J = 15.1 Hz, 1H), 0.67 – 1.77 (m, 19H), 1.64 (s,
1.1, 29.6, 25.8, 24.2, 21.6, 20.3, 19.8, 16.3, 15.6, 14.8; ESIMS:
+
m/z 835.75 [100%, (M+Na) ], HRMS-ESI: calculated for
C H N O [M+H] 813.5538, found 813.5503.
3H), 1.24 (s, 3H), 1.13 (s, 3H), 0.95 (s, 3H), 0.92 (s, 3H), 0.77 (s,
+
13
3H); C NMR (101 MHz, DMSO-d ): δ (ppm) 176.1, 151.5,
50
68
8
2
6
1
41.9, 136.9, 128.3, 120.6, 118.4, 117.9, 111.1, 109.9, 105.5,
Preparation of triazole 24d: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 23 with
azide 9d. Yield: 85%; white solid; mp 112 – 114 °C; H NMR
82.6, 72.5, 55.6, 53.8, 50.3, 49.6, 46.9, 42.6, 41.0, 40.8, 39.6,
38.5, 38.0, 37.6, 34.6, 33.9, 32.7, 31.1, 30.9, 29.6, 28.5, 26.1,
23.3, 21.8, 19.7, 19.5, 16.9, 16.3, 15.0; ESIMS: m/z calculated
1
+
(
400 MHz, CDCl ): δ (ppm) 9.96 (s, 1H), 8.39 (d, J = 2.3 Hz,
for C H N O (M-H) 563.40, found 563.50.
3
39 52
2
1
7
4
4
3
2
1
0
1
1
5
3
2
H), 8.26 (d, J = 2.4 Hz, 1H), 7.95 (s, 1H), 7.59 (s, 1H), 7.28 –
.39 (m, 3H), 7.21 – 7.25 (m, 2H), 6.37 (s, 1H), 4.75 (s, 1H),
.61 (s, 1H), 4.60 – 4.55 (m, 2H), 4.50 (dd, J = 5.4, 15.1 Hz, 1H),
.44 (dd, J = 5.4, 15.1 Hz, 1H), 3.82 – 3.65 (m, 2H), 3.33 (s, 2H),
.13 (dt, J = 4.3, 11.2 Hz, 1H), 3.01 (d, J = 16.6 Hz, 1H), 2.51 –
.26 (m, 6H), 2.14 (s, 6H), 2.06 (s, 3H), 0.76 – 1.97 (m, 19H),
.69 (s, 3H), 1.28 (s, 3H), 1.26 (s, 3H), 0.98 (s, 3H), 0.88 (s, 3H),
Preparation of triazole 27a: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 26 with
azide 9a. Yield: 80%; tan color solid; mp 133 – 136 °C; H
1
NMR (400 MHz, CDCl ): δ (ppm) 8.01 (s, 1H), 7.48 – 7.76 (m,
3
4H), 7.34 – 7.42 (m, 3H), 7.27 – 7.45 (m, 2H), 6.96 – 7.07 (m,
2H), 6.26 (m, 1H), 5.30 (s, 2H), 4.71 (s, 1H), 4.56 (s, 1H), 4.45 –
4.53 (m, 1H), 4.39 (dd, J = 5.9, 14.9 Hz, 1H), 3.79 (s, 3H), 3.12
(dt, J = 6.2, 12.1 Hz, 1H), 2.74 (d, J = 14.9 Hz, 1H), 2.33 – 2.43
(m, 1H), 2.05 (d, J = 14.8 Hz, 1H), 0.68 – 1.98 (m, 19H), 1.64 (s,
3H), 1.20 (s, 3H), 1.08 (s, 3H), 0.93 (s, 3H), 0.76 (s, 3H), 0.73 (s,
13
.78 (s, 3H); C NMR (101 MHz, CDCl ): δ (ppm) 176.4, 168.4,
3
59.6, 150.8, 150.7, 144.9, 142.2, 141.4, 140.1, 135.6, 130.5,
28.9, 128.2, 127.9, 122.9, 109.4, 56.1, 55.6, 53.9, 53.0, 52.5,
0.0, 49.2, 48.8, 48.6, 46.6, 45.0, 42.4, 41.6, 40.6, 39.9, 39.4,
8.2, 37.7, 36.7, 34.7, 33.4, 33.3, 31.4, 30.9, 29.3, 25.6, 24.0,
13
3H); C NMR (101 MHz, CDCl ): δ (ppm) 175.3, 165.9, 149.9,
3
1.4, 20.0, 19.5, 16.1, 15.4, 14.7; ESIMS: m/z calculated for
144.9, 143.9, 139.8, 135.1, 132.5, 128.9, 128.7, 127.9, 127.3,
123.8, 122.1, 119.8, 117.8, 116.8, 109.3, 108.4, 105.9, 54.6, 52.2,
+
C H N O (M+H) 858.61, found 858.75.
52
75
9
2
5
1.5, 49.1, 49.0, 48.4, 45.8, 41.4, 39.7, 37.2, 37.2, 36.9, 36.2,
Preparation of triazole 24e: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 23 with
azide 9e. Yield: 87%; cream color solid; mp 132 – 135 °C; H
33.8, 33.1, 32.5, 29.8, 29.8, 28.7, 28.4, 24.7, 22.1, 20.4, 18.4,
+
18.2, 15.3, 14.6, 13.6; ESIMS: m/z 782.85 [100%, (M+H) ];
1
+
HRMS-ESI: calculated for C H N O [M+H] 782.5004, found
50
63
5
3
NMR (400 MHz, CDCl ): δ (ppm) 9.79 (s, 1H), 8.33 (s, 1H),
782.4986.
3
8
(
3
.20 (s, 1H), 7.85 (s, 1H), 7.58 (s, 1H), 7.15 – 7.30 (m, 5H), 6.37
s, 1H), 4.69 (s, 1H), 4.55 (s, 1H), 4.34 – 4.51 (m, 4H), 3.69 –
.91 (m, 2H), 3.28 (s, 2H), 3.00 – 3.14 (m, 1H), 2.95 (d, J = 16.4
Hz, 1H), 2.14 – 2.52 (m, 13H), 0.68 – 1.93 (m, 19H), 1.63 (s,
Preparation of triazole 27f: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 26 with
1
azide 9f. Yield: 81%; tan color solid; mp 167 – 169 °C; H NMR
3
3
1
1
4
3
1
H), 1.21 (s, 3H), 1.20 (s, 3H), 0.92 (s, 3H), 0.79 (s, 3H), 0.72 (s,
(400 MHz, CDCl ): δ (ppm) 7.75 (s, 1H), 7.64 (d, J = 15.4 Hz,
3
13
H); C NMR (101 MHz, CDCl ): δ (ppm) 176.7, 168.9, 159.8,
1H), 7.61 (s, 1H), 7.44 - 7.54 (m, 2H), 7.33 – 7.41 (m, 4H), 7.28
– 7.32 (m, 1H), 7.00 – 7.13 (m, 2H), 6.38 (d, J = 15.7 Hz, 1H),
6.29 – 6.36 (m, 1H), 6.20 – 6.28 (m, 1H), 4.75 (s, 1H), 4.61 (s,
1H), 4.37 – 4.57 (m, 4H), 3.87 – 3.96 (m, 2H), 3.05 – 3.20 (m,
1H), 2.80 (d, J = 15.1 Hz, 1H), 2.33 – 2.50 (m, 1H), 2.11 (d, J =
15.1 Hz, 1H), 0.76 – 1.96 (m, 19H), 1.68 (s, 3H), 1.26 (s, 3H),
3
50.9, 150.8, 145.6, 142.5, 141.7, 141.0, 135.4, 129.4, 129.2,
28.5, 128.3, 123.4, 109.8, 55.8, 54.9, 54.8, 53.2, 52.2, 50.2,
9.9, 48.9, 48.8, 46.9, 45.8, 42.7, 40.8, 39.7, 39.4, 38.4, 38.0,
6.9, 34.9, 33.7, 33.5, 31.7, 31.1, 29.6, 25.8, 24.3, 21.6, 20.3,
9.7, 16.4, 15.6, 14.8; ESIMS: m/z calculated for C H N O
52
73
9
2
+
13
(M+H) 856.60, found 856.70.
1.17 (s, 3H), 0.99 (s, 3H), 0.85 (s, 3H), 0.83 (s, 3H); C NMR
(
101 MHz, CDCl ): δ (ppm) 177.0, 166.7, 150.9, 145.7, 141.9,
3
Preparation of triazole 24f: Procedure similar to that of 19a.
This compound was prepared by the reaction of alkyne 23 with
azide 9f. Yield: 88%; white solid; mp 150 – 152 °C; H NMR
141.2, 136.4, 134.8, 130.1, 129.1, 128.5, 128.1, 123.6, 121.1,
120.2, 119.0, 118.1, 110.6, 109.9, 107.0, 55.9, 53.5, 50.3, 49.9,
49.6, 47.0, 42.7, 41.0, 39.8, 38.5, 38.2, 37.5, 35.0, 34.4, 33.8,
33.7, 31.0, 29.7, 25.9, 23.4, 21.7, 19.6, 19.4, 16.6, 15.9, 14.9;
1
(
400 MHz, CDCl ): δ (ppm) 8.39 (s, 1H), 8.26 (s, 1H), 7.63 (d, J
3
+
=
3
(
15.6 Hz, 1H), 7.61 (s, 1H), 7.45 – 7.52 (m, 2H), 7.30 – 7.39 (m,
H), 6.26 – 6.42 (m, 3H), 4.74 (s, 1H), 4.60 (s, 1H), 4.37 – 4.56
m, 4H), 3.84 – 3.95 (m, 2H), 3.05 – 3.17 (m, 1H), 3.00 (d, J =
ESIMS: m/z calculated for C H N O (M+Na) 803.50, found
50
64
6
2
803.55.
1
3
3
1
1
4
3
6.5 Hz, 1H), 2.34 – 2.49 (m, 2H), 0.72 – 1.99 (m, 19H), 1.67 (s,
H), 1.26 (s, 3H), 1.25 (s, 3H), 0.97 (s, 3H), 0.84 (s, 3H), 0.77 (s,
Preparation of N-2-azidoethyl betulonamide 28: The title
compound was prepared by the reaction of betulonic acid 17 (850
mg, 1.87 mmol) and 2-azidoethylamine (241 mg, 2.80 mmol)
using the general amide-coupling procedure A to furnish the
13
H); C NMR (101 MHz, CDCl ): δ (ppm) 176.9, 166.7, 159.8,
3
50.9, 150.8, 145.7, 142.5, 141.9, 141.7, 134.8, 130.1, 129.0,
28.1, 123.7, 120.3, 109.8, 55.8, 53.2, 50.2, 49.9, 49.0, 48.8,
6.9, 42.7, 40.8, 39.8, 39.7, 38.4, 38.0, 36.9, 35.0, 33.6, 33.5,
1
amide 28 (722 mg, 74%) as a white solid. Mp 82 – 84 °C; H
NMR (400 MHz, CDCl ): δ 5.94 (s, 1H), 4.72 (s, 1H), 4.58 (s,
3
1.7, 31.1, 29.6, 25.8, 24.3, 21.7, 20.2, 19.7, 16.4, 15.7, 14.8;
1H), 3.32 – 3.48 (m, 4H), 3.03 – 3.15 (m, 1H), 2.35 – 2.49 (m,
3H), 0.81 – 1.95 (m, 21H), 1.66 (s, 3H), 1.05 (s, 3H), 1.00 (s,
+
ESIMS: m/z calculated for C H N O (M+H) 744.50, found
7
46
61
7
2
13
44.55.
3H), 0.96 (s, 6H), 0.91 (s, 3H); C NMR (101 MHz, CDCl ): δ
3
2
18.4, 176.7, 150.9, 109.7, 55.9, 55.2, 51.4, 50.2, 50.2, 47.5,
Preparation of N-Propargyl indolylbetulinamide 26: The title
compound was prepared by the reaction of compound 25 (300
mg, 0.6 mmol) and propargyl amine (37.5 mg, 0.7 mmol) as per
the general amide-coupling procedure A to yield 224 mg (71%)
46.9, 42.7, 40.9, 39.8, 38.9, 38.4, 38.0, 37.1, 34.4, 33.9, 31.0,
29.6, 26.8, 25.8, 21.7, 21.2, 19.8, 19.7, 16.2, 16.1, 14.8; ESIMS:
+
m/z calculated for C H N O (M-H) 521.39, found 521.20.
32
50
4
2

1
2
Tetrahedron
ACCEPTED MANUSCRIP
+
Preparation of triazole 29a: Procedure similar to that of 19a.
calculated for C H N O [M+Na] 846.5980, found 846.5954.
50 77 7 3
T
This compound was prepared by the reaction of azide 28 with
alkyne 13a. Yield: 79%; off white solid; mp 108 – 111 °C; H
NMR (400 MHz, CDCl ): δ (ppm) 7.86 (s, 1H), 7.61 (s, 1H),
1
Preparation of 30b: Procedure similar to that of 20a. Yield:
1
89%; pale orange solid; mp 129 – 131 °C; H NMR (400 MHz,
3
7
3
.20 – 7.40 (m, 5H), 6.40 (brs, 1H), 4.73 (s, 1H), 4.53 – 4.61 (m,
H), 4.35 – 4.50 (m, 2H), 3.69 – 3.79 (m, 2H), 3.41 (s, 2H), 3.07
CDCl ): δ (ppm) 10.22 (t, J = 5.3 Hz, 1H), 7.95 (s, 1H), 7.60 (s,
3
1H), 7.33 – 7.37 (m, 2H), 7.28 – 7.31 (m, 1H), 7.21 – 7.24 (m,
2H), 6.00 – 6.10 (m, 1H), 4.73 (s, 1H), 4.56 – 4.60 (m, 3H), 4.39
– 4.52 (m, 2H), 3.77 (q, J = 5.7 Hz, 2H), 3.42 (s, 2H), 3.16 (dd, J
= 5.0, 11.2 Hz, 1H), 3.06 (dt, J = 3.8, 11.1 Hz, 1H), 2.24 – 2.61
(m, 10H), 2.29 (s, 3H), 0.65 – 1.92 (m, 23H), 1.66 (s, 3H), 0.94
(m, 1H), 2.27 – 2.63 (m, 12H), 2.14 (s, 3H), 0.83 – 1.99 (m,
2
2H), 1.66 (s, 3H), 1.04 (s, 3H), 1.00 (s, 3H), 0.96 (s, 3H), 0.94
13
(s, 3H), 0.91 (s, 3H); C NMR (101 MHz, CDCl ): δ (ppm)
3
2
1
4
3
2
18.1, 176.9, 168.3, 150.7, 145.7, 139.5, 135.6, 131.3, 128.9,
28.3, 127.8, 123.4, 109.4, 56.1, 55.6, 54.9, 53.3, 53.1, 49.9,
9.8, 47.3, 46.5, 44.9, 42.4, 41.8, 40.7, 39.6, 39.3, 38.2, 37.6,
6.9, 35.3, 34.1, 33.7, 33.4, 30.8, 29.6, 29.4, 26.6, 25.6, 21.5,
0.9, 19.6, 19.4, 15.9, 15.9, 14.5; ESIMS: m/z 822.60 [100%,
13
(s, 6H), 0.90 (s, 3H), 0.80 (s, 3H), 0.74 (s, 3H); C NMR (101
MHz, CDCl ): δ (ppm) 176.9, 168.1, 150.6, 145.2, 140.5, 135.3,
3
129.4, 128.9, 128.3, 128.0, 122.9, 109.5, 78.8, 55.7, 55.4, 54.7,
54.6, 51.9, 50.6, 49.9, 49.6, 46.6, 45.6, 42.4, 40.7, 39.1, 38.8,
38.7, 38.2, 37.6, 37.2, 35.0, 34.4, 33.4, 30.8, 29.6, 29.4, 27.9,
+
+
(M+H) ]; HRMS-ESI: calculated for C H N O [M+H]
50 75 7 3
8
22.6004, found 822.6025.
27.4, 25.6, 20.9, 19.4, 18.3, 16.1, 15.4, 14.6; ESIMS: m/z 822.80
+
[
100%, + (M+H) ]; HRMS-ESI: calculated for C H N O
5
0
75
7
3
Preparation of triazole 29b: Procedure similar to that of 19a.
This compound was prepared by the reaction of azide 28 with
alkyne 13b. Yield: 86%; cream color solid; mp 115 – 117 °C; H
[M+Na] 844.5824, found 844.5885.
1
Preparation of 30c: Procedure similar to that of 20a. Yield:
1
NMR (400 MHz, CDCl ): δ (ppm) 10.23 (m, 1H), 7.94 (s, 1H),
91%; white solid; mp 163 – 166 °C; H NMR (400 MHz,
3
7
=
4
4
.60 (s, 1H), 7.29 – 7.37 (m, 3H), 7.21 – 7.25 (m, 2H), 6.05 (t, J
5.7 Hz, 1H), 4.73 (s, 1H), 4.59 (s, 2H), 4.57 (s, 1H), 4.38 –
.52 (m, 2H), 3.78 (q, J = 5.7 Hz, 2H), 3.41 (s, 2H), 3.07 (dt, J =
CDCl ): δ (ppm) 7.63 (d, J = 15.6 Hz, 1H), 7.59 (s, 1H), 7.46 –
3
7.50 (m, 2H), 7.33 – 7.39 (m, 3H), 6.50 (brs, 1H), 6.41 (d, J =
15.6 Hz, 1H), 6.10 (brs, 1H), 4.72 (s, 1H), 4.62 (d, J = 5.8 Hz,
2H), 4.57 (s, 1H), 4.37 – 4.52 (m, 2H), 3.76 (q, J = 5.6 Hz, 2H),
3.11 – 3.20 (m, 1H), 3.04 (dt, J = 4.0, 12.0 Hz, 1H), 2.42 (dt, J =
3.6, 12.7 Hz, 1H), 0.60 – 1.89 (m, 24H), 1.64 (s, 3H), 0.93 (s,
.3, 11.2 Hz, 1H), 2.31 – 2.62 (m, 10H), 3.69 (s, 3H), 0.87 – 1.66
(m, 22H), 1.66 (s, 3H), 1.04 (s, 3H), 1.00 (s, 3H), 0.95 (s, 6H),
13
0
1
1
4
3
1
.91 (s, 3H); C NMR (101 MHz, CDCl ): δ (ppm) 218.1, 176.9,
3
1
3
68.2, 150.6, 145.3, 140.4, 135.3, 129.5, 128.9, 128.3, 128.0,
22.9, 109.5, 55.6, 54.9, 54.8, 54.6, 52.1, 49.9, 49.9, 49.6, 47.3,
6.5, 45.7, 42.4, 40.6, 39.6, 39.1, 38.2, 37.6, 36.9, 35.0, 34.1,
3.6, 33.4, 30.7, 29.4, 26.6, 25.6, 21.4, 20.9, 19.6, 19.4, 15.9,
3H), 0.92 (s, 3H), 0.89 (s, 3H), 0.79 (s, 3H), 0.73 (s, 3H);
C
NMR (101 MHz, CDCl ): δ (ppm) 177.0, 166.2, 150.6, 144.9,
3
141.3, 134.6, 129.8, 128.8, 127.8, 123.2, 120.4, 109.5, 78.9, 55.6,
55.3, 50.6, 49.9, 49.7, 46.6, 42.4, 40.7, 39.2, 38.8, 38.7, 38.2,
37.6, 37.2, 34.9, 34.4, 33.4, 30.8, 29.4, 28.0, 27.4, 25.5, 20.9,
19.4, 18.3, 16.1, 15.4, 14.6; ESIMS: m/z 710.65 [100%,
+
5.9, 14.5; ESIMS: m/z 820.60 [100%, (M+H) ]; HRMS-ESI:
+
calculated for C H N O [M+H] 820.5848, found 820.5855.
50
73
7
3
+
+
(
M+H) ]; HRMS-ESI: calculated for C H N O [M+Na]
44 63 5 3
Preparation of triazole 29c: Procedure similar to that of 19a.
This compound was prepared by the reaction of azide 28 with
alkyne 13c. Yield: 81%; white solid; mp 155 – 158 °C; H NMR
732.4823, found 732.4836.
1
Cell Viability Assay: Human pancreatic cancer MIAPaCa-2
cells were purchased from ATCC. Cells were maintained in D-
MEM supplemented with 10% FBS, 2.5% horse serum, and 1%
(
400 MHz, CDCl ): δ (ppm) 7.62 (d, J = 15.9 Hz, 1H), 7.60 (s,
3
1
1
4
3
1
3
H), 7.44 – 7.51 (m, 2H), 7.31 – 7.39 (m, 3H), 6.55 – 6.66 (m,
H), 6.43 (d, J = 15.6 Hz, 1H), 6.12 – 6.20 (m, 1H), 4.71 (s, 1H),
.62 (d, J = 5.5 Hz, 2H), 4.57 (s, 1H), 4.39 – 4.53 (m, 2H), 3.68 –
.80 (m, 2H), 2.97 – 3.09 (m, 1H), 2.29 – 2.53 (m, 3H), 0.82 –
.90 (m, 21H), 1.64 (s, 3H), 1.03 (s, 3H), 0.99 (s, 3H), 0.93 (s,
Penicillin Streptomycin in a humidified atmosphere of 5% CO at
2
37 °C. Murine breast cancer 4T1 cells were purchased from
ATCC. Cells were maintained in RPMI-1640 supplemented with
10% FBS and 1% Penicillin Streptomycin in a humidified
13
H), 0.92 (s, 3H), 0.90 (s, 3H); C NMR (101 MHz, CDCl ): δ
atmosphere of 5% CO at 37 °C. Cells were seeded in 96 well
3
2
4
(ppm) 218.5, 177.2, 166.4, 150.8, 145.2, 141.6, 134.8, 130.1,
plates at a density of 5 x 10 cells/mL, 100µL/well incubated for
1
4
3
1
7
29.0, 128.0, 123.5, 120.5, 109.8, 55.8, 55.2, 50.2, 50.1, 49.9,
7.5, 46.8, 42.7, 40.9, 39.8, 39.4, 38.4, 37.8, 37.1, 35.3, 34.4,
3.9, 33.6, 30.9, 29.6, 26.8, 25.8, 21.7, 21.2, 19.8, 19.6, 16.2+,
6.2, 14.7; HRMS-ESI: calculated for C H N O [M+H]
18-24 hours, then exposed to compounds starting at a
concentration of 50 µM and serial diluted. Compounds were
added in duplicate and exposed for 72 hours. Each IC50 was
reported as an average of a minimum of three trials. Compounds
were dissolved in DMSO at a concentration of 50 mM and
diluted into the appropriate media 1:1000. The final
concentration of DMSO was kept below 0.1%. Furthermore,
DMSO was added as a negative control. Cellular viability was
determined using MTT (3-(4, 5-dimethylthiazolyl-2)-2, 5-
diphenyltetrazolium bromide). MTT was dissolved in 1X PBS
solution (5 mg/mL) and 10 µL was added to each well. After 4
hours of incubation, cells were lysed with 100 µL of a SDS
(sodium dodecyl sulfate) solution (100 mg/mL of 0.01 N HCl)
and incubated for an additional 4 hours. The absorbance of each
well was then measured using a microplate reader at 570 nm.
Control wells absorbance was defined as 100% viability and all
of the tested compounds were expressed as percentage relative to
the control.
44
61
5
3
08.4847, found 708.4840.
Preparation of 30a: Procedure similar to that of 20a. Yield:
8
CDCl ): δ (ppm) 10.21 (brs, 1H), 7.92 (s, 1H), 7.61 (s, 1H), 7.33
–
1
6%; pale yellow solid; mp 116 – 118 °C; H NMR (400 MHz,
3
7.43 (m, 2H), 7.28 – 7.32 (m, 1H), 7.19 – 7.25 (m, 2H), 6.21
(brs, 1H), 4.73 (s, 1H), 4.58 (s, 1H), 4.53 – 4.57 (m, 2H), 4.33 –
4
5
5
3
3
1
7
4
3
1
.51 (m, 2H), 3.74 (q, J = 5.5 Hz, 2H), 3.38 (s, 2H), 3.16 (dd, J =
.0, 11.2 Hz, 1H), 3.07 (dt, J = 3.7, 11.2 Hz, 1H), 2.36 – 2.51 (m,
H), 2.25 (brs, 6H), 2.12 (s, 3H), 0.64 – 1.91 (m, 24H), 1.66 (s,
H), 0.94 (s, 3H), 0.93 (s, 3H), 0.91 (s, 3H), 0.80 (s, 3H), 0.73 (s,
13
H); C NMR (101 MHz, CDCl ): δ (ppm) 176.9, 168.2, 150.8,
3
45.9, 139.7, 135.7, 130.9, 128.9, 128.2, 127.8, 123.2, 109.4,
8.8, 56.2, 55.6, 55.4, 53.8, 52.8, 50.6, 50.0, 49.6, 46.6, 45.1,
2.4, 41.8, 40.7, 39.2, 38.8, 38.7, 38.2, 37.5, 37.2, 35.3, 34.4,
3.4, 30.8, 29.6, 29.4, 27.9, 27.4, 25.6, 20.9, 19.4, 18.3, 16.1,
+
5.4, 14.7; ESIMS: m/z 824.75 [100%, (M+H) ]; HRMS-ESI:

1
3
Acknowledgments
ACCEPTED MANU(b)
S
M
C
il
A.; Kokpol, U. J. Agri. Food Chem. 1994, 42, 1561-1562.
c) Huang, F. Y.; Chung, B. Y.; Bentley, M. D.; Alford, A. R. J.
R
es
I
,
P
D
T
. H.; Tunsuwan, K.; Chittawong, V.; Hedin, P.
(
We thank the Departments of Chemistry and Biochemistry
and of Biomedical and Translational Sciences at Rowan
University. We also thank the Honors Concentration at Rowan
University for the undergraduate student support (CMH). We
sincerely thank Prof. Venkatram Mereddy (University of
Minnesota) for the helpful discussions. We are thankful to
Channel Therapeutics, Serenity Enterprises, and Avante Pharma
for their financial research support.
Agri. Food Chem. 1995, 43, 2513-2516.
9
1
1
.
Auclair, N.; Kaboorani, A.; Riedl, B.; Landry, V. Industrial
Crops and Prod. 2015, 76, 530-537.
0. Gorbunova, M. N.; Krainova, G. F.; Tolmacheva, I. A.;
Grishko, V. V. Russ. J. Appl. Chem. 2012, 85, 1137-1141.
1. (a) Zhao, J.; Schlaad, H.; Weidner, S.; Antonietti, M. Polym.
Chem. 2012, 3, 1763-1768.
(
b) Jeromenok, J.; Bohlmann, W.; Antonietti, M.; Weber, J.
Macromol. Rapid Commun. 2011, 32, 1846-1851.
2. For general reviews refer:
Supplementary Data
1
(
a) Basavaiah, D.; Reddy, B. S.; Badsara, S. S. Chem. Rev.
Supplementary data associated with this article can be found
in the online version, at XXX.
2010, 110, 5447–5674.
(b) Declerck, V.; Martinez, J.; Lamaty F. Chem. Rev. 2009, 109,
1
-48.
c) Basavaiah, D.; Rao, A. J.; Satyanarayana, T. Chem. Rev.
003, 103, 811-891.
3. For general reviews refer:
(
2
References
1
1
1
2
3
.
.
.
(a) Jager, S.; Trojan, H.; Kopp, T.; Laszczyk, M. N.; Scheffler,
A. Molecules 2009, 14, 2016-2031. (b) Hayek, E. W. H.; Jordis,
U.; Moche, W.; Sauter, F. Phytochem. 1989, 28, 2229-2242.
(a) Jain, A.; Srivastava, S. K. Indian J. Pharm. Sci. 1984, 46,
(a) Banfi, L.; Riva, R. Org. Reactions 2005, 65, 1-141.
(b) Domling, A.; Ugi, I. Angew Chem. Int. Ed. 2000, 39, 3168-
3
210.
4. For general reviews refer:
a) Kacprzak, K.; Skiera, I.; Piasecka, M.; Paryzek, Z. Chem.
Rev. 2016, 116, 5689-5743.
b) Tiwari, V. K.; Mishra, B. B.; Mishra, K. B.; Mishra, N.;
Singh, A. S.; Chen, X. Chem. Rev. 2016, 116, 3086-3240.
c) Alonso, F.; Moglie, Y.; Radivoy, G. Acc. Chem. Res.
015, 48, 2516-2528.
d) Kolb, H. C.; Finn, M. G.; Sharpless, K. B. Angew. Chem.
1
1
61-162. (b) Batta, A. K.; Rangaswami, S. Phytochem. 1973,
2, 214-16.
(
For general reviews refer:
a) Zhang, D. M.; Xu, H. G.; Wang, L.; Li, Y. J.; Sun, P. H.;
(
(
Wu, X. M.; Wang, G. J.; Chen, W. M.; Ye, W. C. Med. Res.
Rev. 2015, 35, 1127-1155.
(
2
(
(
b) Seyed, M. A.; Jantan, I.; Vijayaraghavan, K.; Nasir, S.;
Bukhari, A. Chem. Biol. Drug Design 2015, 87, 517-536.
c) Lee, S. Y.; Kim, H. H.; Park, S. U.; EXCLI J. 2015, 14, 199-
03.
d) Periasamy, G.; Teketelew, G.; Gebrelibanos, M.; Sintayehu,
Int. Ed. 2001, 40, 2004-2021.
(
2
(
1
5. (a) Alam, M. A.; Arora, K.; Gurrapu, S.; Jonnalagadda, S. K.;
Nelson, G. L.; Kiprof, P.; Jonnalagadda, S. C.; Mereddy, V. R.
Tetrahedron. 2016, 72, 3795-3801.
B.; Gebrehiwot, M.; Karim, A.; Geremedhin, G. Arch. Appl.
Sci. Res. 2014, 6, 47-58.
(
b) Suman, P.; Patel, B. P.; Kasibotla, A. V.; Solano, L. N.;
Jonnalagadda, S. C. J. Organomet. Chem. 2015, 798, 125-131.
c) Kumar, J. S.; Alam, M. A.; Gurrapu, S.; Nelson, G. L.;
(
e) Jonnalagadda, S. C.; Corsello, M. A.; Sleet, C. E. Anti-
Cancer Agents Med. Chem. 2013, 13, 1477-1499.
f) Mullauer F. B.; Kessler J. H.; Medema J. P. Anti Cancer
Drugs 2010, 21, 215-227.
(
Williams, M.; Corsello, M. A.; Johnson, J. L.; Jonnalagadda, S.
C.; Mereddy. V. R. J. Het. Chem. 2013, 50, 814-820.
(
(d) Gunasekera, D. S.; Gerold, D. A.; Aalderks, N. A.;
(g) Fulda, S. Int. J. Mol. Sci. 2008, 9, 1096-1107.
Jonnalagadda, S. C.; Kiprof, P.; Zhdankin, V. V.; Reddy, M. V.
R. Tetrahedron 2007, 63, 9401-9405.
(h) Sami, A.; Taru, M.; Salme, K.; Jari, Y. K. Eur. J.
Pharmaceutical Sci. 2006, 29, 1-13.
i) Tolstikov, G. A.; Flekhter, O. B.; Shultz, E. E.; Baltina, L.
A.; Tolstikov, A. G. Chem. Sus. Dev. 2005, 13, 1-29.
j) Yu, D.; Wild, C. T.; Martin, D. E.; Morris-Natschke, S. L.;
Chen, C-H.; Allaway, G. P.; Lee, K-H. Exp. Op. Invest. Drugs.
005, 14, 681-693.
k) Yogeeswari, P.; Sriram, D. Curr. Med. Chem. 2005, 12,
57-666.
(a) Fulda, S.; Kroemer, G. Drug Discovery Today 2009, 14,
85-890.
b) Schmidt, M. L.; Kuzmanoff, K. L.; Ling-Indeck, L.;
Pezzuto, J. M. Eur. J. Cancer 1997, 33, 2007–2010.
c) Pisha, E.; Chai, H.; Lee, I. S.; Chagwedera, T. E.;
(
e) Reddy, V. J.; Chandra, J. S.; Reddy, M. V. R. Org. Biomol.
Chem. 2007, 5, 889-891.
f) Ramachandran, P. V.; Prabhudas, B.; Chandra, J. S.; Reddy,
(
(
(
M. V. R.; Brown, H. C. Tetrahedron Lett. 2004, 45, 1011-1013.
6. (a) Tekkam, S.; Alam, M. A.; Just, M. J.; Johnson, J. L.;
Jonnalagadda, S. C.; Mereddy, V. R. Anti-Cancer Agents in
Med. Chem. 2013, 13, 1514-1530.
1
2
(
6
(
b) Kumar, J. S.; Bashian, C. M.; Corsello, M. A.;
Jonnalagadda, S. C.; Mereddy, V. R. Tetrahedron Lett. 2010,
1, 4482-4485;
c) Jonnalagadda, S. C.; Verga, S. R.; Patel, P. D.; Reddy, A.
4
.
8
(
5
(
V.; Tekkam, S.; Scott, P. M.; Mereddy, V. R. Appl. Organomet.
Chem. 2010, 24, 294-300.
(
Farnsworth, N. R.; Cordell, G. A.; Beecher, C. W. W.; Fong, H.
H. S.; Kinghorn, A. D.; Brown, D. M.; Wani, M. C.; Wall, M.
E.; Hieken, T. J.; Das Gupta, T. K.; Pezzuto, J. M. Nature Med.
1
7. (a) Kumar, J. S.; Jonnalagadda, S. C.; Mereddy, V. R.
Tetrahedron Lett. 2010, 51, 779-782.
(
b) Jonnalagadda, S. C.; Cruz, J.; Connell, R. J.; Scott, P.;
1
995, 1, 1046–1051.
Mereddy, V. R. Tetrahedron Lett. 2009, 50, 4314-4317.
8. Gurrapu, S.; Walsh, W. J.; Brooks, J. M.; Jonnalagadda, S. C.;
Mereddy, V. R. Nat. Prod., Ind. J. 2012, 8, 115-120.
9. (a) Xiao, S.; Wang, Q.; Si, L.; Zhou, X.; Zhang, Y.; Zhang, L.;
Zhou, D. Eur. J. Med. Chem. 2016, 124, 1-9.
5
.
Zuco, V.; Supino, R.; Righetti, S. C.; Cleris, L.; Marchesi, E.;
Gambacorti-Passerini, C.; Formelli, F. Cancer Lett. 2002, 175,
1
1
1
7–25.
6
7
.
.
Yi, J.; Zhu, R.; Wu, J.; Wu, J.; Xia, W.; Zhu, L.; Jiang, W.;
Xiang, S.; Tan, Z. Pharmacological Rep. 2016, 68, 95-100.
(a) Zhao, H.; Zheng, Q.; Hu, X.; Shen, H.; Li, F. Life Sci. 2016,
(
b) Spivak, A. Y.; Gubaidullin, R. R.; Galimshina, Z. R.;
Nedopekina, D. A.; Odinokov, V. N. Tetrahedron 2016, 72,
249-1256.
c) Khan, I.; Guru, S. K.; Rath, S. K.; Chintakindi, P. K.; Singh,
B.; Koul, S.; Bhushan, S.; Sangwan, P. L. Eur. J. Med. Chem.
016, 108, 104-116.
1
44, 185-193.
1
(
(b) Zhao, H.; Liu, Z.; Liu, W.; Han, X.; Zhao, M. Intl.
Immunopharmacol. 2016, 30, 50-56.
(a) Lugemwa, F. N.; Huang, F. Y.; Bentley, M. D.; Mendel, M.
J.; Alford, A. R. J. Agri. Food Chem. 1990, 38, 493-496.
8
.
2

1
4
Tetrahedron
ACCEPTED MANUSCRIPT
(
d) Bhunia, D.; Pallavi, P. M. C.; Bonam, S. R.; Reddy, S. A.;
(b) Soural, M.; Hodon, J.; Dickinson, N. J.; Sidova, V.; Gurska,
Verma, Y.; Halmuthur, M. S. K. Arch. Pharm. Chem. Life Sci.
015, 348, 689-703.
e) Chakraborty, B.; Dutta, D.; Mukherjee, S.; Das, S.; Maiti, N.
C.; Das, P.; Chowdhury, C. Eur. J. Med. Chem. 2015, 102, 93-
05.
f) Thi, T. A. D.; Tuyet, N. T. K.; The, C. P.; Nguyen, H. T.;
S.; Dzubak, P.; Hajduch, M.; Sarek, J.; Urban, M. Bioconjugate
Chem. 2015, 26, 2563-2570.
(c) Zhang, H.; Wang, Y.; Zhu, P.; Liu, J.; Xu, S.; Yao, H.;
Jiang, J.; Ye, W.; Wu, X.; Xu, J. Eur. J. Med. Chem. 2015, 97,
235-244.
(d) Havikko, R.; Nasereddin, A.; Sacerdoti-Sierra, N.;
Kopelyanskiy, D.; Alakurtti, S.; Tikka, M.; Jaffe, C. L.; Yli-
Kauhluoma, J. MedChemCommun, 2014, 5, 445-451.
(e) Xu, J.; Li, Z.; Luo, J.; Yang, F.; Liu, T.; Liu, M.; Qiu, W-
W.; Tang, J. J. Med. Chem. 2012, 55, 3122-3134.
2
(
1
(
Thi, C. B.; Phuong, H. T.; Boi, L. V.; Nguyen, T. V.; D’hooghe,
M. Tetrahedron Lett. 2015, 56, 218-224.
(
1
(
g) Shi, W.; Tang, N.; Yan, W-D. J. Asian Nat. Prod. Res. 2015,
7, 159-169.
h) Bori, I. D.; Hung, H. Y.; Qian, K.; Chen, C. H.; Morris-
Natschke, S. L.; Lee, K. H. Tetrahedron Lett. 2012, 53, 1987-
989.
i) Csuk, R.; Barthel, A.; Sczepek, R.; Siewert, B.; Schwarz, S.
Arch. Pharm. Chem. Life Sci. 2011, 344, 37-49.
0. Beld, J.; Cang, H.; Burkart, M. D. Angew. Chem. Int. Ed. 2014,
3, 14456-14461.
(f) Urban, M.; Sarek, J.; Kvasnica, M.; Tislerova, I.; Hajduch,
M. J. Nat. Prod. 2007, 70, 526-532.
23. (a) Kazakova, O. B.; Khusutdinova, E. F.; Suponitskii, K. Y. J.
Str. Chem. 2011, 52, 1030-1032.
1
(
(b) Kumar, V.; Rani, N.; Aggarwal, P.; Sanna, V. K.; Singh, A.
T.; Jaggi, M.; Joshi, N.; Sharma, P. K.; Irchhaiya, R.; Burman,
A. C. Bioorg. Med. Chem. Lett. 2008, 18, 5058-5062.
(c) Mukherjee, R.; Srivastava, S. K.; Siddiqui, M. J. A.; Jaggi,
M.; Singh, A. T.; Vardhan, A.; Singh, M. K.; Rajendran, P.;
Jajoo, H. K.; Burman, A. C.; Kumar, V.; Rani, N.; Agarwal, S.
K. PCT Int. Appl. (2006), WO2006085334 A2 20060817.
2
2
5
1. Suman, P.; Srikanth, D.; Rajukumar, K.; Murthy, T. R.;
Dayakar, Ch.; Kishor, K.; Addlagatta, A.; Kalivendi, S. V.;
Raju, B. C. Eur. J. Med. Chem. 2015, 90, 603-619.
2
2. (a) Vik, M.; Micolova, P.; Urban, M.; Kvasnica, M.; Saman, D.;
Saren, J. J. Radioanal. Nucl. Chem. 2016, 308, 733-739.