Synthesis of novel polyesteramine dendrimers by divergent and convergent methods

Article abstract of DOI:10.1016/j.tet.2013.06.024

Novel dendrimers having an adamantane structure as a core were synthesized such that even low generation dendrimers had a globular structure. Moreover we tried to give them biodegradable function by using ester bonds. Synthesis of the dendrimers, particularly at higher generations, proved difficult via a stepwise procedure, and thus a convergent route was used in which the adamantane core is coupled to the dendritic segments in the final step. We achieved the synthesis of two separate dendrimers with convergent methods till the third generations. The convergent dendrimers were synthesized in good yields compared with divergent one and both dendrimers were found to have narrow polydispersities by GPC analysis.

Full text of DOI:10.1016/j.tet.2013.06.024

Tetrahedron 69 (2013) 6810e6820
Contents lists available at SciVerse ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Synthesis of novel polyesteramine dendrimers by divergent and
convergent methods
*
Hitoshi Akiyama, Kazuyuki Miyashita, Yoshiyuki Hari, Satoshi Obika , Takeshi Imanishi
Graduate School of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Novel dendrimers having an adamantane structure as a core were synthesized such that even low
generation dendrimers had a globular structure. Moreover we tried to give them biodegradable function
by using ester bonds. Synthesis of the dendrimers, particularly at higher generations, proved difficult via
a stepwise procedure, and thus a convergent route was used in which the adamantane core is coupled to
the dendritic segments in the final step. We achieved the synthesis of two separate dendrimers with
convergent methods till the third generations. The convergent dendrimers were synthesized in good
yields compared with divergent one and both dendrimers were found to have narrow polydispersities by
GPC analysis.
Received 20 April 2013
Received in revised form 1 June 2013
Accepted 8 June 2013
Available online 14 June 2013
Keywords:
Dendrimer
Polyester
Adamantine
Click chemistry
Divergent
Ó 2013 Elsevier Ltd. All rights reserved.
Convergent
1. Introduction
degradable to natural metabolites in vivo. Other types of polyester
dendrimers have been synthesized [27e29] and have shown an
Dendrimers are very interesting macromolecules with highly
branched structures and globular shapes. Molecular sizes of den-
drimers are increased stepwise via repeated reaction sequences.
Since the first dendrimers were synthesized by Tomalia et al. [1e3],
many kinds of dendrimers have been synthesized [4], used not only
for chemical applications but also for biomedical applications
[5e11]. For example, commercially available polyamidoamine
(PAMAM) and polypropylenimine (PPI) dendrimers are widely used
as drugs [6], gene delivery systems [7], and MRI contrast agents [9].
Additionally, these dendrimers provide a high gene transfer effi-
ciency into mammalian cells [12e15]. This transfer efficiency is
considered to be a result of the many interior tertiary amines,
which exist in the dendrimer, leading to an effect known as a pro-
ton sponge [16]. Moreover, these dendrimers have many functional
groups such as amino groups and hydroxyl groups on their pe-
riphery [12e15,17], and modification of these surface groups with
various molecules offers the chance for other potential applications
[18e23]. However, for medical applications, dendrimers must be
less toxic and more biodegradable than such dendrimers. Recently
polyester dendrimers called ‘biodendrimers’ have been reported
[24e26], which have building blocks known to be biocompatible or
antitumor effect [29]. Furthermore, a robust and biodegradable
PEGylated dendrimer based on a polyester-polyamide hybrid core
has been synthesized and biodistribution and chemotherapy study
in tumored mice have been evaluated [30]. However, there are few
reports on polyester dendrimers including primary and tertiary
amines. To form complexes with plasmid DNA, antisense oligonu-
cleotide or siRNA and other biological molecules, it is necessary for
the dendrimers to have primary amines. These amino groups
would not only allow complex formation, but would also interact
with cellular membranes and enable conjugation with various
ligands.
In this study, we designed novel polyester dendrimers XeZ
named ‘polyesteramine dendrimers’ (Fig. 1).
As the core of the dendrimer, we selected an adamantane
structure. Typically, planar or linear molecules, such as ammonia,
ethylenediamine, 1,4-diaminobutane, benzene derivatives, lactic
acid, succinic acid, adipic acid, and ethylene glycol, have been used
for the dendrimer core [4,24e26,28]. These dendrimers maintain
a planar structure in lower generations. Adamantane, on the other
hand, has a three-dimensional structure, and dendrimers having an
adamantane core are expected to have a more globular structure
than dendrimers such as PAMAM even in lower generations. In
terms of synthesis, PAMAM dendrimers are synthesized by a typical
stepwise and iterative two-step reaction sequence [1e3], consisting
of amidation of methyl acrylate with ethylenediamine and Michael
* Corresponding author. Tel.: þ81 6 6879 8200; fax: þ81 6 6879 8204; e-mail
address: obika@phs.osaka-u.ac.jp (S. Obika).
0040-4020/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tet.2013.06.024

H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
6811
BocHN
BocHN
BocHN
NHBoc
O
BocHN
O
O
O
O
O
O
O
BocHN
O
NHBoc
O
O
NHBoc
O
BocHN
N
O
O
N
O
O
N
O
O
BocHN
N
N
N
O
NH
O
N
O
O
N
N
O
O
O
O
O
O
O
O
N
NH
O
O
N
O
O
N
O
O
NHBoc
NHBoc
O
BocHN
O
O
O
O
O
O
O
O
BocHN
O
N
N
N
N
N
N
O
O
O
O
O
O
O
O
H
O
O
O
O
O
HN
N
N
N
N
O
O
NHBoc
N
N
O
N
O
O
NHBoc
O
O
BocHN
O
O
NHBoc
O
O
BocHN
O
O
O
O
O
NHBoc
O
NHBoc
BocHN
NHBoc
NHBoc
X
Z
Y
Fig. 1. Structures of polyester dendrimers X, Y, and Z.
addition of primary amines with methyl acrylate. But it is known
that this method sometimes leads to a lot of structural defects and
also requires a long reaction time, which is a critical impediment
for obtaining dendrimers with a uniform molecular weight, par-
ticularly in higher generations [1,2]. Separation of dendrimers
having primary amines in the periphery is also a difficult task. In
order to resolve these problems, we designed a novel dendrimer
having a three-dimensional adamantane core, and synthesized
dendrimers via two separate convergent routes employing ami-
dation and Huisgen [3þ2] cycloaddition reaction as the key cou-
pling reactions, respectively.
of 3 by treatment with trifluoroacetic acid (TFA) smoothly took
place, and the desired glycinoyloxy derivative 4 was isolated as
a tetratrifluoroacetate salt in 91% yield.
At first, we examined the usual stepwise elongation method for
the synthesis of dendrimer 7 as shown in Scheme 2. Michael re-
action of 4 with 2-Boc-aminoethylacrylate 1, prepared from 2-Boc-
aminoethanol and acryloyl chloride, proceeded to give the first
generation dendrimer 5 in 32% yield accompanied by the deacy-
lated product 6 in 32% yield. Although we examined the reaction
under various conditions, it was not possible to prevent formation
of the deacylated product 6. Next, deprotection of dendrimer 5 by
TFA followed by Michael reaction was carried out. However, un-
fortunately, a complex mixture was given. MALDI-TOF-MS analysis
of the crude product showed the existence of a number of in-
completely reacted products (Fig. 2). The desired second generation
dendrimer 7 was also detected by the spectrum, but could not be
isolated from the mixture.
2. Results
1,3,5,7-Tetrakis(aminoacetoxy)adamantane core 4 was synthe-
sized as shown in Scheme 1. 1,3,5,7-Tetrakis(bromoacetoxy)ada-
mantane, prepared according to a literature procedure, was treated
with NaN3 to give azidoacetoxy derivative 2 in 73% yield. Although
several attempts to obtain 4 by direct reduction of 2 resulted in
a complex mixture, Boc-protected derivative 3 was successfully
obtained by reduction of 2 using triphenylphosphine and simul-
taneous Boc-protection in 76% yield. Deprotection of the Boc group
These results indicated that it was going to be difficult to obtain
higher generation dendrimers having a uniform molecular weight
by the present stepwise method. Therefore, we selected a conver-
gent method for the synthesis of the higher generation dendrimers.
We planned for the adamantane core to be coupled with dendritic
N3
Br
O
O
O
O
N3
O
O
Br
O
O
NaN3
O
O
DMF, 12h
N3
Br
O
O
O
O
O
O
N3
Br
2
NH2
NHBoc
O
O
O
O
H2N
O
O
BocHN
O
(1) P(Ph)3, H2O, THF, 2h
(2) (Boc)2O, Et3N, 12h
TFA, 2h
O
O
O
O
NH2
NHBoc
O
O
O
O
O
H2N
BocHN
CF3COOHx4
3
4
Scheme 1. Synthesis of two types of adamantane core.

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H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
Scheme 2. Synthesis of novel dendrimers by divergent method.
Fig. 2. MALDI-TOF-MS spectra of crude Boc-G2 (7).
segments by amide formation. The segments were to be synthe-
sized according to a usual procedure (deprotection and Michael
reaction) as shown in Scheme 3. Michael reaction of GlyeOBn with
acrylate 1 afforded bisadduct 8, which corresponds to the segment
for the first generation, in 93% yield. The segment 9 for the second
generation was prepared from 8 by deprotection of the Boc group
followed by Michael reaction in 37% yield. Similarly, the segment 10
for the third generation was obtained in 22% yield.

H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
6813
Scheme 3. Synthesis of benzyl ester glycine dendrons.
The coupling reaction between the adamantane core 4 with the
segment was carried out as shown in Scheme 4. After removal of
the benzyl ester of 8 under reductive conditions, the resultant
carboxylic acid was coupled with the adamantane core 4 using
PyBOP as a condensing agent to give 11 in 87% yield; PyBOP is
known to be superior to DCC, TFFH, and a number of other com-
mercially available peptide coupling reagents [31]. The segments
for the higher generation dendrimers, 9 and 10, were also coupled
with 4 according to the same procedure to give 12 and 13 in 36%
and 22% yields, respectively.
Recently, Huisgen [3þ2] dipolar cycloaddition reaction has
gained much attention in general synthetic chemistry [32], and has
also been applied to the synthesis of dendrimers [33e38]. As it is
obvious that azido-intermediate 2 would work as a substrate for
the Huisgen reaction with a proper alkyne derivative, we synthe-
sized novel segments having an alkyne structure as shown in
Scheme 5. In this case, propargyl amine was employed for the
starting material, which reacted quantitatively with acrylate 1 to
give 14 as the first generation segment. Segment 15 for the second
generation, and segment 16 for the third generation were also
obtained in 24% and 14% yields, respectively, via sequential re-
actions (deprotection by TFA treatment and Michael reaction with
acrylate).
As shown in Scheme 6, dendrimers 17, 18, and 19, were obtained
by coupling of azido-derivative 2 with segments 14, 15, and 16,
respectively, under the typical conditions of Huisgen reaction. The
yield for dendrimers of the higher generation decreased, but was
nonetheless satisfactory.
Molecular weight data determined by MALDI-TOF-MASS and gel
permeation chromatography (GPC) and polydispersity indices
(PDIs) are summarized in Table 1. The mass spectrometry data
agrees with the calculated molecular weights. All dendrimers show
narrow PDIs, indicating that these compounds were obtained in
pure form, thus confirming the effectiveness of a convergent route
for the synthesis of dendrimers.
3. Discussion
We aimed to synthesize biodegradable polyester dendrimers
including primary and tertiary amino groups for biochemical and
medical applications. We chose adamantane as a dendrimer core to
produce novel dendrimers that were more globular in lower gen-
erations than commercially available dendrimers and many other
reported dendrimers. The number-average molecular weight (Mn)
and weight-average molecular weight (Mw), which were de-
termined by GPC measurements, deviate from the mass spectrom-
etry data as the generation number increases (Table 1). This result is
consistent with the previous data of dendrimers adopting a more
globular structure [25,26]. In this study, amide glycine dendrimers
11e13 (AG Boc-G1e3) and click chemistry dendrimers 17e19 (CC
Boc-G1e3) show less Mw than the expected one even at the low
generation numbers (G2 and G3). These results suggest that the
novel dendrimers possessing an adamantane core have a more
globular structure even in lower generations. We constructed mo-
lecular models of G1, AG G1, and CC G1, which are compared with
PAMAM G1 (Fig. 3). Although PAMAM G1 is planar, the novel G1, AG
G1, and CC G1 dendrimers are more steric due to a tetrahedral core.
We expect these novel dendrimers to grow up more spherically.
It is known that the transfection activity of PAMAM dendrimers
increases with higher generations [12,13]. This enhanced activity is
largely considered to be a result of the spherical structure of the
dendrimers. That is, the more spherical the dendrimer structures,
the better their ability as a gene carrier. However, it is also the case
that the higher the dendrimer generation, the greater the toxicity
of the dendrimer [8]. If the dendrimers can be degraded and me-
tabolized at physiological conditions after drug delivery, they

6814
H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
Scheme 4. Synthesis of amide glycine dendrimers.

H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
6815
Scheme 5. Synthesis of click dendrons.
would have greater application as non-viral carriers. Recently,
a variety of biodegradable polymers have been used as non-viral
carriers for plasmid DNA delivery [39]. Park et al. reported that
a poly(amino ester) including primary and tertiary amines and
esters exhibited relatively slow biodegradability, as the DNA/poly-
mer complex was maintained for 7 days [40]. By contrast, in syn-
thesis of 1,3,5,7-tetrakis(aminoacetoxy)adamantane bearing free
primary amines, generation of primary amino groups led to readily
self-degradation. In the synthesis of divergent dendrimer Boc-G1 5,
the generation of primary amino groups also led to the appearance
of incomplete Boc-G1 6. Fife et al. reported that the existence of
intramolecular neighboring amino groups effectively catalyzed
ester hydrolysis [41]. Thus, this phenomenon would be due to high
degradability of 1,3,5,7-tetrakis(aminoacetoxy)adamantane bear-
ing neighboring amines. From these results, we considered that the
emergence of free primary amino groups is an obstacle to synthesis
of polyesteramine dendrimers because of degradation of the ada-
mantane core. Thus, it is very important to limit generation of free
primary amines in the synthesis of polyesteramine dendrimers.
Using two separate convergent methods, we were able to produce
polyesteramine dendrimers until the third generation. The yields of
the two kinds of dendrimers synthesized by a convergent approach
drop with higher generations, which is consistent with previous
observations [31,36]. This could be due to significant retention of
the polar dendrimers in the silica column, steric hindrance of the
higher generation dendrons, and partial degradability.
generation dendrons and dendrimers with MALDI-TOF-MASS, we
found that they were not attained completely in higher genera-
tions. Though the low yields must be improved, these methods
provide interesting new polyesteramine dendrimers and insight
into possible modifications or changes of the periphery and core of
such dendrimers. We are currently working toward the character-
ization of the dendrimers given here and the synthesis of a variety
of surface-modified dendrimers. Polyesteramine dendrimers offer
many potential platforms in areas such as medicine, catalysis,
photonics, and nanotechnology.
5. Experimental
5.1. General
All solvents were dried and freshly distilled prior to use.
All chemicals were purchased from chemical suppliers. For column
chromatography, Fuji Silysia silica gel PSQ-100B (0.100 mm) and FL-
100D (0.100 mm) was used. 1H NMR (270, 300 or 400 MHz) and 13
C
NMR (67 or 75 MHz) spectra were recorded on JEOL JNM-EX270,
JEOL JNM-AL300, and JEOL JNM-ECS400 spectrometers, re-
spectively. IR spectra were recorded on a JASCO FT/IR-200 and
JASCO FT/IR-4200 spectrometer. FAB Mass spectra were measured
on a JEOL JMS-600 or JEOL JMS-700 mass spectrometer. MALDI-
TOF-Mass spectra were recorded on a Bruker Daltonics Autoflex II
TOF/TOF mass spectrometer. Gel permeation chromatography
(GPC) was performed using THF as the eluent on a SHIMADZU
column and refractive index detector. Polystyrene standards (820,
2460, 4100, 12,400, and 18,100) were used for calibration.
4. Conclusion
In summary, we have presented divergent and convergent
procedures for the synthesis of novel polyesteramine dendrimers.
Convergent approaches were more efficient than the divergent
method in synthesizing higher generation dendrimers. By moni-
toring Michael addition reactions in synthesis of the higher
5.2. Synthesis of 2-Boc-aminoethylacrylate 1
The dendritic unit 1 was prepared by modification of a pub-
lished procedure [42]. 2-Aminoethanol (25.4 g, 0.41 mol) was

6816
H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
Scheme 6. Synthesis of click chemistry dendrimers.

H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
6817
(s, 8H). 13C NMR (CDCl3)
d
¼43.0, 50.5, 79.1, 167.0. MS (FAB): m/z 555
Table 1
MALDI-TOF-MS and GPC data for novel polyesteramine dendrimers
[MþNa]þ. HRMS (FAB): m/z calcd for C18H20N12O8 [MþNa]þ:
555.1527; found: 555.1417. Anal. Calcd for C18H20N12O8: C, 40.61; H,
3.79; N, 31.57. Found: C, 40.58; H, 3.78; N, 31.22.
Dendrimer
GPC
Mw/Mn
MALDI-TOF-MS
Mn
Mw
calcd Mw
Found
3
5
1167
2623
2625
4536
8550
2985
5114
8568
1178
2689
2701
4745
9066
3150
5486
8912
1.01
1.03
1.03
1.05
1.06
1.06
1.07
1.04
829
2150
2379
5022
10,308
2475
852 [MþNa]þ
2151 [MþH]þ
2379 [MþH]þ
5022 [MþH]þ
10,304 [MþH]þ
2475 [MþH]þ
5119 [MþH]þ
10,404 [MþH]þ
5.3.2. Synthesis of 1,3,5,7-tetrakis(n-Boc-aminoacetoxy)adamantane
3. The adamantane compound 2 (2.4 g, 4.43 mmol) was dissolved
in a 3:1 solvent ratio of THF/H2O (29.6 ml), and triphenylphosphine
(5.1 g, 19.48 mmol) was added. After stirring for 2 h, (Boc)2O (4.3 g,
19.48 mmol) and triethylamine (3.7 ml, 26.57 mmol) were added to
the reaction mixture and stirred overnight. The reaction mixture
was extracted with ethyl acetate and dried over Na2SO4. Purifica-
11
12
13
17
18
19
5118
10,404
Fig. 3. Molecular models of PAMAM G1, G1, AG G1, and CC G1 constructed by Spartan ’06.
dissolved in CHCl3 (500 ml) in a flask equipped with a magnetic
stirrer, and (Boc)2O (100.0 g, 0.45 mol) was added. After stirring for
30 min, the concentrated solution was diluted with CH2Cl2
(500 ml), and triethylamine (113.6 ml, 0.82 mol) and acryloyl
chloride (44.5 g, 0.49 mol) were added. After stirring for 1 h under
a nitrogen atmosphere, satd NaHCO3 aq solution (200 ml) was
added to the reaction mixture. The aqueous layer was extracted
with CH2Cl2 and dried over Na2SO4. Purification of the concentrated
compound was carried out by column chromatography on silica gel
using ethyl acetate and hexane as eluent. A pale yellowish oil;
tion of the concentrated compound was carried out by column
chromatography on silica gel using ethyl acetate and hexane as
eluent. A white foam; 2.8 g, 76%. FTIR:
n
(cmꢀ1) 3362, 2978, 2934,
1755, 1707. 1H NMR (CD3Cl)
d
¼1.45 (s, 36H), 2.53 (s, 12H), 3.82 (d,
8H, J¼6 Hz), 4.93 (t, 4H, J¼5 Hz). 13C NMR (CDCl3)
¼28.3, 42.8, 43.1,
d
78.6, 80.1, 155.6, 169.0. MS (FAB): m/z 851 [MþNa]þ. HRMS (FAB):
m/z calcd for C38H60N4O16 [MþNa]þ: 851.4004; found: 851.3887.
MS (MALDI): m/z calcd for C38H60N4O16 [MþNa]þ: 851.400; found:
851.746.
74.9 g, 85%. FTIR:
n
(cmꢀ1) 3371, 3095, 3043, 2983, 2938, 1986, 1706.
¼1.37 (s, 9H), 3.33e3.38 (m, 2H), 4.15 (t, 2H,
5.3.3. Synthesis of 1,3,5,7-tetrakis(aminoacetoxy)adamantane tetra-
trifluoroacetate 4. Compound 3 (2.0 g, 2.38 mmol) was added toTFA
(7.5 ml) and stirred for 2 h. The concentrated compound was rep-
recipitated with ethyl acetate and lyophilized. A white solid; 1.9 g,
1H NMR (CDCl3):
d
J¼5 Hz), 4.98 (br, 1H), 5.78 (dd, 1H, J¼1, 10 Hz), 6.06 (dd, 1H, J¼10,
17 Hz), 6.36 (dd, 1H, J¼1, 17 Hz). 13C NMR (CDCl3):
¼28.2, 39.5,
d
63.6, 79.3, 127.9, 131.1, 155.7, 165.9. MS (FAB): m/z 216 [MþH]þ.
HRMS (FAB): m/z calcd for C10H17N1O4 [MþH]þ: 216.1158; found:
216.1226. Anal. Calcd for C10H17NO4: C, 55.80; H, 7.96; N, 6.51.
Found: C, 55.93; H, 7.87; N, 6.47.
91%. FTIR: n
(cmꢀ1) 3362, 2978, 2934, 1755, 1707. 1H NMR (D2O)
d
¼2.64 (s, 12H), 3.87 (s, 8H). 13C NMR (CD3OD)
¼41.5, 43.8, 80.9,
d
167.2. MS (FAB): m/z 429 [MþH]þ. HRMS (FAB): m/z calcd for
C18H28N4O8 [MþH]þ: 429.1907; found: 429.1967. Anal. Calcd for
C26H32F12N4O16$H2O: C, 34.60; H, 3.80; N, 6.21. Found: C, 34.44; H,
3.97; N, 5.95.
5.3. Synthesis of adamantane core
5.3.1. Synthesis of 1,3,5,7-tetrakis(azidoacetoxy)adamantane 2. 1,3,5,7-
Tetrakis(bromoacetoxy)adamantane was synthesized from ada-
mantane in three steps according to a literature method [43]. 1,3,5,7-
Tetrakis(bromoacetoxy)adamantane (200.0 mg, 0.29 mmol) was
dissolved in DMF (2.9 ml) and sodium azide (152.0 mg, 2.34 mmol)
was added. After stirring for 2 h under a nitrogen atmosphere, water
was added to the reaction mixture together with CH2Cl2. The aqueous
layer was extracted with CH2Cl2 and dried over Na2SO4. Purification
of the concentrated compound was carried out by column chroma-
tography on silica gel using CHCl3. A white solid; 112.7 mg, 73%. FTIR:
5.4. Synthesis of dendrimers by divergent method
5.4.1. Synthesis of Boc-G1 5. Compound 4 (519.8 mg, 0.59 mmol)
was dissolved in CH3CN (5.9 ml), and 1 (5.1 g, 23.51 mmol) and
triethylamine (650 ml, 4.70 mmol) were added. After stirring for 7
days at 45 ꢁC under a nitrogen atmosphere, the reaction mixture
was extracted with ethyl acetate and dried over Na2SO4. Purifica-
tion of the concentrated compound was carried out by column
chromatography on silica gel using ethyl acetate and hexane as
eluent. A pale yellowish foam; 403.8 mg, 32%. FTIR:
n
(cmꢀ1) 3380,
n
(cmꢀ1) 2923, 2208, 2108, 1739. 1H NMR (CDCl3)
d¼2.62 (s,12H), 3.80
2977, 1713. 1H NMR (CDCl3)
d
¼1.36 (s, 72H), 2.36e2.43 (m, 28H),

6818
H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
2.89 (t, 16H, J¼6 Hz), 3.26e3.32 (m, 24H), 4.06 (t, 16H, J¼5 Hz), 5.14
acetone as eluent. A brownish gum; 212 mg, 22%. FTIR:
n
(cmꢀ1
¼1.35 (s,
)
(br, 8H). 13C NMR (CDCl3)
d
¼28.3, 33.4, 39.4, 43.3, 49.3, 54.8, 63.5,
3726, 3593, 3371, 2977, 2837, 2013, 1729. 1H NMR (CDCl3)
d
78.0, 79.2, 155.6, 169.6, 171.9. MS (FAB): m/z 2150 [MþH]þ. MS
(MALDI): m/z calcd for C98H164N12O40 [MþH]þ: 2150.117; found:
2150.650.
72H), 2.38 (t, 28H, J¼6 Hz), 2.63 (t, 12H, J¼5 Hz), 2.74 (t, 24H,
J¼6 Hz), 2.89 (t, 4H, J¼7 Hz), 3.27e3.32 (m, 16H), 3.37 (s, 2H),
4.01e4.07 (m, 28H), 5.04 (s, 2H), 5.19 (br, 8H), 7.27 (s, 5H). 13C NMR
(CDCl3)
d
¼28.3, 32.5, 32.7, 33.2, 39.4, 49.3, 49.4, 49.6, 51.7, 51.8, 54.5,
5.4.2. Synthesis of Boc-G2 7. Compound 5 (112 mg, 0.05 mmol) was
added to TFA (2.0 ml) and stirred for 2 h. The concentrated com-
pound was reprecipitated with ethyl acetate and lyopholized. The
deprotected compound (100 mg, 0.04 mmol) was dissolved in
62.0, 62.1, 63.4, 66.0, 79.1, 127.9, 128.0, 128.3, 135.4, 155.5, 170.6,
171.7, 171.8, 171.8. MS (MALDI): m/z calcd for C119H201N15O46
[MþH]þ: 2578.943; found: 2576.640.
iPrOH (0.4 ml), and 1 (1.5 g, 7.07 mmol) and triethylamine (123
ml,
5.6. Synthesis of amide glycine dendrimers
0.88 mmol) were added. After stirring for 7 days at 45 ꢁC under
a nitrogen atmosphere, the reaction mixture was extracted with
ethyl acetate and dried over Na2SO4. Purification of the concen-
trated compound was carried out by column chromatography on
silica gel using ethyl acetate and acetone as eluent. A yellowish
gum; trace.
5.6.1. Synthesis of AG Boc-G1 11. Compound 8 (150 mg, 0.25 mmol)
was dissolved in CH3CN (2.5 ml) and PdOH/C (150 mg) was added.
After stirring for 2 h under a hydrogen atmosphere, the reaction
mixture was filtered. The concentrated compound was dissolved in
CH3CN (2.5 ml), and 4 (37 mg, 0.042 mmol), PyBOP (109 mg,
0.21 mmol), and triethylamine (47 ul, 0.34 mmol) were added. After
stirring for 1 day at room temperature under a nitrogen atmo-
sphere, the reaction mixture was extracted with ethyl acetate and
dried over Na2SO4. Purification of the concentrated compound was
carried out by column chromatography on silica gel using ethyl
acetate and acetone as eluent. A pale yellowish foam; 87 mg, 87%.
5.5. Synthesis of benzyl ester glycine dendrons
5.5.1. Synthesis of BnGD Boc-G1 8. Glycine benzyl ester p-toluene-
sulfonate (2.0 g, 5.92 mmol) was dissolved in CH3CN (10 ml), and 1
(3.8 g, 17.78 mmol) and triethylamine (1.6 ml, 11.86 mmol) were
added. After stirring for 4 days at 60 ꢁC under a nitrogen atmo-
sphere, the reaction mixture was extracted with ethyl acetate and
dried over Na2SO4. Purification of the concentrated compound was
carried out by column chromatography on silica gel using ethyl
FTIR:
n
(cmꢀ1) 3331, 2974, 2927, 1712. 1H NMR (CDCl3)
d
¼1.43 (s,
72H), 2.49e2.53 (m, 28H), 2.80 (t, 16H, J¼6 Hz), 3.13 (s, 8H),
3.33e3.40 (m, 16H), 3.93 (d, 8H, J¼5 Hz), 4.12 (t, 16H, J¼5 Hz), 5.28
(br, 8H), 8.04 (t, 4H, J¼6 Hz). 13C NMR (CDCl3)
¼28.5, 32.1, 39.5,
d
acetate and hexane as eluent. A pale yellowish oil; 3.3 g, 93%. FTIR:
(cmꢀ1) 3730, 3592, 3368, 3064, 2977, 1958, 1712. 1H NMR (CDCl3)
¼1.43 (s, 18H), 2.46 (t, 4H, J¼7 Hz), 2.98 (t, 4H, J¼7 Hz), 3.33e3.39
(m, 4H), 3.46 (s, 2H), 4.12 (t, 4H, J¼5 Hz), 5.13 (m, 4H), 7.34 (s, 5H).
13C NMR (CDCl3)
n
41.4, 43.1, 49.6, 58.3, 63.9, 78.5, 79.4, 155.8, 168.2, 171.3, 172.2. MS
(MALDI): m/z calcd for C106H176N16O44 [MþH]þ: 2379.613; found:
2378.987.
d
d
¼28.4, 33.4, 39.6, 49.6, 54.5, 63.7, 66.3, 79.4,
5.6.2. Synthesis of AG Boc-G2 12. Compound 9 (188 mg,
128.2, 128.3, 128.5, 135.4, 155.7, 170.8, 172.1. MS (FAB): m/z 596
[MþH]þ. HRMS (FAB): m/z calcd for C29H45N3O10 [MþH]þ:
596.3105; found: 596.3177.
0.15 mmol) was dissolved in CH3CN (1.5 ml) and PdOH/C (190 mg)
was added. After stirring for 2 h under a hydrogen atmosphere,
the reaction mixture was filtered. The concentrated compound
was dissolved in CH3CN (1.5 ml), and 4 (22 mg, 0.025 mmol),
5.5.2. Synthesis of BnGD Boc-G2 9. Compound
8
(721 mg,
PyBOP (97 mg, 0.19 mmol), and triethylamine (28 ml, 0.20 mmol)
1.21 mmol) was added to TFA (3 ml) and stirred for 2 h. The
concentrated compound was lyopholized. The deprotected com-
pound (916 mg, 1.24 mmol) was dissolved in CH3CN (14.1 ml), and
1 (6.1 g, 28.31 mmol) and triethylamine (1.2 ml, 8.49 mmol) were
added. After stirring for 5 days at 45 ꢁC under a nitrogen atmo-
sphere, the reaction mixture was extracted with ethyl acetate and
dried over Na2SO4. Purification of the concentrated compound
was carried out by column chromatography on silica gel using
ethyl acetate and hexane as eluent. A yellowish gum; 577 mg,
were added. After stirring overnight at room temperature under
a nitrogen atmosphere, the reaction mixture was extracted with
ethyl acetate and dried over Na2SO4. Purification of the concen-
trated compound was carried out by column chromatography on
silica gel using ethyl acetate and acetone as eluent. A yellowish
gum; 45 mg, 36%. FTIR:
(CDCl3)
n
(cmꢀ1) 3364, 2926, 2854, 1713. 1H NMR
d
¼1.42 (s, 144H), 2.47e2.51 (m, 60H), 2.74e2.85 (m, 64H),
3.14 (s, 8H), 3.33e3.38 (m, 32H), 3.91 (d, 8H, J¼4 Hz), 4.12 (t, 48H,
J¼5 Hz), 5.26 (br, 16H), 7.96 (t, 4H, J¼5 Hz). 13C NMR (CDCl3)
37%. FTIR:
(CDCl3)
n
(cmꢀ1) 3730, 3372, 2976, 2011, 1957, 1715. 1H NMR
d
¼28.4, 31.7, 32.6, 39.5, 49.3, 49.8, 52.0, 57.9, 63.7, 78.5, 79.3, 155.7,
d
¼1.37 (s, 36H), 2.39 (t, 12H, J¼7 Hz), 2.64 (t, 4H, J¼6 Hz),
171.3, 172.0. MS (MALDI): m/z calcd for C226H384N32O92 [MþH]þ:
2.75 (t, 8H, J¼7 Hz), 2.92 (t, 4H, J¼7 Hz), 3.27e3.33 (m, 8H), 3.40
5022.628; found: 5021.661.
(s, 2H), 4.02e4.08 (m, 12H), 5.06 (s, 2H), 5.17 (br, 4H), 7.28 (s, 5H).
13C NMR (CDCl3)
d
¼28.3, 32.7, 33.2, 39.4, 49.3, 49.6, 51.8, 54.4,
5.6.3. Synthesis of AG Boc-G3 13. Compound 10 (162 mg,
0.063 mmol) was dissolved in CH3CN (0.63 ml) and PdOH/C
(160 mg) was added. After stirring for 4 h under a hydrogen at-
mosphere, the reaction mixture was filtered. The concentrated
compound was dissolved in CH3CN (0.63 ml), and 4 (9 mg,
62.0, 63.5, 66.0, 79.2, 128.0, 128.0, 128.3, 135.3, 155.5, 170.6, 171.8,
171.9. MS (FAB): m/z 1256.6 [MþH]þ. HRMS (FAB): m/z calcd
for C59H97N7O22 [MþH]þ: 1256.6687; found: 1256.6769. MS
(MALDI): m/z calcd for C59H97N7O22 [MþH]þ: 1256.669; found:
1256.432.
0.010 mmol), PyBOP (41 mg, 0.079 mmol), and triethylamine (12 ml,
0.083 mmol) were added. After stirring for 1 day at room temper-
ature under a nitrogen atmosphere, the reaction mixture was
extracted with ethyl acetate and dried over Na2SO4. Purification of
the concentrated compound was carried out by column chroma-
tography on silica gel using ethyl acetate and acetone as eluent.
5.5.3. Synthesis of BnGD Boc-G3 10. Compound
9 (535 mg,
0.43 mmol) was added to TFA (2 ml) and stirred for 2 h. The con-
centrated compound was lyopholized. The deprotected compound
(631 mg, 0.38 mmol) was dissolved in CH3CN (3.8 ml), and 1 (3.3 g,
15.24 mmol) and triethylamine (740
m
l, 5.33 mmol) were added.
A brownish gum; 24 mg, 22%. FTIR:
NMR (CDCl3)
n
(cmꢀ1) 3364, 2925, 1701. 1H
After stirring for 7 days at 45 ꢁC under a nitrogen atmosphere, the
reaction mixture was extracted with ethyl acetate and dried over
Na2SO4. Purification of the concentrated compound was carried out
by column chromatography on silica gel using ethyl acetate and
d
¼1.43 (s, 288H), 2.46 (t, 124H, J¼6 Hz), 2.72 (t, 48H,
J¼6 Hz), 2.82 (t, 112H, J¼6 Hz), 3.15 (s, 8H), 3.33e3.39 (m, 64H), 3.91
(s, 8H), 4.09e4.14 (m, 112H), 5.23 (br, 32H), 7.94 (br, 4H). 13C NMR
(CDCl3)
d
¼28.4, 32.7, 39.6, 49.7, 52.0, 62.2, 63.6, 79.4, 155.9, 172.3.

H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
6819
MS (MALDI): m/z calcd for C466H800N64O188 [MþH]þ: 10,308.657;
concentrated compound was carried out by column chromatogra-
phy on silica gel using ethyl acetate and acetone as eluent. A pale
found: 10,303.944.
yellowish foam; 184 mg, 82%. FTIR:
NMR (CDCl3)
¼1.34 (s, 72H), 2.41e2.45 (m, 28H), 2.69 (t, 16H,
J¼6 Hz), 3.23e3.29 (m, 16H), 3.75 (s, 8H), 4.03 (t, 16H, J¼5 Hz), 5.06
(s, 8H), 5.30 (br, 8H), 7.58 (s, 4H). 13C NMR (CDCl3)
n
(cmꢀ1) 3367, 2928, 1701. 1H
5.7. Synthesis of click dendrons
d
5.7.1. Synthesis of CD Boc-G1 14. Propargyl amine hydrochloride
(915 mg, 10.0 mmol) was dissolved in CH3CN (10 ml), and 1 (6.5 g,
30.0 mmol) and triethylamine (2.8 ml, 20.00 mmol) were added.
After stirring for 3 days at 45 ꢁC under a nitrogen atmosphere, the
reaction mixture was extracted with ethyl acetate and dried over
Na2SO4. Purification of the concentrated compound was carried out
by column chromatography on silica gel using ethyl acetate and
d¼28.3, 32.5,
39.4, 42.6, 47.9, 48.7, 50.9, 53.8, 63.4, 79.1, 124.1, 144.0, 155.6, 164.8,
172.0. MS (MALDI): m/z calcd for C110H176N24O40 [MþH]þ:
2475.712; found: 2474.737.
5.8.2. Synthesis of CC Boc-G2 18. Compounds
2
(25 mg,
0.046 mmol) and 15 (318 mg, 0.28 mmol) were dissolved in a 4:1
solvent ratio of THF/H2O (2.8 ml), and CuSO4$5H2O (14 mg,
0.055 mmol) and sodium ascorbate (22 mg, 0.11 mmol) were
added. After stirring overnight, the reaction mixture was extracted
with ethyl acetate and dried over Na2SO4. Purification of the con-
centrated compound was carried out by column chromatography
on silica gel using ethyl acetate and acetone as eluent. A yellowish
hexane as eluent. A pale yellowish oil; 5.3 g, quant. FTIR:
3955, 3728, 3592, 3356, 2976, 2104, 2013, 1706. 1H NMR (CDCl3)
¼1.34 (s, 18H), 2.16 (t, 1H, J¼2 Hz), 2.38 (t, 4H, J¼7 Hz), 2.74 (t, 4H,
n )
(cmꢀ1
d
J¼7 Hz), 3.23e3.30 (m, 4H), 3.35 (d, 2H, J¼2 Hz), 4.04 (t, 4H,
J¼5 Hz), 5.21 (br, 2H). 13C NMR (CDCl3)
¼28.7, 33.2, 39.8, 41.9, 49.2,
d
63.8, 74.0, 77.9, 79.5,156.0,172.2. MS (FAB): m/z 486 [MþH]þ. HRMS
(FAB): m/z calcd for C23H39N3O8 [MþH]þ: 486.2737; found:
486.2823.
gum; 85 mg, 36%. FTIR:
(CDCl3)
n
(cmꢀ1) 3371, 2976, 2110, 1731. 1H NMR
¼1.42 (s, 144H), 2.42e2.53 (m, 60H), 2.70 (t, 16H, J¼6 Hz),
d
2.81 (t, 48H, J¼6 Hz), 3.33e3.39 (m, 32H), 3.82 (s, 8H), 4.08e4.13
5.7.2. Synthesis of CD Boc-G2 15. Compound 14 (1.2 g, 2.54 mmol)
was added to TFA (5.0 ml) and stirred for 2 h. The concentrated
compound was lyopholized. The deprotected compound (510 mg,
0.81 mmol) was dissolved in CH3CN (8.1 ml), and 1 (3.5 g,
(m, 48H), 5.13 (s, 8H), 5.22 (br, 16H), 7.64 (s, 4H). 13C NMR (CDCl3)
d
¼28.4, 32.5, 32.9, 39.6, 42.9, 48.1, 48.7, 49.8, 52.0, 62.2, 63.7, 79.4,
124.1, 155.7, 164.9, 172.1. MS (MALDI): m/z calcd for C230H384N40O88
[MþH]þ: 5118.727; found: 5118.790.
16.26 mmol) and triethylamine (676 ml, 4.87 mmol) were added.
After stirring for 4 days at 45 ꢁC under a nitrogen atmosphere, the
reaction mixture was extracted with ethyl acetate and dried over
Na2SO4. Purification of the concentrated compound was carried out
by column chromatography on silica gel using ethyl acetate and
5.8.3. Synthesis of CC Boc-G3 19. Compounds 2 (8 mg, 0.014 mmol)
and 16 (221 mg, 0.089 mmol) were dissolved in a 4:1 solvent ratio
of THF/H2O (890
ml), and CuSO4$5H2O (4 mg, 0.018 mmol) and
sodium ascorbate (7 mg, 0.036 mmol) were added. After stirring
overnight, the reaction mixture was extracted with ethyl acetate
and dried over Na2SO4. Purification of the concentrated compound
was carried out by column chromatography on silica gel using ethyl
hexane as eluent. A yellowish gum; 223 mg, 24%. FTIR:
n
(cmꢀ1
¼1.40 (s, 36H),
)
3725, 3372, 2976, 2103, 2014, 1711. 1H NMR (CDCl3)
d
2.19 (t, 1H, J¼2 Hz), 2.43 (t,12H, J¼7 Hz), 2.69 (t, 4H, J¼6 Hz), 2.79 (t,
12H, J¼7 Hz), 3.31e3.37 (m, 10H), 4.06e4.12 (m, 12H), 5.14 (br, 4H).
acetate and acetone as eluent. A brownish gum; 30 mg, 21%. FTIR:
n
13C NMR (CDCl3)
d
¼28.1, 32.6, 32.6, 39.3, 41.5, 48.6, 49.5, 51.7, 61.9,
(cmꢀ1) 3363, 2925, 1703. 1H NMR (CDCl3)
d
¼1.43 (s, 288H),
63.4, 73.2, 77.6, 79.1, 155.4, 171.6, 171.8. MS (FAB): m/z 1146.6
[MþH]þ. HRMS (FAB): m/z calcd for C53H91N7O20 [MþH]þ:
1146.6319; found: 1146.6372. MS (MALDI): m/z calcd for
C53H91N7O20 [MþH]þ: 1146.632; found: 1146.663.
2.42e2.54 (m, 124H), 2.71 (t, 48H, J¼6 Hz), 2.82 (t, 112H, J¼7 Hz),
3.34e3.40 (m, 64H), 3.83 (s, 8H), 4.09e4.15 (m, 112H), 5.14 (s, 8H),
5.24 (br, 32H), 7.64 (s, 4H). 13C NMR (CDCl3)
d
¼28.5, 32.7, 32.9, 39.6,
49.6, 49.8, 52.0, 62.2, 63.7, 79.4, 155.7, 172.1, 172.1. MS (MALDI): m/z
calcd for C470H800N72O184 [MþH]þ: 10,404.756; found: 10,403.867.
5.7.3. Synthesis of CD Boc-G3 16. Compound 15 (196 mg,
0.17 mmol) was added to TFA (3.0 ml) and stirred for 2 h. The
concentrated compound was lyopholized. The deprotected com-
pound was dissolved in CH3CN (1.6 ml), and 1 (1.4 g, 6.56 mmol)
References and notes
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and triethylamine (318 ml, 2.30 mmol) were added. After stirring for
7 days at 45 ꢁC under a nitrogen atmosphere, the reaction mixture
was extracted with ethyl acetate and dried over Na2SO4. Purifica-
tion of the concentrated compound was carried out by column
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n
(cmꢀ1) 3727, 3374,
¼1.42 (s, 72H), 2.22
2976, 2838, 2102, 2014, 1731. 1H NMR (CDCl3)
d
(t, 1H, J¼2 Hz), 2.45 (t, 28H, J¼7 Hz), 2.70 (t, 12H, J¼6 Hz), 2.81 (t,
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2
(50 mg,
0.090 mmol) and 14 (201 mg, 0.41 mmol) were dissolved in a 4:1
solvent ratio of THF/H2O (4.1 ml), and CuSO4$5H2O (21 mg,
0.082 mmol) and sodium ascorbate (33 mg, 0.17 mmol) were
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6820
H. Akiyama et al. / Tetrahedron 69 (2013) 6810e6820
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