Solid-phase construction: High efficiency dendrimer synthesis using AB3 isocyanate-type monomers

Article abstract of DOI:10.1016/S0040-4020(03)00463-0

Solid-phase synthesis is an ideal tool for reactions that require high concentrations and excess reagents and forcing chemical conditions. One such chemistry is that required for dendrimer construction. In this paper the synthesis of a series of symmetric

Full text of DOI:10.1016/S0040-4020(03)00463-0

Tetrahedron 59 (2003) 3945–3953
Solid-phase construction: high efficiency dendrimer synthesis
using AB3 isocyanate-type monomers
Sylvain Lebreton, Siew-Eng How, Monika Buchholz, Boon-Ek Yingyongnarongkul
*
and Mark Bradley
Department of Chemistry, University of Southampton, Southampton, Hampshire SO17 1BJ, UK
Received 10 October 2002; revised 23 December 2002; accepted 10 January 2003
Abstract—Solid-phase synthesis is an ideal tool for reactions that require high concentrations and excess reagents and forcing chemical
conditions. One such chemistry is that required for dendrimer construction. In this paper the synthesis of a series of symmetrical AB3 isocyanate-
type monomers is reported and used for the preparation of tri-branched dendrimers on the solid-phase. This method not only allows isolable
dendrimer but can generate high-loading supports and devices for multivalent presentation. q 2003 Elsevier Science Ltd. All rights reserved.
1. Introduction
dendrimers, as excess reagents can be used to ensure
complete reaction of the surface functional groups and can
be removed easily by simple washing of the resin-bound
dendrimer.6 This methodology can be employed to prepare
dendrimers which can be cleaved from the support via the
use of a linker, or non-cleavable dendrimers which are
covalently bound to a number of polymeric based supports.
The former also allows the preparation of multivalent
constructs which can be used to probe weak non-bonding
interactions (‘cluster effect’)7 while the latter is useful for
increasing single-bead loading capacities. Both of these
approaches have been widely investigated within our group
since we reported the first solid-phase synthesis of PAMAM
dendrimer in 1997.8 The strategy was found to be an
efficient method for enhancing resin loading by at least one
order of magnitude and allowed structure characterization
for compounds cleaved from a single bead.9 However, some
limitations were found to arise due to the formation of
structural defects, especially at high generation, caused by
retro-Michael addition or intramolecular lactam for-
mation.10 These side-reactions, which accumulate through-
out dendrimer growth result in a decrease of terminal
functional groups and the loss of symmetry. The AB3-type
monomers do not suffer from many of these potential
limitations, and thus they became an attractive target for
allowing solid-phase dendrimer synthesis, and allowing a
much more rapid increase in functionality. In this paper, the
synthesis of these monomers and their use in solid-phase
dendrimer synthesis for the preparation of homogeneous
dendrimers is described.
Dendrimers are macromolecules whose synthesis is gener-
ally based on a repetitive sequence of reaction steps, with
each complete sequence creating a new generation of
dendrimer.1 This class of polymer are highly symmetrical
monodisperse molecules consisting of three distinct regions:
a core, from which they generally emanate, successive
branching units and a peripheral multivalent surface. It is a
combination of all these three regions that determine the
physical and chemical properties of the molecule.2 Since the
3
4
¨
early work of Vogtle and Tomalia, research in the area has
evolved to enable the development of engineered dendritic
macromolecules with unique chemical and physical proper-
ties for applications in the fields of medicinal chemistry,
supramolecular chemistry, and catalysis among others.5
Divergent synthesis, based on the growth of the dendrimer
from the central core to the periphery, is probably the most
efficient and rapid procedure to construct dendrimers of high
generation. This method is characterized by a rapid increase
in the number of functional groups on the periphery
However, due to the exponential increase in the number
of terminal functional groups as well as the increasing steric
congestion at the periphery during dendrimer growth,
incomplete reactions can occur, especially at higher
generations, which can produce structural defects.2 In
addition, excess reagents needed to drive reactions to
completion may render purification difficult. Solid-phase
synthesis has proven to be a powerful tool in making
Keywords: solid-phase chemistry; dendrimers; high-loading; multivalency;
isocyanate.
2. Results and discussion
*
Corresponding author. Tel.: þ44-23-8059-3598; fax: þ44-23-8059-6766;
e-mail: mb14@soton.ac.uk
Isocyanate monomers of the AB3-type are advantageous
0040–4020/03/$ - see front matter q 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0040-4020(03)00463-0

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S. Lebreton et al. / Tetrahedron 59 (2003) 3945–3953
Figure 1. AB3 Isocyanate-type monomers.
building blocks both for the solid-phase synthesis of
dendrimers due to their high branching multiplicity and
the presence of a reactive isocyanate group, which allows
fast dendrimer construction without using additional
coupling reagents. Our research focused on the use of
AB3-type monomers 1 and 3a (Fig. 1) developed by
Newkome et al.11 and a series of novel monomers
developed in our group (3b, 2a–b).
method led to high yields while avoiding the use of
triphosgene.
The Boc- and Dde-protected monomers 3a and 3b were
prepared by a similar sequence using a modification of the
procedure reported by Newkome for the synthesis of 3a
(Scheme 2).16 The synthesis started from commercially
available tris-(2-cyanoethyl)nitromethane. After reduction
of the nitrile groups and protection of the resulting amines
the nitro group was reduced by hydrogenation using
commercial Raney Nickel as the catalyst. Treatment of
the amines 9a and 9b with DMAP and Boc2O gave the
isocyanates 3a and 3b in good yields.
The terminal amino groups of 2 and 3 were protected either
with the Boc (tert-butoxycarbonyl) or the Dde-group (4,4-
dimethyl-2,6-dioxocyclohexylidene)ethyl), which is a
highly selective protecting group for primary amines that
can be cleaved under very mild conditions (2% H2NNH2 in
DMF).12
2.2. Non-cleavable dendrimers
2.1. Monomer synthesis
In 2000 the use of monomer 1 for the preparation of high-
loading polystyrene resin beads was reported by our
group.17 Coupling of monomer 1 onto 250–300 mm
aminomethyl polystyrene resin beads was followed by
displacement of the methyl ester groups with propane-1,3-
diamine to afford the first generation dendrimer (Scheme 3).
Repetition of the cycle gave Generation 2.0 dendrimer
functionalized resin with a loading of 36 nmol/bead (85% of
the theoretical maximum loading). These beads were found
to swell to a much greater degree in more polar solvents than
the starting aminomethyl polystyrene resin, this is due to the
polar nature of the dendrimer portion. Dendrimer synthesis
was also carried out on 400–500 mm aminomethyl
The synthesis of monomers 2a13 and 2b started from the
tris-nitrile 4, which was easily prepared by the Michael
addition14 of acrylonitrile to 1,1,1-tris-(hydroxymethyl)
aminomethane (Scheme 1). The amine was protected either
by the Boc or Cbz (benzyloxycarbonyl) group. Reduction of
the nitrile group with the borane–tetrahydrofuran complex
afforded the tris-amines, which were treated crude with
Boc2O (di-tert-butyl dicarbonate) or DdeOH (2-acetyl-
dimedone) to afford the protected tris-amines 6a and 6b.
After removal of the protecting groups the isocyanates 2a
and 2b were prepared using stoichiometric amounts of
DMAP (4-dimethylaminopyridine) and Boc2O.15 This
Scheme 1. Synthesis of 1!3 C-branched tris-based isocyanate monomers. (i) CbzCl, sat. NaHCO3 (91%) or Boc2O, Et3N (quant.); (ii) BH3·THF, dioxane,
558C; (iii) Boc2O, Et3N (37% for the two steps) or DdeOH, DIPEA (42% for the two steps); (iv) 10% Pd/C, H2 (7a 92%) or 20% TFA in CH2Cl2 (7b, 87%);
(v) Boc2O, DMAP (2a 91%, 2b 94%).

S. Lebreton et al. / Tetrahedron 59 (2003) 3945–3953
3947
Scheme 2. Synthesis of 1!3 C-branched isocyanate monomers. (i) BH3·THF, dioxane or THF, reflux; (ii) Boc2O, NEt3, MeOH, reflux (71% for the two steps)
or DdeOH, MeOH (25% for the two steps); (iii) H2, Raney-nickel, EtOH, 808C (9a 81%, 9b 75%); (iv) Boc2O, DMAP, CH2Cl2 (3a 93%, 3b 69%).
Scheme 3. Solid-phase dendrimer synthesis up to Generation 2.0. (i) Monomer 1, 2a,b, or 3a, DIPEA, DMAP, CH2Cl2 and/or DMF; (ii) propane-1,3-diamine,
MeOH or DMSO; (iii) 10–40% TFA/CH2Cl2, then 5% DIPEA/DMF; (iv) 5% H2NNH2/DMF.
polystyrene beads, giving a loading of 116 and 230 nmol/
bead for Generation 1.0 and 2.0, respectively.
0.46 mmol/g).16 The supports were reacted with the
monomer in the presence of DIPEA and a catalytic
amount of DMAP to afford Generation 0.5 resin-bound
dendrimer. Treatment with TFA afforded Generation 1.0
(Scheme 3). This process was repeated to form higher
generations. Each coupling was monitored by a qualitative
ninhydrin test18 and repeated if necessary until a satisfactory
test was obtained. A portion of each full generation was then
coupled to FmocGlyOH for loading determination via a
quantitative Fmoc test.19 The results are summarized in
Table 1.
Although this monomer afforded high-loading beads, one
limitation came from possible macrocyclisation during ester
displacement. The use of AB3-type monomer 3a was
therefore investigated, as cross-coupling side-reactions
with this monomer are prevented. Solid-phase dendrimer
synthesis was carried out on polystyrene aminomethyl
resins (250–300 mm, 1.38 mmol/g and 75–150 mm,
0.92 mmol/g) as well as on TentaGele resin (160 mm,
Table 1. Loading measurements on resin-bound dendrimers
Monomer
Support
Initial loading
Gen. 1.0
Gen. 2.0
Gen. 3.0
Overall loading increase
3a
PS (250–300 mm)
PS (75–150 mm)
TG (160 mm)
18
0.7
0.7
41 (76%)a
1.5 (77%)
1.4 (68%)
96 (78%)
3.6 (79%)
3.2 (77%)
Broken
6.6 (61%)
6.9 (72%)
£5.3
£10.0
£10.2
2a
2b
PS (250–300 mm)
PS (75–150 mm)
TG (160 mm)
18
0.7
0.7
46 (85%)
1.7 (87%)
1.4 (68%)
120 (87%)
4.5 (86%)
3.5 (84%)
Broken
12.5 (94%)
9.0 (72%)
£6.7
£18.9
£13.2
PS (200–250 mm)
13
36 (92%)
90 (83%)
£6.9
Loadings are in nmol/bead.
a
Yields based on theoretical maximum loading.

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S. Lebreton et al. / Tetrahedron 59 (2003) 3945–3953
Scheme 4. Solid-phase dendrimer synthesis up to Generation 3.0.
A 10-fold increase in loading for standard polystyrene resin
and TentaGele was observed, however, a major problem
was encountered with the larger polystyrene beads. Treat-
ment of Generation 1.5 dendrimer-beads with 40% TFA in
DCM, while attempting to form Generation 2.0, resulted in
breakage of the majority of the beads. Observations under
the microscope revealed that the damage was caused by the
fast release of carbon dioxide and isobutylene bubbles
formed during the Boc deprotection with TFA. It was found
that progressively increasing the TFA concentration could
overcome bead breakage, allowing the formation of
Generation 2.0 dendrimer with a loading of nearly
100 nmol/bead. However, these resin beads were not
physically robust enough to accommodate formation of
Generation 3.0 when using the same deprotection pro-
cedure. Kinetic studies based on Fmoc release of Fmoc-
Glycine derivatised resin-bound dendrimers with 20%
piperidine in DMF were carried out on the large polystyrene
beads (250–300 mm). Full deprotection required 22 and
57 min for Generations 1.0 and 2.0, respectively, compared
to 5 min for the aminomethyl resin alone. This significant
drop in reaction kinetics is the consequence of steric
congestion interfering with site accessibility.
addition of subsequent monomer units onto the dendritic
assembly, although this effect was less pronounced in the
case of TentaGele resin. A 19-fold increase in loading was
possible on standard polystyrene resin after the third
generation. The same trend was obtained with 250–
300 mm polystyrene resin, with Generation 2.0 exhibiting
an average of 120 nmol/bead. Kinetic studies showed that
the longer dendritic branches significantly increased reac-
tion kinetics. However, the mechanical instability of the big
polystyrene beads towards Boc deprotection remained, with
the formation of generation 3.0 resulting in broken beads.
Monomer 2b, where the peripheral amino groups were
protected with the Dde protecting group rather than Boc,
was then investigated as a means of loading enhancement.
One advantage of this monomer is that Dde deprotection can
be carried out under mild condition without the release of
any gas. The solid-phase dendrimer synthesis strategy
remained the same (Scheme 3) only the deprotection of
the Dde protected amines was achieved with a 5% hydrazine
in DMF overnight. Slightly smaller resin beads (200–
250 mm) were used. Generation 2.0 exhibited a loading
capacity of 90 nmol/bead, which corresponded to the same
overall loading increase as that with monomer 2a on 250–
300 mm polystyrene resin beads (Table 1).
In order to improve reaction kinetics on the dendrimer
functionalized resins, a new isocyanate monomer was
designed. It was anticipated that increasing the flexibility
of the dendritic assembly should help to maximize site
accessibility, which in turn should result in speeding up the
reaction rates, as well as allowing higher loadings to be
obtained. Therefore monomers 2a13 and 2b (where the
monomer chain length was increased from three to five
atoms) were investigated.
2.3. Cleavable dendrimers
Solid phase dendrimer synthesis was also carried out by
attaching an acid-cleavable polyamine scaffold on poly-
styrene aminomethyl resin (75–150 mm, 0.61 mmol/g).17,20
The resulted amino derivatised resins 10 were then reacted
with the isocyanate monomer 1 and Dde-protected mono-
mers 2b and 3b in CH2Cl2/DMF (1:1) in the presence of
DIPEA and DMAP with microwave assisted heating
(1008C, 60 min) to afford the respective Generation 0.5
resin-bound dendrimers, which were further reacted with
diaminoethane in DMSO and 5% hydrazine in DMF,
respectively, to give Generation 1.0 resin-bound dendri-
mers. The processes were repeated to afford higher
dendrimer generations as outlined in Scheme 4. Each
coupling was monitored by a qualitative ninhydrin test and
The procedure for solid phase dendrimer synthesis remained
identical (Scheme 3). As anticipated increasing the length of
the dendritic branches greatly facilitated the formation of
resin beads with higher loading due to the greater degree of
flexibility that minimized peripheral steric congestion
(Table 1). The length of the spacer between the branching
center and the reactive groups dictated the successful

S. Lebreton et al. / Tetrahedron 59 (2003) 3945–3953
3949
Figure 2. Generation 0.5 and 2.5 dendrimers cleaved as TFA salts. (i) Monomer 1, 2b, or 3b, DIPEA, DMAP, CH2Cl2/DMF (1:1); (ii) ethane-1,2-diamine,
DMSO; (iii) 5% H2NNH2/DMF.
repeated if necessary to drive the reactions to completion.
Cleavage of the Generation 0.5 and 2.5 dendrimers with
TFA/DCM/H2O (9.5:0.25:0.25) gave the respective den-
drimers (11–16) as TFA salts (Fig. 2). Attempts to obtain
molecular ion of the Generation 2.5 products by ESI and
MALDI-TOF were unsuccessful.
reported on the d scale in ppm and are referenced to residual
non-deuterated solvent resonances. Mass spectra were
obtained on a VG Platform single quadrupole mass
spectrometer in electrospray positive mode. IR spectra
were obtained on a Biorad Golden gate FTS 135
spectrometer with neat compounds as oils or solids with
stretches reported in cm21. Melting points are uncorrected.
Microwave assisted heating was carried out by irradiating
the mixture in a Smith Synthesizer at 2.45 GHz (available
from Personal Chemistry). Commercially available reagents
were used without further purification. THF was freshly
distilled under nitrogen from a solution of sodium and
benzophenone. Purifications by column chromatography
were done on silica gel 60 (230–400 mesh) purchased from
Merck. Compounds 1, 3a, and 4 were synthesized according
to literature procedures.11,14
Various ligands were also successfully coupled to the
constructed dendrimer prior to cleavage (results will be
published elsewhere). The respective conjugates displayed
multiple copies of ligands are excellent candidates for
further biological evaluation21 due to their high degree of
define structure and multivalency.
3. Conclusion
4.1.1. [2-(2-Cyanoethoxy)-1,1-bis-(2-cyanoethoxy-
methyl)ethyl]carbamic acid benzyl ester (5a). To a
stirred solution of the amine 4 (2.00 g, 7.13 mmol) in
ethyl acetate (15 mL) was added a saturated aqueous
solution of NaHCO3 (10 mL). Benzyl chloroformate
(1.59 g, 9.32 mmol) was added and the reaction mixture
was stirred for 2 h. Water (80 mL) was added and the
reaction mixture was extracted with ethyl acetate
(2£80 mL). The combined organic phases were washed
with water (2£100 mL). The organic layer was dried over
MgSO4, concentrated in vacuo and purified by column
chromatography (ethyl acetate/hexane 6:4) to give com-
pound 5a (1.69 g, 58%) as a colorless oil.
AB3-type monomers have proved to be very efficient for
rapid dendrimer synthesis on solid supports. Non-
cleavable dendrimers constructed on resins allow signifi-
cant loading amplification. These resins have been
successfully tested in a number of syntheses in our
group and are used extensively in the area of single-bead
screening. Synthesis and cleavage of these dendrimeric
architectures showed that the reactions had proceeded as
anticipated. The strategy allows multivalent constructs to
be synthesized which subsequently can be screened for
biological activity.
4. Experimental
1
Rf¼0.21 (ethyl acetate/hexane 6:4); H NMR (300 MHz,
CDCl3) d 2.58 (t, J¼5.9 Hz, 6H), 3.67 (t, J¼5.9 Hz, 6H),
3.79 (s, 6H), 5.06 (s, 2H), 5.14 (s, 1H), 7.30–7.38 (m, 5H);
13C NMR (75 MHz, CDCl3) d 18.9, 58.9, 65.9, 66.7, 69.4,
118.1, 128.4, 128.7, 136.7, 155.2; IR (neat) n 2251, 1716,
1236, 1105; HRMS (ESþ) calcd for C21H26O5N4Na
[MþNa]þ: 437.1795, found: 437.1800.
4.1. General information
NMR spectra were recorded using a Bruker AC-300
spectrometer operating at 300 MHz for 1H and 75 MHz
for 13C, or a Bruker DPX 400 spectrometer operating at
400 MHz for 1H and 100 MHz for 13C. Chemical shifts are

3950
S. Lebreton et al. / Tetrahedron 59 (2003) 3945–3953
4.1.2. [2-(2-Cyanoethoxy)-1,1-bis-(2-cyanoethoxy-
methyl)-ethyl]carbamic acid tert-butyl ester (5b). To a
solution of amine 4 (1.32 g, 4.70 mmol) in CH2Cl2 (5 mL)
and MeOH (5 mL) was added triethyl amine (5.64 mmol)
and Boc2O (1.23 g, 5.64 mmol). The reaction mixture was
stirred overnight and poured into water (100 mL) and
extracted with CH2Cl2 (2£100 mL). The combined organic
layers were washed with water (200 mL), dried over
Na2SO4 and evaporated to dryness. The crude product was
purified by column chromatography (CH2Cl2/MeOH 98:2)
to obtain 5b (1.85 g, 100%) as a colorless oil.
BH3·THF complex (1 M solution in THF, 39 mL, 39 mmol)
in dioxane (10 mL) was stirred at 558C for 5 h. After work-
up the crude product was dissolved in MeOH (15 mL) and
DIPEA (7.76 mmol) was added. A solution of DdeOH
(3.53 g, 19.41 mmol) in MeOH (2 mL) was added and the
reaction mixture was stirred at 458C overnight. The reaction
mixture was poured into brine and extracted with CH2Cl2
(2£250 mL). The combined organic layers were washed
with water (400 mL), dried over Na2SO4 and evaporated to
dryness. The crude product was purified by column
chromatography (CH2Cl2/MeOH 97:3) to give 6b (2.41 g,
42% for two steps) as a colorless oil.
Rf¼0.62 (CH2Cl2/MeOH 14:1); 1H NMR (400 MHz,
CDCl3) d 1.48 (s, 9H), 2.65 (t, J¼6.0 Hz, 6H), 3.74 (t,
J¼6.0 Hz, 6H), 3.82 (s, 6H); 13C NMR (100 MHz, CDCl3) d
19.0, 28.5, 58.7, 66.0, 69.6, 79.9, 118.1, 155.0; IR (neat) n
2251, 1706, 1106; HRMS (ESþ) calcd. for C18H28N4O5Na
[MþNa]þ: 403.1952, found: 403.1941.
Rf¼0.44 (CH2Cl2/MeOH 14:1); 1H NMR (400 MHz,
CDCl3) d 0.96 (s, 18H), 1.33 (s, 9H), 1.85 (quint,
J¼6.0 Hz, 6H), 2.29 (s, 12H), 2.49 (s, 9H), 3.41 (t,
J¼6.0 Hz, 6H), 3.46 (t, J¼6.0 Hz, 6H), 3.59 (s, 6H),
13.37 (br s, 3H); 13C NMR (100 MHz, CDCl3) d 19.3, 29.8,
29.9, 30.8, 31.4, 42.1, 54.4, 60.0, 69.8, 71.3, 80.6, 109.4,
156.4, 175.0, 199.3; IR (neat) n 1710, 1636; HRMS (ESþ)
calcd for C48H77N4O11 [MþH]þ: 885.5583, found:
885.5586.
4.2. General procedure A (reduction of tris-nitriles with
BH3·THF complex)
To a stirred solution of tris-nitrile was slowly added
BH3·THF complex. The solution was refluxed (times
given below). After cooling, 2 M HCl was added until pH
1–2 was reached. The mixture was then neutralized with
NaOH, and solvent was removed in vacuo. Yields and
spectroscopic data are given below.
4.2.3. {3-[2-Amino-3-(3-tert-butoxycarbonylaminopro-
poxy)-2-(3-tert-butoxycarbonyl aminopropoxy-methyl)-
propoxy]propyl} carbamic acid tert-butyl ester (7a).
Tris-carbamate 6a (1.31 g, 1.80 mmol) and 10% Pd/C
(1.00 g) in EtOH (20 mL) was stirred under an atmosphere
of hydrogen for 18 h. The product was filtrated over celite
and washed thoroughly with EtOH before removal of the
solvent in vacuo. The crude product was purified by column
chromatography (CH2Cl2/MeOH 95:5) to give 7a (0.98 g,
92%) as a colorless oil.
4.2.1. {3-[2-Benzyloxycarbonylamino-3-(3-tert-butoxy-
carbonylamino-propoxy)-2-(3-tert-butoxycarbonyl-
aminopropoxymethyl)propoxy]propyl}carbamic acid
tert-butyl ester (6a). According to general procedure A,
carbamate 5a (2.13 g, 5.14 mmol) and BH3·THF (1.0 M
solution in THF, 25.7 mL, 25.7 mmol) in dry dioxane
(20 mL) were stirred for 3 h. After the work-up the crude
tris-amine was dissolved in methanol (20 mL). Triethyl
amine (0.323 g, 23.13 mmol) and Boc2O (5.05 g,
23.13 mmol) were added and the reaction mixture was
refluxed at 858C for 4 h. The solvent was then removed in
vacuo. The whitish gel obtained was extracted with ethyl
acetate (2£100 mL). The combined organic phases were
then washed with water (2£100 mL). The organic phase was
dried over MgSO4 and concentrated in vacuo and the crude
product was purified by column chromatography (hexane/
ethyl acetate 1:1) to obtain 6a (1.37 g, 37% for the two
steps) as a colorless oil.
Rf¼0.52 (CH2Cl2/MeOH/NH3 9:1:0.1); 1H NMR
(300 MHz, CDCl3) d 1.43 (s, 27H), 1.74 (quint, J¼5.9 Hz,
6H), 1.92 (br s, 2H), 3.20 (dt, J¼5.9, 5.9 Hz, 6H), 3.33 (s,
6H), 3.50 (t, J¼5.9 Hz, 6H), 5.00 (br s, 3H); 13C NMR
(75 MHz, CDCl3) d 28.6, 29.7, 38.7, 56.2, 69.9, 73.1, 79.1,
156.1; IR (neat) n 3347, 1688, 1167, 1103; HRMS (ESþ)
calcd. for C28H57O9N4 [MþH]þ: 593.4120, found:
593.4117.
4.2.4. [2-{3-1-(4,4,-Dimethyl-2,6-dioxocyclohexyl-
idene)ethylamino]propoxy}-1,1-bis-{3-[1-(4,4-dimethyl-
2,6-dioxocyclohexylidene)ethylamino]propoxymethyl}-
ethyl]amine (7b). The protected amine 6b (0.18 g,
0.20 mmol) in 20% TFA in CH2Cl2 (5 mL) was stirred for
30 min. The solvent was removed in vacuo and CH2Cl2
(100 mL) was added to the crude product and washed with
saturated aqueous NaHCO3 solution (100 mL) and water
(100 mL). The organic layer was dried over Na2SO4 and the
solvent was removed in vacuo and the crude product was
purified by column chromatography (CH2Cl2/MeOH/Et3N
94:6:1) to afford 7b (0.14 g, 87%) as a colorless oil.
1
Rf¼0.19 (hexane/ethyl acetate 6:4); H NMR (300 MHz,
CDCl3) d 1.43 (s, 27H), 1.72 (quint, J¼5.9 Hz, 6H), 3.18
(dt, J¼5.9 Hz, 6H), 3.49 (t, J¼5.9 Hz, 6H), 3.66 (s, 6H),
4.93 (br s, 3H), 5.06 (s, 2H), 5.35 (s, 1H), 7.29–7.36
(m, 5H); 13C NMR (75 MHz, CDCl3) d 28.6, 29.7, 38.4,
58.8, 66.4, 69.7, 70.1, 79.1, 128.2, 128.6, 136.5, 155.3,
156.1; IR (neat) n 1689, 1247, 1166, 1102; HRMS (FABþ)
calcd for C36H63O11N4 [MþH]þ: 727.4497, found:
727.4493.
Rf¼0.34 (CH2Cl2/MeOH 14:1); 1H NMR (400 MHz,
CDCl3) d 1.04 (s, 18H), 2.00 (m, 6H), 2.37 (s, 12H), 2.59
(s, 9H), 3.58 (m, 6H), 3.64 (t, J¼5.3 Hz, 6H), 3.76 (s, 6H),
13.29 (br s, 3H); 13C NMR (100 MHz, CDCl3) d 18.3, 28.6,
29.1, 30.5, 42.6, 53.1, 59.3, 69.9, 70.9, 108.4, 174.0, 198.3;
IR (neat) n 1676; HRMS (ESþ) calcd for C43H69N4O9
[MþH]þ: 785.5059, found: 785.5045.
4.2.2. [2-{3-[1-(4,4-Dimethyl-2,6-dioxocyclohexyl-
idene)ethylamino]propoxy}-1,1-bis-{3-[1-(4,4-dimethyl-
2,6-dioxocyclohexylidene)ethylamino]propoxymethyl}-
ethyl] carbamic acid tert-butyl ester (6b). According to
general procedure A, compound 5b (2.46 g, 6.47 mmol) and

S. Lebreton et al. / Tetrahedron 59 (2003) 3945–3953
3951
4.2.5. {3-[3-(3-tert-Butoxycarbonylaminopropoxy)-2-(3-
tert-butoxycarbonylaminopropoxymethyl)-2-isocyan-
atopropoxy]propyl} carbamic acid tert-butyl ester (2a).
The amine 7a (4.54 g, 7.66 mmol) was stirred in THF
(15 mL) at 2138C for 5 min. DMAP (1.03 g, 8.42 mmol)
was added and the reaction mixture was stirred for another
5 min before a solution of Boc2O (2.34 g, 10.72 mmol) in
THF (2 mL) was added dropwise. The mixture was stirred at
2138C for 30 min, then concentrated in vacuo. The
resulting residue was purified by column chromatography
(SiO2, ethyl acetate/hexane 1:1) to give 2a (4.29 g, 91%) as
a yellowish oil.
4.2.8. 4-Amino-1,7-bis{N-[1-(4,4-dimethyl-2,6-dioxo-
cyclohexylidene)ethyl]amino}-4-{3-N-[1-(4,4-dimethyl-
2,6-dioxocyclohexylidene)ethylamino]propyl}heptane
(9b). A mixture of nitro compound 8b (3.44 g, 4.74 mmol)
and Raney Nickel (1.0 g) in ethanol (100 mL) was heated at
608C for 2 days. After cooling the reaction mixture was
filtered over celite and the solvent was removed in vacuo.
The residue was dissolved in ether (100 mL) and then
washed with 20% aqueous AcOH (4£20 mL). The aqueous
phase was basified to pH 13–14 and extracted with CH2Cl2
(4£50 mL). The organic phase was dried (MgSO4) and
concentrated in vacuo to give the amine 9b (2.47 g, 75%) as
a yellow oil.
1
Rf¼0.35 (ethyl acetate/hexane 1:1); H NMR (300 MHz,
CDCl3) d 1.41 (s, 27H), 1.75 (quint, J¼5.9 Hz, 6H), 3.20
(dt, J¼5.9, 5.9 Hz, 6H), 3.45 (s, 6H), 3.53 (t, J¼5.9 Hz, 6H),
4.95 (br s, 3H); 13C NMR (75 MHz, CDCl3) d 28.6, 29.7,
38.5, 64.1, 70.0, 71.6, 79.1, 127.8, 156.2; IR (neat) n 2247,
1704, 1167, 1104; HRMS (ESþ) calcd. for C29H54O10N4Na
[MþNa]þ: 641.3732, found: 641.3733.
1H NMR (300 MHz, CDCl3) d 1.03 (s, 18H), 1.43–1.49 (m,
6H), 1.76–1.68 (m, 8H), 2.36 (s, 12H), 2.57 (s, 9H), 3.40
(td, J¼5.9, 6.1 Hz, 6H), 13.5 (br s, 3H); 13C NMR (75 MHz,
CDCl3) d 18.2, 23.6, 28.4, 30.3, 37.2, 43.9, 53.0, 108.0,
173.6; IR (neat) n 1630, 1565, 1460, 1330; HRMS (ESþ)
calcd. for C40H62N4O6Na [MþNa]þ: 717.4562, found:
717.4588.
4.2.6. [2-{3-{1-(4,4-Dimethyl-2,6-dioxocyclohexylidene)-
ethylamino]-propoxy}-1,1-bis-{3-[1-(4,4-dimethyl-2,6-
dioxocyclohexylidene)ethylamino]propoxymethyl}-
ethyl]isocyanate (2b). Amine 7b (1.41 g, 1.79 mmol) and
DMAP (262 mg, 2.15 mmol) was dissolved in THF (20 mL)
and stirred at 2138C for 5–10 min. To this solution was
added dropwise a solution of Boc2O (508 mg, 2.33 mmol) in
THF (5 mL). The reaction mixture was stirred for 30 min.
The solvent was removed in vacuo. The crude product was
purified by column chromatography (CH2Cl2/MeOH 98:2)
to obtain 2b (1.37 g, 94%) as a colorless oil.
4.2.9. 4-Isocyanato-1,7-bis{N-[1-(4,4-dimethyl-2,6-dioxo-
cyclohexylidene)ethyl]amino}-4-{3-N-[1-(4,4-dimethyl-
2,6-dioxocyclohexylidene)ethylamino]propyl}heptane
(3b). To a mixture of amine 9b (1.22 g, 1.76 mmol) and
DMAP (0.215 g, 1.76 mmol) in DCM (15 mL) was added
dropwise a solution of Boc2O (0.538 g, 2.46 mmol) in
CH2Cl2 (7 mL). The resulting mixture was stirred for 1 h.
The solvent was evaporated in vacuo and the crude product
was purified by column chromatography (CH2Cl2/MeOH
95:5) to yield 3b (0.876 g, 69%) as a colorless oil.
Rf¼0.49 (CH2Cl2/MeOH 14:1); 1H NMR (400 MHz,
CDCl3) d 1.06 (s, 18H), 1.98 (m, 6H), 2.39 (s, 12H), 2.60
(s, 9H), 3.52 (s, 6H), 3.55 (m, 6H), 3.61 (s, 6H), 13.50 (br s,
3H); 13C NMR (CDCl3, 100 MHz) d 18.2, 28.7, 29.7, 30.5,
40.6, 53.3, 64.3, 68.7, 72.0, 108.3, 127.9, 174.1, 198.2; IR
Rf¼0.26 (CH2Cl2/MeOH 95:5); 1H NMR (300 MHz,
CDCl3) d 1.01 (s, 18H), 1.65–1.85 (m, 12H), 2.34 (s,
12H), 2.55 (s, 9H), 3.38–3.42 (m, 6H), 13.5 (br s, 3H); 13C
NMR (75 MHz, CDCl3) d 18.0, 23.6, 28.2, 30.1, 36.5, 43.2,
53.5, 62.5, 107.9, 122.8, 173.5; IR (neat) n 2255, 1635,
1570; HRMS (ESþ) calcd for C41H60N4O7Na [MþNa]þ:
743.4354, found: 743.4390.
(neat)
n
2242, 1636; HRMS (ESþ) calcd for
C44H66N4O10Na [MþNa]þ: 833.4671, found: 833.4659.
4.2.7. 4-Nitro-1,7-bis{N-[1-(4,4-dimethyl-2,6-dioxocyclo-
hexylidene)ethyl]amino}-4-{3-N-[1-(4,4-dimethyl-2,6-
dioxocyclohexylidene)ethyl]amino}propylheptane (8b).
According to general procedure A, tris-(2-cyanoethyl)nitro-
methane (5.00 g, 22.7 mmol) and BH3·THF complex (1 M
solution in THF, 120 mL, 120 mmol,) in THF (50 mL) were
refluxed for 4 h. After work-up, the crude product was
dissolved in MeOH (50 mL) and a solution of DdeOH
(15.0 g, 82 mmol) in CH2Cl2 (10 mL) was added. The
reaction mixture was stirred overnight and the solvent was
evaporated in vacuo. The crude product was purified by
column chromatography (ethyl acetate/MeOH 9:1) to yield
8b (4.03 g, 25%) as a white solid.
4.3. General procedure for the solid-phase synthesis of
Generation n.5 dendrimers (n50, 1 or 2)
Isocyanate monomers 1, 2 or 3 (1.5 equiv.), DMAP
(0.1 equiv.) and DIPEA (1.5 equiv.) were dissolved in
CH2Cl2/DMF (1:1). The resulting solutions were added to
preswollen aminomethyl polystyrene resin or resin-bound
dendrimer generations n.0 (n¼0, 1 or 2). The mixtures were
shaken for 1–5 days or irradiated with microwaves at 1008C
for 60 min and reaction completion monitored by a
qualitative ninhydrin test and FTIR. Couplings were
repeated until a satisfactory ninhydrin test was obtained.
The resins were filtered and washed thoroughly with DMF,
DMF/THF (1:1), THF, MeOH, and CH2Cl2 and dried in
vacuo to yield the corresponding resin-bound dendrimers.
1
Rf¼0.24 (CH2Cl2/MeOH 95:5); mp: 165–1678C; H NMR
(400 MHz, CDCl3) d 1.05 (s, 18H), 1.64–1.72 (m, 6H),
2.06–2.11 (m, 6H), 2.37 (s, 12H), 2.58 (s, 9H), 3.46 (td,
J¼5.7, 6.4 Hz, 6H), 13.59 (br s, 3H); 13C NMR (100 MHz,
CDCl3) d 18.2, 23.9, 28.6, 30.5, 33.1, 43.2, 53.2, 93.2,
108.4, 174.0, 197.8; IR (neat) n 1629, 1560, 1535, 1335;
HRMS (ESþ) calcd for C40H60N4O8Na [MþNa]þ:
747.4303, found: 747.4338.
4.4. Coupling of resin-bound dendrimer Generation n.5
with diaminoethane (n50 or 1)
Diaminoethane (20 equiv.) in DMSO was added to the
resin-bound dendrimer Generation n.5 (n¼0 or 1) syn-
thesised from monomer 1 and the mixtures shaken for 3–7

3952
S. Lebreton et al. / Tetrahedron 59 (2003) 3945–3953
days. Reaction completion was monitored by FT-IR. The
resins were filtered and washed thoroughly with DMF,
DMF/THF (1:1), THF, MeOH, and CH2Cl2 and dried in
vacuo to yield the resin-bound dendrimers.
(400 MHz, CD3CN) d 1.03 (s, 36H), 1.84 (t, J¼6.0 Hz, 4H),
1.91 (quint, J¼6.0 Hz, 6H), 2.41 (s, 24H), 2.56 (s, 18H),
2.95 (m, 4H), 3.27 (t, J¼6.0 Hz, 4H), 3.52 (t, J¼6.0 Hz,
12H), 3.58 (m, 12H), 3.64 (s, 12H), 13.26 (s, 6H); 13C NMR
(100 MHz, CD3CN) d 17.6, 26.9, 28.3, 29.6, 35.2, 41.0,
44.2, 51.0, 59.0, 68.3, 69.7, 106.9, 115.1, 158.4, 159.9,
175.5, 198.0; IR (neat) n 1731, 1637, 1569, 1106; MS
4.5. General procedure for the deprotection of Boc
group
(ESþ) m/z 585.4 [(Mþ3H)3þ, 30%], 877.4 [(Mþ2H)2þ
,
To the respective preswollen resin-bound dendrimer
Generation n.5 (n¼0, 1 or 2) in CH2Cl2 was added 10%
TFA in CH2Cl2 (5£30 min) with occasional stirring and
washed with CH2Cl2. The same procedure was repeated
with 25% TFA in CH2Cl2 and finally 40% TFA in CH2Cl2
(2£15 min). The resin was washed with CH2Cl2, DMF, 4%
DIPEA in DMF, DMF, THF, THF/CH2Cl2, and CH2Cl2,
before being dried in vacuo at 408C.
60%].
4.7.4. Dendrimer 14. Cleavage from 50 mg of resin
afforded 14 (36 mg, 5%) as a yellowish oil. 1H NMR
(400 MHz, CD3CN) d 1.04 (s, 324H), 1.70 (s, 52H), 1.93
(m, 108H), 2.45 (s, 216H), 2.56 (s, 162H), 2.97 (s, 4H), 3.20
(s, 4H), 3.54 (m, 156H), 3.60 (m, 156H), 3.65 (s, 156H),
13.22 (s, 54H); 13C NMR (100 MHz, CD3CN) d 18.0, 26.9,
28.2, 29.7, 37.1, 41.3, 46.5, 50.4, 59.4, 68.3, 69.8, 106.9,
114.5, 158.0, 159.2, 175.1, 198.2; IR (neat) n 3329, 1777,
1670, 1562, 1148.
4.6. General procedure for the deprotection of Dde
group
To the respective preswollen resin-bound dendrimer
Generation n.5 (n¼0 or 1) in DMF were added 5%
N2H4.H2O in DMF and the mixtures shaken for 4 h.
Progress of reactions was monitored by the ninhydrin test.
The resins were filtered and washed thoroughly with DMF,
DMF/THF (1:1), THF, MeOH, and CH2Cl2 and dried in
vacuo to yield the corresponding resin-bound dendrimers.
4.7.5. Dendrimer 15. Cleavage from 50 mg of resin
afforded 15 (20 mg, 62%) as a colorless oil. 1H NMR
(400 MHz, CD3CN) d 1.79 (m, 4H), 2.52 (t, J¼6.0 Hz,
12H), 2.91 (br s, 4H), 3.23 (t, J¼6.0 Hz, 4H), 3.56 (s, 12H),
3.64 (s, 18H), 3.65 (t, J¼6.0 Hz, 12H); 13C NMR
(100 MHz, CD3CN) d 26.8, 34.2, 34.9, 43.8, 50.9, 58.9,
66.5, 69.3, 159.5, 171.8; IR (neat) n 3381, 1737, 1653, 1200;
MS (ESþ) m/z 471.8 [(Mþ2H)2þ, 65%], 942.4 [(MþH)þ,
100%].
4.7. General procedure for the acidolytic liberation of
dendrimers from the solid support
4.7.6. Dendrimer 16. Cleavage from 50 mg of resin
afforded 16 (54 mg, 15%) as a colorless oil. 1H NMR
(400 MHz, CD3CN) d 1.85 (s, 4H), 2.50 (s, 48H), 2.57 (t,
J¼6.0 Hz, 108H), 3.04 (s, 4H), 3.21–3.35 (m, 100H), 3.59
(s, 108H), 3.62 (s, 48H), 3.67 (s, 162H), 3.69 (t, J¼6.0 Hz,
156H); 13C NMR (100 MHz, CD3CN) d 27.0, 34.1, 35.7,
39.2, 39.7, 45.0, 51.0, 59.3, 66.6, 67.0, 69.4, 114.9, 158.0,
159.1, 171.8, 173.2; IR (neat) n 3350, 1737, 1163.
The dendrimers of Generation n.5 (n¼0, 1 or 2) attached to
the polyamine scaffold 10 were washed with CH2Cl2 before
the addition of TFA/CH2Cl2/H2O (9.5:0.25:0.25). The
mixtures were shaken for 3 h. The solutions were filtered
and the resins were washed with CH2Cl2 and dried in vacuo
to yield dendrimers of Generation n.5 (n¼0, 1 or 2) as TFA
salts. The yield of products was calculated based on the
molecular weight of fully protonated TFA salts relative to
initial loading of aminomethyl resin (0.61 mmol/g).
Acknowledgements
4.7.1. Dendrimer 11. Cleavage from 50 mg of resin
afforded 11 (26 mg, 36%) as a yellowish oil. 1H NMR
(400 MHz, CD3CN) d 0.98 (s, 36H), 1.59 (m, 12H), 1.68 (m,
12H), 1.77 (m, 4H), 2.34 (s, 24H), 2.50 (s, 18H), 2.91 (m,
4H), 3.18 (m, 4H), 3.42 (m, 12H), 13.32 (s, 6H); 13C NMR
(100 MHz, CD3CN) d 17.4, 22.4, 26.6, 27.0, 29.5, 32.0,
35.3, 43.3, 44.0, 51.4, 56.9, 106.9, 115.2, 158.4, 163.4,
174.0, 198.2; IR (neat) n 1775,1564, 1152; MS (ESþ) m/z
525.3 [(Mþ3H)3þ, 100%], 787.1 [(Mþ2H)2þ, 87%],
1572.7 [(MþH)þ, 5%].
The authors would like to thank the BBSRC, the Deutsche
Forschungsgemeinschaft, the University of Malaysia Sabah,
the Ministry of Science, Technology and Environment,
Malaysia, and Ramkhamhaeng University, Thailand, for
funding. We thank Personal Chemistry for the use of the
Smith Synthesiser.
References
4.7.2. Dendrimer 12. Cleavage from 50 mg of resin
afforded 12 (32 mg, 5%) as a yellowish oil. 1H NMR
(300 MHz, CD3CN) d 0.98 (s, 324H), 1.59 (m, 156H), 1.68
(m, 160H), 2.34 (s, 216H), 2.50 (s, 162H), 2.91 (m, 4H),
3.18 (m, 4H), 3.42 (m, 156H), 13.22 (s, 54H); 13C NMR
(100 MHz, CDCl3) d 17.1, 22.5, 24.6, 27.2, 29.1,31.9, 43.5,
51.7, 57.5, 106.8, 164.0, 174.0, 197.0; IR (neat) n 3329,
1775, 1562, 1152.
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