Electronic supplementary information for Chem. Commun.
This journal is © the Royal Society of Chemistry 2004.
Self-Immolative Dendrimer Biodegradability
by Multi-Enzymatic Triggering
Roey J. Amir, and Doron Shabat*
School of Chemistry, Raymond and Beverly Sackler Faculty of Exact Sciences,
Tel-Aviv University, Tel Aviv 69978 Israel.
Electronic Supplementary Information
Experimental
General methods. Thin layer chromatography (TLC): silica gel plates Merck 60 F 254: compounds were
visualized by irradiation with UV light and/or by treatment with a solution of 25 g phosphomolybdic acid,
10 g Ce(SO4)2·H2O, 60 mL conc. H2SO4 and 940 mL H2O followed by heating and/or by staining with a
solution of 12 g 2,4-dinitrophenylhydrazine in 60 mL conc. H2SO4, 80 mL H2O and 200 mL 95% EtOH
followed by heating. – Flash chromatography (FC): silica gel Merck 60 (particle size 0.040-0.063 mm),
eluent given in parentheses. 1H NMR spectra were measured using Bruker Avance operated at 200 MHz.
The chemical shifts are expressed in  relative to TMS ( = 0 ppm) and the coupling constants J in Hz. The
spectra were recorded in CDCl3 as solvent at room temperature unless stated otherwise. All general
reagents, including salts and solvents, were purchased from Aldrich (Milwaukee, MN). PGA was
purchased from Sigma.
Abbreviations. Boc- t-butoxycarbonyl, CDI- Carbonyl diimidazol, DCM- Dichloromethane, DMAPDimethyl aminopyridine, DMF- Dimethylformamide, EtOAc- Ethyl acetate, Et3N- Triethyl amine, Hex- nHexane, MeOH- Methanol, PBS- Phosphate buffer saline, PNP- 4-Nitrophenyl, THF – tetrahydrofuran.
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Synthesis of Dendrons 1-3:
Compound 4
Commercially available N-Boc-N-methylethylenediamine (100 mg, 0.574 mmol) and Et 3N (160 L, 1.15
mmol) were dissolved in 10 mL DCM. The solution was cooled to 0 0C and phenylacetyl chloride (84 L,
0.63 mmol) was added dropwise. The reaction was allowed to warm to RT, diluted by EtOAc (100 ml) and
washed with brine. The organic layer was dried over magnesium sulfate, and the solvent was removed
under reduced pressure. The crude product was used without further purification.
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H NMR (200MHz,CDCl3):  = 7.33-7.23 (5H, m); 3.53 (2H, s); 3.36-3.32 (4H, m); 2.80 (3H, s); 1.43 (9H,
s).
Compound 1
Compound 4 was deprotected with 2 ml TFA to remove the Boc group. The excess of the acid was
removed under reduced pressure and the residue was dissolved in 2 ml DMF. Bis(4-nitrophenyl) carbonate
(262 mg, 0.86 mmol) and 0.5 ml Et3N were added and the solution was stirred for 10 minutes. After
completion the mixture was diluted with EtOAc (100 ml) and washed with brine. The organic layer was
dried over magnesium sulfate, and the solvent was removed under reduced pressure. The crude product was
purified by column chromatography on silica gel (EtOAc:Hex=3:1) to give pure compound 1 in the form of
pale yellow oil (164 mg, 80% overall yield for two steps).
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H NMR (200MHz,CDCl3):  = 8.28 (2H, d, J = 9 Hz); 7.32-7.21 (7H, m); 3.58-3.43 (6H, m); 3.08 (3H, s).
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C NMR (200MHz,CDCl3):  = 171.6, 156.2, 154.0, 144.8, 134.7, 129.3, 128.9, 127.3, 125.0, 122.2, 48.7,
43.7, 37.9, 35.3. MS (FAB) Calcd for C18H19N3O5 358.1 [MH] +, found 358.2.
Compound 5
Commercially available phenylacetic acid (3 gr, 22 mmol) was dissolved in THF (60 ml). CDI (3.6 gr 22
mmol) was added and the release of CO2 was observed. The reaction was monitored by TLC
(EtOAc:Hex=1:1) for the complete disappearance of starting materials. The activated phenylacetyl
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Electronic supplementary information for Chem. Commun.
This journal is © the Royal Society of Chemistry 2004.
imidazol amid1 was then added dropwise to a stirred solution of diethylenetriamine (1.2 ml, 11 mmol) in
THF (40 ml). After completion the solvent was removed under reduced pressure. The residue was dissolved
in DCM and washed with water. The organic layer was dried over magnesium sulfate, and the solvent was
removed under reduced pressure. The crude product was used for the next step without further purification
(2.8 gr, 75%).
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H NMR (200MHz,CDCl3):  = 7.34-7.23 (10H, m); 5.82 (2H, bs); 3.55 (4H, s); 3.23 (4H, q, j = 5.8 Hz);
2.62 (4H, t, j = 5.8 Hz). 13C NMR (200MHz,CDCl3):  = 171.3, 135.1, 129.4, 129.0, 127.3, 48.3, 43.8,
39.4.
Compound 2
Compound 5 (100 mg, 0.29 mmol) was dissolved in DMF (3 ml). Et3N (122 l, 0.88 mmol) was added,
followed by the addition of bis(4-nitrophenyl)-carbonate (134 mg, 0.44 mmol) and the solution was stirred
for 10 minutes. After completion the mixture was diluted with EtOAc (100 ml) and washed with brine. The
organic layer was dried over magnesium sulfate, and the solvent was removed under reduced pressure. The
crude product was purified by column chromatography on silica gel (EtOAc) to give pure compound 2 in
the form of pale yellow oil (112 mg, 76%).
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H NMR (200MHz,CDCl3):  = 8.23 (2H, d, J = 9 Hz); 7.35-7.20 (12H, m); 6.51 (1H, bs); 6.06 (1H, bs);
3.60-3.32 (12H, m). 13C NMR (200MHz,CDCl3):  = 172.0, 156.0, 154.0, 144.9, 134.9, 129.4,128.9,127.3,
125.1, 122.2, 48.8, 43.6, 38.7. MS (FAB) Calcd for C27H28N4O6 505.2 [MH] +, found 505.1.
Compound 6
Compound 5 (2.8 gr , 8.2 mmol) was dissolved in DMF (100 ml). Et3N (2.8 ml, 20 mmol) was added,
followed by the addition of carbonic acid 4-hydroxymethyl-phenyl ester 4-nitrophenyl ester (2.9 gr, 10
mmol) and DMAP (200 mg, 1.6 mmol). The reaction was monitored by TLC (EtOAc: MeOH=9:1). After
completion, the mixture was diluted with EtOAc (500 ml) and washed with saturated NH 4Cl and brine. The
organic layer was dried over magnesium sulfate and the solvent was removed under reduced pressure. The
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crude product was purified by column chromatography on silica gel (EtOAc: MeOH=9:1) to give pure
compound 6 in the form of pale yellow oil (3.0 gr, 76%).
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H NMR (200MHz,CDCl3):  = 7.36 (2H, d, J = 8.6 Hz); 7.29-7.21 (10H, m); 7.02 (2H, d, J = 8.6 Hz);
6.43 (1H, bs); 6.13 (1H, bs); 4.69 (2H, s); 3.54-3.37 (12H, m).
C NMR (200MHz,CDCl3):  = 172.1,
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155.2, 153.6, 150.1, 135.1, 129.2, 128.6, 127.9, 127.8, 127.0, 63.8, 48.1, 43.2, 38.4.
Compound 7
Compound 6 (2.3 gr, 4.7 mmol) was dissolved in EtOAc (20 ml). PNP-chloroformate (1.9 gr, 9.4 mmol)
was added, followed by the addition of DMAP (1.1 gr, 9.4 mmol). The reaction was monitored by TLC
(EtOAc: MeOH=9:1). After completion the mixture was diluted with EtOAc (400 ml) and washed with
saturated NH4Cl and brine. The organic layer was dried over magnesium sulfate, and the solvent was
removed under reduced pressure. The crude product was purified by column chromatography on silica gel
(EtOAc: MeOH=9:1) to give pure compound 7 in the form of pale yellow oil (1.4 gr, 46%).
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H NMR (200MHz,CDCl3):  = 8.26 (2H, d, J = 9.1 Hz); 7.45 (2H, d, J = 8.5 Hz); 7.36 (2H, d, J = 9.1
Hz); 7.30-7.20 (10H, m); 7.10 (2H, d, J = 8.5 Hz); 6.58 (1H, bs); 6.24 (1H, bs); 4.28 (2H, s); 3.53-3.37
(12H, m). 13C NMR (200MHz,CDCl3):  = 171.8, 155.4, 154.9, 152.3, 151.6, 145.2, 135.0, 131.4, 129.9,
129.2, 128.7, 127.0, 125.2, 121.9, 121.8, 70.2, 48.3, 43.3, 38.6.
Compound 3
Diethylenetriamine (32 l, 0.30 mmol) and Et3N (164 l, 1.2 mmol) were dissolved in DMF (3 ml).
Compound 7 (388 mg, 0.60 mmol) in DMF (7 ml) was added dropwise, and the solution was stirred for 10
minutes. The reaction was monitored by TLC (EtOAc: MeOH=9:1). After completion, bis(4-nitrophenyl)
carbonate (180 mg, 0.60 mmol) was added, and the reaction mixture was stirred for 1 hr in RT. The
solution was diluted with EtOAc (200 ml) and washed with brine. The organic layer was dried over
magnesium sulfate, and the solvent was removed under reduced pressure. The crude product was purified
by column chromatography on silica gel (EtOAc: MeOH= 9:1) to give pure compound 3 in the form of
white powder (206 mg, 54%).
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Electronic supplementary information for Chem. Commun.
This journal is © the Royal Society of Chemistry 2004.
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H NMR (200MHz,CDCl3):  = 8.06 (2H, d, J = 9.0 Hz); 7.35-7.10 (26H, m); 6.94 (4H, d, J = 8.4 Hz);
6.57 (2H, bs); 6.29 (2H, bs); 5.58 (1H, bs); 5.44 (1H, bs); 5.05 2H, s), 5.02 (2H, s); 3.50 (8H, s) 3.40-3.20
(24H, m). 13C NMR (200MHz,CDCl3):  = 172.0, 156.7, 156.1, 155.1, 153.9, 151.0, 144.7, 135.0, 133.8,
129.4, 128.8, 127.2, 125.0, 122.3, 121.7, 121.2, 66.0, 48.7, 48.3, 43.5, 38.6, 32.6; HRMS (MALDI) Calcd
for C69H74N10O16 1321.5177 [MNa] +, found 1321.5169.
Reporter Release Analysis – General Protocol.
Dendrons 1 and 2 (2 L of 10 mM stock solution in DMSO) were dissolved in 98 L of PBS 7.4 to give a
final concentration of 200 M. Dendron 3 (2 L of 10 mM stock solution in DMSO:Chremephor EL(1:1))
was dissolved in 98 l of PBS 7.4 to give a final concentration of 200 M. All solutions were kept at 370C.
PGA stock solution in PBS 7.4 was used to activate the dendrons.
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p.nitrophenol
G0
G1
0.9
Absorbance
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0
250
300
350
400
450
500
 [nm]
Figure S1: UV-Visible spectra of p-nitrophenol and of Dendrons 1and 2 in PBS 7.4 at concentrations used
for the kinetic measurements.
References:
1.
S. P. Rannard, N. J. Davis. Organic Letters 2000, 2(14), 2117-2120.
2.
A. P. Krapcho, C. S. Kuell. Synthetic Communications 1990, 20(16), 2559-2564.
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