High Molecular Weight Polyacrylamides by ATRP: Enabling
Advancements in Water-Based Applications
Eric A. Appel, Jesús del Barrio, Xian Jun Loh, Joseph Dyson, and Oren A. Scherman
Melville Laboratory for Polymer Synthesis, Department of Chemistry, University of Cambridge,
Lensfield Road, Cambridge CB2 1EW, United Kingdom
S.1 Instrumentation and Materials
1 H NMR (400 MHz)
spectra was recorded using a Bruker Avance QNP 400. Chemical shifts are recorded in
ppm (δ ) in D2 O with the internal reference set to δ 4.79 ppm. Gel permeation chromatography (GPC) was
carried out in either tetrahydrofuran (THF) or in water (H2 O). Aqueous GPC was performed on a Shodex
glucose column with a Shimadzu SPD-M20A prominence diode array detector, Optilab refractive index
detector and dynamic light scattering detector (both Wyatt). Samples were filtered over 0.2 µ m PVDF filters before injection using a 0.6 mL / min flow rate. THF GPC was performed on two Jordi 5µ m DVB
columns connected in series with a SPD-M20A prominence diode array detector and refractive index detector (both Shimadzu) calibrated in relation to poly(methyl methacrylate) standards. Samples were filtered
over 0.45 µ m PTFE filters before injection using a 1.0 mL / min flow rate.
N, N’-dimethylacrylamide (DMAm), N-hydroxyethylacrylamide (HEAm) was purchased from Aldrich and
purified on a basic alumina column to remove the inhibitor and stored over 4Å molecular sieves. Nisopropylacrylamide (NIPAm) was purchased from Aldrich and recrystallized from a mixture of toluene
and hexane. All other materials were purchased from Sigma-Aldrich and used as received.
S.2 General Synthetic Protocols
General synthesis of Poly(N-isopropylacrylamide). N-Isopropylacrylamide (0.49 g, 4.3 mmol) and methyl
2-chloropropanoate (MCP) (1.1 mg, 0.98 µ L, 8.7 µ mol, Target DP = [M]/[I] = 500) were dissolved in a 1:4
(v/v) mixture of deionized water and ethanol (0.75 mL) and the solution was degassed by bubbling nitrogen
for 5 min. CuCl (0.9 mg, 8.7 µ mol) and Me6 TREN (2.0 mg, 2.4 µ L, 8.7 µ mol) were dissolved in a 1:4
(v/v) mixture of deionized water and ethanol (0.25 mL) and the solution was degassed by bubbling nitrogen for 5 min. The monomer and initiator solution was then added via syringe into the copper and ligand
solution and the mixture was allowed to stir at 20 o C for 30 min. Aliquots were removed periodically to
analyze conversion. The reaction mixture was then diluted with ethanol (5 mL) and precipitated into cold
diethyl ether. The resulting residue was redissolved in chloroform, dried over magnesium sulfate, filtered
and precipitated from cold diethyl ether. The solid was filtered and dried under vacuum yielding the title
compound as a white solid (0.45 g, 93 %). 1 H-NMR Spectroscopy (D2 O, 400 MHz) δ (ppm) = 3.67-3.50
(1H, (CH3 )2 -CH-NH-), 2.2-0.8 (polymer backbone and (CH3 )2 -CH-NH-). GPC (H2 O): Mn (PDI) = 56.0
kDa (1.06).
S1
(a)
(b)
PDI
PDI
PDI
Mn (kDa)
Mn
PDI
Mn (kDa)
Mn
[M]o/[I]o
Conversion (%)
Figure S1: (a) Mn and PDI versus conversion for the CuCl/Me6 -TREN catalyzed SET-LRP of Nisopropylacylamide (NIPAm) with [NIPAm]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1. (b) Plot demonstrating the effect of [NIPAm]o /[MCP]o on Mn and PDI for the polymerization of NIPAm.
Table S1: Polymerization of N-isopropylacrylamide using the CuCl/Me6 -TREN catalyst system.
Entry
1
2
3
2
[M]o /[I]o Solvent (EtOH:H2 O) Mn (kDa)a
50
50:50
17
50
70:30
14
100
50:50
16
1000
50:50
192
a
determined by GPC using H2 O as eluent.
100
2
Mn from GPC (kDa)
PDI
50
1.4
1.5
1.3
PDI
1
40
40
Mn (kDa)
60
Ln([M]o/[M])
Conversion (%)
1.5
60
Conversion
Ln([M]o/[M])
80
PDIa
1.10
1.17
1.08
1.22
30
1.2
20
0.5
20
1.1
10
kp = 0.0201 min-1
0
0
10
20
30
40
50
60
Ieff = 98%
0
0
70
0
10
20
30
40
50
1
Mn Theoretical (kDa)
Time (min)
Figure S2: (a) Conversion and ln([M]o /[M]) versus time for the CuCl/Me6 -TREN catalyzed SET-LRP of Nisopropylacylamide (NIPAm) with [NIPAm]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1. (b) Plot demonstrating the effect of initiator efficiency for the polymerization of NIPAm.
S2
General synthesis of Poly(N, N’-Dimethylacrylamide). N, N’-Dimethylacrylamide (0.43 g, 4.3 mmol) and
methyl 2-chloropropanoate (MCP) (1.1 mg, 0.98 µ L, 8.7 µ mol, Target DP = [M]/[I] = 500) were dissolved
in a 1:4 (v/v) mixture of deionized water and ethanol (0.75 mL) and the solution was degassed by bubbling
nitrogen for 5 min. CuCl (0.9 mg, 8.7 µ mol) and Me6 TREN (2.0 mg, 2.4 µ L, 8.7 µ mol) were dissolved
in a 1:4 (v/v) mixture of deionized water and ethanol (0.25 mL) and the solution was degassed by bubbling
nitrogen for 5 min. The monomer and initiator solution was then added via syringe into the copper and
ligand solution and the mixture was allowed to stir at 20 o C for 30 min. Aliquots were removed periodically
to analyze conversion. The reaction mixture was then diluted with ethanol (5 mL) and precipitated into cold
diethyl ether. The resulting residue was redissolved in chloroform, dried over magnesium sulfate, filtered
and precipitated from cold diethyl ether. The solid was filtered and dried under vacuum yielding the title
compound as a white solid (0.40 g, 94 %). 1 H-NMR Spectroscopy (D2 O, 400 MHz) δ (ppm) = 3.50-3.45
(6H, (CH3 )2 N-COO-CH-CH2 ), 2.45-2.00 (1H, (CH3 )2 N-COO-CH-CH2 ), 1.95-1.35 (2H, H2 N-COO-CHCH2 ). GPC (H2 O): Mn (PDI) = 40.6 kDa (1.08).
(a)
(b)
Mn
Mn
PDI
PDI
Mn (kDa)
PDI
Mn (kDa)
PDI
[M]o/[I]o
Conversion (%)
Figure S3: (a) Mn and PDI versus conversion for the CuCl/Me6 -TREN catalyzed SET-LRP of N, N’dimethylacrylamide (DMAm) with [DMA]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1. (b) Plot demonstrating the effect of [DMAm]o /[MCP]o on Mn and PDI for the polymerization of DMAm.
S3
1.6
60
Conversion
Ln([M]o/[M])
Mn from GPC (kDa)
PDI
1.4
40
1.4
1
30
0.8
0.6
20
0.4
10
30
1.3
20
1.2
PDI
40
Mn (kDa)
1.2
Ln([M]o/[M])
Conversion (%)
50
1.5
50
10
0.2
kp = 0.0474 min-1
0
0
5
10
15
0
0
1.1
Ieff = 78%
0
20
5
10
15
20
25
30
1
Mn Theoretical (kDa)
Time (min)
Figure S4: (a) Conversion and ln([M]o /[M]) versus time for the CuCl/Me6 -TREN catalyzed SET-LRP of
N, N’-dimethylacylamide (DMAm) with [DMAm]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1. (b) Plot
demonstrating the effect of initiator efficiency for the polymerization of DMAm.
S4
General synthesis of Poly(N-hydroxyethylacrylamide). N-Hydroxyethylacrylamide (0.5 g, 4.3 mmol)
and methyl 2-chloropropanoate (MCP) (1.1 mg, 0.98 µ L, 8.7 µ mol, Target DP = [M]/[I] = 500) were dissolved in a 1:1 (v/v) mixture of deionized water and ethanol (0.75 mL) and the solution was degassed by
bubbling nitrogen for 5 min. CuCl (0.9 mg, 8.7 µ mol) and Me6 TREN (2.0 mg, 2.4 µ L, 8.7 µ mol) were
dissolved in a 1:1 (v/v) mixture of deionized water and ethanol (0.25 mL) and the solution was degassed by
bubbling nitrogen for 5 min. The monomer and initiator solution was then added via syringe into the copper
and ligand solution and the mixture was allowed to stir at 20 o C for 30 min. Aliquots were removed periodically to analyze conversion. The reaction mixture was then diluted with ethanol (5 mL) and precipitated into
acetone. The solid was filtered and dried under vacuum yielding the title compound as a white solid (0.48 g,
96 %). 1 H-NMR Spectroscopy (D2 O, 400 MHz) δ (ppm) = 3.50-3.32 (2H, HO-CH2 -CH2 ), 3.25-2.87 (2H,
HEAm, HO-CH2 -CH2 ), 2.2-0.8 (polymer backbone). GPC (H2 O): Mn (PDI) = 59.2 kDa (1.10).
(b)
PDI
PDI
Mn (kDa)
Mn
Mn (kDa)
Mn
PDI
PDI
(a)
[M]o/[I]o
Conversion (%)
Figure S5: (a) Mn and PDI versus conversion for the CuCl/Me6 -TREN catalyzed SET-LRP of Nhydroxyethylacylamide (HEAm) with [HEAm]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1. (b) Plot
demonstrating the effect of [HEAm]o /[MCP]o on Mn and PDI for the polymerization of HEAm.
Table S2: Polymerization of N-hydroxyethylacrylamide using the CuCl/Me6 -TREN catalyst system.
Entry
1
2
a
[M]o /[I]o Solvent (EtOH:H2 O) Conversion (%)a Mn (kDa)b PDIb
5000
80:20
36
180
1.68
5000
30:70
85
405
1.75
1
b
determined by H NMR. determined by GPC using H2 O as eluent.
S5
Mn from GPC (kDa)
PDI
50
0.8
1.4
40
0.4
20
0.2
10
kp = 0.1537 min
0
0
1
2
3
4
1.3
PDI
30
40
Mn (kDa)
0.6
Ln([M]o/[M])
Conversion (%)
50
1.5
60
1
Conversion
Ln([M]o/[M])
30
1.2
20
1.1
10
-1
0
0
5
Ieff = 82%
1
0
Time (min)
10
20
30
40
50
Mn Theoretical (kDa)
Figure S6: (a) Conversion and ln([M]o /[M]) versus time for the CuCl/Me6 -TREN catalyzed SET-LRP of
N-hydroxyethylacylamide (HEAm) with [HEAm]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1. (b) Plot
demonstrating the effect of initiator efficiency for the polymerization of HEAm.
S6
General synthesis of Poly(acrylamide). Acrylamide (0.31 g, 4.3 mmol) and methyl 2-chloropropanoate
(MCP) (1.1 mg, 0.98 µ L, 8.7 µ mol, Target DP = [M]/[I] = 500) were dissolved in a 7:3 (v/v) mixture of
deionized water and ethanol (0.75 mL) and the solution was degassed by bubbling nitrogen for 5 min. CuCl
(0.9 mg, 8.7 µ mol) and Me6 TREN (2.0 mg, 2.4 µ L, 8.7 µ mol) were dissolved in a 7:3 (v/v) mixture of
deionized water and ethanol (0.25 mL) and the solution was degassed by bubbling nitrogen for 5 min. The
monomer and initiator solution was then added via syringe into the copper and ligand solution and the mixture was allowed to stir at 20 o C for 30 min. Aliquots were removed periodically to analyze conversion.
The reaction mixture was then diluted with ethanol (5 mL) and precipitated into acetone. The solid was
filtered and dried under vacuum yielding the title compound as a white solid (0.29 g, 95 %). 1 H-NMR Spectroscopy (D2 O, 400 MHz) δ (ppm) = 2.45-2.00 (H2 N-COO-CH-CH2 ), 1.95-1.35 (H2 N-COO-CH-CH2 ).
GPC (DMF): Mn (PDI) = 47.9 kDa (1.16).
(a)
(b)
Mn
Mn
PDI
Conversion (%)
PDI
Mn (kDa)
PDI
Mn (kDa)
PDI
[M]o/[I]o
Figure S7: (a) Mn and PDI versus conversion for the CuCl/Me6 -TREN catalyzed SET-LRP of acylamide (Am) with [Am]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1. (b) Plot demonstrating the effect
of [Am]o /[MCP]o on Mn and PDI for the polymerization of Am.
S7
30
50
40
Mn from GPC (kDa)
PDI
25
1.4
0.8
0.4
20
20
1.3
PDI
0.6
Mn (kDa)
30
Ln([M]o/[M])
Conversion (%)
1.5
1
Conversion
Ln([M]o/[M])
15
1.2
10
0.2
10
1.1
5
kp = 0.1298 min
0
0
0
1
2
3
4
Ieff = 80%
-1
0
5
1
0
Time (min)
5
10
15
20
Mn Theoretical (kDa)
Figure S8: (a) Conversion and ln([M]o /[M]) versus time for the CuCl/Me6 -TREN catalyzed SET-LRP of
acylamide (Am) with [Am]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1. (b) Plot demonstrating the effect
of initiator efficiency for the polymerization of Am.
S8
General synthesis of Poly(NIPAM-r-HEAm) 500:500. N-Isopropylacrylamide (0.49 g, 4.3 mmol), Nhydroxyethylacrylamide (0.5 g, 4.3 mmol), and methyl 2-chloropropanoate (MCP) (1.1 mg, 0.98 µ L, 8.7
µ mol, total DP=[M]/[I]=1000) were dissolved in a 1:1 (v/v) mixture of deionized water and ethanol (2 mL)
and the solution was degassed by bubbling nitrogen for 5 min. CuCl (0.9 mg, 8.7 µ mol) and Me6 TREN
(2.0 mg, 2.4 µ L, 8.7 µ mol) were dissolved in a 1:4 (v/v) mixture of deionized water and ethanol (1.0 mL)
and the solution was degassed by bubbling nitrogen for 5 min. The monomer and initiator solution was
then added via syringe into the copper and ligand solution and the mixture was allowed to stir at 20 o C for
30 min. Aliquots were removed periodically to analyze conversion. The reaction mixture was then diluted
with ethanol (5 mL) and precipitated into cold acetone. The solid was filtered and dried under vacuum
yielding the title compound as a white solid (0.40 g, 94 %). The composition of the final copolymers was
determined by 1 H-NMR using integration of the signals for the single proton α to the amide functionality
in NIPAm and the two protons α to the hydroxyl functionality in HEAm (Figure S9 in the supporting
information). 1 H-NMR Spectroscopy (D2 O, 400 MHz) δ (ppm) = 3.67-3.50 (1H, NIPAm, (CH3 )2 -CH-NH), 3.50-3.32 (2.2H, HEAm, HO-CH2 -CH2 ), 3.25-2.87 (2.2H, HEAm, HO-CH2 -CH2 ), 2.2-0.8 (HEAm and
NIPAm polymer backbone and NIPAm (CH3 )2 -CH-NH-)). GPC (H2 O): Mn (PDI) = 66.0 kDa (1.16).
3.0
2.5
2.0
1.5
1.0
0.5
ppm
2.2
3.5
2.2
4.0
1.0
4.5
Figure S9: 1 H-NMR spectrum of poly(NIPAM-r-HEAm) (500:500) in D2 O.
S9
Table S3: Random copolymerization of acrylamides initiated with MCP and using the CuCl/Me6 -TREN
catalyst system.
Entry Monomer Aa Monomer Ba Target B (%) Experimental B (%)b Mn (kDa)c PDIc
1d
NIPAm
DMAm
10
17.7
85
1.22
NIPAm
DMAm
20
32.0
66
1.17
2d
e
NIPAm
HEAm
10
12.7
56
1.15
3
NIPAm
HEAm
20
22.2
47
1.18
4e
e
NIPAm
HEAm
30
31.7
56
1.17
5
e
NIPAm
HEAm
50
52.4
66
1.16
6
HEAm
Am
50
52.5
47
1.16
7e
a
NIPAm = N-isopropylacrylamide, HEAm = N-hydroxyethylacrylamide, DMAm = N, N’-dimethylacrylamide and Am = acrylamide. b determined by 1 H NMR. c determined by GPC using H2 O as eluent. d [M]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 1000:1:1:1 e
[M]o :[MCP]o :[CuCl]o :[Me6 -TREN]o = 500:1:1:1
S10
S.3 Characterization of Poly(N-Isopropylacrylamide) Thermoresponsive Behavior
Figure S10: Plot demonstrating the effect of poly(NIPAm) molecular weight on the observed LCST for
both heating and cooling cycles of the polymers in water. The legend refers to the DP of the poly(NIPAm)
polymers.
Figure S11: Plot demonstrating the effect of poly(NIPAm) molecular weight on the observed difference in LCST between heating and cooling of the polymers in water. The plot demonstrates a logarithmic relationship between the Mn in kDa and the change in LCST according to the following equation:
∆LCST = 1.3ln(Mn -5.4).
S11
LCSTheating (oC)
Figure S12: Plot demonstrating the effect of HEAm loading on the LCST of poly(NIPAm-r-HEAm) copolymers and the hysteresis observed between LCSTheat ing and LCSTcooling . The legend refers to the relative
loadings in mol % of NIPAm:HEAm in the copolymer.
HEAm Composition (%)
Figure S13: Plot demonstrating the change in LCSTheat ing of poly(NIPAm-r-HEAm) copolymers as a function of copolymer composition.
S12
S.3 DOX Release from Poly(N-Isopropylacrylamide) Particles
Preparation of Doxorubicin (DOX) loaded Poly(NIPAm) particles. An aqueous solution of PNIPAm
(10% w/v) and DOX (0.5% w/v) was kept at 4 ◦ C for 12 hours to allow the polymer to dissolve. The polymer/drug solution was then added dropwise into 50 mL of silicon oil kept at 40 ◦ C. The polymeric particles
were washed with hexane 5 times and air dried overnight.
DOX release (%)
Release study of DOX-loaded particles. The release profile of DOX from DOX-loaded particles was
determined by dialysis. Poly(NIPAm)-DOX particles (10 mg) were dissolved in water to form a 1 mg/mL
Poly(NIPAm)-DOX particle solution containing 45 µ g DOX. This solution was loaded into MWCO 100 kDa
dialysis tubing and dialyzed against 7 mL of phosphate buffered saline (PBS, pH 7.4) in the dark at 37 ◦ C.
At specified time points, 1 mL of the dialysis buffer was collected and replaced with equal volume of fresh
PBS. The concentrations of DOX present in the dialysate were determined by measuring absorbance at
480 nm. The concentration of DOX released from the micelles was expressed as a percentage of the total
DOX concentration and plotted as a function of time.
In order to study the effect of temperature on the kinetics of release, a similar experiment was carried
out but with the temperature maintained at 4 ◦ C.
DP 100, k = 5.57 % min-1
DP 1000, k = 3.16 % min-1
DP 5000, k = 2.32 % min-1
Time (min)
Figure S14: Plot of the release of DOX from poly(NIPAm) particles.
S13
Figure S15: Plot demonstrating the effect of poly(NIPAm) molecular weight on the observed release rate
constant of DOX from polymer particles in aqueous buffer at 37 o C. The plot demonstrates a logarithmic
relationship between the Mn in kDa and the release kinetics in %/min according to the following equation:
Release Rate = -0.85ln(Mn )+9.33.
S14