b-Cyclodextrin-Appended Giant Amphiphile
FULL PAPER
Fluorescence spectroscopy: Fluorescence spectra were measured on a
Perkin–Elmer LS 55 fluorescence spectrophotometer by using a 50 mL
quartz cuvette.
Both samples from the experiment and the control were
analysed by TEM to confirm the presence of the vesicles
and to verify their integrity after the numerous centrifuga-
tion steps.
TEM: TEM images were obtained by using a JEOL JEM 1010 micro-
scope (60 kV) equipped with a CCD camera. Samples were prepared by
pouring a drop of the aggregate suspension onto a carbon-coated copper
grid, which was allowed to dry in air. The structures were visualised with-
out further treatment. The aggregate solution was prepared by desolving
amphiphile 1 (1mg) in distilled THF (1mL). The resulting solution was
then slowly injected into ultrapure (Millipore) water (1mL). The sample
was shaken by hand to obtain a homogeneous mixture. Measurements of
the sealed sample were done after 5 min, 1d, 3 d and 6 d.
Conclusion
We have shown that PS-appended bCDs form well-defined
vesicular structures. The solvent accessibility of the bCDs
was proven by the subsequent addition of dansyl (10) and 1-
adamantol (11) apolar derivatives. We subsequently utilised
the host–guest interaction of bCD with adamantane to deco-
rate the polymersomes with PEG–adamantane-functional-
ised HRP. Activity studies confirmed that HRP remained
catalytically active and was successfully bound to the poly-
mersomes. This opens new routes for the construction of
polymersomes decorated with multiple enzymes that can act
in concert.
SEM: SEM was performed on a JEOL JSM-6330F instrument by using
the same samples as prepared for TEM. Before measurement a 1.5 nm
layer of Pd/Au was sputtered on the grids with a Cressington 208 HR
sputter coater fitted with a Cressington layer-thickness controller.
GPC: Molecular weight distributions were measured with a Shimadzu
size-exclusion column equipped with a guard column and a PL gel 5 mm
mixed
D column (Polymer Laboratories) with differential refractive
index and UV (l=254 nm and l=330 nm) detection using THF as an
eluent (1mLmin ꢀ1 at 358C). PS standards were used for calibration.
Compound 1: Nitrogen was bubbled through distilled THF for 1h prior
to use. Compounds 2 (0.05 g, 0.03 mmol) and 3 (0.24 g, 0.03 mmol) were
dissolved in THF (3 mL) under nitrogen. A solution of CuBr (0.005 g,
0.03 mmol) and PMDTA (0.007 mL, 0.03 mmol) in THF (2 mL) was
made free of air by bubbling N2 through it for 30 min and it was added to
the solution of 2 and 3. The mixture was stirred under a nitrogen atmos-
phere for 48 h. The solvent was removed by evaporation and the solid
obtained was purified by column chromatography (CHCl3!CHCl3/
MeOH 4:1v/v). Product 1 was obtained as a white solid (0.12 g, 41%).
1H NMR (400 MHz, CDCl3): d=1.25–2.10 (brm; aliphatic protons of the
PS tail) 3.15–4.00 (m, 118H), 7.09 ppm (brm; aromatic protons of the PS
tail); IR: n˜ =3023, 3053, 3088 (CH2); 2841, 2919 (CH); 1938, 1873, 1804,
1739 (C=O); 1600 (C=C); 1160 (O-Me); 693, 771 cmꢀ1 (monosubsti-
tuted benzenes); the vibration of the N3 group of compound 3 had disap-
peared in compound 1.
Experimental Section
Materials: THF was purified by distillation under nitrogen from sodium/
benzophenone. CH2Cl2 was distilled under nitrogen from CaH2. Com-
pounds 2 and 3 were prepared according to literature procedures.[7,11,12]
All other chemicals were purchased from Aldrich, Fluka or Acros and
used as received. Analytical thin-layer chromatography (TLC) was per-
formed on Merck precoated silica gel 60 F-254 plates (layer thickness
0.25 mm) with visualisation by UV irradiation at l=254 nm and/or l=
366 nm and/or staining with phosphomolybdic acid reagent or KMnO4.
Silica gel chromatography was performed by using Acros (0.035–
0.070 mm, pore diameter ca. 6 nm) silica gel. Ultra pure water (R>18
106 W) was used for the aggregation studies in aqueous solutions. Dialysis
membranes were purchased from Spectrum Laboratories and Dialyser
Tubes (Maxi, Midi or Mini) were purchased from Novagem. Peroxidase
from horseradish type VI (E.C.1.11.1.7) was purchased from Sigma.
1-[2-(1-Adamantyl)acetoxy]succinimide (13): 1-Adamantylacetic acid
(0.30 g, 1.5 mmol) and N-hydroxysuccinimide (0.19 g, 1.6 mmol) were dis-
solved in distilled CH2Cl2 under a nitrogen atmosphere. The mixture was
cooled to 08C and EDCI (0.33 g, 1.7 mmol) was added. The mixture was
stirred under a nitrogen atmosphere for 20 h. The solvent was removed
and the solid was redissolved in EtOAc and extracted with NH4Cl (2),
dried over anhydrous sodium sulfate, filtered and evaporated to dryness.
Product 13 was obtained as a white solid (0.38 g, 87%). M.p. 1348C;
1H NMR (400 MHz, CDCl3): d=1.67–1.74 (d, 12H), 2.01 (s, 3H), 2.33 (s,
2H), 2.82 ppm (s, 4H); 13C NMR (75 MHz, CDCl3): d=169.2 (CO succ),
166.2 (CO), 45.3 (adamantane-CH2-CO), 41.9, 36.5 (CH2 from adaman-
NMR spectroscopy: 1H and 13C NMR spectra were recorded on Bruker
DPX300 or Varian inova 400 spectrometers at room temperature.
1H NMR chemical shifts are reported in ppm (d) relative to tetramethyl-
silane (d=0.00) as an internal standard when measured in CDCl3, other-
wise the residual solvent peak was used as a reference.
tane), 33.1(quaternary C), 28.5 (CH from adamantane), 25.6 ppm (CH
2
succ).
Multiplicities are reported as follows: s (singlet), d (doublet), t (triplet), q
(quartet), dd (doublet of doublets), dt (doublet of triplets) or m (multip-
let). Broad peaks are indicated by the addition of br. Coupling constants
are reported as a J value in Hertz (Hz). The number of protons (n) for a
given resonance is indicated as nH, and is based on spectral integration
values. 13C NMR chemical shifts (d) are reported in ppm relative to
CDCl3 (d=77.0). Succ refers to the succinimide group.
3-{2-[2-(1-Adamantyl)acetamido]ethyl[octahexaconta(oxyethylene)]}pro-
panoic acid (14): Compound 12 (0.20 g, 0.06 mmol) was dissolved in dis-
tilled CH2Cl2 (20 mL) under a N2 atmosphere followed by the addition of
triethylamine (19 mL, 0.13 mmol). The mixture was stirred for 10 min and
compound 13 (0.03 g, 0.10 mmol) was added. The reaction was stirred for
20 h and the resulting mixture extracted with a saturated aqueous solu-
tion of NaHCO3 (2) and HCl (1m), dried over anhydrous sodium sul-
fate, filtered and evaporated to dryness. The product was precipitated in
Et2O and filtered. The solid was dissolved in CHCl3. The solvent was re-
moved in vacuo and the product was dried under high vacuum to give
compound 14 as a white powder (0.18 g, 50%). 1H NMR (300 MHz,
CDCl3): d=3.35–3.85 (brm, 276H), 2.58 (t, 2H), 1.95 (s, 3H), 1.92 (s,
2H), 1.57–1.70 ppm (m, 12H); 13C NMR (75 MHz, CDCl3): d=172.4
(CO), 170.5 (CO), 70.0 (CH2 repeating unit), 66.2, 65.3, (CH2), 51.0 (ada-
mantane-CH2-CO), 42.1, 36.3 (CH2 from adamantane), 28.1(CH from
adamantane), 38.5 (O-CH2), 36.3 (CH2-COOH), 32.2 ppm (quaternary
C).
Mass spectrometry: Electrospray LC/MS analysis was performed using a
Shimadzu LC/MS 2010A system. Matrix-assisted laser desorption/ionisa-
tion time-of-flight (MALDI-TOF) spectra were measured on a Bruker
Biflex III spectrometer and samples were prepared from solutions in
MeOH by using indoleacrylic acid (IAA) (20 mgmLꢀ1) as a matrix.
Infrared spectroscopy: IR spectra were recorded on an ATI Matson Gen-
esis Series FTIR spectrometer with a fitted ATR cell. Frequencies (n˜) are
given in cmꢀ1
.
UV/Vis spectroscopy: UV/Vis spectra were recorded on a Varian Cary
50 spectrophotometer by using a quartz cuvette. For catalysis experi-
ments, the UV/Vis spectra were recorded on an Elisa Ryder spectropho-
tometer.
1-(3-{2-[2-(1-Adamantyl)acetamido]ethyl[octahexaconta(oxyethylene)]}-
propoxy) succinimide (15): Compound 14 (0.18 g, 0.06 mmol) and N-hy-
Chem. Eur. J. 2008, 14, 9914 – 9920
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
9919