Nanofibers from oxazolidi-2-one containing hybrid foldamers: what is the right molecular size?

Article abstract of DOI:10.1002/chem.200900185

A series of oligomers of the type Boc-(L-Phe-D-Oxd)_n-OBn (Boc = tert-butoxycarbonyl; Oxd = 4-methyl-5carboxy oxazolidin-2-one; Bn = benzyl) were prepared for n = 2-5. The shortest oligomer, Boc-(L-Phe-D-Oxd)₂-OBn, aggregates and form

Full text of DOI:10.1002/chem.200900185

FULL PAPER
DOI: 10.1002/chem.200900185
Nanofibers from Oxazolidi-2-one Containing Hybrid Foldamers: What is the
Right Molecular Size?
Gaetano Angelici,[a] Giuseppe Falini,[a] Hans-Jçrg Hofmann,[b] Daniel Huster,[c]
Magda Monari,[a] and Claudia Tomasini*[a]
Dedicated to the Centenary of the Italian Chemical Society
Abstract: A series of oligomers of the
type Boc-(l-Phe-d-Oxd)n-OBn (Boc=
tert-butoxycarbonyl; Oxd=4-methyl-5-
carboxy oxazolidin-2-one; Bn=benzyl)
were prepared for n=2–5. The shortest
ture in which the oligopeptide units are
connected to each other by only one
intermolecular hydrogen bond. The
longer oligomers exhibit structural het-
erogeneity. They start to organize into
secondary structures by the formation
of intramolecular hydrogen bonds at
the pentamer level. Microscopy and
diffraction of the oligomers indicated a
crystalline character for only the short-
er ones.
Keywords: foldamers · nanofibers ·
oxazolidin-2-ones · self-assembly ·
anti-parallel beta sheet
oligomer,
Boc-(l-Phe-d-Oxd)2-OBn,
aggregates and forms a fiber-like mate-
rial with an anti-parallel b-sheet struc-
Introduction
with the aim of interfering with the pathological aggregation
of proteins.[3] Synthetic moieties with a uniquely designed
backbone are able to stabilize the aggregation of peptides
and proteins by means of noncovalent interactions, such as
hydrogen bonds and stacking interactions.[4]
Short synthetic oligomers that tend to assume an ordered
conformation have been defined as “foldamers”.[5] Among
the numerous foldamer classes,[6] the study of hybrid fol-
damers consisting of different constituents is gaining more
and more interest.[7] This is because of the endless confor-
mational variants that can be obtained by simply changing
their order and/or their total configuration. In particular, a-
and b-amino acids or their synthetic analogues, as well as
other homologous amino acids, have been introduced into
short polypeptide chains to obtain compounds with a range
of different structures.
Following these lines, we have recently described the syn-
thesis and conformational analysis of hybrid foldamers con-
taining the pseudoproline group Oxd (Oxd=4-methyl-5-car-
boxy oxazolidin-2-one).[8] The Oxd moiety can be easily ob-
tained from threonine.[9] The carbonyl group of the cycle in-
troduces a constraint that enforces the pseudo-peptide bond
to always have the trans conformation, thus favoring secon-
dary structure formation even in sequences with only a
small number of residues. The oligomers of the Boc-(l-Ala-
The self-assembly of short oligomers that form fibers and fi-
brils is of particular interest in understanding the develop-
ment of neurodegenerative diseases, such as Alzheimerꢀs
disease, diabetes, scrapie (in sheep), bovine spongiform en-
cephalopathy (BSE; in cattle), and Creutzfeld–Jakobꢀs dis-
ease (in humans). The diseases are induced by amyloid pep-
tides or proteins that tend to accumulate and aggregate in
extracellular space.[1] This process is thermodynamically fa-
vorable because b-sheet layers are the most stable proteina-
ceous superstructures in nature, as recently demonstrated.[2]
Oligopeptides could therefore be designed and prepared
[a] Dr. G. Angelici, Prof. Dr. G. Falini, Prof. Dr. M. Monari,
Prof. Dr. C. Tomasini
Dipartimento di Chimica “G. Ciamician”
Alma Mater Studiorum Universitꢁ di Bologna
Via Selmi 21, 40126 Bologna (Italy)
Fax : (+39)051-2099456
E-mail: claudia.tomasini@unibo.it
[b] Prof. Dr. H.-J. Hofmann
Institute of Biochemistry, University of Leipzig
Brꢂderstraße 34, 04103 Leipzig (Germany)
[c] Prof. Dr. D. Huster
Institute of Medical Physics and Biophysics, University of Leipzig
Hꢃrtelstr. 16-18, 04107 Leipzig (Germany)
d-Oxd)n-OBn
(Boc=tert-butoxycarbonyl;
Bn=benzyl)
series, which act as a-peptide analogues, showed a strong
tendency to form helices with 10-membered hydrogen-
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/chem.200900185.
Chem. Eur. J. 2009, 15, 8037 – 8048
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8037

bonded rings in solution. The oligomers of the Boc-(l-b3-
hPhg-d-Oxd)n-OBn series, which are a,b-hybrid peptides,
however, preferred the formation of helices with 11-mem-
bered hydrogen-bonded turns for nꢀ5. Unfortunately, these
molecules could not be crystallized and their structures
could not be determined in the solid state. Thus, the l-Ala
residue was replaced by l-Phe to facilitate crystallization.
Herein, we report on a series of oligomers of hybrid pep-
tides with the general formula Boc-(l-Phe-d-Oxd)n-OBn, in
which n=2–5. Recently, we have described the structure of
the parent molecule Boc-l-Phe-d-Oxd-OBn in solution and
in the solid state.[10] This dipeptide spontaneously forms
fibers consisting of infinite linear chains in which the paral-
lel dipeptide units are connected by only a single hydrogen
bond. In addition, the l-Phe residue exhibits b-sheet-like
backbone torsion angles. Given this, we thought it would be
of interest to investigate the behavior of oligomers of this
compound and to determine factors such as the influence of
the chain length on structure formation. In particular, the in-
creasing possibilities to form intramolecular hydrogen bonds
in the longer oligomers may compete with the formation of
intermolecular hydrogen bonds, thus influencing the type of
secondary structure that is formed.
(l-Phe-d-Oxd)-OH 1 and H-l-Phe-d-Oxd-OBn·TFA 2, re-
spectively (Scheme 1).
Scheme 1. i) H2, Pd/C (10%), MeOH, RT, 16 h; ii) TFA (18 equiv), dry
CH2Cl2, RT, 2 h; iii) HATU (1.1 equiv), Et3N (3 equiv), dry CH3CN,
40 min, RT.
Compounds 1 and 2 were then coupled by use of O-(7-
azabenzotriazol-1-yl)-N,N,N’,N’-tetramethyluronium hexa-
fluorophosphate (HATU) and triethylamine in dry acetoni-
trile under inert atmosphere providing 3 in satisfactory
yield. Deprotection of the carbobenzoxy group of 3 with H2
in methanol in the presence of Pd/C (5%) gave the corre-
sponding acid 4. Repetition of these steps afforded the com-
plete foldamer series up to Boc-(l-Phe-d-Oxd)5-OH (10).
All the deprotection steps could be performed with excel-
lent yields. The yields of the coupling steps were 65–81%.
Compound 5 was obtained in low yield due to the poor solu-
bility of both compounds 4 and 5. The oligomers 3, 5, 7, and
9 were purified by flash chromatography using a mixture of
cyclohexane and ethyl acetate as eluent.
Results and Discussion
Synthesis: The starting material, Boc-l-Phe-d-Oxd-OBn, for
the preparation of the oligomer series Boc-(l-Phe-d-Oxd)n-
OR (n=2–5; R=H, OBn) 3–10 was prepared in high yield
by the addition of Boc-l-Phe-OH to d-Oxd-OBn in the
presence of O-(benzotriazol-1-yl)tetramethyluronium hexa-
fluorophosphate (HBTU) and 1,8-diazabicycloACTHNURTGNEU[GN 5.4.0]undec-
7-ene (DBU) in dry acetonitrile, as reported previously.[11]
The d-Oxd-OBn moiety, which was utilized by our research
group several times to prepare different types of foldam-
ers,[4,5] can be easily synthesized from d-Thr.[9] The selective
deprotection of the C-terminal benzyl ester of Boc-l-Phe-d-
Oxd-OBn with H2 in methanol in the presence of Pd/C
(5%) or of the N-terminal Boc moiety with anhydrous tri-
fluoroacetic acid (TFA) in dichloromethane generated Boc-
Conformational analysis in solution: Information on the pre-
ferred conformation of the oligomers in solution was ob-
1
tained by the analysis of FTIR, H NMR, and circular di-
chroism (CD) absorption spectra. The FTIR spectra were
recorded as 3 mm solutions in dichloromethane. At this con-
centration aggregation of the oligomers is negligible.[12] The
ꢁ
FTIR absorption spectra for the N H and C=O stretching
Abstract in Italian: Eꢀ stata sintetizzata la serie di oligomeri
regions for all synthesized compounds are shown in
Figure 1. The spectra allow the detection of non-hydrogen-
bonded amides (above 3400 cmꢁ1) and hydrogen-bonded
amides (below 3400 cmꢁ1). For compound 3 only one band
centered at 3430 cmꢁ1 was observed. For compounds 7 and
9, a band centered at about 3280 cmꢁ1 appears, whereas
compound 5 exhibits two bands, one above and the other
below 3400cmꢁ1.< Thus, we can deduce that compound 3
Boc-(l-Phe-d-Oxd)n-OBn
(Boc=ter-butossi
carbonile;
Oxd=4-metil-5-carbossiossazolidin-2-one;
Bn=benzile)
contenenti l’unitꢁ l-Phe-d-Oxd ripetuta 2–5 volte. L’oligo-
mero piꢂ corto, Boc-(l-Phe-d-Oxd)2-OBn, si aggrega in fase
solida e forma un materiale fibroso avente una struttura a fo-
glietto b antiparallelo, in cui le unitꢁ oligopeptidiche sono
legate fra loro da un solo legame idrogeno. Gli oligomeri su-
periori mostrano invece un disordine strutturale ed incomin-
ciano ad organizzarsi a livello del pentamero. Analisi me-
diante microscopia ottica ed elettronica indicano che il solo
dimero ha carattere cristallino.
ꢁ
does not form intramolecular N H···O=C hydrogen bonds,
whereas equilibria between unfolded and folded structures
result for compounds 5, 7, and 9. In particular, compound 9
shows a high tendency to form intramolecular hydrogen
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Nanofibers Containing Hybrid Foldamers
FULL PAPER
ꢁ
ꢁ
imide CO N bond outside of the ring system is forced into
the trans conformation.
ꢁ
The occurrence of intramolecular N H···O=C bonds was
determined by investigation of the [D6]DMSO dependence
of the HN proton chemical shifts. DMSO is a strong hydro-
gen-bond acceptor and if bound to a free HN proton a con-
siderable downfield shift of the proton signal can be expect-
ed. The titration of DMSO with compounds 3 and 5 con-
firmed that the amide hydrogen atoms are poorly intramo-
lecular chelated in 5 and not chelated in 3. (Figure S5). Un-
fortunately, it was impossible to perform the titration for the
longer oligomers due to their low solubility in CDCl3, so the
presence of intramolecular hydrogen bonds in all the oligo-
mers was determined by recording 1H NMR spectra in
[D6]DMSO (3 mm solution) at different temperatures
(Figure 2). It is well known that small temperature coeffi-
cients of ꢂ3 ppbKꢁ1 in CDCl3 indicate the presence of hy-
drogen-bonded amide protons,[14] whereas protons that par-
ticipate in an equilibrium between hydrogen-bonded and
non-hydrogen-bonded states show larger temperature coeffi-
cients. In [D6]DMSO, the criterion for the temperature coef-
ficients is a bit higher. Thus, the values for the amide pro-
tons that form hydrogen bonds with DMSO are 4 ppbKꢁ1 or
smaller.[15] In Figure 2 it can be seen that the variation of
the chemical shifts for compounds 3 and 5 (Figure 2a and b)
are always larger than 4 ppbKꢁ1, whereas the four amide
and Boc protons of compound 9 (Figure 2d) have tempera-
ture coefficients smaller than 4 ppbKꢁ1, which is consistent
with intramolecular hydrogen-bonded amide protons.
Finally, NOESY experiments were recorded on a 10 mm
solution of compound 9 in [D6]DMSO to obtain additional
structural information to confirm the results from the pre-
ceding experiments (Figure S6 in the Supporting Informa-
tion). We found NiH!Ni+1H cross-peaks between the HN
amide hydrogen atoms (d=8.91, 8.93, 8.99, and 9.11 ppm)
and the HN carbamate hydrogen (dꢃ7.25 ppm) that account
for the formation of a folded structure.[16] Further cross-
peaks consistent with a folded conformation were recorded
(Figure S6b in the Supporting Information) and their signal
assignment is reported in Figure 3.[17] Interestingly, we found
a cross-peak between the HN amide proton and one of the
two phenylalanine Hb atoms (dꢃ2.6 ppm), but not the other
(dꢃ3.3–3.4 ppm). In addition, cross-peaks were also ob-
served between the HN amide hydrogen atoms and the phe-
nylalanine Ha (dꢃ6 ppm), the oxazolidin-2-one CHN and
CHO hydrogen atoms (dꢃ4.5 ppm), and the methyl hydro-
gen atoms (dꢃ1.3 ppm).
ꢁ
Figure 1. FTIR absorption spectra of the N H (top) and C=O (bottom)
stretching regions for 3 mm solutions of oligomers 3 (c), 5 (d), 7
(g), and 9 (a) in pure CH2Cl2 at room temperature.
bonds, as indicated by the strong stretching band at
3270 cmꢁ1. A similar trend can be observed for the CO
stretching bands with a band centered at about 1670 cmꢁ1
only observed for compounds 7 and 9. These preliminary re-
sults suggest that at least four l-Phe-d-Oxd units are re-
quired to obtain a folded structure, as was also the case for
the oligomers in the l-Ala-d-Oxd series.[9]
Additional information on the folding properties of these
compounds was obtained by means of H NMR spectroscop-
CD spectroscopy did not provide conclusive results due to
the strong absorption of the phenyl groups around
200 nm,[18] which partially covered the variation of the CD
spectra and the reversal of the Cotton effect associated with
the chain length increase (Figure S7 in the Supporting Infor-
mation).
1
ic experiments in solution. The analysis of the Ha protons of
the phenylalanine units shows that the chemical shifts are
always at d=5.5–6.0 ppm in these compounds (see the Ex-
perimental Section). The signal is more deshielded than that
usually observed for the Ha proton of the amino acid units.
This indicates the formation of non-conventional, weak
CH···O=C intramolecular hydrogen bonds[13] because the
Self-assembly of the oligomers in the solid state: Taking into
account the dependence of the oligomeric structures on the
chain length, evidenced by the different spectroscopic meth-
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C. Tomasini et al.
Figure 2. Variation in the chemical shifts d of the amide HN protons and of the Boc-NH ( ) proton of the oligomers 3 (a); 5 (b); 7 (c), and 9 (d) as a
&
~
!
*
^
function of the temperature for 3 mm samples measured in [D6]DMSO. ( : NH-1, : NH-2, : NH-3, : NH-4)
troscopy, microscopy, X-ray dif-
fraction, and solid-state NMR
spectroscopic techniques.
Given that fiber formation
occurred in the parent molecule
Boc-l-Phe-d-Oxd-OBn,[10] we
could predict that the shorter
oligomers will preferentially
form fibers, since competitive
intramolecular hydrogen bonds
are weak, or even absent. First,
the oligomer series was exam-
ined by solid-state FTIR (1%
Figure 3. Results for the NOESY experiments of Boc-(l-Phe-d-Oxd)5-OBn
9 performed on a 10 mm
[D6]DMSO solution (400 MHz spectrometer; mixing time 0.5 s; the hydrogen indicated by the arrow does not
couple with the HN hydrogen).
ꢁ
mixture in solid KBr). The N
H and C=O stretching regions
of the FTIR absorption spectra
ods in solution, we further investigated the structural behav-
ior of the oligomers in the solid state. The relationships be-
tween inter- and intramolecular hydrogen bonds might play
an important role in the formation of fibers. Thus, the oligo-
mers 3, 5, 7, and 9 were analyzed by solid-state FTIR spec-
of oligomers 3, 5, 7, and 9 are shown in Figure 4. From the
NH stretching we can see that all the oligomers have chelat-
ed hydrogen atoms although the stretching bands have dif-
ferent appearances; compounds 3 and 5 show a double band
with peaks at 3350 and 3320 cmꢁ1, whereas 7 and 9 exhibit
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Nanofibers Containing Hybrid Foldamers
FULL PAPER
Figure 5. Optical micrograph of fiber-like material of 3 formed after the
slow evaporation of a 1:1 cyclohexane/ethyl acetate solution. Scale bar
50 mm.
show any crystalline habit and birefringence under crossed
polarizers was always absent.
The oligomers were also analyzed by SEM and TEM and
the important results are shown in Figure 6. Under SEM
analysis the filamentous crystals of compound 3 can be seen
to align in parallel clusters (Figure 6a) and at higher mag-
nification (Figure 6b) the filaments were seen to be 2–5 mm
wide and <1 mm thick ribbon-like structures. Oligomer 5
did not form a precipitate of crystalline habit, however, on
its surface filamentous structures are visible (Figure 6c).
The oligomers 7 and 9 form materials in which typical fea-
tures of crystals or fibers could not be observed. Precipitates
from oligomers 5, 7, and 9 were also morphologically ana-
lyzed by TEM. Only oligomer 5 showed the presence of a fi-
brous material dispersed through an amorphous precipitate
(Figure 6d). These fibers have a ribbon-like structure with a
width of 10–15 nm and appear very thin. They are branched
and tend to form aggregates.
A deeper inspection of the behavior in the solid state was
obtained by analysis with X-ray powder diffraction (Fig-
ure S13 in the Supporting Information). Oligomer 3 showed
ꢁ
Figure 4. FTIR absorption spectra of the N H (top) and C=O (bottom)
stretching regions for a 1% solid mixture of oligomers 3 (c), 5 (d),
7 (g), and 9 (a) with KBr at room temperature.
only a broad band centered at 3270 and 3400 cmꢁ1 respec-
tively. A similar outcome was observed in the CO region,
where the two shorter oligomers show similar bands with
multiple peaks, whereas the two longer oligomers have only
two very broad bands.
Compounds 3, 5, 7, and 9 were then analyzed by micro-
scopy techniques. All samples were prepared by overnight
evaporation of a solution containing 50 mg of each com-
pound in a 1:1 cyclohexane/ethyl acetate solution (1 mL).
For compound 3, the formation of a white solid composed
of microscopic fiber-like crystals was observed by optical mi-
croscopy (Figure 5 and Figure S12 in the Supporting Infor-
mation). These crystals formed a highly crystalline material,
as shown by their high birefringence, and clearly showed a
preferential direction of growth. The crystals were found to
reach millimeter lengths and are a few micrometers wide.
The optical observation of oligomers 5, 7, and 9 did not
Figure 6. SEM (a, b) pictures of oligomer 3. SEM (c) and TEM (d) pic-
tures of oligomer 5.
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C. Tomasini et al.
a diffraction pattern characterized by the presence of sharp
peaks (average full-width half maximum (FWHM)=0.28)
indicating that the material is highly crystalline and in agree-
ment with the morphological observations. The diffraction
pattern of oligomer 5 shows several broad diffraction peaks
(average FWHM=0.388) indicating a reduced crystallinity.
These peaks correspond to lattice distances of 17, 14, 8.8,
7.4, 5.0, 4.1, and 3.4 ꢅ. The diffraction patterns obtained
from oligomers 7 and 9 exhibit only two broad diffraction
peaks corresponding to lattice distances of 14.0 and 5.0 ꢅ.
These peaks are also present in the diffraction pattern of 5.
It is important to note that the peaks at 14.0 and 5.0 ꢅ
could correspond to intersheet and interstrand separations,
respectively, in an amyloid-like cross-b-structure.[19,20]
The aggregation state of the oligomers was also investigat-
ed by differential scanning calorimetry (DSC). The heating
profile (Figures S8–S11) of compound 3 showed only a
sharp endothermic peak at 473 K, whereas that of 5 showed
a broad endothermic peak at
Figure 7. X-ray structure and numbering scheme of oligomer 3.
455 K, followed by an exother-
mic peak at 450 K and, finally,
an endothermic peak at 489 K.
The heating profile of com-
pound 7 did not show any ther-
mal event associated with the
Table 1. Selected backbone torsion angles[a] for oligomer 3.
C13-C14-N2-C15 (l-Phe1)
N2-C14-C13-N1
C8-C9-N1-C13 (Oxd1)
N1-C9-C8-N3
ꢁ134(2) (f1)
153(2) (y1)
51(2) (f2)
C27-C7-N3-C8 (l-Phe2)
N3-C7-C27-N4
C32-C30-N4-C27 (Oxd2)
N4-C30-C32-O11
ꢁ133(2) (f3)
160(2) (y3)
57(2) (f4)
ꢁ148(2) (y2)
33(2) (y4)
foldamer with an increase of [a] In degrees, see Figure 7 for numbering.
temperature, whereas com-
pound 9 showed a weak endo-
thermic peak at 463 K.
nomer and dimer units of l-Phe-d-Oxd provide the border-
line cases for a parallel and an antiparallel b-sheet structure,
respectively, realized by only one intermolecular hydrogen
bond between the basic units.
The experimental data clearly indicates a reduction in the
degree of order of the oligomers on an increase in chain
length in the solid state. Clearly, the intermolecular interac-
tions become weaker and are less specific, favoring a disor-
dered aggregation of the molecules in the precipitate. Inter-
estingly, oligomer 3 showed a different diffraction pattern
(Figure S14 in the Supporting Information), which corre-
sponded to altered polymorphs when the precipitation con-
ditions were changed (solvent and rate of precipitation).
Very slow crystallization from deuterated chloroform afford-
ed single crystals suitable for crystallographic determination
of the molecular structure.
Further structural information for compounds 3, 5, 7, and
9 (containing 10% U-13C enriched Phe) was obtained from
solid-state 13C cross polarization (CP) magic-angle spinning
(MAS) NMR spectroscopy (Figure S15 in the Supporting In-
formation). Distinct chemical shifts and line width changes
of the amino acid signals can be observed for the individual
foldamers. The most prominent are the Ca and Cb signals of
the 13C labeled Phe at approximately 55 and 40 ppm, respec-
tively. The chemical shift data are summarized in Table 2.
The most intriguing observations are the differences in line
widths between the oligomers. Compound 3 features narrow
lines (in the order of 1 ppm), whereas significantly larger
line widths are observed for the higher oligomers. In con-
trast, the isotropic chemical shift changes between the oligo-
mers are moderate. Interestingly, compound 3 shows two
signals of equal height for the Ca atoms of l-Phe. Increased
line widths of NMR signals are indicative of structural heter-
ogeneity or molecular dynamics. Therefore, we measured
the motionally averaged 13C–1H dipolar couplings of the Ca
and Cb signals in the foldamers using the DIPSHIFT pulse
sequence. Typically, the experiment provides amplitude in-
formation about the fast motions with correlation times in
the nanosecond time window. As demonstrated recently,
however, also the intermediate timescale region with corre-
lation times in the microsecond range can be observed.[21]
The X-ray molecular structure of compound 3 (Figure 7)
exhibits some similarities with the structure already reported
for the monomer unit Boc-l-Phe-d-Oxd-OBn.[10] The most
important backbone torsion angles are given in Table 1.
Only a single intermolecular NH···CO hydrogen bond
exists between the dimeric units in the crystal packing of 3
(Figure 8), which is established between the symmetry
ꢁ
equivalent amide groups N3ACTHNURGTNE(UNG H3N) C8(O2) (H3N···O2’ 2.01,
ꢁ
N3···O2’ 2.85(2) ꢅ, N3 H3N···O2’ 1648, symmetry operation
(I): x+1/2, ꢁy+1/2, ꢁz+1) of the neighboring units. These
single hydrogen bonds generate an infinite antiparallel b-
sheet structure running along the crystallographic a axis.
There are no stacking interactions between the phenyl rings
of adjacent peptides in the chain. Interestingly, the parent
monomer Boc-l-Phe-d-Oxd-OBn establishes
alignment of the oligopeptide units. Thus, the blocked mo-
a parallel
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Nanofibers Containing Hybrid Foldamers
FULL PAPER
energy differences between var-
ious conformers could be re-
sponsible for the polymorphism
observed in the oligomer 3.
The two most stable con-
formers of the monomer, C1
and C2, appear to be connected
in the X-ray structure of the
dimer
Boc-(l-Phe-d-Oxd)2-
OBn. The values of the torsion
angles for the two conformers
obtained at both approximation
levels of ab initio MO theory
are in good agreement with the
experimental data obtained for
the dimer in Table 1. In both
conformers, the l-Phe residue is
in a b-sheet conformation. The
conformation of the two d-Oxd
moieties is different and corre-
sponds to a poly-proline II- and
Figure 8. View down the crystallographic b axis of the crystal packing diagram of 3 showing the intermolecular
NH···CO hydrogen bonds running along the a axis.
Table 2. Summary of the 13C NMR data of the foldamer series Boc-(l-
Phe-d-Oxd)n-OBn at 308C.
Table 3. Torsion angles[a] and stabilization energies[b] for the most stable
conformers of the Boc-l-Phe-d-Oxd-OBn monomer unit and the dimer 3
according to ab initio MO theory.
Ca
Cb
n
d
dCH
S
d
dCH
S
Conformer
l-Phe
d-Oxd
y[d]
DE[c]
1
Dn =
1
Dn =
2
2
[ppm] [ppm]
[kHz]
[ppm] [ppm]
[kHz]
f[c]
y[d]
f[c]
2
56.3/
53.9
55.0
54.3
53.0
1.0
9.9
0.86 40.8
0.9
12.8
1.0
C1
ꢁ117.9
ꢁ112.9
ꢁ121.3
ꢁ114.7
ꢁ84.1
ꢁ98.3
72.9
69.6
68.8
70.1
ꢁ116.6
ꢁ120.0
ꢁ106.9
ꢁ120.3
159.8
157.9
158.7
159.9
155.4
154.2
60.3
68.5
68.7
68.2
73.9
73.3
75.0
73.8
76.9
68.4
67.5
53.5
65.1
51.6
67.4
ꢁ164.3
ꢁ161.2
15.6
0.0[e]
0.0
4.3
2.8
4.3
4.6
7.4
9.7
7.6
3
4
5
2.9
5.5
5.6
10.2
10.0
10.0
0.89 40.4
0.87 39.4
0.87 38.0
2.7
7.0
5.5
12.6
12.6
12.2
0.98
0.98
0.95
C2
14.8
C3
ꢁ7.4
4.3
C4
0.3
59.7
ꢁ1.4
The results of this experiment are also given in Table 2. All
dipolar couplings measured are relatively close to the rigid
limit value. Order parameters between 0.86 and 0.89 are ob-
served for Ca and between 0.95 and 1.0 for Cb. On the fast
timescale the foldamers are very rigid performing only very
small amplitude motions, similar to those observed for fibrils
of large proteins.[22] In addition, there are no intermediate
timescale motions. All rotor echoes determined in the DIP-
SHIFT experiments reached more than 92% of the signal
intensity, indicating the absence of slower motions. It can,
therefore, be concluded that the increase in the line width
of the higher order foldamers is due to structural heteroge-
neity.
C5
166.1
162.1
145.0
135.4
143.0
139.5
ꢁ161.3
ꢁ158.2
ꢁ141.6
18.3
10.9
dimer 3[f]
ꢁ137.6
16.6
[a] In degrees. [b] In kJmolꢁ1. [c] HF/6-31G*. [d] B3LYP/6-311+G
ACHTUNGTRENNUNG
[e] ET(HF/6-31G*)=ꢁ1635.101363 a.u.;
ET(B3LYP/6-311+GACHTUNGTRENNUNG
ꢁ1645.490761 a.u. [f] Backbone torsion angles for the dimer 3: first two
rows HF/6-31G* data for the two monomer units, third and fourth rows
B3LYP/6-311+GACTHNUGRTNEUNG(2d,p) data.
a distorted left-handed 310-helical (bIIIꢀ-turn) conformation.
Quantum chemical calculations on the dimer Boc-(l-Phe-d-
Oxd)2-OBn itself and a section of the crystal structure with
two dimer units confirm the preference of the conformation-
al arrangement found in the solid state (Table 3, Figure S16
in the Supporting Information).
Theoretical calculations: To get further information on the
preferred secondary structures in the oligomer series, quan-
tum chemical calculations were performed employing ab
initio molecular orbital (MO) theory at various approxima-
tion levels (HF/6-31G*, B3LYP/6-311+G
(2d,p)). A system-
The association of two Boc-(l-Phe-d-Oxd)2-OBn units re-
sults in an antiparallel b-sheet structure in which the two
peptides are connected by only one intermolecular hydrogen
bond. This bond results between the peptidic CO group of
the first d-Oxd moiety of one peptide molecule and the NH
atic conformational analysis of the blocked monomer Boc-l-
Phe-d-Oxd-OBn was performed to understand the fiber
structure of this basic unit[10] and resulted in several con-
formers within a close energy range (Table 3). The small
Chem. Eur. J. 2009, 15, 8037 – 8048
ꢄ 2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
8043

C. Tomasini et al.
group of the second l-Phe residue in the second peptide
(Figure S16 in the Supporting Information). As already
mentioned, the crystal structure of the blocked monomer
Boc-l-Phe-d-Oxd-OBn represents the borderline case for a
parallel b-sheet structure with only one hydrogen bond con-
necting the peptide molecules.[10] The dimer Boc-(l-Phe-d-
Oxd)2-OBn represents the borderline case of an antiparallel
b-sheet structure with only one hydrogen bond between the
peptide molecules.
The chemical shifts determined by NMR spectroscopy can
be employed for the determination of the secondary struc-
ture type.[23,24] For l-Phe residues, the difference of the 13C
shifts of the Ca and Cb carbon atoms is a well-established in-
dicator. A value of d=21.9 ppm suggests that l-Phe is in an
a-helical conformation. Values of d=17.0 and 14.6 ppm cor-
respond to random coil and b-sheet conformations, respec-
tively. Thus, the shift differences of d=13–15 ppm obtained
from the data in Table 2 support the proposed b-sheet con-
formations of the l-Phe residues in all oligomers confirmed
by the experimental structure of oligomer 3. The theoretical
estimation of the 13C shift differences at the B3LYP/6-311+
GACHTUNGTRENNUNG(2d,p) level of ab initio MO theory for 3 provides values
of d=16.1 and 14.5 ppm for the two l-Phe residues. The
values at the HF/6-31G* level are d=14.0 and 13.9 ppm, re-
spectively. These data are in good agreement with the exper-
imental data (Table 2).
Figure 9. HF/6-31G* structures of Boc-(l-Phe-d-Oxd)5-OBn pentamer in
the poly-bII-turn (a) and poly-bI’-turn (b) arrangements. (For simplifica-
tion, the terminal groups were replaced by acetyl and N-methyl amide
groups.).
The experimental results provided no convincing evidence
for ordered structures in the trimer 5 and tetramer 7. For
the pentamer 9, both IR and NMR spectroscopies indicate
that all peptidic hydrogen atoms are involved in intramolec-
ular hydrogen bonds. This stimulates ideas on the corre-
sponding secondary structure. Due to the restriction of the
torsion angle f of the d-Oxd constituent to about 608, only
a small number of conformational alternatives have to be
considered, which additionally fulfill a regular intramolecu-
lar hydrogen-bonding pattern. Looking at the conformer
pool of the blocked monomer Boc-l-Phe-d-Oxd-OBn in
Table 3, the two conformers C3 and C4, which can be con-
sidered as b-turn structures,[25] could be candidates for a reg-
ular structure in the pentamer.
tion of a bII-turn in sequences of alternating d,l-peptides
was already found by Di Blasio et al. in the Boc-(d-alloiso-
leucyl-l-isoleucyl)3-methyl ester.[26] However, this shorter tri-
peptide, which exhibits the bII-turn between the residues l-
Ile2 and d-aIle3, has no regular structure as was also found
for our l-Phe-d-Oxd trimer and tetramer. Possibly, the regu-
lar structure is only enforced with the greater number of in-
tramolecular hydrogen bonds in the longer pentamer.
As shown in Table 2, the NMR spectroscopic data showed
chemical shift differences between the Ca and Cb carbon
atoms of the l-Phe residues in the pentamer of about d=
15.0 ppm, which agree with those for the dimer, in which the
l-Phe constituents are in a b-sheet conformation. The calcu-
lations of the shift differences for several poly-bII-turn pen-
tamers with a different orientation of the phenyl rings pro-
vide average values of d=14.1 ppm for the l-Phe residues
in the most stable conformer at the HF/6-31G* level and
The stabilities of both conformers are not so different
from those of the two most stable ones realized in the dimer
structure. Conformer C3 corresponds to a bII-turn; the con-
former C4 to a distorted bI’ACTHNURGTNE(NUG bIII’)-turn. The structure of a
pentamer with the latter conformer as the basic unit is close
to a left-handed 310-helix induced by the d-Oxd constituent.
The quantum chemical calculations on pentameric structures
constructed from the two monomers confirmed the rather
perfect poly-bI’- and poly-bII-turn structures (Figure 9). The
average backbone torsion angles for the b-turn moiety in
d=13.0 ppm at the B3LYP/6-311+GACTHUNTGRNEUNG(2d,p) level of ab initio
MO theory. Clearly, the data for the l-Phe residues in this
type II b-turn arrangement are similar to those in a b-sheet
conformation. Thus, the regular poly-bII-turn structure for
the pentamer is supported by the NMR spectroscopic data.
The corresponding values for the most stable poly-bI’-turn
pentamer were d=21.0 ppm at the HF/6-31G* and d=
the poly-bII conformer are fi+1 =ꢁ52.78, yi+1 =132.08, fi+2
=
76.28, and yi+2 =12.48. The corresponding values for the
poly-bIꢀ conformer are fi+1 =52.58, yi+1 =39.78, fi+2 =63.98,
and yi+2 =12.88. The detailed geometries are given in the
Supporting Information.
21.8 ppm at the B3LYP/6-311+GACTHNUGTRNEUNG(2d,p) level and are dis-
tinctly different. Thus, this structure cannot be excluded
only by its lower stability, but also by the deviating NMR
spectroscopy data.
The poly-bII-turn oligomer was found to be 31.7 kJmolꢁ1
more stable than the poly-bI’-turn oligomer. In fact, forma-
8044
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Chem. Eur. J. 2009, 15, 8037 – 8048

Nanofibers Containing Hybrid Foldamers
FULL PAPER
5.17 (AB, 2J
ACTHNUGTRNEUNG(H,H)=12.0 Hz, 2H; OCH2Ph), 5.51–5.58 (m, 1H; CHN-
CH2Ph), 7.20–7.41 (m, 10H; 2ꢆPh), 7.73 ppm (m, 3H; NH3).
Conclusion
Boc-(l-Phe-d-Oxd)2-OBn (3): A mixture of compound 2 (1 mmol) and
Et3N (3 mmol, 0.44 mL) in dry acetonitrile (10 mL) was added to a
stirred solution of 1 (1 mmol, 0.39 g) and HATU (1 mmol, 0.38 g) in dry
acetonitrile (10 mL) under an inert atmosphere at room temperature.
The solution was stirred for 40 min under an inert atmosphere before the
acetonitrile was removed under reduced pressure and replaced with ethyl
acetate. The mixture was washed with brine, 1n aqueous HCl (3ꢆ30 mL)
and finally 5% aqueous NaHCO3 (1ꢆ30 mL). The organic layer was
dried over sodium sulfate and concentrated in vacuo. The product was
obtained pure after silica gel chromatography (cyclohexane/ethyl acetate
8:2) (0.72 g, 95%). M.p. 1848C (dec.); [a]2D0 =+62.9 (c=1.0 in CH2Cl2);
Employing several experimental techniques, we have dem-
onstrated that the structural behavior of the oligomers of
the Boc-(l-Phe-d-Oxd)n-OBn series for n=2–5 is strongly
dependent on the chain length. In the dimer, only one inter-
molecular hydrogen bond between neighboring peptides is
sufficient to form a fiber-like material with an anti-parallel
b-sheet structure, thus showing that the dimer has “the right
molecular size” to form nanofibers. This result is comple-
mentary to our previously published crystal structure of the
blocked parent monomer Boc-l-Phe-d-Oxd-OBn,[10] which
represents the borderline case for a parallel b-sheet struc-
ture with only one hydrogen bond connecting the peptide
molecules.
1H NMR (300 MHz, CDCl3, 258C): d=1.31 (d, 3J
ACTHNUTRGNE(NUG H,H)=6.6 Hz, 3H;
Me), 1.32–1.40 (m, 9H; Me+tBu), 2.80–3.00 (m, 1H; CHN-CHHPh),
3.00–3.30 (m, 3H; CHN-CH2-Ph+CHN-CHH-Ph), 4.22–4.35 (m, 2H; 2ꢆ
CHN), 4.38–4.51 (m, 2H; 2ꢆCHO), 5.08 (brs, 1H; NH), 5.21 (s, 2H;
OCH2Ph), 5.63–5.70 (q, 3J
ACTHNUTRGNEUGN(H,H)=6.6 Hz, 1H; CHN-CH2Ph), 5.89–6.02
(brs, 1H; CHN-CH2Ph), 7.24–7.38 ppm (m, 16H; NH+3xPh); 13C NMR
(75 MHz, CDCl3, 258C): d=20.7, 20.9, 28.2, 37.8, 53.3, 54.1, 61.8, 62.7,
68.0, 73.6, 74.7, 80.3, 127.1, 128.3, 128.4, 128.5, 128.6, 129.4, 134.6, 135.5,
In the higher oligomers, there is a stronger tendency to
form intra- instead of intermolecular hydrogen bonds, with
the production of amorphous materials. The formation of or-
dered secondary structures increases and seems to be realiz-
ed at the pentamer level.
151.2, 151.6, 166.8, 167.3, 171.1, 172.7 ppm; IR (CH2Cl2, 3 mm): n˜ =3432,
ꢁ1
ꢁ
3346 (N H), 1792, 1756, 1701 cm (C=O); elemental analysis calcd (%)
for C40H44N4O11: C 63.48, H 5.86, N 7.40; found: C 63.53, H 5.80, N 7.45.
Boc-(l-Phe-d-Oxd)2-OH (4): Compound 4 was obtained in quantative
yield from 3 using the synthetic procedure described for compound 1
(0.65 g, 98.5%). M.p. 1508C; [a]2D0 =+45 (c=0.3 in MeOH); 1H NMR
(300 MHz, [D6]DMSO, 258C): d=1.27 (m, 12H; tBu+Me), 1.48 (d,
3J
CHN-CH2Ph), 4.36 (d, 3J
4.2 Hz, 1H; CHN), 4.83 (m, 2H; 2ꢆCHO), 5.46 (m, 1H; CHN-CH2Ph),
5.89 (m, 1H; CHN-CH2Ph), 7.07 (d, 3J
(H,H)=9 Hz, 1H; NH), 7.20–7.33
(m, 10H; Ph), 8.94 ppm (d, 3J(H,H)=9 Hz, 1H; NH); 13C NMR
ACHTUNGTRENNUNG
Experimental Section
N
ACHTUNGTRENNUNG
General: The melting points of the compounds were determined in open
capillaries and are uncorrected. High-quality infrared spectra (64 scans)
were obtained at 2 cmꢁ1 resolution by using a 1 mm NaCl solution cell
and a Nicolet 210 FT-infrared spectrometer. All spectra were obtained in
3 mm solutions in dry CH2Cl2 at 297 K or as 1% solid mixture with dry
KBr. All compounds were dried in vacuo and all of the sample prepara-
tions were performed in a nitrogen atmosphere. Routine NMR spectra
were recorded with spectrometers at 400, 300, or 200 MHz (1H NMR)
and at 100, 75, or 50 MHz (13C NMR). High-quality 1H NMR spectra
were recorded with a Varian Inova 600 spectrometer. The measurements
were carried out in CDCl3, [D6]acetone, or [D6]DMSO. The proton sig-
nals were assigned by gCOSY spectra. Chemical shifts are reported in d
values relative to the solvent peak.
AHCTUNGTRENNUNG
AHCTUNGTRENNUNG
(75 MHz, CD3CN, 258C): d=20.7, 21.1, 28.5, 39.1, 53.6, 55.0, 62.3, 63.1,
75.5, 76.0, 127.7, 128.0, 129.3, 129.4, 130.4, 130.5, 137.5, 137.9, 153.1,
ꢁ
153.3, 168.4, 169.6, 172.2 ppm; IR (Nujol): n˜ =3543, 3322 (N H), 1778,
1708, 1668, 1665 cmꢁ1 (C=O); elemental analysis calcd (%) for
C33H38N4O11: C 59.45, H 5.75, N 8.40; found: C 59.50, H 5.73, N 8.41.
Boc-(l-Phe-d-Oxd)3-OBn (5): Insoluble compound 4 (1 mmol, 0.66 g) in
dry acetonitrile (80 mL) was sonicated for 5 min and solid HATU
(1 mmol, 0.38 g) was added. The mixture was stirred under an inert at-
mosphere for 10 min before the addition of a mixture of 2 (1 mmol) and
Et3N (3 mmol, 0.44 mL) in dry acetonitrile (10 mL) at room temperature.
The solution was stirred for 2 h under an inert atmosphere until complete
dissolution of the reagents, then the acetonitrile was removed under re-
duced pressure and replaced by ethyl acetate. The mixture was washed
with brine, 1n aqueous HCl (3ꢆ30 mL) and 5% aqueous NaHCO3 (1ꢆ
30 mL), dried over sodium sulfate, and concentrated in vacuo. The prod-
uct was obtained pure after silica gel chromatography (cyclohexane/ethyl
acetate 8:2 ! 6:4) (0.42 g, 41%). M.p.=1788C (dec.); [a]2D0 =+48.0 (c=
Boc-l-Phe-d-Oxd-OH (1): Palladium on charcoal (10%) was added to a
solution of Boc-l-Phe-d-Oxd-OBn (2 mmol, 0.96 g) in methanol (40 mL)
and the mixture was stirred under a hydrogen atmosphere for 2 h. The
catalyst was then removed by filtration using a Celite pad and the solu-
tion was concentrated. The corresponding acid 1 was obtained pure in
99% yield without further purification. M.p. 127–1298C; [a]2D0 =+41.0
(c=1.1 in MeOH); 1H NMR (300 MHz, CD3OD, 258C): d=1.33 (d,
1
3J
2J
2J
N
ACHTUNGTRENNUNG
0.9 in CH2Cl2); H NMR (300 MHz, [D6]acetone, 258C): d=1.20–1.46 (m,
12H; tBu+Me), 1.52 (d, 3J(H,H)=6.3 Hz, 3H; Me), 1.59 (d, 3J
ACTHNUGTRENNUGN ACHTUNGTRENNUNG
ACHTUNGTRENNUNG
ACHTUNGTRENNUNG
(dd, 3J
G
G
ACHTUNGTERN(NUNG H,H)=13.5 Hz, 1H; CHN-CH2Ph), 3.27 (dd,
CHN), 4.73 (dq, 1H; 3J
N
G
3J
3J
3J
ACHTUNGTRENNUNG
N
ACHTUNGTRENNUNG
N
ACHTUNGTRENNUNG
129.6, 130.9, 138.4, 153.9, 157.6, 171.4, 174.4 ppm; IR (Nujol): n˜ =3367
ꢁ1
ꢁ
(d, 3J
A
(N H), 1786, 1713, 1691 cm (C=O); elemental analysis calcd (%) for
OCH2Ph+1 NH), 5.70 (m, 1H; CHN-CH2Ph), 5.94 (m, 1H; CHN-
CH2Ph), 6.15 (m, 1H; CHN-CH2Ph), 7.20–7.50 (m, 20H; Ph), 8.06 ppm
(m, 2H; NH); 13C NMR (75 MHz, CDCl3, 258C): d=20.4, 20.9, 28.1,
29.7, 37.5, 37.8, 53.3, 54.1, 61.7, 62.4, 68.2, 73.8, 74.8, 80.5, 127.2, 128.4,
C19H24N2O7: C 58.16, H 6.16, N 7.14; found: C 58.21, H 6.22, N 7.10.
H-l-Phe-d-Oxd-OBn·TFA (2):
A solution of Boc-l-Phe-d-Oxd-OBn
(2 mmol, 0.96 g) and TFA (36 mmol, 2.78 mL) in dry dichloromethane
(20 mL) was stirred at room temperature for 4 h. The volatile compounds
were then removed under reduced pressure and 2 was obtained pure in
quantitative yield without further purification (0.98 g, 99%). 1H NMR
128.7, 129.5, 134.7, 135.7, 151.0, 151.6, 167.0, 167.8, 171.3, 173.0 ppm; IR
1
ꢁ
(CH2Cl2, 3 mm): n˜ =3428, 3348 (N H), 1791, 1715, 1692 cm (C=O); anal.
(200 MHz, CDCl3, 258C): d=1.48 (d, 3J
ACHTUNGTRENNUNG
calcd. (%) for C54H58N6O15: C 62.90, H 5.67, N 8.15, found: C 62.94, H
5.72, N 8.18.
(dd, 3J
G
ACHTUNGTRENUNG
3J
3J
R
Boc-(l-Phe-d-Oxd)3-OH (6): Compound 6 was prepared in quantitative
yield from 5 following the synthetic procedure described for compound 1
3
ACHTUNGTNER(NUNG H,H)=3.2, 6.6 Hz, 1H; CHO),
Chem. Eur. J. 2009, 15, 8037 – 8048
ꢄ 2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
8045

C. Tomasini et al.
(0.38 g, 98%). M.p. 1558C (dec.); [a]2D0 =+33 (c=0.9 in MeOH);
1H NMR (400 MHz, CD3CN, 258C): d =1.08–1.30 (m, 18H; 3ꢆMe+
tBu), 2.6 5–3.28 (m, 6H; 3ꢆCH2-CHPh), 3.68–4.78 (m, 6H; 3ꢆCHO+
3ꢆCHN), 5.51–5.79 (m, 1H; CHN-CH2Ph), 5.80–6.02 (m, 2H; 2ꢆCHN-
CH2Ph), 6.25 (brs, 1H; NH), 7.20–7.45 ppm (m, 17H; 2ꢆNH+3ꢆPh);
13C NMR (100 Mz, CD3CN, 258C): d=20.7, 21.2, 28.5, 38.4, 38.9, 53.8,
54.4, 62.8, 63.1, 75.7, 76.0, 76.2, 76.5, 127.8, 128.0, 129.4, 130.3, 130.5,
the Supporting Information were obtained by the slow evaporation of
0.02m solutions of 3 in the different solvents, as reported in the figure.
Imaging: The fiber-like material was observed by optical microscopy
(OM), SEM, TEM. The OM images were collected by using a Leica opti-
cal microscope equipped with a CCD camera. Sample SEM images were
collected on a glass coverslip after coating with gold and observed by
using Philips 515 scanning electron microscope. TEM observations were
carried out by using a Philips CM 100 instrument (80 kV). The powdered
samples were dispersed in water and a few droplets of the slurry deposit-
ed on holey-carbon foils supported on conventional copper microgrids.
The images were recorded by using a CCD digital camera.
137.5, 137.7, 137.9, 153.2, 168.4, 171.1, 171.9, 172.4 ppm; IR (Nujol): n˜ =
ꢁ1
ꢁ
3307 (N H), 1775, 1721, 1684 cm (C=O); elemental analysis calcd (%)
for C47H52N6O15: C 59.99, H 5.57, N 8.93, found: C 60.05, H 5.62, N 9.01.
Boc-(l-Phe-d-Oxd)4-OBn (7): Compound 7 was prepared from com-
pounds 2 and 6 using the synthetic procedure described for 3 (0.34 g,
65%). M.p. 1238C (dec.); [a]2D0 =+55.0 (c=1.0 in CH2Cl2); 1H NMR
X-ray diffraction analyses: Powder X-ray diffraction patterns were col-
lected by using a PanAnalytical X’Pert Pro equipped with X’Celerator
detector powder diffractometer using CuKa radiation (l=1.5418 ꢅ) gen-
erated at 40 kV and 40 mA. The instrument was configured with a 1/48 di-
vergence and 1/48 antiscattering slits. A standard quartz sample holder
1 mm deep, 20 mm high, and 15 mm wide was used. The diffraction pat-
terns were collected within the 2q range from 28 to 608 with a step size
(D2q) of 0.028 and a counting time of 30 s.
(300 MHz, [D6]acetone, 258C): d = 1.01 (d, 3J
ACTHNUTRGNEUNG(H,H)=6.6 Hz, 3H; Me),
1.27–1.47 (m, 15H; tBu+2 Me), 1.52 (d, 3H; J=6.3 Hz, Me), 2.77–3.31
(m, 8H; 4ꢆCHN-CH2Ph), 4.24 (m, 2H; 2ꢆCHN), 4.54 (m, 2H; 2ꢆ
CHN), 4.65–4.85 (m, 4H; 4ꢆCHO), 5.24 (m, 2H; OCH2Ph), 5.67–5.78
(m, 2H; 2ꢆCHN-CH2Ph), 5.90–6.13 (m, 2H; 2ꢆCHN-CH2Ph), 7.17–7.50
(m, 25H; 5ꢆPh), 7.94–8.10 ppm (m, 3H; 3ꢆNH); 13C NMR (CDCl3,
100 MHz): d=20.7, 21.3, 28.5, 37.9, 53.3, 54.1, 62.2, 62.8, 63.0, 68.4, 73.6,
74.0, 74.3, 127.5, 128.6, 128.7, 129.0, 129.4, 129.7, 129.8, 130.2, 134.8,
Single-crystal X-ray diffraction: Intensity data for compound 3 were col-
lected by an Oxford Diffraction Excalibur diffractometer using CuKa ra-
diation (l=1.5418 ꢅ). The diffractometer was equipped with a cryo-cool-
ing device used to set the temperature at 100 K. Data collection was per-
formed with the program CrysAlis CCD[27] Data reduction was carried
out using the program CrysAlis RED.[28] Absorption correction was ap-
plied by using the ABSPACK[29] program. The structure was solved using
the SIR-2004 package[30] and was subsequently refined on the F2 values
by the full-matrix least-squares program SHELXL-97.[31] All non-hydro-
gen atoms were refined isotropically to have a good data/parameters
ratio. The methyl, methylene, and methine hydrogen atoms were placed
in calculated positions, constrained to ride on their parent atoms, and re-
fined isotropically with Uiso(H)=1.2 or 1.5 Ueq(C). The absolute struc-
ture configuration was not determined from X-ray data, but was known
from the synthetic route. Crystallographic data and refinement parame-
ters are reported in Table S1 in the Supporting Information. CCDC-
717012 contains the supplementary crystallographic data for 3. These
data can be obtained free of charge from The Cambridge Crystallograph-
ic Data Centre via www.ccdc.cam.ac.uk/data_request/cif.
135.8, 137.5, 151.7, 154.8, 167.5, 168.0, 171.6 ppm; IR (CH2Cl2, 3 mm): n˜ =
ꢁ1
ꢁ
3424, 3351, 3299 (N H), 1791, 1711, 1679 cm (C=O); elemental analysis
calcd (%) for C68H72N8O19: C 62.57, H 5.56, N 8.58; found: C 62.52, H
5.49, N 8.66.
Boc-(l-Phe-d-Oxd)4-OH (8): Compound 8 was prepared in quantitative
yield from 7 using the synthetic procedure described for compound 1
(0.31 g, 98%). M.p. 1188C (dec.); [a]2D0 =+39.0 (c=0.4 in MeOH);
1H NMR (400 MHz, CD3CN, 258C): d=1.30–1.38 (m, 18H; 3ꢆMe+
tBu), 1.53 (d, 3J
ACHTUNGTRENNUNG(H,H)=6.0 Hz, 3H; Me), 2.73–3.36 (m, 8H; 4ꢆCHN-
CH2Ph), 3.68–4.81 (m, 8H; 4ꢆCHO+4ꢆCHN), 5.61 (brs, 2H; NH+
CHN-CH2Ph), 5.85–6.11 (m, 3H; 3ꢆCHN-CH2Ph), 7.25–7.61 ppm (m,
23H; 3ꢆNH+4ꢆPh); 13C NMR (100 Mz, CD3CN, 258C): d=20.6, 20.8,
21.0, 21.1, 28.4, 38.4, 39.0, 39.2, 39.4, 53.5, 53.6, 62.6, 63.1, 75.8, 75.9, 76.1,
76.2, 76.4, 80.0, 127.7, 128.0, 129.0, 129.4, 129.5, 130.4, 130.6, 137.4, 137.5,
153.2, 153.3, 153.5, 168.5, 171.9, 172.2, 173.3 ppm; IR (Nujol): n˜ =3299
(N H), 1786, 1668 cmꢁ1 (C=O); elemental analysis calcd (%) for
ꢁ
C61H66N8O19: C 60.29, H 5.47, N 9.22; found: C 60.20, H 5.55, N 9.24.
Boc-(l-Phe-d-Oxd)5-OBn (9): Compound 9 was prepared from com-
pounds 2 and 8 by using the synthetic procedure described for 3 (0.31 g,
78%). M.p. 2038C (dec.); [a]2D0 =+3.4 (c=1.0 in CH2Cl2); 1H NMR
(400Mz, [D6]acetone, 258C): d=0.95–1.58 (m, 21H; tBu+4 Me), 1.61 (d,
Solid-state NMR spectroscopy: For the solid-state NMR spectroscopic
studies, powdered samples were filled into 4 mm MAS rotors with a
Teflon insert providing a volume of approximately 50 mL. All 13C MAS
NMR experiments were carried out on a Bruker Avance 750 NMR
(Bruker Biospin, Rheinstetten, Germany) spectrometer operating at a
resonance frequency of 749.7 MHz for 1H and 188.5 MHz for 13C. A
double-resonance MAS probe equipped with a 4 mm spinning module
was used with a MAS frequency of 7 kHz at 308C. The 13C 908 pulse
length was typically 5 ms. The CP MAS spectra were recorded with a 1H
908 pulse length of 4 ms and a CP contact time of 700 ms. The 1H radio-
frequency field strength during heteronuclear decoupling using TPPM[32]
was about 65 kHz. The 13C chemical shifts were referenced to the 13C=O
signal of 13C-labelled Gly at 176.45 ppm as an external standard. All ppm
values are relative to TMS.[33] The strength of 13C–1H dipolar couplings
was measured in DIPSHIFT experiments[34] at a MAS frequency of
5 kHz. The MREV-8 sequence was used for 1H–1H homonuclear decou-
pling applying a decoupling field of 100 kHz.[35] Since the dipolar-induced
signal decay is periodic with the rotor period, the signal was acquired
only over one rotor period in the indirect dimension. The dipolar de-
phased signal was extracted for each resolved peak and fitted over one-
rotor-period time domain to yield the coupling strength and to determine
3J
ACHTUNGTRENNUNG(H,H)=6.9 Hz, 3H; Me), 2.88- 3.45 (m, 10H; 5ꢆCHN-CH2Ph), 3.96–
4.62 (m, 5H; 5ꢆCHN), 4.72–4.94 (m, 5H; 5ꢆCHO), 5.34 (m, 3H;
OCH2Ph+NH), 5.70–6.37 (m, 5H; 5ꢆCHN-CH2Ph), 7.27–7.50 (m, 30H;
Ph), 7.78–8.13 ppm (m, 4H; 4ꢆNH); 13C NMR (100 MHz, CDCl3, 258C):
d=20.7, 21.3, 28.5, 37.9, 53.3, 54.1, 62.0, 62.2, 62.8, 63.0, 68.4, 73.6, 74.0,
74.3, 127.5, 128.6, 128.7, 129.0, 129.4, 129.7, 129.8, 130.2, 134.8, 135.8,
137.5, 151.7, 154.8, 167.5, 168.0, 171.6 ppm; IR (CH2Cl2, 3 mm): n˜ =3350,
ꢁ1
ꢁ
3286 (N H), 1789, 1737, 1707, 1673 cm (C=O); elemental analysis calcd
(%) for C82H86N10O23: C 62.35, H 5.49, N 8.87; found: C 62.30, H 5.48, N
8.90.
Boc-(l-Phe-d-Oxd)5-OH (10): Compound 10 was prepared in quantita-
tive yield from 9 by using the synthetic procedure described for com-
pound
MeOH); 1H NMR (300 MHz, CD3OD, 258C): d=1.20–1.48 (m, 21H; 4ꢆ
Me+tBu), 1.53 (d, 3J
(H,H)=6.0 Hz, 3H; Me), 2.68–3.18 (m, 10H; 5ꢆ
1
(0.28 g, 98%). M.p. 1748C (dec.); [a]2D0 =+25.0 (c=0.3 in
ACHTUNGTRENNUNG
CH-CH2Ph), 3.62–4.98 (m, 10H; 5ꢆCHO+5ꢆCHN), 5.69 (brs, 2H;
NH+CHN-CH2Ph), 5.92–6.14 (m, 4H; 4ꢆCHN-CH2Ph), 7.18–7.42 ppm
(m, 25H; 5ꢆPh); 13C NMR (100 Mz, CD3OD, 258C): d=21.2, 21.9, 28.9,
38.5, 38.8, 39.0, 39.2, 40.0, 54.7, 54.9, 55.2, 55.6, 55.7, 56.9, 63.0, 64.1, 76.7,
78.6, 79.1, 128.3, 129.7, 129.8, 130.0, 130.5, 130.7, 130.8, 130.9, 131.2,
138.4, 154.0, 161.6, 170.0, 170.2, 172.7, 173.2 ppm; IR (Nujol): n˜ =3269
ꢁ
the C H order parameter.
Theoretical calculations: All theoretical calculations were performed em-
ploying the Gaussian 03 program package (Gaussian, Wallingford, CT
06492, USA). A systematic conformational analysis for the monomer
unit was done at the HF/6-31G* level of ab initio MO theory. The con-
(N H), 1786, 1664 cmꢁ1 (C=O); elemental analysis calcd (%) for
ꢁ
C75H80N10O23: C 60.44, H 5.40, N 9.42; found: C 60.48, H 5.41, N 9.40.
formers were reoptimized at the B3LYP/6-311+GACTHNUTRGNEUG(N 2d,p) level. The start-
ing point for the optimization of the dimer of 3 was a corresponding sec-
tion of the X-ray structure. The chemical shifts were calculated employ-
Fiber-like aggregate precipitation: All compounds were obtained by the
slow evaporation of a 0.02m solution in a 1:1 mixture of cyclohexane and
ethyl acetate. All of the polymorphs of 3 that are shown in Figure S14 in
8046
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Chem. Eur. J. 2009, 15, 8037 – 8048

Nanofibers Containing Hybrid Foldamers
FULL PAPER
ing the GIAO approximation. Further numerical details are given in the
Supporting Information.
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Acknowledgements
G.A., G.F., M.M., and C.T. are grateful to Ministero dell’Universitꢁ e
della Ricerca Scientifica (PRIN 2006) and Universitꢁ di Bologna (Funds
for selected topics) for financial support. H.-J.H. and D.H. are obliged to
DFG (SFB 610, HO 2346/1-3) for funding. Furthermore, we thank Dr. A.
Guerri for the single-crystal X-ray data collection. D.H. thanks the DFG
(German Research Foundation) and the Experimental Physics Institutes
of the University of Leipzig for measuring time the Avance 750 MHz
NMR spectrometer. C.T. is grateful to Fondazione del Monte di Bologna
e di Ravenna for financial support.
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Published online: April 9, 2009
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