Oligomers of β2- and of β3-homoproline: What are the secondary structures of β-peptides lacking H-bonds?

Article abstract of DOI:10.1002/(SICI)1522-2675(19991006)82:10<1539::AID-HLCA1539>3.0.CO;2-E

To study the role of H-bonds in stabilizing β-peptidic secondary structures, we have synthesized β-oligopeptides (up to the octadecamer 12) consisting of β²- and β³-homoproline, i.e., β-peptides lacking amide protons. The enantiomer purity of the building block β²-homoproline (nipecotic acid, 4) was determined by HPLC analysis of the N-(2,4- dinitrophenyl) derivative 5 on a Chiralcel-OD column (cf. Fig. 2). The CD spectra of the all-(S)-β²- and all-(S)-β³-HPro-containing, β-peptides display novel and intensive CD patterns which may be indicative of a secondary structure (cf. Fig. 3). It is noteworthy that a distinct CD pattern was observed with the β³-HPro derivatives containing as few as three residues (7a). The crystal structure of a N-deprotected β³-HPro-tripeptide 7c is presented (cf. Figs. 4 and 5), and a model for the structure of β- peptides consisting of β³-HPro is discussed (cf. Figs. 6 and 7).

Full text of DOI:10.1002/(SICI)1522-2675(19991006)82:10<1539::AID-HLCA1539>3.0.CO;2-E

Helvetica Chimica Acta ± Vol. 82 (1999)
1539
Oligomers of b2- and of b3-Homoproline: What are the Secondary Structures
of b-Peptides Lacking H-Bonds?
1
2
by Stefan Abele ), Kurt Vögtli ), and Dieter Seebach*
Laboratorium für Organische Chemie der Eidgenössischen Technischen Hochschule, ETH-Zentrum,
Universitätstrasse 16, CH-8092 Zürich
Dedicated to Professor Helmut Ringsdorf on the occasion of his 70th birthday
To study the role of H-bonds in stabilizing b-peptidic secondary structures, we have synthesized b-
oligopeptides (up to the octadecamer 12) consisting of b2- and b3-homoproline, i.e., b-peptides lacking amide
protons. The enantiomer purity of the building block b2-homoproline (nipecotic acid, 4) was determined by
HPLC analysis of the N-(2,4-dinitrophenyl) derivative 5 on a Chiralcel-OD column (cf. Fig. 2). The CD spectra
of the all-(S)-b2- and all-(S)-b3-HPro-containing b-peptides display novel and intensive CD patterns which may
be indicative of a secondary structure (cf. Fig. 3). It is noteworthy that a distinct CD pattern was observed with
the b3-HPro derivatives containing as few as three residues (7a). The crystal structure of a N-deprotected
b3-HPro-tripeptide 7c is presented (cf. Figs. 4 and 5), and a model for the structure of b-peptides consisting of
b3-HPro is discussed (cf. Figs. 6 and 7).
1. Introduction. ± A major goal of bio-organic chemistry is the synthesis of
oligomers with unnatural backbones that combine the structural diversity and
functional characteristics of biopolymers (polypeptides, polynucleotides, polysacchar-
ides, polyisoprenoides, and poly(hydroxyalkanoates) [1]) with the stability of synthetic
polymers. Recent results demonstrate the potential of b-peptides as oligomers
consisting of the homologues of simple proteinogenic amino acids [2]; secondary
structural elements of proteins (helices [3 ± 5], sheets [3][6], turns [7] or steps [8], and
hairpins [6][9]) are formed by b-peptides as well, and they are of higher stability even
with short chain lengths [5][10]. A further asset is that the secondary structures can be
predicted by conformational analysis and by theoretical calculations [11 ± 13]. More-
over, b-peptides represent an ideal combination of being stable to mammalian
proteases [14][15], and yet biologically active, for instance as mimics of a-peptide
hormones [16].
The following question arose in the course of rational b-peptide design which we are
currently pursuing: does a b-peptidic chain without backbone H-bonds fold into stable
secondary structures? Considering the importance of H-bonds in stabilizing a-peptidic
structures [17], the answer may be no. However, there are several pieces of evidence
indicating that stable secondary b-peptide structures may be possible without H-bonds:
i) The preferred backbone conformation around the central C(a) C(b) bond has been
identified as a major contributor to the stabilization of b-peptide secondary struc-
1
)
)
Part of the Ph. D. thesis of S.A., Dissertation ETH-Zürich, No. 13203, 1999.
Part of the Master Thesis (Diplomarbeit) of K.V., ETH-Zürich, 1999.
2

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Helvetica Chimica Acta ± Vol. 82 (1999)
tures3) [5], and, in N-alkylated b-peptides, this staggering effect is still present. ii)
Recent molecular-dynamics calculations suggest that spontaneous folding to give the b-
peptidic 314 helix occurs prior to formation of the H-bonds4) [11]. iii) ꢀPeptoidsꢁ
(oligomers of N-substituted glycine [18]) containing chirality centers at the a-position
of their side chains also form secondary structures in solution5), despite their lack of
amide protons and thus of H-bonding within the backbone [19 ± 21].
A fully N-methylated b-hexapeptide consisting of N-methyl-b-homoalanine has
already been synthesized by us, but the lack of crystallinity and the presence of
rotamers prevented structural analysis [22]. We have, therefore, turned to the
homologues of proline: the constraints, which determine the allowed values of the
backbone dihedral angles in b-peptides consisting of b2- or b3-homoproline, should be
fundamentally different from those operating in non-N-alkylated b-peptides of acyclic
b-amino acids, i.e., distinct backbone torsion angles F, V, and Y (Fig. 1,a) are enforced,
and this might compensate for the lack of H-bonding.6)7)
Homologation of l-proline by inserting a CH2 group between the carbonyl C-atom
and the C(a)-atom leads to (S)-b3-homoproline, while insertion of CH2 group between
the C(a)-atom and the N-atom affords (R)-b2-homoproline (nipecotic acid; see
Fig. 1,b). Whereas the former transformation can be realized by classical Arndt-Eistert
homologation, the latter can not be effected in a simple step.
Fig. 1. a) Torsion angles as defined in b-amino acids. b) Relationship between proline and b3- and b2-
homoproline. Arrows indicate the formal sites of homologation. The b2-HPro-containing b-peptides described
herein happen to consist of the homologs of d-proline.
3
)
)
)
)
)
Actually, the failure to find a ꢀmelting pointꢁ in temperature-dependent NMR and CD spectra of MeOH
solutions of b-peptides [10] is compatible with this non-cooperative source of stability.
There is only a small energy difference between an intramolecular and an intermolecular H-bond in
MeOH [10]!
NMR Analysis indicated that the major conformation of a peptoid pentamer in MeOH consists of a (P)-
31-helix [19].
4
5
6
7
Cf. the b-peptides having a backbone enforced by incorporated rings, studied by Gellman and co-workers
[4][9].
It is well known that proline, the only proteinogenic amino acid with a secondary amino group, imparts
special conformations on a peptidic backbone by virtue of its pyrrolidine ring and of its fully substituted
amide N-atom. As a consequence, the energy difference between the cis- and trans-conformation of the
prolyl peptidyl bond is decreased, accounting for 10 ± 30% of cis-amide bond (w ˆ 08, or ap-
conformation) in Pro-containing peptides [17]. This leads to unique structures of poly-proline [23] and
of Pro-rich proteins (see collagen triple helix [24 ± 26]).

Helvetica Chimica Acta ± Vol. 82 (1999)
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2. Preparation of b3- and b2-Homoproline Derivatives. ± Boc-Protected (S)-b3-
HPro OH has previously been prepared by Arndt-Eistert [14][27 ± 29] or C1
homologation with cyanide [30] of l-proline8)9).
We chose the classical Arndt-Eistert homologation for the preparation of the
required building blocks. Thus, the Boc-protected diazo ketones (R)- and (S)-1
were prepared according to published procedures [34][35]. Wolff rearrangement of the
diazo ketones (R)- and (S)-1 in either THF/BnOH or THF/H2O afforded the Boc-
protected b3-homoproline benzyl esters, (R)- and (S)-2a, and the Boc-protected b3-
homoprolines, (R)- and (S)-2b, in high yields (cf. [22] and see Exper. Part).
The (R)- and (S)-ethyl nipecotate were prepared by classical resolution [36]10).
Only one diastereoisomeric salt (the like-salt) 3 precipitated upon treatment of
commercially available rac-ethyl nipecotate (rac-4a) with either enantiomer of tartaric
acid in EtOH. After two or three recrystallizations, (R,R,R)-3 and (S,S,S)-3 were
obtained in diastereoisomerically pure form, as determined by comparison of the
optical rotation and the melting points with literature values [36]. Basic extraction
(pH 13) afforded (R)- 4a and (S)-4a11)12). Both enantiomers of ethyl nipecotate 4a
showed optical rotations similar to reported values. However, even at high concen-
trations, the value of the optical rotation ([a]rD:t: ꢀ 1.3) is too small to allow accurate
determination of the enantiomer purity13). Consequently, other methods for deter-
mining the enantiomer purity were tested14).
)
The synthesis of H-(S)-b3-HPro-OH by Arndt-Eistert homologation of Z-l-Pro was first reported in 1975
[31]. However the published value and the sign of optical rotation are different from those reported by an
industrial group [28], and by others [32][33].
8
9
10
11
12
13
)
)
)
)
)
However, both the Boc-protected diazo ketones 1, derived from d- and l-proline, and the benzyl-ester
derivatives 2 of H-b3-HPro-OH, described herein, are new compounds.
An esterase-catalyzed hydrolysis of rac-N-acetyl methyl nipecotate provides the products with poor ee
values (22 ± 24%) [37].
The absolute configuration of ( )-nipecotic acid was established by CD spectroscopy [38] and by chemical
correlation [39].
This step has to be carried out rapidly at 08, and the pH must be controlled, because complete
saponification may occur.
It is noteworthy that enantiomerically pure nipecotic acid is often encountered in peptidomimetics [40 ±
42]; the ee value was always determined by polarimetry! In independent work, (R)- and (S)-nipecotic acids
were incorporated into a b-tetrapeptide, promoting hairpin formation [9].
14
)
i) Derivatization of 4a with (S)- and (R)-a-methoxy-a-(trifluoromethyl)phenylacetyl chloride (ˆ 3,3,3-
trifluoro-2-methoxy-2-phenylpropanoyl chloride; MTPA-Cl) to give the corresponding diastereoisomeric

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Helvetica Chimica Acta ± Vol. 82 (1999)
Eventually, reaction of rac-4a with Sangerꢁs reagent (1-fluoro-2,4-dinitrobenzene,
DNPF [49][50]) in alkaline solution, to give the 2,4-dinitrophenyl (DNP) derivative of
ethyl nipecotate rac-5, and chromatography on Chiralcel-OD (Daicel)15) was found to
provide an excellent analysis of the enantiomer ratio (er; see Fig. 2). Integration of the
peaks in the corresponding chromatograms revealed that (R)-4a and (S)-4a had been
obtained by the resolution procedure with an er value of 98.9 :1.1 and 99.6 :0.4,
respectively. With this method at hand, the diastereioisomer purity of the l-salts
(R,R,R)-3 and (S,S,S)-3 was determined by liberating a small quantity of 4a, the
enantiomer purity of which was determined as outlined in Fig. 216 17).
)
The following transformations were effected with the (S)-enantiomer of 4a: it was
Boc-protected to give the ester 4b as a colorless oil in 74% yield. The subsequent
saponification step was considered crucial, because there was the risk of racemization
(cf. the epimerization of b2,3- [5] and b2-amino-acid derivatives [52]). The mildest
procedure for saponification was the hydrolysis with 2.5 equiv. of LiOH in MeOH/H2O
at r.t. for 2 ± 3 days providing, after recrystallization, Boc-protected (S)-nipecotic acid
4c in 90% yield. The enantiomeric purity of 4c was determined by transformation to
amides, the so-called Mosher amides, for 19F- and 1H-NMR spectroscopic analysis [43]. ii) The use of
a,a,a'a'-tetraphenyl-1,3-dioxolane-4,5-dimethanol (TADDOL) as chiral shift reagent for 13C- and 1H-NMR
spectroscopy [44][45]. The 13C-NMR spectrum (100 MHz) of a solution of rac-4a and TADDOL (1:2) in
CDCl3 showed non-equivalence of two signals after several hours. The chemical-shift differences were ca.
24 and 45 ppm, but the peaks showed tailing, preventing a complete separation. This enantioselective shift
effect could allow for a quick, yet not accurate, determination of enantiomer purity. iii) Derivatization of
rac-4a with 2,2,3,3,3-pentafluoropropanoyl chloride and i-PrOH [46 ± 48] to give the corresponding
isopropyl N-(2,2,3,3,3-pentafluoropropionyl)nipecotate for GC analysis (a-CD, b-CD, g-CD, and Chirasil-
Val). All of these methods failed to elucidate the enantiomer ratio.
14
)
15
16
17
)
)
)
Recently, rac-ethyl 4-hydroxypiperidine-3-carboxylate was successfully separated as N-(2,4-dinitrophenyl)
derivative on a Chiralpak AD HPLC column [51].
In this case, the enantiomer purity of (S)-4a was 97.0% after two recrystallizations of (S,S,S)-3. A third
recrystallization increased the enantiomer purity to 99.6%.
It should be noted that polarimetry with the enantiomers of 5 is suitable for a first determination of the
enantiomer purity by virtue of the high values of optical rotation (‡ 165 and 165 for (S)- and (R)-5,
resp.; see Exper. Part).

Helvetica Chimica Acta ± Vol. 82 (1999)
1543
Fig. 2. HPLC Traces of N-(2,4-dinitrophenyl) derivatives 5 (Chiralcel OD, mobile phase: i-PrOH/hexane
35 :165; see GP 7 in Exper. Part). The arrows indicate the peak of the minor enantiomer. The retention times
were 20.3 and 25.0 min for the (R)- and (S)-enantiomer, respectively. The er value was determined by
integration of the corresponding peak: (R)-5: 98.9 :1.1; (S)-5: 99.6 :0.4.
the DNP derivative 5 (see Exper. Part); (S)-4a with an er value of 99.6 :0.4 was
employed in the subsequent syntheses18).
3. Preparation of b3- and b2-HPro-Peptides. ± Both the all-(S)-b-peptides (isotactic)
and the b-peptides containing an alternating sequence of (S)- and (R)-b3-homoproline
(syndiotactic) were prepared in solution, using the N-[3-(dimethylamino)propyl]-N'-
ethylcarbodiimide hydrochloride/1-hydroxy-1H-benzotriazole (EDC/HOBt) proce-
dure [3][5][7]. The benzyl-ester derivative (S)-2a was Boc-deprotected (TFA,
CH2Cl2), and the resulting TFA salt was employed for coupling with the Boc-protected
amino acid (S)-2b to give the fully protected dipeptide 6. After N-deprotection,
another coupling step with (S)-2b followed to give the fully protected all-(S)-b-
tripeptide 7a as a white foam in 91% yield. Benzyl-ester cleavage (H2, Pd/C) provided
the b-tripeptide acid 7b and Boc deprotection yielded the TFA salt 7c. Likewise, fully
protected (S)/(R)-b-dipeptide 8a was prepared by coupling the TFA salt derived from
(R)-2a with the acid (S)-2b. Benzyl-ester cleavage (H2, Pd/C) yielded the b-dipeptide
acid 8b, which was used for coupling with the Boc-deprotected dipeptide benzyl ester
derived from 8a, to provide (S)/(R)-b-tetrapeptide derivative 9 in 81% yield.
Fragment condensation of the tripeptide derivatives 7c and 7b then furnished
protected isotactic b-hexapeptide 10a (85%), which was converted to the peptide acid
10b by hydrogenation (H2, Pd/C). The fully protected peptide 10a was soluble in
various protic and aprotic solvents (CHCl3, CH2Cl2, MeOH, AcOEt, Et2O); its good
solubility and the high yields in the coupling steps leading to it provided motivation for
the synthesis of higher oligomers. Thus, the TFA salt obtained from hexapeptide 10a
was used for coupling with hexapeptide acid 10b to give the fully protected b-
18
)
Assuming that the Boc protection (without base) occurred without racemization, the er value has been
slightly reduced to 97.9:2.1 during saponification. Alternatively, saponification with the same amounts of
base in refluxing MeOH/H2O 3:1 for 3 h produced 4c with an er of 97.3:2.7.

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Helvetica Chimica Acta ± Vol. 82 (1999)
dodecapeptide 11 as a white foam in 79% yield after purification by precipitation from
CH2Cl2/hexane. After Boc deprotection, this dodecamer was further coupled with the
peptide acid 10b to give the fully protected b-octadecapeptide 12 as a white powder.
This is the longest b-peptide synthesized to date19).
To increase the crystallinity of the b-peptides, the Ac and p-nitrobenzoyl group
were introduced at the N-terminus of the b-hexapeptide. The TFA salt from 10a was
used for acylation with either Ac2O or p-nitrobenzoylchloride to provide the b-peptide
esters 13 and 14. In the syndiotactic series, a second fragment coupling step of the N-
deprotected form of b-tetrapeptide 9 with the peptide acid 8b furnished the fully
protected b-hexapeptide 15 (white powder).
19
)
A highly insoluble b-pentadecapeptide was reported in [52].

Helvetica Chimica Acta ± Vol. 82 (1999)
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The synthesis of the all-(S)-b2-HPro-peptides began with the acylation of
enantiomerically pure ethyl nipecotate (S)-4a with Boc-protected (S)-b2-homoproline
(S)-4c to give the dipeptide derivative 16 (waxy solid). After Boc deprotection, a
further coupling with the b-amino acid (S)-4c gave fully protected b-tripeptide 17a
(white waxy solid). The following saponification was performed applying the same mild
procedure as for the saponification of the monomeric building block 4b. Thus,
treatment of 17a with LiOH in a MeOH/H2O solution provided the tripeptide acid 17b
after precipitation from AcOEt/pentane. Final fragment coupling, with the peptide acid
17b and the TFA salt derived from 17a, gave the fully protected b-hexapeptide 18
(colorless ꢀglassꢁ, 79% yield).
4. Structure Analysis. ± 4.1. Circular Dichroism Spectroscopy. Circular dichroism
(CD) spectroscopy is a low-resolution method that provides a first indication for the
presence of secondary structures [53][54]. Compared to NMR spectroscopy, it has a
very short ꢀtime scaleꢁ (UV vs. radar frequency!), and, thus, an (unstable) chiral
conformer present in a very small amount but with a large DeR/l value may contribute
very strongly to the spectrum of the ensemble of all molecules present.
In comparison with the oligomers of homologues of non-cyclic l-amino acids
(Fig. 3,a, shows the typical CD pattern associated with a 314 helix), the b3- and b2-HPro
oligomers give rise to a quite different Cotton effect (Fig. 3,b and d). The all-(S)-b3-
HPro-peptides have a characteristic, very intense minimum at 202 nm and a maximum
at 223 nm, with a zero cross-over at 212 nm (Fig. 3,b)20). The absolute mean-residue
molar ellipticity at 202 nm decreases with growing chain length (7a with 3 residues:
4.40 ´ 104 vs. 12 with 18 residues: 1.92 ´ 104). The same is true for the mean-residue
molar ellipticity at 223 nm. This suggests that the secondary structure of longer peptide
chains of this type is destabilized. However, the high molar ellipticities (for instance
3.46 ´ 105 for 12 at 202 nm) still imply that a secondary structure is present in
MeOH21). In sharp contrast to the CD spectra of the b3-HPro oligomers consisting of
20
)
There is a conspicuous (mirror-image-type) analogy between the CD spectra of our b-HPro-containing b-
peptides and the CD spectra of Gellmanꢁs b2,3-hexapeptide derivative consisting of trans-2-aminocyclo-
pentanecarboxylic-acid building blocks [4]. However, we think that it is highly unlikely that the b-HPro-
peptides adopt the same conformation (a 2.5 helix consisting of twelve-membered H-bonded rings) as the
cyclopentane derivatives.
21
)
The fully protected b-hexapeptide 10a feature the same CD pattern in CF3CH2OH and in aqueous
buffered solution (pH5.7), albeit with lower intensity, as compared to the CD spectra measured in MeOH.

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Helvetica Chimica Acta ± Vol. 82 (1999)
homochiral [55] b-amino acids, the CD spectra of the b-peptides containing ± in an
alternating fashion ± (S)- and (R)-b3-HPro building blocks (9 and 15) show virtually no
Cotton effect (Fig. 3,c), indicating that these compounds may be devoid of an ordered
secondary structure22) ± candidates for random coils in the world of b-peptides!? For
both substitution patterns, the CD spectra did not change substantially with different
groups at C- and N-termini. The b-peptides composed of (S)-b2-HPro building blocks
show weaker Cotton effects, but the overall CD pattern is similar to that of the b3-HPro-
peptides (Fig. 3,d). However, the mean residue molar ellipticity of b-hexapeptide
derivative 18 (‡ 4.07 ´ 103) is nearly three times larger than that of b-tripeptide
derivative 17a (‡ 1.38 ´ 103) at ca. 230 nm. Thus, the secondary structure may be
stabilized by a longer b-peptide chain, in this case. The CD curve of non-structured
protected b-dipeptide 16 is included to show that the measured Cotton effects of the
higher oligomers are due to a distinct chiral supramolecular arrangement of the
b-peptide chain.
4.2. X-Ray Crystal-Structure Analysis. The TFA salt 7c, which had been isolated as a
colorless oil, solidified after two-weeks storage in a freezer. Suitable crystals were
separated and the structure was solved. Four molecules of CF3CO2H are incorporated
into the crystal (Fig. 4). The dihedral angles are given in the Table. Inspection of this
structure revealed some interesting features (Fig. 5): i) The Ph group is parallel (at van-
der-Waals distance) to the plane formed by the amide group of the first two b-amino
acid residues23). ii) The substituents at C(a) of the second and third pyrrolidine rings
are in a pseudo-axial position, a direct consequence of allylic 1,3-strain (A1,3-strain).
The exocyclic amide group pushes the neighboring substituent out of its plane into an
axial position of the ring. This is in agreement with numerous structures of Pro-
containing compounds, as well as with a variety of X-ray structures of simple N-
acylated five-membered heterocycles [60]. iii) The pyrrolidine rings exhibit the twist
conformation24), and the N-atoms are not pyramidalized in the crystal structure of 7c.
5. Modelling of a Possible Secondary Structure of R-(b3-HPro)n-OR and
Conclusion. ± As with N-methyl b-peptides [22], the NMR spectra of the b-HPro
oligomers show that rotamers, i.e., conformers with cis- and trans-amide bonds, are
present, similar to the situation with proline-containing a-peptides [17][23][64].
Structure determination by 2D-NMR techniques is thus complicated. The large
coupling constant JAX of the ABX system from the CO CH2 CH units in the b3-HPro
oligomers is ca. 8 Hz (not quite large enough for a coupling between antiperiplanar
protons). Still, this value, the crystal structure of 7c, and the general rules of
conformational analysis [17] provided guidance for the construction of a model for the
structure of b-peptides composed of (S)-b3-homoproline. The amide bond was fixed in
22
)
)
)
The absence of a Cotton effect does not preclude the existence of a stable secondary structure a priori; for
instance, g-peptides which adopt a helical structure according to 2D-NMR analysis do not show any
Cotton effect [56].
A similar, more twisted geometry of an a-dipeptide derivative has been reported [57]. For a-peptides with
NH protons, various examples are known where this bond points to the center of the aromatic p-system of
Ph groups (p-type interaction) [58][59].
23
24
For a definition of five-ring twist and envelope conformations, see [61 ± 63].

Helvetica Chimica Acta ± Vol. 82 (1999)
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Fig. 3. CD Spectra of b-peptides. a) Typical CD pattern of a 314 b-peptide helix [3]. b) All-(S)-b-tri- (7a), -hexa-
(10a), -dodeca- (11), and -octadecapeptide (12), consisting of (S)-b3-homoproline. c) (S)/(R)-b-tetra- (9) and
hexapeptide (15) with alternating (S)- and (R)-b3-homoproline residues. d) All-(S)-b2-di- (16), -tri- (17a) and
-hexapeptide (18) composed of (S)-b2-homoproline residues. All b-peptides were measured at 0.2 mm in MeOH
at r.t. The molar ellipticity [V] is represented in 10 dg ´ cm2 ´ mol 1, and is not normalized.
the normal trans-conformation. The angle F (Fig. 6,a), enforced by the pyrrolidine
ring, is ca. 728 according to the X-ray structure. An antiperiplanar conformation
around the C(a) C(b) bond (Vˆ 1808; Fig. 6,b), as encountered in the central residue
of the crystal structure of 7c, is suggested. The angle Y was chosen to be 1808 so that
the large substituents at the carbonyl C-atom and at C(a) are antiperiplanar (the CˆO
group lies between the g-CH2 group and the C(b)H atom; see Fig. 6,c). The resulting
model is a right-handed 103 helix with three pitches to bring residue (i ‡ 10) above
residue i (Fig. 7). The model unveils consecutive fully extended chain segments
(N C(b) C(a) CO N), which are twisted by 728 (Fig. 7,c).

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Helvetica Chimica Acta ± Vol. 82 (1999)
Fig. 4. X-Ray crystal structure of b-tripeptide derivative 7c. The peptide crystallized with four CF3CO2H
molecules. Two of these form H-bonds with amide-carbonyl O-atoms, and one CF3CO2 is the counterion of the
terminal ammonium group; the distance O ´´´ O or N ´´´ O is indicated in [Š]. H-Atoms have been omitted for
clarity.
Table. Torsion Angles in the Crystal Structure of b-Tripeptide Derivative 7c. The torsion angles are defined in
Figs. 1, a and b.
Residuea)
F [8]
V [8]
Y [8]
175.6
82.9
‡ 88.2
1
2
3
±
‡ 59.2
‡ 171.9
66.8
73.6
71.6
a) Numbering starts from the N-terminus.
Fig. 5. Two views of the crystal structure of the TFA salt 7c. a) Projection showing the quasi-parallel arrangement
of the Ph ring with the amide plane. The distance is indicated in [Š]. b) View in which the ap-conformation
around the C(a) C(b) bond of the central b-amino acid is visible.

Helvetica Chimica Acta ± Vol. 82 (1999)
1549
Fig. 6. Dihedral angles F, V, and Y used for the construction of a model for the structure of R-(b3-HPro)n-OR.
See discussion in the accompanying text and Fig. 7.
Fig. 7. Model consisting of (S)-b3-HPro, constructed with the torsion angles F ˆ 728, Vˆ Yˆ 1808. a) Side
view of a 103 helix. b) Top view of a 103 helix. Two of the pyrrolidine rings are in juxtaposition. c) Characteristic
fully extended chain segment (N C(b) C(a) CO N) identified in this structure. Model constructed with
MacMoMo (program by Prof. Dr. M. Dobler, ETH-Zürich).
Although one of the major forces that direct the self-assembly of biopolymers, i.e.,
the formation of a H-bonding network, is lacking in b-HPro-peptides, the present
results indicate that this class of b-peptides may still adopt distinct folding patterns due
to the high intrinsic folding propensity of the b-peptide backbone; this is strongly
suggested by the characteristic and intensive CD curves of the all-(S)-b2- and all-(S)-b3-
HPro-peptides (Fig. 3)25). Efforts to crystallize larger b-peptides, such as the b-
hexapeptide 10a, and to investigate its structure by 2D-NMR spectroscopy, are in
progress. The high solubility of b-HPro-peptides will facilitate the synthesis of even
larger b-peptides with defined structures by the incorporation of b-homoproline
residues.
We gratefully acknowledge generous supply of d-Pro from Degussa AG, Hanau. We would like to thank Dr.
P. Seiler (Laboratorium für Organische Chemie, ETH-Zurich) for the determination of the X-ray crystal
structure of 7c. A. Lerchner has carried out some of the experiments described herein, as part of the advanced
laboratory course in organic chemistry. Financial support from Novartis Pharma and Agro, Basel, is gratefully
acknowledged.
It is remarkable that the N-deprotected b3-HPro-tripeptide 7c, of which an X-ray crystal structure is
available (Figs. 4 and 5), gives rise to the same, albeit less intensive, CD pattern as the fully protected
derivative 7a (cf. Fig. 3,b, and Exper. Part).
25
)

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Helvetica Chimica Acta ± Vol. 82 (1999)
Experimental Part
1. General. Abbreviations: Boc2O: di(tert-butyl) dicarbonate, DMAP: 4-(dimethylamino)pyridine, EDC:
1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride, er: enantiomer ratio, HOBt: 1-hydroxy-1H-
benzotriazole, h.v.: high vacuum (0.01 ± 0.1 Torr), b-HXaa (b-homoamino acid) [3][5], TFA: trifluoroacetic
acid, CHCl3 employed for the coupling reactions was filtered through Al2O3 (Alumina Woelm N, activity I) to
remove EtOH. Et3N was distilled from CaH2 and stored under Ar. Solvents for chromatography and workup
procedures were distilled from Sikkon (anh. CaSO4; Fluka). Boc-Pro was purchased from Senn, d-Pro was a gift
from Degussa. All other reagents were used as received from Fluka or Quantum Biotechnologies, Montreuil
(EDC). TLC: Merck silica gel 60 F254 plates; detection with UV, I2 (30 g of I2, 20 g of KI, 200 ml of EtOH,
200 ml of H2O), anisaldehyde (9.2 ml of anisaldehyde, 3.75 ml of AcOH, 12.5 ml of conc. H2SO4, 350 ml of
EtOH) or ninhydrine (0.6 g of ninhydrine, 2 ml of HOAc, 13 ml of H2O, 285 ml of BuOH). FC: Fluka silica gel
60 (40 ± 63 mm); at ca. 0.3 bar. Anal. HPLC: Knauer HPLC system (pump type 64, EuroChrom 2000 integration
package, degaser, UV detector (variable-wavelength monitor)). M.p.: Büchi-510 apparatus; uncorrected. UV
Spectra: Uvikon 860 Kontron Instruments (1-cm rectangular cell) at r.t., lmax in nm. Optical rotations: Perkin-
Elmer 241 polarimeter (10 cm, 1-ml cell) at r.t. CD Spectra: Jasco J-710 between 190 and 250 nm in a 1-mm
rectangular cell at r.t. The optical system was flushed with N2 at a flow rate of ca. 10 l ´ min 1. Band width 1.0 nm,
resolution 0.5 nm, sensitivity 100 mdeg, response 0.5 s, speed 50 nm ´ min 1. All spectra were the average of five
scans and were corrected for the corresponding solvent spectrum. Peptide concentration: 0.2 mm in MeOH. The
molar ellipticity V is reported in deg ´ cm2 ´ dmol 1. Smoothing was performed with the software provided by
Jasco. IR Spectra: Perkin-Elmer-782 spectrophotometer. NMR Spectra: Bruker AMX 500 (1H: 500 MHz, 13C:
125 MHz), AMX 400 (1H: 400 MHz, 13C: 100 MHz); chemical shifts d in ppm downfield from internal Me4Si
(ˆ 0 ppm); J values in Hz; some compounds show the presence of rotamers which are indicated. MS: VG Tribrid
(EI), Hitachi Perkin-Elmer RHU-6M (FAB, in a 3-nitrobenzyl alcohol matrix), or Finnigan MAT TSQ 7000
(ESI) spectrometer; in m/z (% of basis peak). Elemental analyses were performed by the Microanalytical
Laboratory of the Laboratorium für Organische Chemie, ETH-Zürich.
2. Ester Hydrolysis: General Procedure 1 (GP 1). A soln. of the fully protected amino acid in MeOH (0.2m)
was treated with a soln. of LiOH (2.5 equiv.) in H2O (MeOH/H2O 3 :1 (v/v)) at r.t. After stirring at r.t. for 1 ±
3 d, the mixture was diluted with H2O (for small-scale reactions) and extracted with Et2O (2Â). The soln. was
adjusted to pH 2 at 08 with 10% HCl and extracted with Et2O (3Â). The org. phase was washed with H2O, dried
(MgSO4), and concentrated under reduced pressure.
3. Benzyl-Ester Deprotection: General Procedure 2 (GP 2). The benzyl ester was dissolved in the
appropriate solvent (0.1m), and a catalytic amount of 10% Pd/C was added. The apparatus was evacuated and
flushed with H2 (3Â), and the mixture was stirred under H2 for 18 h. Subsequent filtration through Celite and
concentration under reduced pressure yielded the crude carboxylic acid, which was identified by NMR and
FAB-MS and used without further purification.
4. Boc Deprotection: General Procedure 3 (GP 3). Similarly to the reported procedure [3][5], the Boc-
protected amino acid was dissolved in CH2Cl2 (0.5m) and cooled to 08. An equal volume of TFA was added, and
the mixture was allowed to warm slowly to r.t. and stirred for further 1.5 h. Concentration under reduced
pressure, co-evaporation with CH2Cl2, and drying of the residue under h.v. yielded the crude TFA salt, which
was identified by NMR and FAB-MS and used without further purification.
5. Peptide Coupling with EDC: General Procedure 4 (GP 4). The appropriate TFA salt was dissolved in
CHCl3 (0.5m) and cooled to 08. This soln. was treated successively with Et3N (4 equiv.), HOBt (1.2 equiv.), a
soln. of the Boc-protected fragment (1 equiv.) in CHCl3 (0.25m), and EDC (1.2 equiv.). The mixture was
allowed to warm to r.t. After TLC displayed complete reaction (12 h ± 3 d), the mixture was subsequently
diluted with CHCl3, followed by thorough washing with 1n HCl, sat. aq. NaHCO3 (3Â), and NaCl solns. (1Â).
The org. phase was dried (MgSO4) and the concentrated under reduced pressure. FC or recrystallization yielded
the pure peptide.
6. HPLC Analysis of (R)- or (S)-5: a) Derivatization of 4a with 1-Fluoro-2,4-dinitrobenzene: General
Procedure 5 (GP 5). To a soln. of 4a in H2O (0.5m) was added NaHCO3 (1.2 equiv.) and a soln. of 1-fluoro-2,4-
dinitrobenzene (1.2 equiv.) in EtOH (0.35m) at 08. After 1 h, EtOH was evaporated and the pH adjusted to 2
with 1n HCl, and the residue was extracted with Et2O (2Â). The Et2O phase was filtered through a Buchner
funnel (G4) packed with silica gel on a MgSO4 layer and evaporated to yield crude 5. The yellow oil was
dissolved in i-PrOH/hexane 35 : 165 (1 mg/ml) and injected onto the HPLC system according to GP 7.

Helvetica Chimica Acta ± Vol. 82 (1999)
1551
26
b) Derivatization of 4c: General Procedure 6 (GP 6). A soln. of 4c (5 mg, 0.022 mmol) in HCl/EtOH
)
(1 ml, 4m) was heated to 1108 for 1.5 h in a Wheats V-Vial (with Teflon-faced rubber septum) to yield 4a ´ HCl.
The HCl salt was further derivatized according to GP 5.
c) HPLC for Determination of Enantiomer Ratio of 5: General Procedure 7 (GP 7). HPLC Analyses were
performed on a Daicel Chiralcel OD column (4.6 Â 250 mm, 10 mm) by using an isocratic eluent of i-PrOH/
hexane 35 : 165 at a flow rate of 1 ml/min with UV detection at 390 nm at 258, tR in min.
7. b3-Peptides. tert-Butyl (R)-2-(Diazoacetyl)pyrrolidine-1-carboxylate (Boc-(R)-Pro-CHN2; (R)-1). Boc-
d-Pro-OH (13.99 g, 65.0 mmol) was transformed with CH2N2 according to [35]. FC (AcOEt/pentane 1:3)
yielded (R)-1 (8.70 g, 56%). Yellow, waxy solid. M.p. 47 ± 488. Rf (AcOEt/pentane 1 :3) 0.25. [a]rD:t: ˆ‡ 146 (c ˆ
1.0, CHCl3). IR (CHCl3): 3120w, 3008m, 2980m, 2881w, 2110s, 1690s, 1646m, 1477w, 1454w, 1394s, 1367s, 1323m,
1163m, 1123m. 1H-NMR (400 MHz, CDCl3; values for rotamers in italics): 1.44, 1.48 (s, t-Bu); 1.84 ± 2.27
(m, CH2CH2); 3.36 ± 3.56 (m, CH2N); 4.24 (br., NCH); 5.44 (br., CHN2). 13C-NMR (100 MHz, CDCl3; values
for rotamers in italics): 23.71, 24.41 (CH2); 28.38, 28.44 (Me); 29.67, 31.27, 46.79, 47.10 (CH2); 52.01, 53.11,
63.60, 64.48 (CH); 80.13, 80.45, 109.16, 154.14, 154.82, 195.09, 196.06 (C). EI-MS: 170 (16.2), 114 (52.4), 70
(93.8), 57 (100). Anal. calc. for C11H17N3O4 (239.27): C 55.22, H 7.16, N 17.56; found: C 55.38, H 7.22, N 17.36.
tert-Butyl (S)-2-(Diazoacetyl)pyrrolidine-1-carboxylate (Boc-(S)-Pro CHN2; (S)-1). Boc-l-Pro OH
(26.9 g, 125 mmol) was transformed with CH2N2 according to [35]. FC (AcOEt/pentane 1 :3) yielded (S)-1
(22.9 g, 77%). Yellow, waxy solid. Rf (AcOEt/pentane 1:3) 0.25. [a]Dr:t:
spectroscopic data: corresponding to (R)-1.
ˆ
145 (c ˆ 1.0, CHCl3). Other
tert-Butyl (S)-2-[2-(Benzyloxy)-2-oxoethyl]pyrrolidine-1-carboxylate (Boc-(S)-b3-HPro-OBn; (S)-2a).
Compound (S)-1 (6.00 g, 25.0 mmol) was rearranged with CF3COOAg (10%) in BnOH/THF 15 :85 (v/v)
according to [22]. After removal of BnOH by distillation (0.5 mbar, 688), FC (Et2O/pentane 1:2) yielded (S)-2a
(5.40 g, 68%). Colorless oil. Rf (Et2O/pentane 1:2) 0.32. [a]Dr:t:
ˆ
35.7 (c ˆ 1.0, CHCl3). IR (CHCl3): 3007w,
1
2977m, 2882w, 1730m, 1684s, 1477w, 1455w, 1403s, 1367m, 1304w, 1166m, 1125m. H-NMR (400 MHz, CDCl3;
values for rotamers in italics): 1.45 (s, t-Bu); 1.70 ± 1.89 (m, 3 CH); 2.00 ± 2.09 (m, 1 CH); 2.36 (dd, J ˆ 15.1, 9.8,
COCH); 2.82 ± 3.04 (br., COCH); 3.30 ± 3.34 (br., CH2N); 4.12, 4.20 (s, NCH); 5.12 (s, PhCH2); 7.35
(m, 5 arom. H). 13C-NMR (100 MHz, CDCl3; values for rotamers in italics): 22.80, 23.53 (CH2); 28.50 (Me);
30.56, 31.27, 38.53, 39.37, 46.20, 46.59 (CH2); 54.05 (CH); 66.21 (CH2); 79.27, 79.62 (C); 128.21, 128.54 (CH);
‡
‡
135.92, 154.27, 171.36 (C). EI-MS: 320 (< 1, [M ‡ 1] ), 319 (< 1, M ), 218 (100), 128 (67.2), 91 (89.0), 70
(45.3). Anal. calc. for C18H25NO4 (319.40): C 67.69, H 7.89, N 4.39; found: C 67.83, H 7.89, N 4.42.
tert-Butyl (R)-2-[2-(Benzyloxy)-2-oxoethyl]pyrrolidine-1-carboxylate (Boc-(R)-b3-HPro-OBn; (R)-2a).
Compound (R)-1 (3.00 g, 12.5 mmol) was rearranged with CF3COOAg (10%) in BnOH/THF 15 :85 (v/v)
according to [22]. After removal of BnOH by distillation (0.5 mbar, 688), FC (Et2O/pentane 1:2) yielded (R)-
2a (2.99 g, 75%). Colorless oil. [a]rD:t: ˆ‡ 36.3 (c ˆ 1.0, CHCl3). Other spectroscopic data: corresponding to
(S)-2a.
(R)-1-[(tert-Butoxy)carbonyl]pyrrolidine-2-acetic Acid (Boc-(R)-b3-HPro-OH; (R)-2b). Compound
(R)-1 (5.20 g, 21.7 mmol) was rearranged with CF3COOAg (10%) in H2O/THF 10 :90 (v/v) according to
[22]. Recrystallization (CH2Cl2/hexane) yielded (R)-2b (3.76 g, 76%). White powder. M.p. 99 ± 1008. Rf
(CH2Cl2/MeOH 20 :1) 0.30. [a]rD:t: ˆ‡ 40.6 (c ˆ 1.9, DMF). IR (CHCl3): 2980m, 2881w, 1711s, 1684s, 1477w,
1403s, 1368m, 1286w, 1168s, 1127m, 927w, 860w. 1H-NMR (400 MHz, CD3OCD3): 1.44 (s, t-Bu); 1.77 ± 1.96
(m, 3 CH); 2.01 ± 2.10 (m, 1 CH); 2.26 ± 2.37 (m, COCH); 2.75 ± 2.95 (m, COCH); 3.31 (m, CH2N); 4.05 ± 4.11
(m, NCH). 13C-NMR (100 MHz, CD3OCD3; values for rotamers in italics): 28.28, 24.06 (CH2); 28.63 (Me);
31.19, 32.00, 38.48, 39.55, 46.90, 47.25 (CH2); 54.91 (CH); 79.28, 154.45, 154.76, 172.85 (C). FAB-MS: 481 (13.4,
‡
‡
‡
‡
[2M ‡ Na] ), 459 (14.7, [2M ‡ 1] ), 252 (28.6, [M ‡ Na] ), 230 (100, [M ‡ 1] ), 174 (92.1), 130 (55.6). Anal.
calc. for C11H19NO4 (229.28): C 57.63, H 8.35, N 6.11; found: C 57.51, H 8.34, N 6.04.
(S)-1-[(tert-Butoxy)carbonyl]pyrrolidine-2-acetic Acid (Boc-(S)-b3-HPro-OH; (S)-2b). Compound (S)-1
(20.34 g, 85.0 mmol) was rearranged with CF3COOAg (10%) in H2O/THF 10 :90 (v/v) according to [22].
Recrystallization (CH2Cl2/hexane) yielded (S)-2b (11.86 g, 61%). White powder. [a]Dr:t:
ˆ
40.5 (c ˆ 1.9, DMF)
([27]: [a]rD:t:
corresponding to (R)-2b.
ˆ
41.6 (c ˆ 1.9, DMF); [28]: [a]Dr:t:
ˆ
39.5 (c ˆ 1.9, DMF)). Other spectroscopic data:
Boc-(S)-b3-HPro-(S)-b3-HPro-OBn (6). Compound (S)-2a (639 mg, 2.0 mmol) was Boc-deprotected
according to GP 3. The resulting TFA salt was coupled with (S)-2b (459 mg, 2.0 mmol) according to GP 4 for
26
)
Freshly prepared according to [47] by the slow addition of EtOH (0.5 ml) to AcCl (0.85 ml) at 08 and
dilution to 1 ml.

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Helvetica Chimica Acta ± Vol. 82 (1999)
2.5 d. FC (AcOEt/pentane 1:1) yielded 6 (540 mg, 63%). White, waxy solid. M.p. 68 ± 698. Rf (AcOEt/pentane
1:1) 0.31. [a]Dr:t:
ˆ
60.8 (c ˆ 1.0, CHCl3). IR (CHCl3): 3007m, 2978m, 2880w, 1730m, 1680s, 1634m, 1454m,
1401s, 1366m, 1305w, 1168m, 1124w, 907w. 1H-NMR (400 MHz, CDCl3): 1.46 (s, t-Bu); 1.81 ± 2.55 (m, 8 CH,
2 COCH); 2.98 ± 3.03 (m, 2 COCH); 3.31 ± 3.68 (m, 4 CHN); 4.05 ± 4.13 (m, NCH); 4.39 ± 4.45 (m, NCH);
5.07 ± 5.14 (m, PhCH2); 7.30 ± 7.39 (m, 5 arom. H). 13C-NMR (100 MHz, CDCl3; values for rotamers in italics):
21.54, 23.47, 24.00 (CH2); 28.58 (Me); 29.91, 30.12, 31.20, 37.55, 39.24, 45.23, 46.28 (CH2); 53.74, 54.24, 54.57
(CH); 66.25, 66.63 (CH2); 79.10, 79.51 (C); 128.20, 128.32, 128.54, 128.67 (CH); 135.92, 154.40, 169.89, 171.31
‡
‡
(C). FAB-MS: 453 (5.9, [M ‡ Na] ), 431 (66.5, [M ‡ 1] ), 331 (100), 329 (35.0). Anal. calc. for C24H34N2O5
(430.54): C 66.95, H 7.96, N 6.51; found: C 66.76, H 7.88, N 6.56.
Boc-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-OBn (7a). Compound 6 (5.73 g, 13.3 mmol) was Boc-depro-
tected according to GP 3. The resulting TFA salt was coupled with (S)-2b (3.05 g, 13.3 mmol) according to GP 4
for 16 h. FC (MeOH/CH2Cl2 1:22 ! 1 : 10) yielded 7a (6.58 g, 91%). Colorless, highly viscous oil. Rf (CH2Cl2/
MeOH 22 : 1) 0.29. [a]Dr:t:
ˆ
69.6 (c ˆ 1.0, CHCl3). UV (0.2 mm, MeOH): lmax 213 nm. CD (0.2 mm, MeOH):
1.32 ´ 105 (202 nm), ‡ 4.09 ´ 104 (222 nm). IR (CHCl3): 3007m, 2977m, 2879w, 1729m, 1681s, 1632s, 1402s,
1366m, 1168m, 1124w, 907w. 1H-NMR (400 MHz, CDCl3): 1.46 (s, t-Bu); 1.74 ± 2.36 (m, 16 CH); 2.94 ± 3.02
(m, 2 COCH); 3.32 ± 3.68 (m, 6 NCH); 4.13 ± 4.15 (m, NCH); 4.27 ± 4.30 (m, NCH); 4.38 ± 4.45 (m, NCH);
5.07 ± 5.14 (m, PhCH2); 7.29 ± 7.36 (m, 5 arom. H). 13C-NMR (100 MHz, CDCl3; values for rotamers in italics):
23.82, 23.93, 24.02 (CH2); 28.57 (Me); 29.36, 29.92, 30.57, 37.61, 38.30, 39.17, 46.64, 47.18, 47.29 (CH2); 53.72,
54.13 (CH); 66.24, 66.57 (CH2); 79.13, 79.47 (C); 128.17, 128.32, 128.52, 128.62 (CH); 135.93, 154.39, 169.60,
‡
169.75, 171.30 (C). FAB-MS: 542 (42.9, [M ‡ 1] ), 443 (30.5), 442 (100), 91 (100). Anal. calc. for C30H43N3O6
(541.69): C 66.52, H 8.00, N 7.76; found: C 66.66, H 7.86, N 7.63.
Boc-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-OH (7b). Compound 7a (3.20 g, 5.9 mmol) was debenzylated
in AcOEt according to GP 2 to yield 7b (2.65 g, quant.). White powder. M.p. 63 ± 688. Rf (CH2Cl2/MeOH 12 :1)
0.28. [a]rD:t:
ˆ
56.5 (c ˆ 1.0, CHCl3). IR (CHCl3): 3007m, 2979m, 2880w, 1728m, 1681s, 1629s, 1451m, 1401s,
1367m, 1170m, 1125w, 907w. 1H-NMR (400 MHz, CD3OD): 1.45 (s, t-Bu); 1.80 ± 2.11 (m, 12 CH); 2.22 ± 2.43
(m, 3 COCH); 2.77 ± 3.03 (m, 3COCH); 3.28 ± 3.66 (m, 6 NCH); 4.10 ± 4.15 (m, NCH); 4.30 ± 4.35
(br., 2 NCH). 13C-NMR (100 MHz, CD3OD; values for rotamers in italics): 22.41, 24.54, 24.67 (CH2); 28.85
(Me); 30.81, 30.99, 32.17, 38.03, 39.02, 40.03, 46.50, 47.33, 47.72 (CH2); 55.45, 55.52, 55.59 (CH); 73.93, 80.70,
‡
‡
81.36, 156.16, 171.92, 175.04 (C). FAB-MS: 474 (16.2, [M ‡ Na] ), 452 (24.4, [M ‡ 1] ), 352 (100), 241 (48.0).
Anal. calc. for C23H37N3O6 (451.56): C 61.18, H 8.26, N 9.31; found: C 61.09, H 8.16, N 9.11.
TFA ´ H-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-OBn (7c). Compound 7a (2.59 g. 4.8 mmol) was Boc-
deprotected according to GP 3 to yield 7c (3.41 g, quant.). Colorless oil, which crystallized after 15 d upon
storage at 208: colorless crystals, suitable for X-ray analysis. CD (0.2 mm, MeOH): 2.83 ´ 104 (203 nm), 8.80 ´
103 (223nm). 1H-NMR (400 MHz, CDCl3): 1.78 ± 2.26 (m, 12CH); 2.43 (dd, J ˆ 14.3, 7.4, COCH); 2.51 (dd, J ˆ
15.5, 8.4, COCH); 2.70 (dd, J ˆ 14.3, 6.8, COCH); 2.83 ± 2.92 (m, 3 COCH); 3.35 ± 3.61 (m, 6 NCH); 3.88 ± 3.94
(m, NCH); 4.33 ± 4.38 (m, NCH); 4.49 ± 4.52 (m, NCH); 5.10 ± 5.17 (m, PhCH2); 7.33 ± 7.42 (m, 5 arom. H);
8.55 (br., NH); 8.91 (br., NH). 13C-NMR (100 MHz, CDCl3; values for rotamers in italics): 23.23, 23.36, 23.61,
29.25, 30.07, 30.19, 34.23, 37.37, 38.25, 45.58, 47.17, 47.89 (CH2); 53.92, 55.32, 57.57 (CH); 66.78, 67.03 (CH2);
115.23 (q, J ˆ 282.9, CF3); 128.25, 128.34, 128.42, 128.62, 128.71 (CH); 135.20, 135.52 (C); 160.20 (q, J ˆ 40.0,
‡
‡
‡
CCF3); 169.90, 171.28, 171.31 (C). FAB-MS: 883 (12.1, [2M] ), 443 (36.1, [M ‡ 1] ), 442 (100, M ).
Boc-(S)-b3-HPro-(R)-b3-HPro-OBn (8a). Compound (R)-2a (3.23 g, 10.1 mmol) was Boc-deprotected
according to GP 3. The resulting TFA salt was coupled with (S)-2b (2.42 g, 10.1 mmol) according to GP 4 for
2 d. FC (AcOEt/pentane 1:1) yielded 8a (2.82 g, 62%). Colorless oil. Rf (AcOEt/pentane 1:1) 0.26. [a]Dr:t:
ˆ
6.02 (c ˆ 1.0, CHCl3). IR (CHCl3): 3007m, 2978m, 2879w, 1730m, 1681s, 1635m, 1456m, 1401s, 1367m,
1168m, 1125w, 907w. 1H-NMR (400 MHz, CDCl3; values for rotamers in italics): 1.43, 1.46 (s, t-Bu); 1.81 ± 2.21
(m, 10 CH); 2.72 ± 3.04 (m, 2 COCH); 3.32 ± 3.57 (m, 4 NCH); 4.15 (m, NCH); 4.41 (m, NCH); 5.07 ± 5.14
(m, PhCH2); 7.30 ± 7.38 (m, 5 arom. H). 13C-NMR (100 MHz, CDCl3; values for rotamers in italics): 23.52,
23.95 (CH2); 28.59 (Me); 30.08, 31.26, 37.69, 39.00, 45.42, 47.19 (CH2); 53.82, 54.28 (CH); 66.25 (CH2); 79.09,
‡
79.49 (C); 128.21, 128.28, 128.53 (CH); 135.94, 154.41, 169.73, 171.30 (C). FAB-MS: 430 (3.8, M ), 329 (64.0),
91 (100). Anal. calc. for C24H34N2O5 (430.54): C 66.95, H 7.96, N 6.51; found: C 66.88, H 7.86, N 6.56.
Boc-(S)-b3-HPro-(R)-b3-HPro-OH (8b). Compound 8a (3.30g, 7.7 mmol) was debenzylated according to
GP 2 in AcOEt to yield 8b (2.65 g, quant.). White powder. M.p. 140 ± 1418. Rf (CH2Cl2/MeOH 9 :1) 0.35.
[a]rD:t:
ˆ
38.4 (c ˆ 1.0, CHCl3). IR (CHCl3): 3007m, 2980m, 2880w, 1729m, 1681s, 1627m, 1455m, 1401s, 1367m,
1169m, 1126w, 904w. 1H-NMR (400 MHz, CD3OD): 1.46 (s, t-Bu); 1.83-2.13 (m, 8 CH); 2.34 ± 2.51
(m, 2 COCH); 2.68 ± 2.89 (m, 2 COCH); 3.28 ± 3.60 (m, 4 NCH); 4.10 ± 4.22 (m, NCH). 13C-NMR (100 MHz,
CD3OD; values for rotamers in italics): 22.42, 24.64 (CH2); 28.84 (Me); 31.12, 38.12, 39.86, 40.37, 46.60, 47.37,

Helvetica Chimica Acta ± Vol. 82 (1999)
1553
‡
47.76 (CH2); 55.51, 56.35 (CH); 80.74, 81.35, 156.22, 172.02, 175.08 (C). FAB-MS: 703 (3.8, [M ‡ Na] ), 681
‡
‡
‡
(7.7, [2M ‡ 1] ), 363 (11.4, [M ‡ Na] ), 341 (100, [M ‡ 1] ), 241 (95.8). Anal. calc. for C17H28N2O5 (340.42):
C 59.98, H 8.29, N 8.23; found: C 60.10, H 8.12, N 8.22.
Boc-(S)-b3-HPro-(R)-b3-HPro-(S)-b3-HPro-(R)-b3-HPro-OBn (9). Compound 8a (1.68 g, 3.9 mmol) was
Boc-deprotected according to GP 3. The resulting TFA salt was coupled with 8b (1.33 g, 3.9 mmol) according to
GP 4 for 16 h. FC (MeOH/CH2Cl2 1 :17) yielded 9 (2.08 g, 81%). White solid. M.p. 156 ± 1598. Rf (CH2Cl2/
MeOH 17:1) 0.30. [a]Dr:t:
ˆ
2.12 (c ˆ 1.0, CHCl3). CD (0.2 mm, MeOH): ‡ 9.86 ´ 103 (214 nm). IR (CHCl3):
3007m, 2979m, 2879w, 1730m, 1681m, 1631s, 1495s, 1366m, 1169m, 1124w, 1103w, 907w. 1H-NMR (400 MHz,
CDCl3; values for rotamers in italics): 1.42, 1.46 (s, t-Bu); 1.70 ± 2.21 (m, 19 CH); 2.40 (dd, J ˆ 15.3, 9.2, COCH);
2.71 ± 3.02 (m, 4 COCH); 3.35 ± 3.66 (m, 8 NCH); 4.07 ± 4.18 (m, NCH); 4.31 ± 4.41 (m, 3 NCH); 5.07 ± 5.15
(m, PhCH2); 7.29 ± 7.38 (m, 5 arom. H). 13C-NMR (100 MHz, CDCl3; values for rotamers in italics): 21.65,
23.83, 23.91, 23.98 (CH2); 29.59 (Me); 29.71, 29.76, 29.84, 30.07, 30.12, 30.61, 31.59, 37.03, 37.63, 37.69, 37.83,
38.18, 38.40, 39.46, 45.57, 47.05, 47.17, 47.26 (CH2); 53.87, 53.99, 54.13, 54.22, 54.57 (CH); 66.26, 66.37 (CH2);
79.16, 79.59, 128.20, 128.24, 128.28, 128.38, 128.54, 128.57, 128.62, 135.94, 154.42, 169.61, 171.36 (C). FAB-MS:
‡
653 (100, [M ‡ 1] ), 554 (21.0), 553 (57.4). Anal. calc. for C36H52N4O7 (652.83): C 66.23, H 8.03, N 8.58; found:
C 66.05, H 7.96, N 8.60.
Boc-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-OBn (10a). Com-
pound 7a (2.60 g, 4.8 mmol) was Boc-deprotected according to GP 3. The resulting TFA salt was coupled
with 7b (2.16 g, 4.8 mmol) according to GP 4 for 2.5 d. FC (MeOH/CH2Cl2 1:15) yielded 10a (3.54 g, 85%).
White foam. M.p. 65 ± 688. Rf (CH2Cl2/MeOH 15 :1) 0.28. [a]Dr:t:
ˆ
84.8 (c ˆ 1.0, CHCl3). UV (0.2 mm, MeOH):
lmax 217 nm. CD (0.2 mm, MeOH): 1.90 ´ 105 (202 nm), ‡ 9.02 ´ 104 (222 nm). M.p. 65 ± 688. IR (CHCl3):
3007m, 2978m, 2879w, 1730w, 1680m, 1632s, 1425s, 1366w, 1170m, 1123w, 1097w, 907w. 1H-NMR (400 MHz,
CDCl3): 1.46 (s, t-Bu); 1.72 ± 2.15 (m, 30 CH); 2.33 (dd, J ˆ 15.4, 9.5, COCH); 2.96 ± 3.05 (m, 5 COCH); 3.36 ±
3.66 (m, 12 NCH); 4.11 ± 4.18 (m, NCH); 4.29 ± 4.44 (m, 5 NCH); 5.07 ± 5.30 (m, PhCH2); 7.29 ± 7.37 (m, 5 ar-
om. H). 13C-NMR (100 MHz, CDCl3; values for rotamers in italics): 21.57, 23.48, 23.80, 23.93, 24.01 (CH2);
28.58 (Me), 29.40, 29.83, 29.86, 29.92, 30.61, 37.62, 38.11, 38.17, 38.31, 39.11, 45.25, 46.65, 47.11, 47.15, 47.23,
47.28, 47.37 (CH2); 53.71, 54.03, 54.06, 54.11, 54.25, 54.49 (CH); 66.22, 66.55 (CH2); 79.14, 79.48 (C); 128.15,
128.31, 128.38, 128.52, 128.60 (CH); 135.68, 135.96, 154.42, 169.51, 169.62, 169.71, 169.74, 171.32 (C). FAB-MS:
‡
‡
‡
898 (13.1, [M ‡ Na] ), 876 (27.1, [M ‡ 1] ), 875 (52.9, M ), 775 (100). Anal. calc. for C48H70N6O9 (875.12):
C 65.43, H 8.69, N 9.54; found: C 65.47, H 8.87, N 9.45.
Boc-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-OH (10b). Com-
pound 10a (1.01 g, 1.2 mmol) was debenzylated in AcOEt according to GP 2 to yield 10b (1.02 g, quant.).
White foam. M.p. 69 ± 738. Rf (CH2Cl2/MeOH 7:1) 0.24. [a]Dr:t:
ˆ
67.3 (c ˆ 1.0, CHCl3). CD (0.2 mm, MeOH):
8.14 ´ 104 (204 nm), ‡ 5.20 ´ 104 (224 nm). IR (CHCl3): 3307m, 2979m, 2879w, 1728w, 1681m, 1630s, 1423s,
1366w, 1173w, 1048w, 881w. 1H-NMR (400 MHz, CD3OD): 1.45 (s, t-Bu); 1.83 ± 2.16 (m, 24 CH); 2.23 ± 2.38
(m, 6 COCH); 2.46 ± 2.59 (m, COCH); 2.77 ± 3.03 (m, 5 COCH); 3.30 ± 3.71 (m, 12 NCH); 4.09 ± 4.14
(m, NCH); 4.31 ± 4.49 (m, 5 NCH). 13C-NMR (100 MHz, CD3OD; values for rotamers in italics): 22.41,
24.33, 24.55, 24.67 (CH2); 28.78, 28.86 (Me); 30.85, 31.01, 31.42, 31.69, 32.16, 38.07, 39.01, 40.03, 46.50, 47.36,
47.72 (CH2); 55.45, 55.51, 55.60, 56.11, 56.22, 57.81 (CH); 80.69, 81.35, 156.16, 171.02, 171.14, 171.92, 172.03,
‡
‡
‡
174.40, 175.11 (C). FAB-MS: 808 (39.1, [M ‡ Na] ), 807 (91.7), 786 (42.1, [M ‡ 1] ), 785 (100, M ), 685
(60.5).
Boc-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b
3HProS)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-OBn (11). Compound 10a (166 mg, 0.19 mmol) was
Boc-deprotected according to GP 3. The resulting TFA salt was coupled with 10b (150 mg, 0.19 mmol)
according to GP 4 for 40 h. Recrystallization (CH2Cl2/hexane) yielded 11 (233mg, 79%). White foam. M.p. 958
(dec.). Rf (CH2Cl2/MeOH 9 :1) 0.30. [a]Dr:t:
ˆ
87.4 (c ˆ 1.0, CHCl3). CD (0.1 mm, MeOH):
2.63 ´ 105
(203 nm), ‡ 1.82 ´ 105 (223 nm). IR (CHCl3): 3005m, 2878w, 1729m, 1631s, 1525m, 1426s, 1352m, 857w.
1H-NMR (400 MHz, CDCl3): 1.46 (s, t-Bu); 1.74 ± 2.17 (m, 60 CH); 2.33 (dd, J ˆ 15.3, 9.5, COCH); 2.96 ± 3.02
(m, 11 COCH); 3.36 ± 3.67 (m, 24 NCH); 4.15 ± 4.49 (m, 12 NCH); 5.07 ± 5.14 (m, PhCH2); 7.28 ± 7.38 (m, 5 ar-
om. H). 13C-NMR (100 MHz, CDCl3): 21.53, 22.66, 23.78, 24.01 (CH2); 28.58 (Me); 29.39, 29.71, 29.88, 29.91,
31.59, 37.61, 38.10, 38.16, 38.30, 47.12, 47.28 (CH2); 53.44, 53.71, 54.05, 54.10, 54.23, 54.52 (CH); 66.23 (CH2);
79.16 (C); 128.16, 128.32, 128.40, 128.52, 128.61 (CH); 135.95, 154.44, 168.64, 169.32, 169.64, 169.74, 171.35 (C).
‡
‡
‡
FAB-MS: 1565 (4.1, [M ‡ Na] ), 1543 (100, [M ‡ 1] ). ESI-MS (pos. mode): 1580.3 ([M ‡ K] ), 1564.4 ([M ‡
‡
‡
Na] ), 1542.3 ([M ‡ 1] ).
Boc-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-
HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-

1554
Helvetica Chimica Acta ± Vol. 82 (1999)
(S)-b3-HPro-(S)-b3-HPro-OBn (12). Compound 11 (62 mg, 0.040 mmol) was Boc-deprotected according to
GP 3. The resulting TFA salt was coupled with 10b (31 mg, 0.040 mmol) according to GP 4 for 3 d.
Recrystallization (CH2Cl2/pentane) yielded 12 (48 mg, 54%). White powder. M.p. 2288 (dec.). Rf (CH2Cl2/
MeOH 9 :1) 0.22. [a]Dr:t:
ˆ
74.0 (c ˆ 1.0, CHCl3). CD (0.2 mm, MeOH): 3.46 ´ 105 (202nm), ‡ 2.09 ´ 105
(223 nm). IR (CHCl3): 3006m, 2878w, 1728w, 1680w, 1632s, 1425m, 1360w, 1323w. 1H-NMR (500 MHz, CDCl3):
1.46 (s, t-Bu); 1.83 ± 2.18 (m, 90 CH); 2.33 (dd, J ˆ 15.4, 9.5, COCH); 2.92 ± 3.02 (m, 17 COCH); 3.33 ± 3.65
(m, 36 NCH); 4.12 ± 4.52 (m, 18 NCH); 5.07 ± 5.13 (m, PhCH2); 7.29 ± 7.37 (m, 5 arom. H). 13C-NMR
(125 MHz, CDCl3): 23.78, 24.01 (CH2); 28.60 (Me); 29.42, 29.89, 37.62, 38.12, 38.40, 47.13, 47.29 (CH2);
53.44, 53.73, 54.06, 54.12, 54.47 (CH); 66.24 (C); 128.16, 128.31, 128.52 (CH); 135.96, 154.45, 169.67, 169.76,
‡
‡
171.34 (C). FAB-MS: 2208 (100, M ), 1104 (52.2). ESI-MS (pos. mode): 2231.8 ([M ‡ Na] ).
Ac-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-OBn (13). Fully pro-
tected b3-hexapeptide 10a (0.197 g, 0.225 mmol) was Boc-deprotected according to GP 3. The resulting TFA salt
was dissolved in CH2Cl2 (2.3 ml) and treated at 08 with Ac2O (0.03 ml, 0.293 mmol) and (i-Pr)2EtN (0.12 ml,
0.675 mmol). After 2.5 h, the soln. was diluted with CH2Cl2 and washed with 1n HCl, sat. aq. NaHCO3, and
NaCl solns. The org. phase was dried (MgSO4) and evaporated under reduced pressure. FC (MeOH/CH2Cl2
1:12) yielded 13 (101 mg, 55%). White foam. M.p. 63 ± 678. Rf (CH2Cl2/MeOH 12 :1) 0.26. [a]Dr:t:
ˆ
77.9 (c ˆ
1.0, CHCl3). CD (0.2 mm, MeOH): 1.14 ´ 105 (203 nm), ‡ 6.42 ´ 104 (223 nm). IR (CHCl3): 3004m, 2878w,
1730w, 1631s, 1503w, 1422m, 1359w, 1325w. 1H-NMR (400 MHz, CDCl3): 1.73 ± 2.18 (m, 33 CH); 2.32 (dd, J ˆ
15.4, 9.5, COCH); 2.96 ± 3.09 (m, 5 COCH); 3.29 ± 3.73 (m, 12 NCH); 4.29 ± 4.44 (m, 6 NCH); 5.07 ± 5.14
(m, PhCH2); 7.28 ± 7.37 (m, 5 arom. H). 13C-NMR (100 MHz, CDCl3; values for rotamers in italics): 14.05 (C);
22.21, 22.33 (CH2); 23.03 (Me); 23.76, 23.90, 23.99, 29.38, 29.72, 29.81, 29.84, 29.90, 30.04, 31.63, 37.59, 37.94,
38.11, 38.15, 38.29, 39.63, 45.25, 45.47. 47.13, 47.21, 47.27, 47.88 (CH2); 53.70, 54.01, 54.05, 54.09, 54.16, 54.39,
54.48, 55.32 (CH); 66.22 (CH2); 128.15, 128.30, 128.38, 128.51, 128.60 (CH); 135.94, 168.64, 169.33, 169.57,
‡
‡
‡
169.59, 169.61, 169.64, 169.73, 171.32 (C). FAB-MS: 839 (11.8, [M ‡ Na] ), 818 (45, [M ‡ 1] ), 817 (100, M ).
(4-NO2C6H4)CO (S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-(S)-b3-HPro-OBn
(14). Fully protected b3-hexapeptide 10a (0.197 g, 0.225 mmol) was Boc-deproteced according to GP 3. The
resulting TFA salt was dissolved in CH2Cl2 (1 ml) and treated at 08 with Et3N (0.28 ml, 0.675 mmol), p-
nitrobenzoyl chloride (50 mg, 0.270 mmol) and DMAP (2.8 mg, 0.023 mmol). The yellow suspension was stirred
for 13 h at r.t. After dilution with CH2Cl2, the mixture was washed with sat. aq. NH4Cl, NaHCO3, and NaCl
solns. The org. phase was dried (MgSO4) and evaporated under reduced pressure. FC (MeOH/CH2Cl2 1:12)
yielded 14 (166 mg, 80%). Yellow foam. M.p. 1488 (dec.). Rf (CH2Cl2/MeOH 12 :1) 0.34. [a]Dr:t:
ˆ
44.1 (c ˆ 1.0,
CHCl3). CD (0.2 mm, MeOH): 1.23 ´ 105 (204 nm), ‡ 6.06 ´ 104 (225 nm). IR (CHCl3): 3005m, 2878w, 1730w,
1632s, 1525m, 1426s, 1352m, 1045w. 1H-NMR (400 MHz, CDCl3): 1.73 ± 2.08 (m, 27 CH); 2.12 (dd, J ˆ 15.0,
10.5, COCH); 2.33 (dd, J ˆ 15.4, 9.4, COCH); 2.45 (dd, J ˆ 15.1, 9.4, COCH); 2.96 ± 3.03 (m, 5 COCH); 3.12
(dd, J ˆ 15.1, 3.1, COCH); 3.35 ± 3.67 (m, 12 NCH); 4.37 ± 4.56 (m, 6 NCH); 5.07 ± 5.14 (m, PhCH2); 7.28 ± 7.38
(m, 5 arom. H); 7.67 ± 7.72 (m, 2 arom. H); 8.25 ± 8.29 (m, 2 arom. H). 13C-NMR (100 MHz, CDCl3; values for
rotamers in italics): 21.54, 23.77, 23.79, 23.99, 24.92, 29.38, 29.69, 29.83, 29.90, 29.94, 30.40, 37.59, 38.01, 38.08,
38.14, 38.22, 38.29, 47.11, 47.14, 47.21, 47.26, 50.23 (CH2); 53.70, 54.02, 54.10, 54.18, 54.98 (CH); 66.22 (CH2);
123.63, 123.71, 128.15, 128.23, 128.30, 128.38, 128.51, 128.60 (CH); 135.94, 143.07, 148.53, 167.58, 169.09, 169.47,
‡
‡
‡
169.56, 169.61, 169.71, 171.33 (C). FAB-MS: 946 (17.4, [M ‡ Na] ), 925 (49.0, [M ‡ 1] ), 924 (100, M ).
Boc-(S)-b3-HPro-(R)-b3-HPro-(S)-b3-HPro-(R)-b3-HPro-(S)-b3-HPro-(R)-b3-HPro-OBn (15). Fully
protected b3-tetrapeptide 9 (1.89 g, 2.9 mmol) was Boc-deprotected according to GP 3. The resulting TFA
salt was coupled with 8b (0.98 g, 2.9 mmol) according to GP 4 for 3 d. FC (MeOH/CH2Cl2 1:15) yielded 15
(632 mg, 25%). White powder. M.p. 2468 (dec.). Rf (CH2Cl2/MeOH 17:1) 0.29. [a]Dr:t:
ˆ
1.68 (c ˆ 1.0, CHCl3).
CD (0.2 mm, MeOH): ‡ 1.06 ´ 104 (210 nm). IR (CHCl3): 3004m, 2976m, 2880w, 1730w, 1682m, 1631s, 1421s,
1366w, 1171m, 1123w, 1103w, 906w. 1H-NMR (500 MHz, CDCl3; values for rotamers in italics): 1.42, 1.46 (s, t-
Bu); 1.73 ± 2.19 (m, 30 CH); 2.39 (dd, J ˆ 15.3, 9.2, COCH); 2.75 ± 3.03 (m, 6 COCH); 3.36 ± 3.65 (m, COCH);
4.08 ± 4.18 (m, NCH); 4.29 ± 4.41 (m, 5 NCH); 5.07 ± 5.15 (m, PhCH2); 5.07 ± 5.14 (m, 5 arom. H). 13C-NMR
(125 MHz, CDCl3; values for rotamers in italics): 21.63, 21.66, 23.75, 23.83, 23.85, 23.89, 23.92, 24.01 (CH2);
28.60 (Me); 29.71, 29.75, 29.82, 30.14, 30.65, 30.69, 31.62, 37.08, 37.64, 37.69, 38.16, 38.22, 38.41, 38.46, 39.49,
46.55, 47.01, 47.11, 47.17, 47.22, 47.26 (CH2); 53.85, 53.97, 54.13, 54.20, 54.22, 54.51, 54.63 (CH); 66.22, 66.33
(CH2); 79.10, 79.56 (C); 128.18, 128.23, 128.26, 128.52, 128.56 (CH); 135.97, 154.39, 169.56, 171.33 (C). FAB-
‡
‡
‡
MS: 898 (14.2, [M ‡ Na] ), 877 (49.3, [M ‡ 1] ), 876 (100, M ). Anal. calc. for C48H70N6O9 (875.12): C 65.88,
H 8.06, N 9.60; found: C 65.62, H 8.01, N 9.58.
8. b2-Peptides. Ethyl (S)-Piperidine-3-carboxylate (S,S)-Hydrogen Tartrate ((S)-Ethyl Nipecotate (S,S)-
Hydrogen Tartrate; (S,S,S)-3). According to [36], rac-ethyl nipecotate (28.0 g, 0.178 mol) was resolved with

Helvetica Chimica Acta ± Vol. 82 (1999)
1555
(S,S)-tartaric acid (26.7 g, 0.178 mol) to yield (S,S,S)-3 (15.04 g, 27%) after recrystallization from EtOH (3Â).
M.p. 156 ± 1578 ([36]: M.p. 155 ± 1568). [a]Dr:t:
51.0 (c ˆ 2.0, (NH4)6Mo7O24 (0.2% aq. soln.))).
ˆ
46.3 (c ˆ 2.0, (NH4)6Mo7O24 (0.2% aq. soln.)) ([36]: [a]Dr:t:
ˆ
Ethyl (R)-Piperidine-3-carboxylate (R,R)-Hydrogen Tartrate ((R)-Ethyl Nipecotate (R,R)-Hydrogen
Tartrate; (R,R,R)-3). According to [36], rac-ethyl nipecotate (11.28 g, 71.6 mmol) was resolved with (R,R)-
tartaric acid (14.5 g, 67.0 mmol) to yield (R,R,R)-3 (5.44 g, 25%) after recrystallization from EtOH (3Â). M.p.
156 ± 1588 ([36]: M.p. 155 ± 1568). [a]Dr:t: ˆ‡ 52.3 (c ˆ 2.0, (NH4)6Mo7O24 (0.2% aq. soln.)) ([36]: [a]rD:t: ˆ‡ 51.0
(c ˆ 2.0, (NH4)6Mo7O24 (0.2% aq. soln.))).
Ethyl (S)-Piperidine-3-carboxylate ((S)-Ethyl Nipecotate; H-(S)-b2-HPro-OEt; (S)-4a). Similarly to [36],
(S,S,S)-3 (30 g, 97.6 mmol) was dissolved at 08 in sat. aq. NaCl soln. (50 ml). At this temp., the pH was carefully
adjusted to 13, and the aq. phase was extracted rapidly with Et2O (3Â). The Et2O phases were washed with H2O,
dried (MgSO4), and evaporated to yield (S)-4a (10.0 g, 65%). Yellowish oil. [a]rD:t: ˆ‡ 1.38 (c ˆ 5.0, H2O) ([36]:
[a]rD:t: ˆ‡ 1.6 (c ˆ 5.0, H2O)). The er was determined by derivatization according to GP 5 and subsequent HPLC
analysis according to GP 7: er 99.6 : 0.4.
Ethyl (R)-Piperidine-3-carboxylate ((R)-Ethyl Nipecotate; H-(R)-b2-HPro-OEt; (R)-4a). Similarly to
[36], (R,R,R)-3 (19.5 g, 64 mmol) was dissolved at 08 in sat. aq. NaCl soln. (30 ml). At this temp., the pH was
carefully adjusted to 13, and the aq. phase was extracted rapidly with Et2O (3Â). The Et2O phases were washed
with H2O, dried (MgSO4), and evaporated to yield (R)-4a (4.5 g, 45%). Yellowish oil. [a]Dr:t:
ˆ
1.26 (c ˆ 5.0,
H2O) ([36]: [a]Dr:t:
subsequent HPLC analysis according to GP 7: er 98.9 : 1.1.
ˆ
1.8 (c ˆ 5.0, H2O)). The er was determined by derivatization according to GP 5 and
Ethyl (S)-1-[(tert-Butoxy)carbonyl]piperidine-3-carboxylate (Boc-(S)-b2-HPro-OEt; (S)-4b). Compound
(S)-4a (6.5 g, 41.3 mmol, er 99.6 :0.4) and Boc2O (9.5 g, 43.5 mmol) were dissolved in CH2Cl2 (80 ml). After
stirring at r.t. for 16 h, the soln. was washed with sat. aq. NH4Cl and NaCl solns., dried (MgSO4), and evaporated
under reduced pressure. FC (Et2O/pentane 1:6 ! 1 : 1) yielded (S)-4b (7.9 g, 74%). Colorless oil. Rf (Et2O/
pentane 1 : 6) 0.29. [a]rD:t: ˆ‡ 50.7 (c ˆ 0.95, CHCl3). IR (CHCl3): 3008m, 2979m, 1725s, 1683s, 1476w, 1426s,
1393w, 1367m, 1170s, 1151s, 1043w, 928w, 880w, 856w. 1H-NMR (400 MHz, CDCl3): 1.26 (t, J ˆ 7.1, Me); 1.46 ±
1.55 (m, t-Bu, CH); 1.56 ± 1.74 (m, 3 CH); 2.01 ± 2.07 (m, CH); 2.39 ± 2.46 (m, CH); 2.80 (ddd, J ˆ 13.3, 11.3, 3.1,
NCH); 2.97 (br., NCH); 3.92 (br. d, J ˆ 12.9, NCH); 3.94 ± 4.35 (m, NCH); 4.13 (q, J ˆ 7.1, CH2O). 13C-NMR
(100 MHz, CDCl3): 14.20 (Me); 24.30, 24.37 (CH2); 28.43 (Me); 41.46 (CH); 44.00, 45.65, 60.50 (CH2); 79.67,
‡
‡
154.70, 173.49 (C). FAB-MS: 281 (< 1, [M ‡ Na] ), 257 (< 1, M ), 200 (18.2), 156 (38.1), 128 (38.1), 86 (51.8),
84 (100), 57 (27.1), 49 (44.3). Anal. calc. for C13H23NO4 (257.33): C 60.68, H 9.01, N 5.44; found: C 60.71,
H 8.98, N 5.45.
(S)-1-[(tert-Butoxy)carbonyl]piperidine-3-carboxylic Acid (Boc-(S)-b2-HPro-OH; (S)-4c). Compound
(S)-4b (6.2 g, 24.1 mmol) was saponified according to GP 1 with LiOH (1.44 g, 60.25 mmol) in MeOH (130 ml)
and H2O (43 ml) for 3 d at r.t. Recrystallization (Et2O/pentane) yielded (S)-4c (5.01 g, 90%). Derivatization
according to GP 6 and HPLC according to GP 7: er 97.9 :2.1. White powder. M.p. 165 ± 1678. Rf (MeOH/CH2Cl2
1:10) 0.29. [a]Dr:t: ˆ‡ 50.5 (c ˆ 1.0, CHCl3). IR (CHCl3): 2980w, 2865m, 1709s, 1684s, 1467w, 1426m, 1367m,
1269m, 1173m, 1150s, 1040w, 1003w, 936w, 873w, 858w. 1H-NMR (400 MHz, CDCl3): 1.41 ± 1.56 (m, t-Bu, CH);
1.60 ± 1.76 (m, 2 CH); 2.05 ± 2.10 (m, CH); 2.45 ± 2.53 (m, COCH); 2.83 ± 2.89 (m, NCH); 3.05 (br., NCH);
3.86 ± 3.91 (m, NCH); 4.12 (br., NCH); 7.27 (br., CO2H). 13C-NMR (100 MHz, CDCl3): 24.12, 27.18 (CH2);
‡
28.40 (Me); 41.07 (CH); 43.83, 45.50 (CH2); 79.92, 154.72, 178.88 (C). FAB-MS: 688 (9.6, [3 M ‡ 1] ), 459
‡
‡
(16.4, [2, M ‡ 1] ), 230 (15.1, [M ‡ 1] ), 174 (100), 156 (34.9), 154 (25.0), 136 (21.5), 128 (24.9). Anal. calc. for
C11H19NO4 (229.28): C 57.63, H 8.35, N 6.11; found: C 57.50, H 8.16, N 5.97.
Ethyl rac-N-(2,4-Dinitrophenyl)piperidine-3-carboxylate (rac-5). To a soln. of rac-ethyl nipecotate (0.671 g,
4.27 mmol) and NaHCO3 (0.43 g, 5.12 mmol) in H2O (8.5 ml) was added a soln. of 1-fluoro-2,4-dinitrobenzene
(0.95 g, 5.12 mmol) in EtOH (16 ml) at 08, similarly to [50]. After 5 h, the soln. was diluted with Et2O and
washed with 1n HCl (1Â) and sat. NH4Cl soln. (3Â), dried (MgSO4), and evaporated. FC (Et2O/pentane 1 : 1)
yielded rac-5 (1.123 g, 81%). Orange sirup. Rf (Et2O/pentane 1 : 1) 0.31. IR (CHCl3): 3091w, 2961w, 2864w,
1726m, 1606s, 1530s, 1447w, 1336s, 1262m, 1178m, 1150w, 1097m, 1067w, 1030m, 965w, 943w, 916w, 858w.
1H-NMR (400 MHz, CDCl3): 1.25 (t, J ˆ 7.1, Me); 1.71 ± 1.91 (m, 3 CH); 2.10 ± 2.17 (m, 1 CH); 2.72 ± 2.78
(m, COCH); 3.07 ± 3.13 (m, NCH); 3.30 ± 3.39 (m, 2 NCH); 3.57 ± 3.62 (m, NCH); 4.15 (q, J ˆ 7.1, CH2O); 7.19
(d, J ˆ 9.4, 1 arom. H); 8.25 (dd, J ˆ 9.3, 2.7, 1 arom. H); 8.70 (d, J ˆ 2.7, 1 arom. H). 13C-NMR (100 MHz,
CDCl3): 14.16 (Me); 23.93, 26.34 (CH2); 40.95 (CH); 51.50, 52.52, 60.99 (CH2); 119.95, 123.76, 128.17 (CH);
‡
138.17, 138.30, 149.74, 172.61 (C). FAB-MS: 323 (7.1, M ), 306 (100), 278 (29.7), 260 (25.5), 232 (59.9), 216
(45), 203 (67.7), 180 (45.2), 157 (29.4). Anal. calc. for C14H17N3O6 (323.30): C 52.01, H 5.30, N 13.00; found:
C 52.27, H 5.52, N 12.85.

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Helvetica Chimica Acta ± Vol. 82 (1999)
Ethyl (R)-N-(2,4-Dinitrophenyl)piperidine-3-carboxylate ((R)-5). Prepared according to GP 5. HPLC
according to GP 7: tR 20.5. Purification by FC (Et2O/pentane 1:1). [a]Dr:t:
spectroscopic data: corresponding to rac-5.
ˆ
165.0 (c ˆ 0.6, CHCl3). Other
Ethyl (S)-N-(2,4-Dinitrophenyl)piperidine-3-carboxylate ((S)-5). Prepared according to GP 5. HPLC
according to GP 7: tR 25.5. Purification by FC (Et2O/pentane 1:1). [a]rD:t: ˆ‡ 164.8 (c ˆ 0.6, CHCl3). Other
spectroscopic data: corresponding to rac-5.
Boc-(S)-b2-HPro-(S)-b2-HPro-OEt (16). Compound (S)-4a (1.37 g, 8.72 mmol) was coupled with (S)-4c
(2 g, 8.72 mmol) for 16 h according to GP 4. FC (AcOEt/pentane 1:2) yielded 16 (2.8 g, 87%). For anal.
purposes, 16 was dried under h.v. at 358 overnight. Yellowish waxy solid. M.p. 71 ± 728. Rf (AcOEt/pentane 1:2)
0.25. [a]rD:t: ˆ‡ 61.1 (c ˆ 0.365, CHCl3). IR (CHCl3): 3007w, 2945w, 2866w, 1726m, 1681s, 1628s, 1468w, 1444m,
1
1425m, 1367w, 1306w, 1265m, 1177m, 1150s, 1031w, 856w. H-NMR (400 MHz, CDCl3, rotamers!): 1.23 ± 1.30
(m, Me); 1.46 ± 1.51 (m, t-Bu, CH); 1.62 ± 1.88 (m, 6 CH); 2.05 ± 2.08 (m, CH); 2.43 ± 3.12 (m, 4 NCH,
2 COCH); 3.39 (br., 0.5 H, NCH); 3.73 ± 4.21 (m, 5.5 H, NCH, CH2O); 4.59 (br. d, J ˆ 10.3, 0.5 H, NCH,
rotamer). 13C-NMR (100 MHz, CDCl3, rotamers!): 14.19 (Me); 23.98, 24.71, 25.41, 27.35, 27.46, 27.73 (CH2);
28.49 (Me); 38.97, 41.27 (CH); 42.02, 43.65, 45.75, 47.19, 60.62, 60.90 (CH2); 72.51 (CH); 79.63, 154.66, 171.67,
‡
171.99, 172.70, 173.19 (C). FAB-MS: 369 (24.2, [M ‡ 1] ), 327 (18.8), 313 (100), 295 (78.8), 269 (63.9), 267
(64.1), 156 (70.1), 154 (49.6), 147 (76.1), 136 (96.1). Anal. calc. for C19H32N2O5 (368.47): C 61.93, H 8.75,
N 7.60; found: C 61.89, H 8.74, N 7.53.
Boc-(S)-b2-HPro-(S)-b2-HPro-(S)-b2-HPro-OEt (17a). Compound 16 (2.53 g, 6.86 mmol) was Boc-
deprotected according to GP 3. The resulting TFA salt was coupled with (S)-4c (1.57 g, 6.86 mmol) for 16 h
according to GP 4. FC (MeOH/CH2Cl2 1:15) yielded 17a (2.31 g, 70%). White, waxy solid. M.p. 808 (sintering at
50 ± 558). Rf (MeOH/CH2Cl2 1:15) 0.33. [a]Dr:t: ˆ‡ 67.8 (c ˆ 0.515, CHCl3). CD (0.2 mm in MeOH): 2.09 ´ 104
(211 nm), ‡ 4.15 ´ 103 (230 nm). IR (CHCl3): 3008m, 2943w, 2865w, 1726m, 1675m, 1631s, 1443m, 1367w,
1150m, 855w. 1H-NMR (500 MHz, CDCl3, rotamers!): 1.23 ± 1.30 (m, Me); 1.44 ± 2.17 (m, t-Bu, 12 CH); 2.40 ±
3.14 (m, 3 COCH, 6 NCH); 3.33 ± 4.21 (m, CH2O, 5 NCH); 4.49 ± 5.30 (m, NCH). 13C-NMR (125 MHz, CDCl3,
rotamers!): 14.19, 23.54, 24.00, 24.65, 25.31, 26.06, 27.05, 27.19, 27.44, 27.63, 27.76, 28.20, 28.46, 28.50, 28.61,
31.44, 36.48, 38.55, 39.04, 39.73, 41.06, 41.27, 42.01, 42.05, 42.50, 43.69, 44.49, 44.79, 45.86, 46.07, 46.78, 47.31,
48.06, 60.59, 60.70, 60.91, 60.59, 79.66, 154.63, 162.52, 171.26, 171.60, 171.69, 171.84, 172.03, 172.68, 172.95, 173.21
‡
‡
(C). FAB-MS: 502 (2.0, [M ‡ Na] ), 480 (62.2, M ), 380 (100), 269 (30.0).
Boc-(S)-b2-HPro-(S)-b2-HPro(S)-b2-HPro-OH (17b). Compound 17a (1.22 g, 2.54 mmol) was saponified
with LiOH (0.15 g, 6.34 mmol) in MeOH (21 ml) and H2O (7 ml) according to GP 1. Recrystallization (AcOEt/
pentane) yielded 17b (0.574 g, 50%). White powder. M.p. 175 ± 1778. Rf (MeOH/CH2Cl2 1:10) 0.25. [a]Dr:t:
ˆ
‡ 52.6 (c ˆ 0.50, CHCl3). IR (CHCl3): 3006w, 2944w, 2863w, 1719w, 1680m, 1625s, 1444m, 1368w, 1152m,
1
855w. H-NMR (500 MHz, CDCl3, rotamers!): 1.44 ± 2.17 (m, t-Bu, 9 H); 2.46 ± 4.21 (m, 3 COCH, 11 NCH);
4.49 ± 4.63 (m, NCH); 5.96 (br. s, CO2H). 13C-NMR (125 MHz, CDCl3): not shown because of rotamers. FAB-
‡
‡
MS: 474 (18.5, [M ‡ Na] ), 452 (100, [M ‡ 1] ), 352 (71.2), 241 (44.7). Anal. calc. for C23H37N3O6 (451.56):
C 61.18, H 8.26, N 9.31; found: C 60.99, H 8.17, N 9.13.
Boc-(S)-b2-HPro-(S)-b2-HPro-(S)-b2-HPro-(S)-b2-HPro-(S)-b2-HPro-(S)-b2-HPro-OEt (18). Com-
pound 17a (0.63 g, 1.31 mmol) was Boc-deprotected according to GP 3. The resulting TFA salt was coupled
with 17b (0.474 g, 1.048 mmol) for 16 h according to GP 4. FC (CH2Cl2/MeOH 9 :1) yielded 18 (681 mg, 79%).
Colorless glass. M.p. 1168 (sintering at 1048). Rf (CH2Cl2/MeOH 9 :1) 0.35. [a]rD:t: ˆ‡ 96.3 (c ˆ 0.325, CHCl3).
CD (0.2 mm in MeOH): 8.54 ´ 104 (208 nm), ‡ 2.44 ´ 104 (228 nm). IR (CHCl3): 3007m, 2946w, 2860w, 1726w,
1682m, 1631s, 1442m, 1367w, 1149m, 856w. 1H-NMR (500 MHz, CDCl3, rotamers!): 1.23 ± 1.29 (m, Me); 1.39 ±
2.17 (m, t-Bu, 24 CH); 2.40 ± 4.19 (m, 6 COCH, 20 NCH, CH2O); 4.60 (br., 4 NCH). FAB-MS: 1649 (12.8, [2
‡
‡
‡
‡
M ‡ Na] ), 1627 (19.4, [2M] ), 835 (15.9, [M ‡ Na] ), 813 (37.9, [M ‡ 1] ), 713 (100), 306 (26.1), 195 (34.7).
9. X-Ray Crystal-Structure Analysis. Compound 7c (C25H35N9O4 ´ 4 CF3CO2H). Crystals were grown from a
supersaturated TFA soln. at 208. Crystal size 0.30 Â 0.20 Â 0.20 mm. Crystal data at 243 K. Triclinic, space
group P1, 1calc. ˆ 1.427 g cm 3, Z ˆ 1, a ˆ 8.632(2) Š, b ˆ 11.739(2) Š, c ˆ 11.775(2) Š, a ˆ 61.53(3)8, b ˆ
85.08(3)8, g ˆ 88.90(3)8, Vˆ 1044.6(3) Š3. Nonius CAD4 diffractometer, CuKa radiation, l ˆ 1.54178 Š, 3208
unique reflections measured in the range 4.29 < q < 60.018. The structure was solved by direct methods
(SHELXS-86 [65]), and refined by full-matrix least-squares analysis (SHELXL-97 [66]), using an isotropic
2
extinction correction and w ˆ 1/[s2(F02) ‡ (0.2635P)2 ‡ 0.6910P], where P ˆ (F0 ‡ 2Fc2)/3 (heavy atoms
anisotropic, H-atoms isotropic, whereby H-positions are based on stereochemical considerations). Final
R(F) ˆ 0.1036, wR(F2) ˆ 0.2701 for 511 variables and 3208 observations. Crystallographic data (excluding
structure factors) for the structure reported in this paper have been deposited with the Cambridge
Crystallographic Data Centre as deposition No. CCDC-129015 (7c). Copies of the data can be obtained, free

Helvetica Chimica Acta ± Vol. 82 (1999)
1557
of charge, on application to the CCDC, 12 Union Road, Cambridge CB2 1EZ UK (fax: ‡ 44 (1223) 336 033;
e-mail: deposit@ccdc.cam.ac.uk).
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Received June 24, 1999