Synthesis of novel nucleo-β-amino acids and nucleobase-functionalized β-peptides

Article abstract of DOI:10.1002/ejoc.200300269

Four novel β-amino acids bearing the canonical nucleobases guanine, cytosine, adenine, and thymine in the side chain, are synthesized starting from Boc-L-aspartic acid 4-benzyl ester. The syntheses are accomplished in six steps by the nucleophilic substitution of (S)-β -(tert-butoxycarbonylamino)-δ-bromopentanoic acid benzyl ester with the corresponding nucleobase derivative as the key step. The guaninyl and cytosinyl β-amino acids were built into β-peptides that were studied by temperature-dependent CD and UV spectroscopy. Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003.

Full text of DOI:10.1002/ejoc.200300269

FULL PAPER
Synthesis of Novel Nucleo-β-Amino Acids and Nucleobase-Functionalized
β-Peptides
Arndt M. Brückner,[a] Margarita Garcia,[b] Andrew Marsh,[b] Samuel H. Gellman,[c] and
Ulf Diederichsen*[a]
Keywords: Amino acids / Peptides / Foldamers / Nucleobases / Peptide nucleic acids
Four novel β-amino acids bearing the canonical nucleobases
guanine, cytosine, adenine, and thymine in the side chain,
are synthesized starting from Boc-L-aspartic acid 4-benzyl
nucleobase derivative as the key step. The guaninyl and
cytosinyl β-amino acids were built into β-peptides that were
studied by temperature-dependent CD and UV spectroscopy.
ester. The syntheses are accomplished in six steps by the nu-
cleophilic substitution of (S)-β-(tert-butoxycarbonylamino)-δ-
bromopentanoic acid benzyl ester with the corresponding
( Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim,
Germany, 2003)
Introduction
homoalanyl peptide nucleic acids (β-homoalanyl PNA)
form stable pairing complexes with a β-sheet-like backbone
conformation in aqueous media.[9]
There is a growing interest in β-amino acids as building
blocks for natural products and pharmaceutical agents.[1]
Furthermore, oligomers composed of β-amino acids (β-
peptides) are an attractive class of compounds for drug de-
sign: oligomers (‘‘foldamers’’) with low molecular weights
fold into well-ordered secondary structures (helix, sheet,
and turn)[2] and show a remarkable stability towards enzy-
matic degradation.[3] The proper choice of β-amino acid
side-chains leads to β-peptides with interesting biological
properties.[4] Side-chain functionalities can be introduced,
for example, to fine-tune solubility[5] and lipophilicity[6] or
to promote secondary structure stability.[7]
Figure 1. Structures of nucleo-β3-amino acids 1 and 2
β-Peptides with proteinogenic and cyclic nonproteinog-
enic side chains usually fold into helical or sheet-like sec-
ondary structures. The incorporation of nucleobases into
the side chains enables β-peptide structures to be influenced
through base pairing and aromatic π-interactions. Recently,
oligomers have been reported that are composed of six nu-
cleo-β3-amino acid building blocks,[8] like β-amino acid 1,
in which the nucleobases are connected to β-homoalanine
at the γ-position (Figure 1).[9] It was shown that these β-
Here, we report the first step towards an analogous or-
ganization of β-peptide helices through nucleobase pairing.
The syntheses of novel protected nucleo-β3-amino acids like
2 with an ethylene linker between the nucleobase and the
β-amino acid backbone is described. The length of the
linker was chosen to minimize the nucleobase-peptide back-
bone interactions and to allow specific nucleobase interac-
tions. Finally, the β-amino acid monomers were incorpor-
ated into β-peptides by solid-phase peptide synthesis, and
first structural investigations were performed with these oli-
gomers.
[a]
Institut für Organische Chemie, Georg-August-Universität
Göttingen
Tammannstraße 2, 37077 Göttingen, Germany
Fax: (internat.) ϩ49-(0)551-392944
E-mail: udieder@gwdg.de
[b]
Department of Chemistry, University of Warwick
Coventry, CV4 7AL, UK
Results and Discussion
Fax: (internat.) ϩ44-024-76524112
E-mail: a.marsh@warwick.ac.uk
[c]
Department of Chemistry, University of Wisconsin
Several methods for the stereoselective preparation of β-
amino acids have been published.[10] However, many meth-
ods are not transferable to the preparation of nucleo-β-am-
1101 University Avenue, Madison, Wisconsin 53706, USA
Fax: (internat.) ϩ01-608-2654534
E-mail: gellman@chem.wisc.edu
Eur. J. Org. Chem. 2003, 3555Ϫ3561
DOI: 10.1002/ejoc.200300269
 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
3555

A. M. Brückner, M. Garcia, A. Marsh, S. H. Gellman, U. Diederichsen
FULL PAPER
ino acids because of the competition of N9/N7 purine or N1/ alkylation and improves the solubility. Z-protected cytosine
N3 pyrimidine regioisomer formation, the low nucleophilic- and adenine[15b] were used to prevent chain extension at the
ity of the nucleobases and their poor solubility in common exocyclic amino groups during later β-peptide synthesis. 2-
organic solvents. In addition, the ArndtϪEistert homol- Amino-6-chloropurine, a synthon for guanine,[16] gave al-
ogation of aromatic α-amino acids is expected to proceed most exclusively the N9-alkylated regioisomer. It is note-
with racemization.[11]
worthy that all alkylations with bromide 3 proceeded much
Hence, we decided to link the nucleobase or its precursor more cleanly than those carried out for the preparation of
to the desired β-amino acid moiety. The linkage of the nu- nucleo-β3-amino acids of type 1.[9] BocHNCH-
cleobases to the amino acid side-chain is usually ac- (CH2CH2Br)CH2CO2Bzl (3) with a two-carbon linker is
complished by nucleophilic substitution of a halide or alkyl- quite stable under the conditions of nucleophilic substi-
sulfonate or — under milder conditions — by employing a tution, whereas BocHNCH(CH2X)CH2CO2Bzl (X ϭ Br,
Mitsunobu reaction. Since the latter strategy often requires OH, OMs) with a shorter side chain showed a tendency
laborious chromatography bromide 3 was used as the key to decompose by formation of aziridines or 2-oxazolidones
intermediate for the alkylation of all nucleobase precursors. under the conditions of Mitsunobu-type reactions (X ϭ
This compound was synthesized from commercially avail- OH) or nucleophilic substitutions (X ϭ Br, OMs), respec-
able Boc--aspartic acid 4-benzyl ester (4; Scheme 1). First, tively.[17]
the amino-acid backbone was extended by one carbon atom
through an ArndtϪEistert homologation. This involved the
preparation of the diazo ketone 5 as described by Seebach
et al.[12] and its Wolff rearrangement, according to a litera-
ture procedure,[13] to give carboxylic acid 6. Reduction with
borane in THF yielded alcohol 7, which was then converted
into the desired bromide 3 by treatment with PPh3/CBr4.
Bromide 3 was reacted with 1.5Ϫ3.3 equivalents of the nu-
cleobase or its precursor at room temperature in DMF in
the presence of K2CO3 and tetrabutylammonium iodide
(TBAI) to give the products 8Ϫ11 in 44Ϫ92% yield
(Schemes 2 and 3). The yields correspond with the range
of solubility and reactivity of the nucleobase moieties. For
example, 3-benzoyl-protected thymine[14] is much more sol-
uble in polar aprotic solvents and possesses a higher reactiv-
ity towards alkylation than the moderately soluble N4-
(benzyloxycarbonyl)cytosine [cytosine(Z)].[15] The benzoyl
protecting group in the N3-position of thymine excludes N3-
Scheme 2. Synthesis of the pyrimidinyl-nucleo-β3-amino acids 12
and 14 from bromide 3
Scheme 1. Preparation of bromide 3 starting from Boc--aspartic
acid 4-benzyl ester (4); for preparation of diazo ketone 5 see ref.[12]
Scheme 3. Synthesis of the purinyl-nucleo-β3-amino acids 13 and 2
from bromide 3
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Synthesis of Novel Nucleo-β-Amino Acids and Nucleobase-Functionalized β-Peptides
FULL PAPER
Figure 2. Complementary oligomers 15 and 16 prepared by solid-phase peptide synthesis (SPPS)
Finally, the free nucleo-β3-amino acids 12 and 13 with oxide hexafluorophosphate} and N,N-diisopropylethylam-
cytosine(Z) and adenine(Z) in the side chains were obtained ine for activation. Because of the sterically demanding
by saponification of the benzyl esters 8 and 9 with aqueous ACHC and the expected limited accessibility of the ter-
NaOH. The thyminyl-β3-amino acid benzyl ester 10 was sa- minal amino group in a compact helix each coupling step
ponified by catalytic hydrogenation on Pd/C, and the N3- was performed twice at 50 °C to ensure chain elongation
atom was deprotected by treatment with N2H4 in ethanol. yields of at least 97%. After cleavage from the solid support
The guaninyl-β3-amino acid 2 was afforded in excellent with trifluoroacetic acid (TFA)/trifluoromethanesulfonic
yield by treating 11 with refluxing HCl, followed by protec- acid/m-cresol/thioanisole/ethanedithiol, 20:2:2:2:1, the olig-
tion with Boc2O.
The oligomers H-(β-HLys-ApaC-ACHC-β-HLys-ApaG- HPLC, and characterized with electrospray ionization mass
ACHC-β-HLys-ApaC-ACHC-β-HGly)-NH2 (15) and spectrometry (ESI-MS).
H-(β-HLys-ApaG-ACHC-β-HLys-ApaC-ACHC-β-HLys- Temperature-dependent circular dichroism (CD) and UV
omers were precipitated with diethyl ether, purified by
ApaG-ACHC-β-HGly)-NH2 (16, Figure 2) were designed spectroscopy were used for initial studies of the structural
in order to explore self-assembly of β-peptides containing behavior of oligomers 15 and 16. A positive Cotton effect
nucleo-β3-amino acid residues.[18] (R)-β-Amino-δ-(guanin- around 215 nm in the CD spectra of β-peptides has often
9-yl)pentanoic acid (ApaG) and (R)-β-amino-δ-(cytosin-1- been correlated with the formation of a right-handed 14-
yl)pentanoic acid (ApaC) were chosen as base-pair recog- helix.[2,5Ϫ7,13,20] The CD spectra of β-peptides 15 and 16
nition units because they should prefer the formation of (data not shown) and of an equimolar mixture of both (Fig-
very stable triply hydrogen bonded guanine-cytosine ure 3) in 10 m aq. Na2HPO4/H3PO4 (pH, 7.0) show a
WatsonϪCrick base pairs, whereas a doubly hydrogen maximum between 210 and 215 nm, which indicates the ex-
bonded adenine-thymine base pair is expected to lead to pected formation of a 14-helix. The Cotton effect at about
less-stable pairing complexes. ApaG and ApaC were incor- 270 nm is probably due to a preferred conformational
porated into a β-peptide scaffold comprised of (1R,2R)-2- orientation of the nucleobases. Potential pairing of olig-
aminocyclohexanecarboxylic acid (ACHC) and β-(R)- omer 15 with the complementary strand 16 was studied by
homolysine (H-β-HLys-OH).[19] β-Peptides built from
ACHC and β-HLys have been shown to adopt a 14-helical
conformation in aqueous solution.[5a] The 14-helix has ap-
proximately three residues per turn; therefore, the spacing
among nucleo-β3-amino acid residues in oligomers 15 and
16 should lead to alignment of the bases along one side of
the 14-helical conformations adopted by these oligomers.
Oligomers 15 and 16 were prepared by manual solid-
phase peptide synthesis on a 4-methylbenzhydrylamine
(MBHA) polystyrene resin loaded with β-homoglycine (H-
β-HGly-OH). The Boc-strategy was chosen to prevent ag-
gregation and deprotection problems that might result from
secondary structure formation during the synthesis of
longer oligomers, especially with Fmoc protection. All am-
ino acids were coupled in DMF with HATU {1-[bis(dime-
thylamino)methyliumyl]-1H-1,2,3-triazolo[4,5-b]pyridin-3-
Figure 3. CD spectra of an equimolar mixture of oligomers 15 and
16 (10 µ each, 10 m aq. Na2HPO4/H3PO4, pH 7.0)
Eur. J. Org. Chem. 2003, 3555Ϫ3561
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A. M. Brückner, M. Garcia, A. Marsh, S. H. Gellman, U. Diederichsen
FULL PAPER
77.0 ppm), [D6]DMSO (1H: δ ϭ 2.49 ppm, 13C: δ ϭ 39.5 ppm), or
D2O (1H: δ ϭ 4.77 ppm). The 13C NMR spectra with D2O as sol-
vent is referenced to MeOH as an internal standard (δ ϭ
49.9 ppm). Multiplicities of 13C NMR peaks were determined with
the APT pulse sequence. Mass spectra were recorded with a LCG
Finnigan spectrometer or with a Micromass LCT spectrometer.
High-resolution mass spectra were recorded with a Varian MAT
731 instrument. Elemental analysis was carried out on a Leco
CHN2000 with Heraeus Micro U/D. HPLC of the oligomers was
performed on a Pharmacia Äkta basic system with YMC J’sphere
temperature-dependent UV spectroscopy. As known from
DNA chemistry the cooperative destacking of nucleobases
due to separation of double strands can be observed as a
sigmoidal increase of the absorption.[21] The self-pairing
and the 1:1 pairing of β-peptides 15 and 16 were examined.
In all three cases, the stabilities were too low (15: Tm Ͻ 5
°C, Arel. ϭ 3%, 26 µ; 16: Tm Ͻ 5 °C, Arel. 12%, 23 µ,
data not shown; 15 ϩ16: Tm Ͻ 5 °C, Arel. 4%, 10 µ each,
Figure 4) to allow us to draw conclusions regarding pairing
behavior. Preparation of longer oligomers is in progress.
˚
ODS-H80, RP-C18, 150 ϫ 10 mm, 4 µm, 80 A for preparation and
˚
150 ϫ 4.6 mm, 4 µm, 80 A for analytical samples. The oligomer
concentration was calculated by taking the extinction coefficient at
90 °C as being the sum of the extinction coefficients of the nucleo-
β3-amino acids. The CD spectra were recorded with JASCO J-500A
Spectropolarimeter with a JASCO ETC-505S/PTC-423S tempera-
ture controller. The UV melting curves were measured with a JA-
SCO V-550 UV/Vis Spectrophotometer with a JASCO ETC-505S/
ETC-505T temperature controller.
(R)-β-(tert-Butoxycarbonylamino)-δ-(guanin-9-yl)pentanoic Acid (2):
Benzyl ester 11 (1.83 g, 3.85 mmol) was suspended in 1  aq. HCl
(200 mL). The mixture was heated to reflux for 6 h and then evapo-
rated to dryness under reduced pressure. An analytical sample of
the residue (freeze dried from water) contained no organic impurity
in addition to the HCl salt of (R)-β-amino-δ-(guanin-9-yl)pen-
tanoic acid: 1H NMR (300 MHz, D2O, 35 °C): δ ϭ 2.38 [dt,
Figure 4. UV melting curve of an equimolar mixture of oligomers
15 and 16 (10 µ each, 10 mm aq. Na2HPO4/H3PO4, pH 7.0,
270 nm)
2
3
3JH,H ϭ 7 Hz, 7 Hz, 2 H, C(γ)-H], 2.85 [dd, JH,H ϭ 18, JH,H
ϭ
8 Hz, 1 H, C(α)-H], 3.00 [dd, 2JH,H ϭ 18, 3JH,H ϭ 5 Hz, 1 H, C(α)-
H], 3.73 [m, 1 H, C(β)-H], 4.44 [t, 3JH,H ϭ 7 Hz, 2 H, C(δ)-H], 8.93
[s, 1 H, C(8)-H] ppm. 13C NMR (75 MHz, D2O, 35 °C): δ ϭ 32.7
[C(γ)], 36.6 [C(δ)], 42.4 [C(α)], 46.7 [C(β)], 109.0 [C(5)], 138.4
[C(8)], 151.1, 156.0, 156.3, 174.4 (CO2H) ppm. MS (ESI): m/z
(%) ϭ 267.5 (54) [M ϩ H]ϩ, 555.5 (100) [2 M ϩ Na]ϩ. Di-tert-
butyl dicarbonate (1.68 g, 7.70 mmol) was added to a suspension
of this residue in water/1  aq. NaOH/dioxane (1:1:1, 100 mL).
Conclusion
The synthesis of novel nucleo-β3-amino acids with all
four canonical nucleobases linked by an ethylene spacer to
the β-amino acid backbone has been developed. These com- After stirring for 7 h at room temperature a second portion of di-
tert-butyl dicarbonate (1.68 g, 7.70 mmol) was added and stirring
continued overnight. The resultant solution was washed with di-
ethyl ether, neutralized with 1  aq. HCl, and concentrated in va-
cuo. Purification of the residue by chromatography (RP-C18 silica
gel, 100% H2O to H2O/MeOH, 4:1 gradient) provided, after freeze
drying, 2 (1.35 g, 96%) as a colorless solid. M.p. Ͼ 300 °C (dec.).
pounds should serve as versatile building blocks for PNA
analogs, templates for 5Ј-methacryloylnucleoside polymeris-
ation,[22] and biologically active agents. We incorporated
these nucleo-β3-amino acids into water-soluble β-peptides.
Additional investigation of the criteria for specific associ-
ation of β-peptide helices via nucleobase pairing is un-
derway.
Rf ϭ 0.30 (ethyl acetate/MeOH/H2O/acetic acid, 80:14:6:3). [α]2D0
ϭ
ϩ6.5 (c ϭ 0.92, DMSO). IR (KBr): ν˜ ϭ 3410, 3134, 2978, 1696,
1574, 1482, 1401, 1367, 1169, 1059, 781, 694 cmϪ1 1H NMR
.
(300 MHz, [D6]DMSO, 35 °C): δ ϭ 1.37 (s, 9 H, tBu), 1.80Ϫ1.95
[m, 2 H, C(γ)-H], 2.06Ϫ2.22 [m, 2 H, C(α)-H], 3.67Ϫ3.74 [m, 1 H,
C(β)-H], 3.83Ϫ3.95 [m, 2 H, C(δ)-H], 7.06 (3 H, NH2, NH), 7.63
[s, 1 H, C(8)-H] 12.02 (s, br., 1 H, CO2H) ppm. 13C NMR (75 MHz,
[D6]DMSO, 35 °C): δ ϭ 28.2 [C(CH3)3], 35.1 [C(γ)], C(δ) overlap-
ping with residual solvent signal as determined by HMQC, 42.5
[C(α)], 46.0 [C(β)], 77.5 [C(CH3)3], 116.4 [C(5)], 136.9 [C(8)], 151.2,
154.2, 155.0, 157.6, 175.4 (CO2H) ppm. MS (ESI): m/z (%) ϭ 389.5
(100) [M ϩ Na]ϩ, 755.5 (86) [2 M ϩ Na]ϩ.
Experimental Section
General Remarks: All reagents were of analytical grade and used
as supplied. For preparation of Boc-(R,R)-ACHC-OH see ref.[23]
Boc-β-HLys(Z)-OH was prepared from Boc-(R)-Lys(Z)-OH by
ArndtϪEistert homologation.[11,13,24] Boc--β-homoaspartic acid
5-benzyl ester (6) is available from Fluka. Solvents were of the high-
est grade available. Melting points were obtained with a Büchi 501
Dr. Tottoli apparatus or a Gallenkamp melting point apparatus
and are uncorrected. Optical rotations were measured on a
PerkinϪElmer 241 polarimeter. IR spectra were recorded on a
PerkinϪElmer 1600 Series FT-IR using KBr pellets or on a Avatar
Benzyl (S)-β-(tert-Butoxycarbonylamino)-δ-bromopentanoate (3): A
solution of alcohol 7 (4.30 g, 13.3 mmol) and CBr4 (8.83 g,
26.6 mmol) in dry CH2Cl2 (110 mL) was cooled under nitrogen to
320 FT-IR with a ЈGolden GateЈ Attenuated Total Reflection Ϫ20 °C and a solution of PPh3 (7.00 g, 26.7 mmol) in dry CH2Cl2
(ATR) cell attachment. NMR spectra were recorded with Bruker (22 mL) was added dropwise over 30 min. The mixture was allowed
AMX 300, Bruker AC 300, Bruker DPX 300, Varian Unity 300, or to warm to room temperature and stirring continued for 15 h. The
Varian Inova 500 instruments. Chemical shifts are referenced to
solvent was removed under reduced pressure and the residue was
purified by chromatography (silica gel, ethyl acetate/n-hexane, 1:4)
the residual solvent peaks of CDCl3 (1H: δ ϭ 7.24 ppm, 13C: δ ϭ
3558
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Synthesis of Novel Nucleo-β-Amino Acids and Nucleobase-Functionalized β-Peptides
FULL PAPER
to afford 3 (4.05 g, 79%) as a colorless solid. M.p. 81Ϫ83 °C. Rf ϭ
0.19 (n-hexane/ethyl acetate, 4:1). IR (solid): ν˜ ϭ 3365, 2977, 1727,
1678, 1514, 1440, 1274, 1247, 1219, 1161, 1123, 1024, 732, 683
cmϪ1. 1H NMR (300 MHz, CDCl3, 25 °C): δ ϭ 1.43 (s, 9 H, tBu),
1.97Ϫ2.07 [m, 1 H, C(γ)-H], 2.07Ϫ2.24 [m, 1 H, C(γ)-H], 2.64 [d,
171.2 (CH2CO2Bzl) ppm. MS (ESI): m/z (%) ϭ 573.4 (82) [M ϩ
Na]ϩ, 1123.3 (100) [2 M ϩ Na]ϩ, 1673.4 (13) [3 M ϩ Na]ϩ.
C29H34N4O7 (550.2): calcd. 550.2428; found 550.2427 (HRMS).
Benzyl (R)-β-(tert-Butoxycarbonylamino)-δ-[N6-(benzyloxycarbonyl)-
adenin-9-yl]pentanoate (9):
A mixture of bromide 3 (2.56 g,
3
3JH,H ϭ 5 Hz, 2 H, C(α)-H], 3.40 [t, JH,H ϭ 7 Hz, 2 H, C(δ)-H],
6.63 mmol), (N6-benzyloxycarbonyl)adenine (5.00 g, 18.6 mmol),
K2CO3 (2.57 g, 18.6 mmol), and TBAI (254 mg, 688 µmol) in dry
DMF (82 mL) was stirred under nitrogen at room temperature for
46 h. Then, the solvent was removed under reduced pressure, water
(200 mL) was added, and the crude compound was extracted with
ethyl acetate (6 ϫ 100 mL). The combined organic phases were
dried (MgSO4) and the solvent was removed under reduced press-
ure. Purification by column chromatography (silica gel, ethyl acet-
ate) afforded 9 (2.13 g, 56%) as a colorless solid. M.p. 75Ϫ79 °C.
Rf ϭ 0.59 (ethyl acetate/methanol, 9:1). IR (solid): ν˜ ϭ 3340, 2979,
1740, 1698, 1616, 1581, 1455, 1243, 1155, 987, 734 cmϪ1. 1H NMR
(300 MHz, CDCl3, 25 °C): δ ϭ 1.44 (s, 9 H, tBu), 2.03Ϫ2.12 [m, 2
H, C(γ)-H], 2.59 [m, 2 H, C(α)-H], 3.87Ϫ4.00 [m, 1 H, C(β)-H],
4.10Ϫ4.19 [m, 1 H, C(δ)-H], 4.24Ϫ4.33 [m, 1 H, C(δ)-H], 5.08 (s,
2 H, CH2Ph), 5.28 (s, 2 H, CH2Ph), 5.31 (s, br., 1 H, NHBoc),
7.26Ϫ7.40 (m, 10 H, 2 Ph), 8.03 [s, br., 1 H, C(2)-H, C(8)-H], 8.74
[s, 1 H, C(2)-H, C(8)-H], 9.30 (s, br., 1 H, NHCO2Bzl) ppm. 13C
NMR (75 MHz, CDCl3, 25 °C): δ ϭ 28.1 [C(CH3)3], 34.3 [C(γ)],
38.9 [C(α)], 41.1 [C(δ)], 44.8 [C(β)], 66.4 (CH2Ph), 67.4 (CH2Ph),
79.7 [C(CH3)3], 121.8 [C(5)], 128.1Ϫ128.4 (Ph), 135.1 (Cipso, Ph),
135.3 (Cipso, Ph), 143.3 [C(8)], 149.3 [C(6)], 151.0, 151.2, 152.4
[C(2)], 155.3 [C(4)], 170.9 (CH2CO2Bzl) ppm. MS (ESI): m/z (%) ϭ
597.3 (100) [M ϩ Na]ϩ, 1171.6 (43) [2 M ϩ Na]ϩ. C30H34N6O6
(575.3): calcd. 575.2618; found 575.2635 (HRMS).
4.07Ϫ4.09 [m, 1 H, C(β)-H], 5.05 (s, br., 1 H, NH), 5.13 (s, 2 H,
CH2Ph), 7.33Ϫ7.39 (m, 5 H, Ph) ppm. 13C NMR (75 MHz, CDCl3,
25 °C): δ ϭ 28.1 [C(CH3)3], 29.3 [C(δ)], 37.2 [C(γ)], 38.4 [C(α)],
46.4 [C(β)], 66.4 (CH2Ph), 79.8 [C(CH3)3], 128.1Ϫ128.4 (Ph), 135.8
(Cipso, Ph), 155.7 (NHCO2), 171.3 (CO2Bzl) ppm. MS (ESI): m/z
(%) ϭ 408.4/410.1 (100/98) [M ϩ Na]ϩ. C17H24BrNO4·Na (408.1):
calcd. 408.0786; found 408.0789 (HRMS).
Benzyl (R)-β-(tert-Butoxycarbonylamino)-δ-hydroxypentanoate (7):
A 1  solution of borane in THF (51.6 mL, 51.6 mmol) was added
dropwise over 40 min to a solution of Boc--β-homoaspartic acid
5-benzyl ester (6) (2.90 g, 8.60 mmol) in dry THF (10 mL) at Ϫ5
°C under nitrogen and stirring continued at Ϫ5 °C for 5 h. Then,
the reaction was quenched by slowly adding methanol/acetic acid
(9:1, 20 mL). The solvents were removed under reduced pressure.
The residue was dissolved in ethyl acetate (300 mL) and washed
with 1  aq. HCl (2 ϫ 150 mL), sat. NaHCO3 (2 ϫ 150 mL), and
brine (1 ϫ 150 mL). The organic phase was dried (MgSO4), and
the solvent was removed under reduced pressure. The crude com-
pound was purified by column chromatography (silica gel, n-hex-
ane/ethyl acetate, 1:1) to afford 7 (2.03 g, 73%) as a colorless solid.
Rf ϭ 0.17 (n-hexane/ethyl acetate, 1:1). IR (solid): ν˜ ϭ 3390, 3335,
2986, 1728, 1668, 1517, 1449, 1274, 1245, 1219, 1161, 1123, 1027,
1
729 cmϪ1. H NMR (300 MHz, CDCl3, 25 °C): δ ϭ 1.41 (s, 9 H,
tBu), 1.48Ϫ1.60 [m, 1 H, C(γ)-H], 1.72Ϫ1.81 [m, 1 H, C(γ)-H],
Benzyl (R)-β-(tert-Butoxycarbonylamino)-δ-[N3-(benzoyl)thymin-1-
yl]pentanoate (10): A mixture of bromide 3 (2.10 g, 5.44 mmol), 3-
benzoylthymine (2.77 g, 12.0 mmol), K2CO3 (1.66 g, 12.0 mmol),
and TBAI (201 mg, 544 µmol) in dry DMF (66 mL) under nitrogen
was stirred at room temperature for 70 h. The solvent was removed
under reduced pressure, water (200 mL) was added, and the crude
compound was extracted with ethyl acetate (6 ϫ 100 mL). The
combined organic phases were dried (MgSO4) and the solvent was
removed under reduced pressure. Compound 10 was obtained as a
colorless solid (2.69 g, 92%) after column chromatography (silica
gel, n-hexane/ethyl acetate, 2:1, to ethyl acetate, 100% gradient).
M.p. 125Ϫ126 °C. Rf ϭ 0.67 (ethyl acetate/methanol, 9:1). IR
(solid): ν˜ ϭ 3337, 2980, 2931, 1733, 1697, 1673, 1640, 1520, 1437,
2.55 [dd, 2JH,H ϭ 16, 3JH,H ϭ 5 Hz, 1 H, C(α)-H], 2.67 [dd, 2JH,H ϭ
3
16, JH,H ϭ 5 Hz, 1 H, C(α)-H], 3.47 (s, br., 1 H, OH), 3.58Ϫ3.67
[m, 2 H, C(δ)-H], 4.02Ϫ4.04 [m, 1 H, C(β)-H], 5.15 (s, 2 H,
CH2Ph), 5.40 (s, br., 1 H, NH), 7.28Ϫ7.37 (m, 5 H, Ph) ppm. 13C
NMR (75 MHz, CDCl3, 25 °C): δ ϭ 28.1 [C(CH3)3], 37.3 [C(γ)],
38.9 [C(α)], 43.9 [C(β)], 58.4 [C(δ)], 66.3 (CH2Ph), 79.7 [C(CH3)3],
128.2Ϫ128.4 (Ph), 135.3 (Cipso
, Ph), 156.5 (NHCO2), 171.4
(CO2Bzl) ppm. MS (ESI): m/z (%) ϭ 346.1 (20) [M ϩ Na]ϩ, 669.2
(100) [2 M ϩ Na]ϩ. C17H25NO5·Na (346.2): calcd. 346.1630; found
346.1636 (HRMS).
Benzyl (R)-β-(tert-Butoxycarbonylamino)-δ-[N4-(benzyloxycarbonyl)-
cytosin-1-yl]pentanoate (8): A suspension of bromide 3 (1.16 g,
3.00 mmol), N4-(benzyloxycarbonyl)cytosine (2.46 g, 10.0 mmol),
K2CO3 (1.04 g, 7.53 mmol), and TBAI (111 mg, 301 µmol) in dry
DMF (50 mL) was stirred at room temperature for 11 days under
argon. Insoluble material was filtered off and washed with DMF
and ethyl acetate. The filtrate and the washing solutions were
pooled. After absorption on silica gel the crude product was puri-
fied by chromatography (silica gel, ethyl acetate/n-hexane, 3:1 to
5:1) to afford 8 (730 mg, 44%) as a colorless solid. M.p. 55 °C. Rf ϭ
0.26 (ethyl acetate/n-hexane, 3:1). [α]2D0 ϭ ϩ21.9 (c ϭ 0.24, MeOH).
IR (KBr): ν˜ ϭ 3366, 2976, 1738, 1694, 1627, 1503, 1366, 1214,
1
1341, 1247, 1203, 1148, 1055 cmϪ1. H NMR (300 MHz, CDCl3,
25 °C): δ ϭ 1.42 (s, 9 H, tBu), 1.91 [s, 3 H, C(5)-CH3], 1.80Ϫ1.93
3
[m, 2 H, C(γ)-H], 2.55 [d, JH,H ϭ 7 Hz, 2 H, C(α)-H], 3.52Ϫ3.67
[m, 1 H, C(δ)-H], 3.83Ϫ4.01 [m, 2 H, C(β)-H, C(δ)-H], 5.09 (m, 2
H, CH2Ph), 5.18 (m, 1 H, NH), 7.20 [m, 1 H, C(6)-H], 7.28Ϫ7.37
(m, 5 H, Ph), 7.46 (m, 2 H, Phm), 7.61 (m, 1 H, Php), 7.91 (m, 2
H, Pho) ppm. 13C NMR (75 MHz, CDCl3, 25 °C): δ ϭ 12.3 [C(5)-
CH3], 28.3 [C(CH3)3], 33.5 [C(γ)], 39.4 [C(α)], 45.1 [C(β)], 46.4
[C(δ)], 66.6 (CH2Ph), 79.8 [C(CH3)3], 110.5 [C(5)], 128.3Ϫ128.6
(Ph), 129.1 (2 C, COPh), 130.4 (2 C, COPh), 131.6 (COPh), 134.9
(COPh), 135.5 (Cipso, CH2Ph), 140.9 [C(6)], 149.8 (NHCO2), 155.5
[C(4)], 163.2 [C(2)], 169.1 (NHCOPh), 171.0 (CO2Bzl) ppm. MS
(ESI): m/z (%) ϭ 558.2 (40) [M ϩ Na]ϩ, 1093.3 (100) [2 M ϩ Na]ϩ.
C29H33N3O7·Na (558.2): calcd. 558.2216; found 558.2210 (HRMS).
1055, 788, 746, 697 cmϪ1 1H NMR (300 MHz, CDCl3, 25 °C):
.
δ ϭ 1.41 (s, 9 H, tBu), 1.83Ϫ2.06 [m, 2 H, C(γ)-H], 2.57 [d, 3JH,H ϭ
5 Hz, 2 H, C(α)-H], 3.58Ϫ3.76 [m, 1 H, C(δ)-H], 3.84Ϫ4.01 [m, 1
H, C(β)-H], 4.03Ϫ4.17 [m, 1 H, C(δ)-H], 5.08 (s, 2 H, CH2Ph), 5.20
(s, 2 H, CH2Ph), 5.27 (d, 3JH,H ϭ 10 Hz, 1 H, NH), 7.12Ϫ7.22 [m,
1 H, C(5)-H], 7.26Ϫ7.31 (m, 10 H, 2 Ph), 7.83Ϫ7.85 [m, 1 H, C(6)-
H] ppm. 13C NMR (125 MHz, CDCl3, 25 °C): δ ϭ 28.3 [C(CH3)3],
Benzyl (R)-β-(tert-Butoxycarbonylamino)-δ-(2-amino-6-chloropurin-
9-yl)pentanoate (11): Bromide 3 (3.77 g, 9.76 mmol) was added to a
suspension of 2-amino-6-chloropurine (2.48 g, 14.6 mmol), K2CO3
33.2 [C(γ)], 39.4 [C(α)], 45.0 [C(β)], 48.5 [C(δ)], 66.5 (CH2Ph), 67.8 (2.02 g, 14.6 mmol), and TBAI (361 mg, 977 µmol) in dry DMF
(CH2Ph), 79.7 [C(CH3)3], 94.7 [C(5)], 128.1Ϫ128.6 (Ph), 135.1 (100 mL). The mixture was stirred at room temperature for 6 days
(Cipso, Ph), 135.5 (Cipso, Ph), 149.4 [C(6)], 152.4, 155.6, 162.4, 162.5, under argon. After removal of the solvent the residue was absorbed
Eur. J. Org. Chem. 2003, 3555Ϫ3561
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 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
3559

A. M. Brückner, M. Garcia, A. Marsh, S. H. Gellman, U. Diederichsen
FULL PAPER
on silica gel (MeOH). Compound 11 was isolated by chromatogra-
phy (silica gel, ethyl acetate/n-hexane, 2:1, to ethyl acetate, 100%
gradient) as a colorless solid (2.17 g, 47%). M.p. 51Ϫ55 °C. Rf ϭ
0.46 (ethyl acetate/n-hexane, 3:1). [α]2D0 ϭ ϩ31.1 (c ϭ 0.29, MeOH).
[C(5)], 128.2Ϫ128.7 (Ph), 136.7 (Cipso, Ph), 144.6 [C(8)], 149.7
[C(4)], 151.7 [C(2)], 152.3, 152.5, 155.5 [C(6)], 172.6 (CO2H) ppm.
MS (ESI): m/z (%) ϭ 507.2 (100) [M ϩ Na]ϩ, 991.4 (35) [2 M ϩ
Na]ϩ. C23H28N6O6 (507.2): calcd. 507.1968; found 507.1956
IR (KBr): ν˜ ϭ 3332, 3212, 2976, 2362, 1705, 1615, 1561, 1520, (HRMS).
1459, 1410, 1367, 1285, 1248, 1163, 1054, 997, 912, 785, 748, 698,
642 cmϪ1. H NMR (300 MHz, CDCl3, 25 °C): δ ϭ 1.42 (s, 9 H,
tBu), 2.03 [m, 2 H, C(γ)-H], 2.59 [m, 2 H, C(α)-H], 3.88Ϫ4.02 [m,
1 H, C(β)-H], 4.02Ϫ4.18 [m, 2 H, C(δ)-H], 5.07 (s, 2 H, CH2Ph),
5.21 (s, 2 H, NH2), 5.31 (m, 1 H, NH), 7.25Ϫ7.36 (m, 5 H, Ph),
7.81 [s, 1 H, C(8)-H] ppm. 13C NMR (75 MHz, CDCl3, 25 °C):
δ ϭ 28.3 [C(CH3)3], 34.3 [C(γ)], 39.1 [C(α)], 41.0 [C(δ)], 45.3 [C(β)],
66.6 (CH2Ph), 79.9 [C(CH3)3], 125.3 [C(5)], 128.3Ϫ128.6 (Ph),
135.3 (Cipso, Ph), 142.7 [C(8)], 151.2, 153.7, 155.4, 158.9, 171.1
(CH2CO2) ppm. MS (ESI): m/z (%) ϭ 497.4/499.3 (100/32) [M ϩ
Na]ϩ. C22H27ClN6O4 (474.2): calcd. 474.1782; found 474.1788
(HRMS).
1
(R)-β-(tert-Butoxycarbonylamino)-δ-(thymin-1-yl)pentanoic
Acid
(14): Ammonium formate (1.26 g, 20.0 mmol) was added to a solu-
tion of benzyl ester 10 (2.68 g, 5.00 mmol) in MeOH (76 mL). Oxy-
gen was removed (3 ϫ vacuoϪnitrogenϪvacuo), and palladium (10
wt.%) on activated carbon (537 mg, 20 wt.%) was slowly added at
room temperature. The mixture was refluxed for 6 h, cooled to
room temperature and filtered through Celite. The catalyst was
washed with MeOH. The filtrate was evaporated, and the residue
was dissolved in EtOH (40 mL).
A solution of hydrazine
monohydrate (24.3 mL, 501 mmol) in EtOH (25 mL) was added
dropwise and the mixture was heated at reflux for 40 h. After con-
centration of the mixture, the residue was dissolved in water
(R)-β-(tert-Butoxycarbonylamino)-δ-[N4-(benzyloxycarbonyl)- (200 mL) and washed with diethyl ether (2 ϫ 100 mL). The aque-
cytosin-1-yl]pentanoic Acid (12): A mixture of benzyl ester 8
(551 mg, 1.00 mmol), 1  aq. NaOH (1.80 mL), dioxane (5.00 mL),
ous phase was acidified with 1  aq. HCl (pH, 2Ϫ3) and extracted
with ethyl acetate (4 ϫ 150 mL). The organic phase was dried
and water (3.00 mL) was stirred at room temperature for 1 day. (MgSO4), and the solvent was removed under reduced pressure to
After neutralization with 2  aq. HCl and freeze drying, the residue
was subjected to chromatography (RP-C18 silica gel, H2O, 100%,
to H2O/MeOH, 1:4 gradient) to afford 12 (288 mg, 63%) as a color-
less solid. M.p. 131Ϫ133 °C. Rf ϭ 0.44 (ethyl acetate/MeOH/H2O/
acetic acid, 20:2:2:1, sat. with NaCl). [α]2D0 ϭ Ϫ23.6 (c ϭ 0.61,
afford acid 14 (804 mg, 47%) as a colorless solid. M.p. 130Ϫ131
°C. Rf ϭ 0.20 (n-hexane/ethyl acetate/acetic acid, 6:4:0.3). IR
(solid): ν˜ ϭ 3411, 3330, 2976, 1689, 1583, 1355, 1243, 1157 cmϪ1
.
1H NMR (300 MHz, [D6]DMSO, 25 °C): δ ϭ 1.48 (s, 9 H, tBu),
1.71 [s, 3 H, C(5)-CH3], 1.57Ϫ1.80 [m, 2 H, C(γ)-H], 2.28Ϫ2.39 [m,
DMSO). IR (KBr): ν˜ ϭ 1749, 1695, 1630, 1563, 1507, 1370, 1218, 2 H, C(α)-H], 3.48Ϫ3.80 [m, 3 H, C(β)-H, C(δ)-H], 6.81 (m, 1 H,
1057, 785, 747, 697 cmϪ1. 1H NMR (300 MHz, [D6]DMSO, 35 °C): NHBoc), 7.44 [s, 1 H, C(6)-H], 11.19 [s, br., 1 H, NH] ppm. 13C
δ ϭ 1.36 (s, 9 H, tBu), 1.64Ϫ1.87 [m, 2 H, C(γ)-H], 2.22 [m, 2 H, NMR (75 MHz, [D6]DMSO, 25 °C): δ ϭ 12.1 [C(5)-CH3], 28.3
C(α)-H], 3.68 [m, 2 H, C(β)-H, C(δ)-H], 3.79 [m, 1 H, C(δ)-H], 5.17 [C(CH3)3], 33.4 [C(γ)], 39.6 [C(α)], 44.9 [C(δ)], 45.3 [C(β)], 77.8
3
(s, 2 H, CH2Ph), 6.93 [d, JH,H ϭ 7 Hz, 1 H, C(5)-H], 7.29Ϫ7.42 [C(CH3)3], 108.3 [C(5)], 141.6 [C(6)], 150.8 (NHCO2), 155.1 [C(4)],
3
(m, 5 H, Ph), 8.02 [d, JH,H ϭ 7 Hz, 1 H, C(6)-H] ppm. 13C NMR
164.4 [C(2)], 172.4 (CO2H) ppm. MS (ESI): m/z (%) ϭ 340.2 (100)
(75 MHz, [D6]DMSO, 35 °C): δ ϭ 28.2 [C(CH3)3], 33.6 [C(γ)], 41.0 [M Ϫ H]Ϫ, 681.4 (32) [2 M Ϫ H]Ϫ. C15H23N3O6 (340.2): calcd.
[C(α)], 45.6 [C(β)], 47.1 [C(δ)], 66.3 (CH2Ph), 77.5 [C(CH3)3], 93.8
[C(5)], 127.8Ϫ128.4 (Ph), 135.9 (Cipso, Ph), 149.7 [C(6)], 153.2,
154.7, 154.9, 162.5, 173.4 (CO2H) ppm. MS (ESI): m/z (%) ϭ 427.5
(16) [M Ϫ tBu ϩ H ϩ Na]ϩ, 483.5 (100) [M ϩ Na]ϩ, 505.6 (42)
[M ϩ2 Na Ϫ H]ϩ, 943.4 (33) [2 M ϩ Na]ϩ, 965.4 (68) [2 M ϩ2
Na Ϫ H]ϩ, 987.4 (94) [2 M ϩ3 Na Ϫ2 H]ϩ. C22H28N4O7·H2O
(478.5): calcd. C 55.22, H 6.32, N 11.71; found C 55.61, H 6.29,
N 17.35.
340.1509; found 340.1508 (HRMS).
General Procedure for Solid-Phase β-Peptide Synthesis: Oligomers
were prepared by manual solid-phase peptide synthesis in a small
column using 4-methylbenzhydrylamine polystyrene (MBHA-PS)
resin loaded with H-β-HGly-OH (20.0 mg, 12.4 µmol β-homogly-
cine amide). For the first step of the double coupling, an excess of
5 equiv. amino acid (62.0 µmol) was used and activated by 1-
[bis(dimethylamino)methyliumyl]-1H-1,2,3-triazolo[4,5-b]pyridin-
3-oxide hexafluorophosphate (HATU, 21.2 mg, 55.8 µmol), 1-
hydroxy-7-azabenzotriazole [HOAt, 124 µL (62.0 µmol) of a 0.5 
solution in DMF], and N-ethyldiisopropylamine (DIEA, 30.3 µL,
174 µmol) in DMF (500 µL). For the second step of the double
coupling, an excess of 3 equiv. amino acid (37.2 µmol) was used
and activated by HATU (12.7 mg, 33.5 µmol), HOAt/DMF (74.4
µL, 37.2 µmol), and DIEA (19.5 µL, 112 µmol) in DMF (500 µL).
(R)-β-(tert-Butoxycarbonylamino)-δ-[N6-(benzyloxycarbonyl)adenin-
9-yl]pentanoic Acid (13): A solution of 9 (1.06 g, 1.84 mmol) in di-
oxane (9 mL) and 1  aq. NaOH (3.7 mL) was stirred at room
temperature for 22 h. After evaporation to dryness, the residue was
dissolved in water (100 mL) and washed with diethyl ether (2 ϫ
250 mL). The aqueous layer was acidified with 1  aq. HCl (pH,
2Ϫ3) and extracted with ethyl acetate (3 ϫ 100 mL). The combined
organic phases were washed with brine (100 mL) and dried After swelling the loaded resin for 1 h in 2 mL of CH2Cl2, the fol-
(MgSO4). The solvent was removed under reduced pressure to af- lowing procedure was repeated for every amino acid unit: 1) depro-
ford 13 (819 mg, 92%) as a colorless solid. M.p. 164Ϫ165 °C. Rf ϭ tection twice, for 3 min with TFA/m-cresol (95:5, 2 mL); 2) washing
0.39 (ethyl acetate/methanol, 9:1). IR (solid): ν˜ ϭ 3425, 3280, 2975,
first five times with CH2Cl2/DMF (1:1, 2 mL) and then five times
with pyridine (2 mL); 3) double coupling steps, each 1 h gentle
1748, 1703, 1617, 1586, 1495, 1212, 1160, 1004, 740 cmϪ1 1H
.
NMR (300 MHz, [D6]DMSO, 25 °C): δ ϭ 1.26 (s, 1 H, tBu), 1.38 moving at 50 °C; 4) washing with CH2Cl2/DMF (1:1, 3 ϫ 2 mL),
(s, 8 H, tBu), 1.83Ϫ1.95 [m, 1 H, C(γ)-H], 2.03Ϫ2.12 [m, 1 H,
C(γ)-H], 2.36Ϫ2.39 [m, 2 H, C(α)-H], 3.62Ϫ3.81 [m, 1 H, C(β)-H],
DMF/piperidine (95:5, 3 ϫ 2 mL), and CH2Cl2/DMF (1:1, 3 ϫ
2 mL). The resin was washed with TFA (3 ϫ 2 mL) and CH2Cl2 (5
ϫ 2 mL), dried overnight in vacuo, and suspended in m-cresol (200
4.15Ϫ4.30 [m, 2 H, C(δ)-H], 5.20 (s, 2 H, CH2Ph), 6.54 (d, 3JH,H ϭ
3
9 Hz, 0.1 H, NHBoc), 6.92 (d, JH,H ϭ 9 Hz, 0.9 H, NHBoc), µL)/thioanisole (200 µL)/ethanedithiol (100 µL). After stirring for
7.30Ϫ7.47 (m, 5 H, Ph), 8.42 [s, 1 H, C(8)], 8.60 [s, 1 H, C(2)], 30 min at room temperature, TFA (2 mL) was added, and the sus-
10.66 (s, br., 1 H, NHCO2Bzl), 12.18 (1 H, CO2H) ppm. 13C NMR pension was cooled to Ϫ15 °C. Trifluoromethanesulfonic acid (200
(75 MHz, [D6]DMSO, 25 °C): δ ϭ 28.6 [C(CH3)3], 34.4 [C(γ)], 40.7 µL) was added, the mixture was allowed to warm to room tempera-
[C(α)], 40.9 [C(δ)], 45.4 [C(β)], 66.5 (CH2Ph), 78.2 [C(CH3)3], 123.7
ture within 1.5 h, and stirring continued for 1.5 h. The filtrate was
3560
 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
Eur. J. Org. Chem. 2003, 3555Ϫ3561

Synthesis of Novel Nucleo-β-Amino Acids and Nucleobase-Functionalized β-Peptides
FULL PAPER
S. H. Gellman, J. Am. Chem. Soc. 1996, 118, 13071Ϫ13072.
concentrated by freeze drying, and the β-homoalanyl PNA was pre-
cipitated with Et2O (30 mL at Ϫ15 °C) as a colorless solid and
purified by HPLC. The yield of each coupling step was estimated
from HPLC to be higher than 97%.
[7b]
P. I. Arvidsson, M. Rueping, D. Seebach, Chem. Commun.
2001, 649Ϫ650.
The designation of β3-amino acids or β3-peptides was proposed
by Seebach to specify the position of the side chain: T. Hinter-
mann, D. Seebach, Synlett 1997, 437Ϫ438. The preparation of
a nucleo-β2-amino acid has recently been described: T. Yokom-
atsu, K. Takada, A. Yasumoto, Y. Yuasa, S. Shibuya, Hetero-
cycles 2002, 56, 545Ϫ552.
[8]
H-(β-HLys-ApaC-ACHC-β-HLys-ApaG-ACHC-β-HLys-ApaC-
ACHC-β-HGly)-NH2 (15): Analytical HPLC: 23.5 min, gradient:
15Ϫ30% B (B ϭ MeCN/H2O, 9:1 ϩ 0.1% TFA) in 30 min. MS
(ESI): m/z (%) ϭ 519.7 (64) [M ϩ3 H]3ϩ, 778.6 (100) [M ϩ2 H]2ϩ
1552.2 (5) [M ϩ H]ϩ.
,
[9] [9a]
U. Diederichsen, H. W. Schmitt, Angew. Chem. 1998, 110,
[9b]
312Ϫ315; Angew. Chem. Int. Ed. 1998, 37, 302Ϫ305.
U.
Diederichsen, H. W. Schmitt, Eur. J. Org. Chem. 1998,
827Ϫ835. [9c] A. M. Brückner, H. W. Schmitt, U. Diederichsen,
Helv. Chim. Acta 2002, 85, 3855Ϫ3866.
H-(β-HLys-ApaG-ACHC-β-HLys-ApaC-ACHC-β-HLys-ApaG-
ACHC-β-HGly)-NH2 (16): Analytical HPLC: 22.9 min, gradient:
15Ϫ30% B (B ϭ MeCN/H2O, 9:1 ϩ 0.1% TFA) in 30 min. MS
(ESI): m/z (%) ϭ 798.7 (100) [M ϩ2 H]2ϩ, 809.7 (30) [M ϩ H ϩ
Na]2ϩ, 1595.0 (7) [M ϩ H]ϩ.
[10] [10a]
[10b]
D. C. Cole, Tetrahedron 1994, 50, 9517Ϫ9582.
M. B.
Smith, Methods of Non-α-Amino Acid Synthesis, Marcel
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[13]
J. Podlech, D. Seebach, Liebigs Ann. 1995, 1217Ϫ1228.
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79, 913Ϫ941.
Acknowledgments
This work was supported by the Deutsche Forschungsgemeinschaft
and the U. S. National Science Foundation (CHE-0140621). We
are grateful for fellowships by the Fonds der Chemischen Industrie
(A.M.B.) and the Universities of Wisconsin Ϫ Madison and War-
wick (M.G.).
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