High yielding selective access to spirocyclopropanated 5-oxopiperazine-2- carboxylates and 1,4-diazepane-2,5-diones from methyl 2-chloro-2- cyclopropylideneacetate

Article abstract of DOI:10.1039/b809174a

The 2-spirocyclopropanated methyl 5-oxopiperazine-2-carboxylate 3 and the 3-spirocyclopropanated 6-chloro-1,4-diazepane-2,5-dione 4 could both be prepared at choice in 93 and 88% yield, respectively, from methyl 2-chloro-2- cyclopropylideneacetate (1) in a sequence of Michael addition of 3-benzyloxypropylamine, peptide coupling with N-Boc-glycine, Boc-group removal and cyclization. Transformation of the benzyloxypropyl side chain, peptide coupling with N-Boc-(S)-asparagine, deprotection and repeated cyclization led to the octahydro[2H]pyrazino[1,2-a]pyrazinetrione scaffold 10 containing a rigidified mimic of a tripeptide with a DGR motif. The overall yield of 11 after deprotection of 10 (a total of 13 steps in 8 distinct operations) was 30%. The Royal Society of Chemistry 2008.

Full text of DOI:10.1039/b809174a

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PAPER
www.rsc.org/obc | Organic & Biomolecular Chemistry
High yielding selective access to spirocyclopropanated
5-oxopiperazine-2-carboxylates and 1,4-diazepane-2,5-diones
from methyl 2-chloro-2-cyclopropylideneacetate†
Michael Limbach,a Vadim S. Korotkov,a Mazen Es-Sayedb and Armin de Meijere*a
Received 30th May 2008, Accepted 24th June 2008
First published as an Advance Article on the web 13th August 2008
DOI: 10.1039/b809174a
The 2-spirocyclopropanated methyl 5-oxopiperazine-2-carboxylate 3 and the 3-spirocyclopropanated
6-chloro-1,4-diazepane-2,5-dione 4 could both be prepared at choice in 93 and 88% yield, respectively,
from methyl 2-chloro-2-cyclopropylideneacetate (1) in a sequence of Michael addition of
3-benzyloxypropylamine, peptide coupling with N-Boc-glycine, Boc-group removal and cyclization.
Transformation of the benzyloxypropyl side chain, peptide coupling with N-Boc-(S)-asparagine,
deprotection and repeated cyclization led to the octahydro[2H]pyrazino[1,2-a]pyrazinetrione scaffold
10 containing a rigidified mimic of a tripeptide with a DGR motif. The overall yield of 11 after
deprotection of 10 (a total of 13 steps in 8 distinct operations) was 30%.
a non-functionalized precursor of an arginine analogue, added
Introduction
smoothly onto 1 (Scheme 1). Subsequent peptide coupling employ-
ing N,N¢-dicyclohexylcarbodiimide (DCC) with Boc-glycine in
THF, without isolation of the unstable Michael adduct, provided
the acyclic dipeptide 2 in 88% yield. The NMR characterization
of this compound was difficult, due to its dynamic behavior on the
NMR timescale at ambient temperature, and the thermal lability
of the tert-butoxycarbonyl group prevented measurements at an
elevated temperature (90 ◦C). After chromatography and acidic
deprotection (trifluoroacetic acid (TFA) or methanolic HCl) of
Amongst the various previously reported mimics of the biologi-
cally highly relevant RGD (Arg-Gly-Asp) tripeptide motif2 were
several conformationally restricted ones with oxopiperazine3 and
2,5-dioxopiperazine moieties.4 As has previously been demon-
strated, the multifunctional building block methyl 2-chloro-
2-cyclopropylideneacetate (1),5 for which a second-generation
synthesis has recently been developed,6 provides an easy access
to a large variety of heterocycles7 including spirocyclopropanated
octahydro[2H]pyrazino[1,2-a]pyrazines8 with an incorporated
oxopiperazine as well as a dioxopiperazine moiety. Since the yields
of the oxopiperazinecarboxylates as the precursors to the bicyclic
compounds were only 21–22%, due to a predominant formation
of a seven-membered lactam in the first cyclization step, we set
out to determine conditions for a selective preparation of the
six-membered heterocycle. Here we report on the success of this
project and an application to the synthesis of a bicyclic and
thereby rigidified mimic of a tripeptide containing a retro-RGD
motif.
1
2, base-induced cyclization gave, according to H NMR, a 1 : 1
mixture of the six-membered cyclic a-amino acid ester 3 and
the seven-membered 6-chloro-1,4-diazepane-2,5-dione (4) in 40%
yield. The separation of these two compounds by chromatography
on silica gel was difficult because of their nearly identical retention
factors, however, when the crude mixture was treated with
cold Et2O–MeOH, 4 crystallized and could be filtered off as
a colorless solid. The mother liquor contained the oily methyl
oxopiperazinecarboxylate 3, which was easily purified by filtration
over silica gel.
In view of the low overall yield and the low selectivity for
the formation of either products 3 or 4 as well as the tedious
separation, various conditions were screened to find out the best
ones for selective formation of either one of the products (Table 1).
Deprotection of the dipeptide 2 with trifluoroacetic acid (TFA),
aqueous work-up and subjection to basic conditions in a biphasic
system (K2CO3 in CH2Cl2) predominantly yielded the 6-chloro-
1,4-diazepane-2,5-dione 4 (66 vs. 29% of 3). Cyclization in aqueous
NH3 at elevated temperature after N-deprotection of 2 with
trifluoroacetic acid or with methanolic HCl gave even better yields
of 4 (70 and 81%, respectively), because extractive work-up was not
necessary, and 3 was not formed at all. The best yield of 4 (88%),
however, was achieved by deprotection of 2 with methanolic HCl
followed by cyclization at 60 ◦C in methanolic NH3. On the other
hand, initial basic hydrolysis of the ester function in the dipeptide
2, subsequent N-deprotection and ensuing cyclization under basic
conditions (methanolic NH3) followed by reesterification, gave
Results and discussion
As an extremely good Michael acceptor, methyl 2-chloro-2-
cyclopropylideneacetate (1) is known to rapidly undergo addition
of all sorts of nucleophiles including secondary and primary
amines, and the products can further react with nucleophilic
substitution of the chlorine substituent adjacent to the cyclopropyl
group.7 Thus, 3-benzyloxypropylamine, which can be regarded as
aInstitut fu¨r Organische und Biomolekulare Chemie, Georg-August-
Universita¨t Go¨ttingen, Tammannstrasse 2, D-37077, Go¨ttingen, Germany.
E-mail: Armin.deMeijere@chemie.uni-goettingen.de; Fax: + 49(0)551/
399475
bBayerCropScience AG, Agricultural Center Monheim, D-51368, Lev-
erkusen, Germany
† Cyclopropyl Building Blocks for Organic Synthesis. Part 149. For Part
148 see: ref. 1a. Part 147: ref. 1b.
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Scheme 1 Assembly of the 4-oxopiperazinecarboxylate 3 and the cyclopropanated 6-chloro-1,4-diazepane-2,5-dione 4 from methyl 2-chloro-2-
cyclopropylideneacetate 1. For details of the second step see Table 1.
the methyl oxopiperazinecarboxylate 3 selectively and in very high
yield (93%). Under these conditions, 4 is not formed at all, because
the carboxylate ion is not prone to undergo attack by a nucleophile,
yet careful monitoring of the reaction by TLC and NMR is
necessary to obtain best yields. Eventually, a similarly good yield of
3 could be achieved by simply stirring the deprotected (methanolic
HCl) dipeptide 2 in THF with an excess of triethylamine. Though
in this case the diazepane-2,5-dione 4 was also formed in 14% yield,
multigram quantities (up to 6 g) of 3 could easily be obtained in
84% yield according to this protocol.
of the amine with an excess of triethylamine provided 8 in 98%
yield.
Standard peptide coupling of 8 with N-Boc-(S)-Asp-OBn em-
ploying DCC and 2,4,6-collidine as a sterically encumbered base to
prevent racemization yielded the fully protected tripeptide mimic
9 (Scheme 2), which was also difficult to characterize by NMR
spectroscopy because of its dynamic behavior at 20 ◦C. The latter
must be due to several coexisting conformers of this tripeptide,
which only slowly (on the 1H-NMR timescale) convert into each
other. Spectra could not be recorded at elevated temperature
because of its thermal instability.
tert-Butoxycarbonylation of the secondary amine 3 gave 5-Bn in
75% yield after chromatographic separation from approximately
14% of 4 remaining from the cyclization step (Scheme 2). Depro-
tection of the benzyloxy group in 5-Bn by catalytic hydrogenation
Removal of the N-Boc group from 9 was achieved by treatment
with an excess of methanolic HCl, and the resulting amino-
terminal dipeptide fragment in the deprotected 9 was cyclized
by heating the compound in toluene at 90 ◦C to furnish the
fully protected octahydro[2H]oxopyrazino[1,2-a]pyrazinedione 10
containing the rigidified retro-RGD motif with a b-amino acid
analogue of arginine. This unseparable 1 : 1 mixture of diastere-
omers was completely deprotected by catalytic hydrogenation in
methanol over palladium on charcoal. The crude mixture was
R
in methanol and filtration of the crude mixture over Celiteꢀ
gave the primary alcohol 5-H in nearly quantitative yield. The
latter was directly used for the Mitsunobu reaction with 1.5
equivalents of the twofold Z-protected guanidine 69 in the presence
of stoichiometric amounts of triphenylphosphine and diisopropyl
azodicarboxylate (DIAD) at 0 ◦C. The termination of the reaction
was indicated by the disappearance of the intensive orange color
of DIAD, and the fully protected oxopiperazinecarboxylate 7 was
isolated in 87% yield (over two steps and after chromatography).
Removal of the N-Boc group with methanolic HCl and liberation
R
purified by filtration over Celiteꢀ to prevent any loss of the DGR
tripeptide mimic 11, which is extremely soluble in water. Thus, the
1 : 1 mixture of trans- and cis-11 was obtained as a colorless solid
in 99% yield.
Table 1 Optimization of the deprotection–cyclization of the dipeptide 2 to either yield the 5-oxopiperazine-2-carboxylate 3 or the cyclopropanated
6-chloro-1,4-diazepane-2,5-dione 4 (see Scheme 2)
Entry
Conditions
Yield 3 (%)
Yield 4 (%)
1
2
3
4
5
(1) TFA, CH2Cl2; (2) NaHCO3, DMF, 20 ◦
(1) TFA, CH2Cl2; (2) aq. K2CO3, CH2Cl2
C
20
29
—
—
—
20
66
70
81
(1) TFA, CH2Cl2; (2) aq. NH3, 60 ◦
C
(1) HCl, MeOH; (2) aq. NH3, 60 ◦
(1) HCl, MeOH; (2) NH3, MeOH, 60 ◦
C
C
6
7
(1) TFA, CH2Cl2; (2) Na2CO3, DMF, 25 ◦
C
49
84
< 2
—
(1) NaOH, MeOH, 20 ◦C, 8 h; (2) MeOH, HCl, 0 ◦C, 4 h; (3) MeOH,
NH3, 0 → 20 ◦C, 10 h; (4) MeOH, cat. conc. H2SO4, 8 h
8
(1) HCl, MeOH; (2) NEt3, THF
14
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Scheme 2 Conversion of the oxopiperazinecarboxylate 3 to the spirocyclopropanated octahydro[2H]oxopyrazino[1,2-a]pyrazinedione 11.
Conclusion
Experimental section
The overall achievement constitutes a 13-step synthesis (8 dis-
tinct operations) of a conformationally rigidified peptidomimetic
with the retro-RGD sequence (DGR) from methyl 2-chloro-
2-cyclopropylideneacetate (1) in 30% overall yield. The three-
membered ring in the intermediate 2 formed by Michael addition
of a primary amine to 1 and subsequent peptide condensation with
N-Boc-glycine, due to its Thorpe–Ingold effect,10 is instrumental
in favoring the cyclization to an oxopiperazinecarboxylate 3.
The seven-membered 6-chloro-1,4-diazepane-2,5-dione 4, which
can alternatively be obtained with high selectivity from 1, 3-
benzyloxypropylamine and N-Boc-protected glycine, also offers
itself as a multifunctional scaffold for various pharmacophores
and as a precursor to other interesting heterocycles.11
General remarks
All reagents were used as purchased from commercial suppliers
without further purification. All reactions in non-aqueous solvents
were carried out using standard Schlenk techniques under an
atmosphere of dry nitrogen. Solvents were purified and dried
according to conventional methods prior to use; diethyl ether
(Et2O) and tetrahydrofuran (THF) were freshly distilled from
sodium–benzophenone. Solvents are abbreviated as follows: Py =
pyridine, EtOAc = ethyl acetate, MeOH = methanol, nBuOH =
n-butanol, TFA = trifluoroacetic acid. 1H- and 13C-NMR spectra
were recorded at ambient temperature on a Bruker AM 250 or
a Varian UNITY-300 instrument. Chemical shifts d are given in
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ppm relative to residual resonances of solvents (1H: 7.26 ppm for
chloroform; 13C: 77.0 ppm for CDCl3), coupling constants J are
given in Hertz. Characterization of the multiplicity of signals: s =
singlet, br s = broad singlet, d = doublet, t = triplet, m = multiplet.
The multiplicities of signals were determined by the DEPT
(Bruker) or the APT (Varian) technique. IR: Bruker IFS 66. MS:
Finnigan MAT 95. Chromatography: Separations were carried out
on Merck Silica 60 (0.063–0.200 mm, 70–230 mesh ASTM). The
dimensions of the columns are given as “diameter ¥ height of the
50 mmol, 2 M) at 0 ◦C with rewarming to 20 ◦C for 10 h. All
volatiles were removed in vacuo, the residue was dissolved in
MeOH (100 mL), conc. H2SO4 (3 mL) was added, and the mixture
was stirred for 8 h. Removal of the solvent in vacuo, uptake of the
residue in CH2Cl2 (200 mL), extraction with sat. NaHCO3 solution
(3 ¥ 30 mL), drying of the organic phase over Na2SO4, removal of
the solvents in vacuo and chromatographic purification on 200 g of
silica gel (5 ¥ 30 cm, CH2Cl2–MeOH 95 : 5, Rf = 0.33, ninhydrin)
yielded 3.90 g (93%) of 3 as an oil. IR (film): n = 3340 cm-1 (NH),
3040, 2912, 2835, 1750 (CO), 1705 (CO), 1660 (CO), 859, 756. 1H
NMR (250 MHz, CDCl3): d = 0.75–0.96 (m, cPr-H, 1 H), 0.97–
1.05 (m, cPr-H, 1 H), 1.20–1.30 (m, cPr-H, 1 H), 1.34–1.44 (m,
cPr-H, 1 H), 1.57–1.70 (m, 2 CH2, 4 H), 2.10–2.18 (br s, NH, 1 H),
R
silica gel column”. TLC: Macherey–Nagel, TLC plates Alugramꢀ
Sil G/UV254. Detection under UV light at 254 nm, development
with MOPS (10% molybdophosphoric acid, solution in ethanol)
or ninhydrin (100 mL n-BuOH, 3 mL glacial acetic acid, 100 mg
ninhydrin) reagent. Melting points: apparatus according to Dr
Tottoli (Bu¨chi); the values are uncorrected. Elemental analyses:
Mikroanalytisches Laboratorium des Instituts fu¨r Organische und
Biomolekulare Chemie der Universita¨t Go¨ttingen.
=
3.18 (s, CH, 1 H), 3.45 (m, CH2, 2 H), 3.58 (s, NCH2C O, 2 H),
3.68 (s, OCH3, 3 H), 4.49 (s, CH2Bn, 2 H), 7.25–7.33 (s, aryl-H,
5 H). 13C NMR (62.9 MHz, CDCl3): d = 8.8 (CH2, cPr-C), 15.2
(CH2, cPr-C), 28.9 (CH2), 38.8 (CH2), 40.0 (C, cPr-C), 47.9 (CH2),
52.6 (CH3), 62.9 (CH), 67.5 (CH2), 72.8 (CH2, OBn), 127.36 (CH,
aryl-C), 127.40 (CH, aryl-C), 138.1 (C, aryl-C), 172.1 (CO), 172.5
(CO). MS (70 eV, EI); m/z (%): 332 (10) [M+], 273 (10), 241 (25),
Methyl {1-[(3-Benzyloxypropyl)-(2-tert-butoxycarbonylamino-
acetyl)amino]cyclopropyl}chloroacetate (2). To a solution of 3-
benzyloxypropylamine12 (13.2 g, 79.9 mmol) in THF (100 mL) was
added dropwise at 0 ◦C a solution of 16 (10.6 g, 72.3 mmol) in THF
(50 mL), and the mixture was stirred at this temperature for 5 h. At
0 ◦C, pyridine (9.64 mL, 119 mmol), N-tert-butoxycarbonylglycine
(20.9 g, 119 mmol) as well as a solution of DCC (24.7 g, 119 mmol)
in THF (70 mL) were added, and the mixture was stirred with
rewarming to 20 ◦C for 18 h. The precipitate was filtered off,
washed with THF (2 ¥ 50 mL), and the solvent was removed under
reduced pressure. The residue was t◦aken up in CH2Cl2 (300 mL),
the mixture extracted with cold (0 C) 1 N HCl (100 mL), water
(100 mL) and sat. NaHCO3 solution (3 ¥ 100 mL), the organic
extracts were dried over Na2SO4, the solvent was removed under
reduced pressure and the residue subjected to chromatography on
800 g of silica gel (10 ¥ 28 cm, pentane–ethyl acetate 6 : 4; Rf =
0.38, ninhydrin) to yield 29.9 g (88%) of 2 as an oil. IR (film): n =
3451 cm-1 (NH), 3288, 3245, 3045, 1770, 1675 (CO), 1410, 1399,
+
209 (10), 167 (20), 91 (100) [C7H7 ]. C18H24N2O4 (332.4): calcd. C
65.04, H 7.28, N 8.43; found C 64.90, H 7.34, N 8.80%.
Method B. To a solution of 2 (3.68 g, 7.85 mmol) in CH2Cl2
(20 mL) was added dropwise at 0 ◦C TFA (6.50 mL, 87.5 mmol),
and the mixture was stirred with rewarming to 20 ◦C for 18 h. It
was then cooled again to 0 ◦C and poured into a vigorously stirred
mixture of DMF (40 mL) and NaHCO3 (6.00 g, 71.4 mmol).
The suspension was stirred at 20 ◦C for 24 h, diluted with CH2Cl2
(200 mL), the organic phase was extracted with water (2 ¥ 100 mL)
and brine (100 mL), the aq. phase was re-extracted with CH2Cl2
(2 ¥ 50 mL), and the combined organic phases were dried over
Na2SO4. Removal of the solvent in vacuo and crystallization of the
◦
residue from MeOH–Et2O at -20 C yielded 527 mg (20%) of 4
as a colorless solid, mp 103–106 ◦C. Concentration of the mother
liquor in vacuo and chromatographic purification of the residue
on 50 g of silica gel (2 ¥ 30 cm, dichloromethane–methanol 95 : 5,
ninhydrin) yielded 521 mg (20%) of 3 as a colorless oil.
◦
1
702, 688. H NMR (300 MHz, C2D2Cl4, 120 C): d = 0.87–1.42
(m, cPr-H, CH2, 6 H), 1.45 (s, tBu-H, CH2, 11 H), 1.92 (unresolved
quintet, CH2, 2 H), 3.53 (m, CH2, 2 H), 3.81 (s, OCH3, 3 H), 4.02
Method C. A solution of 2 (9.60 g, 20.5 mmol) was stirred in
◦
methanolic HCl (80 mL, 100 mmol, 1.25 M) at 0 C for 4 h. All
=
(br s, CHCl, 1 H), 4.47 (m, NHC O, 1 H), 7.26–7.45 (m, aryl-H,
5 H). 13C NMR (75.5 MHz, C2D2Cl4, 120 ◦C): d = 20.1 (CH2,
cPr-C), 23.7 (CH2, cPr-C), 28.3 (CH3, tBu-C), 29.2 (CH2), 42.0
(C, cPr-C), 43.2 (CH2), 47.8 (CH2), 52.2 (CH3), 64.8 (CH), 67.6
(CH2), 72.8 (C, tBu-C) 79.5 (CH2), 126.4 (CH, aryl-C), 128.3 (CH,
aryl-C), 138.1 (C, aryl-C), 168.3 (CO), 168.5 (CO), 171.8 (CO). MS
volatiles were removed in vacuo, the residue was dissolved in THF
(200 mL), triethylamine (10.0 mL, 72.1 mmol) was added, and
the suspension was stirred at 20 ◦C for 15 h. The precipitate was
filtered off, washed with THF (2 ¥ 50 mL), and all volatiles were
removed in vacuo. Chromatographic purification of the residue on
100 g of silica gel (4 ¥ 30 cm, CH2Cl2–MeOH 95 : 5, ninhydrin)
yielded 5.72 g (84%) of 3 as a colorless oil, along with ca. 14% of 4.
This mixture was used in the next step without further purification.
+
(70 eV, DCI); m/z (%): 486 (75) [M + NH4 ], 469 (35), 452 (30), 435
(10), 163 (65), 123 (100), 106 (90). C23H33ClN2O6 (469.0): calcd. C
58.91, H 7.09, N 5.97, Cl 7.56; found C 58.60, H 6.83, N 5.85, Cl
7.35%.
4-(3-Benzyloxypropyl)-9-chloro-4,7-diazaspiro[2.6]nonane-5,8-
dione (4). Compound 2 (3.24 g, 6.91 mmol) was dissolved in
methanolic HCl (70 mL, 63.7 mmol, 0.91 M) at 0 ◦C, and the
mixture was stirred with rewarming to 20 ◦C for 11 h. All volatiles
were removed in vacuo, the residue was dissolved in methanolic
NH3 (◦80 mL, 373 mmol, 4.66 M, pH ~ 10), and the mixture heated
at 60 C for 4.5 h. The solvents were removed in vacuo, and the
residue subjected to chromatography on 70 g of silica gel (23 ¥
3 cm, CH2Cl2–MeOH 95 : 5, ninhydrin, Rf = 0.30) to furnish the
crude product as an oil, which crystallized upon treatment with
Methyl 4-(3-benzyloxypropyl)-5-oxo-4,7-diazaspiro[2.5]octane-
8-carboxylate (3).
Method A. To a solution of 2 (5.92 g, 12.6 mmol) in MeOH
(70 mL) was added 1 M aq. NaOH (15 mL), and the emulsion was
◦
stirred at 20 C for 8 h. All volatiles were removed in vacuo, the
aq. phase was extracted with CH2Cl2 (3 ¥ 50 mL), the combined
organic phases were dried over Na2SO4, the solvent was removed
in vacuo, and the residue was stirred in methanolic HCl (20 mL,
◦
92 mmol, 4.60 M) at 0 C for 4 h. All volatiles were removed in
◦
vacuo, and the residue was stirred in methanolic NH3 (25 mL,
MeOH–Et2O at -20 C to yield 2.04 g (88%) of 4 as a colorless
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solid, mp 103–106 ◦C. IR (KBr): n = 3250 cm-1 (NH), 3175, 3056,
2837, 2801, 1652 (CO), 801, 745. 1H NMR (250 MHz, CDCl3): d =
1.11–1.23 (m, cPr-H, 1 H), 1.23–1.31 (m, cPr-H, 1 H), 1.35–1.44
(m, cPr-H, 1 H), 1.60–1.70 (m, cPr-H, 1 H), 1.92 (m, CH2, 2 H),
3.30–3.46 (m, CH2, NCH2, 4 H), 3.68–3.90 (m, NCH2CO, CHCl,
3 H), 4.41 (s, CH2Ph, 2 H), 7.24–7.35 (s, aryl-H, 5 H), 7.67 (s,
NH, 1 H). 13C NMR (62.9 MHz, CDCl3): d = 15.1 (CH2, cPr-C),
19.9 (CH2, cPr-C), 29.3 (CH2), 42.0 (C, cPr-C), 47.5 (CH2), 47.8
(CH2), 64.7 (CH), 67.7 (CH2), 73.0 (CH2), 127.7 (CH, aryl-C),
128.4 (CH, aryl-C), 138.1 (C, aryl-C), 168.1 (CO), 169.9 (CO).
MS (70 eV, DCI); m/z (%): 301 (25) [(M - Cl)+], 195 (30), 91 (100)
Bis(benzyloxycarbonyl)guanidine (6). A solution of N,N’-
bis(benzyloxycarbonyl)-S-methylisothiourea13 (2.06 g, 5.75 mmol)
in methanolic NH3 (30 mL, 140 mmol, 4.66 M) was stirred at 20 ◦C
for 3 h. After 30 min, the colorless precipitate was filtered off and
dried in vacuo to yield 1.15 g (61%) of 6 as a solid, mp 146–
◦
1
148 C. H NMR (250 MHz, CDCl3): d = 5.08 (s, CH2, 4 H),
7.24–7.41 (br s, aryl-H, 10 H), 8.23–9.60 (br s, NH2, NH, 3 H). 13
C
NMR (62.9 MHz, CDCl3): d = 67.3 (CH2,), 128.0 (CH, aryl-C),
128.3 (CH, aryl-C), 128.5 (CH, aryl-C), 135.6 (C, Cipso), 158.9 (C,
=
=
N C–N), 165.7 (C, C O). The additional spectroscopic data were
identical with those reported in the literature.9
+
[C7H7 ]. C17H21ClN2O3 (336.8): calcd. C 60.62, H 6.28, N 8.32;
Methyl 4-{3-[bis(benzyloxycarbonyl)guanidino]propyl}-7-tert-
butoxycarbonyl-5-oxo-4,7-diazaspiro[2.5]octane-8-carboxylate (7).
To a suspension of 6 (3.75 g, 11.5 mmol) and triphenylphosphine
(3.02 g, 11.5 mmol) in THF (100 mL) was added at 0 ◦C a
solution of 5-H (3.25 g, 9.50 mmol) in THF (100 mL), and then
slowly within 30 min, DIAD (2.23 mL, 11.5 mmol). The mixture
was stirred until the orange color had disappeared (5 h), water
(10 drops) was added, all volatiles were removed in vacuo and
the residue purified by chromatography on 100 g of silica gel
[3 ¥ 20 cm, pentane–Et2O 1 : 1 → 0 : 1, ninhydrin, Rf = 0.27
(pentane–Et2O 2 : 1)] to yield 5.47 g (87%) of 7 as a foam. IR
(KBr): n = 3393 cm-1 (NH), 3033, 2977, 2952, 1750 (CO), 1717
(CO), 1611 (CO), 1511, 1391, 1251, 1098, 1010, 910, 808, 733. 1H
NMR (300 MHz, CDCl3): d = 0.72–0.88 (m, cPr-H, 2 H), 0.98–
1.18 (m, cPr-H, 1 H), 1.28–1.34 (m, cPr-H, 1 H), 1.37 (s, tBu, 4.4
H), 1.45 (s, tBu, 4.6 H), 1.50–1.80 (m, CH2, 2 H), 2.86–3.12 (m,
1 H), 3.20–3.40 (m, 1 H), 3.50 (s, 1 H), 3.56 (s, 3 H), 3.72 (s,
1 H), 3.84–4.02 (m, 1 H), 4.26–4.52 (m, 1 H), 5.05–5.15 (m,
1 H), 5.09–5.11 (m, 2 H), 5.19–5.21 (m, 2 H), 7.28–7.50 (m, aryl-H,
10 H), 9.14–9.40 (m, 2 H). 13C NMR (125.7 MHz, CDCl3): d =
9.6 (CH2, cPr-C), 13.5 (CH2, cPr-C), 28.0 (CH2), 28.1 (CH3, tBu),
28.2 (CH3, tBu), 39.9 (C, cPr-C), 40.4 (CH2), 46.8 (C), 47.8 (CH2),
52.3 (CH3), 61.9 (CH), 66.9 (CH2), 69.0 (2 ¥ CH2), 128.01 (CH,
aryl-C), 128.03 (CH, aryl-C), 128.36 (CH, aryl-C), 128.42 (CH,
aryl-C), 128.7 (CH, aryl-C), 128.8 (CH, aryl-C), 134.5 (C, aryl-C),
136.7 (C, aryl-C), 155.7 (CN), 160.2 (CO), 163.8 (CO), 168.4 (CO),
172.1 (CO), 170.0 (CO). MS (ESI); m/z (%): 1325 (100) [2 M +
Na]+, 1303 (6) [2 M + H+], 674 (40) [M + Na+], 652 (26) [M + H+].
HRMS (ESI): calcd. for C33H42N5O9 [M + H+] 652.2983; found
652.2977.
found C 60.71, H 6.38, N 8.55%.
Methyl 4-(3-benzyloxypropyl)-7-tert-butoxycarbonyl-5-oxo-4,7-
diazaspiro[2.5]octane-8-carboxylate (5-Bn). To a solution of 3
(5.00 g, 15.0 mmol) in CH2Cl2 (100 mL) were added at 20 ◦C
pyridine (1.34 mL, 16.5 mmol) and Boc2O (3.45 g, 15.8 mmol),
and the mixture was stirred at this temperature for 6 h. Extraction
with cold (0 ◦C) 1 N HCl (30 mL), water (30 mL) and sat. NaHCO3
solution (30 mL), drying of the organic phase over Na2SO4,
removal of all volatiles in vacuo and chromatographic purification
of the residue on 40 g of silica gel (3 ¥ 20 cm, Et2O, Rf = 0.49)
yielded 4.87 g (75%) of 5-Bn as an oil. IR (film): n = 3327 cm-1,
3088, 3063, 2930, 2830, 1750 (CO), 1682 (CO), 894, 741. 1H NMR
(250 MHz, CDCl3): d = 0.70–0.85 (m, cPr-H, 1 H), 0.90–1.00 (m,
cPr-H, 1 H), 1.20–1.35 (m, cPr-H, 2 H), 1.37 (s, tBu, 4.3 H), 1.45 (s,
tBu, 4.7 H), 1.50–1.70 (m, 1 H), 2.75–3.00 (m, 1 H), 3.35–3.40 (m,
2 H), 3.61 (s, CH2N, 2 H), 3.70 (s, CH3O, 3 H), 3.83 (s, 1 H), 3.90–
4.08 (m, 1 H), 4.33–4.55 (m, 1 H), 4.45 (s, 2 H), 7.27–7.38 (m, 5 H).
13C NMR (62.9 MHz, CDCl3): d = 9.6 (CH2, cPr-C), 13.1 (CH2,
cPr-C), 28.1 (CH3, tBu), 28.2 (CH3, tBu), 28.9 (CH2), 39.1 (CH2),
40.1 (C, cPr-C), 47.8 (CH2), 52.3 (CH3), 61.8 (C), 63.3 (CH), 67.5
(CH2), 73.0 (CH2), 127.6 (CH), 128.3 (CH), 138.2 (CH), 163.2 (C),
168.4 (CO), 169.0 (CO), 170.1 (CO), 170.5 (CO). MS (ESI); m/z
(%): 887 (100) [2 M + Na+], 455 (40) [M + Na+]. HRMS (ESI):
calcd. for C23H33N2O6 [M + H+] 433.2339; found 433.2333.
Methyl 4-(3-hydroxypropyl)-7-tert-butoxycarbonyl-5-oxo-4,7-
diazaspiro[2.5]octane-8-carboxylate (5-H). To
genated suspension of Pd/C (500 mg, 10% Pd/C, ~5 mol%) in
MeOH (100 mL) was added a solution of 5-Bn (4.45 g, 10.3 mmol)
a
prehydro-
◦
in MeOH (100 mL), and the suspension was stirred at 20 C for
R
12 h. Filtration over Celiteꢀ, washing of the solids with MeOH
Methyl 4-{3-[bis(benzyloxycarbonyl)guanidino]propyl}-5-oxo-
4,7-diazaspiro[2.5]octane-8-carboxylate (8). Gaseous HCl was
bubbled through a solution of 7 (3.76 g, 5.77 mmol) in MeOH
(100 mL) kept at 0 ◦C, and the mixture was stirred at 20 ◦C for 2 h.
All volatiles were removed in vacuo, the residue was dissolved in
CH2Cl2 (100 mL), triethylamine (1.60 ml, 11.5 mmol) was added,
and the suspension was stirred for 3 h. Removal of all volatiles in
vacuo, re-uptake in THF (100 mL), collection of the precipitate
on a filter, washing with THF (2 ¥ 50 mL), removal of the solvent
from the filtrates in vacuo and chromatographic purification of the
residue on 40 g of silica gel (3 ¥ 20 cm, CH2Cl2–MeOH 10 : 1,
ninhydrin, Rf = 0.47) yielded 3.12 g (98%) of 8 as an oil. IR (film):
n = 3390 cm-1 (NH), 3033, 2952, 1721 (CO), 1650 (CO), 1611,
1511, 1437, 1409, 1379, 1255, 1102, 1007, 911, 807, 733. 1H NMR
(250 MHz, CDCl3): d = 0.72–0.87 (m, cPr-H, 2 H), 1.03–1.15
(m, cPr-H, 1 H), 1.20–1.30 (m, cPr-H, 1 H), 1.50–1.70 (m, 2 H),
2.82–3.05 (m, 1 H), 3.07 (s, 1 H), 3.25–3.35 (m, 2 H), 3.46–3.54
(2 ¥ 50 mL), and removal of the volatiles from the filtrate in vacuo
yielded 3.45 g (98%) of 5-H as an oil (diethyl ether, Rf = 0.10,
ninhydrin). IR (KBr): n = 3433 cm-1, 3095, 2952, 2933, 1749
1
(CO), 1697 (CO), 1394, 1205, 1168, 1008, 951, 745. H NMR
(300 MHz, 125 ◦C, C2D2Cl4): d = 0.60–0.80 (m, cPr-H, 1 H),
0.85–0.97 (m, cPr-H, 1 H), 1.20–1.30 (m, cPr-H, 1 H), 1.30 (s, tBu,
4.5 H), 1.36–1.44 (m, cPr-H, 1 H), 1.38 (s, tBu, 4.5 H), 1.62–1.80
(m, CH2, 2 H), 2.85–3.15 (m, 1 H), 3.33 (s, CH2, 2 H) 3.45 (m,
CH2, 2 H), 3.54 (s, 1 H), 3.64 (s, OCH3, 3 H), 3.86–4.10 (m, 1 H),
4.20–4.45 (m, 1 H). 13C NMR (62.9 MHz, 125 ◦C, C2D2Cl4): d =
9.7 (CH2, cPr-C), 12.8 (CH2, cPr-C), 27.96 (CH3, tBu), 28.1 (CH3,
tBu), 31.3 (CH2), 38.3 (CH2), 39.3 (C, cPr-C), 47.4 (CH2), 52.5
(OCH3), 57.9 (CH2), 61.1 (CH), 169.8 (CO), 170.0 (CO), 170.6
(CO). MS (70 eV, EI); m/z (%): 342 (1) [M+], 286 (14), 227 (17),
183 (8), 57 (100), 41 (24). C16H26N2O6 (342.4): calcd. C 56.13, H
7.65, N 8.18; found C 56.08, H 7.61, N 8.14%.
3820 | Org. Biomol. Chem., 2008, 6, 3816–3822
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(m, 2 H), 3.59 (s, OCH3, 3 H), 3.60–3.92 (m, 2 H), 5.08–5.12 (m,
CH2Ph, 2 H), 5.21 (s, CH2Ph, 2 H), 7.28–7.40 (m, aryl-H, 10 H),
9.18–9.42 (m, NH, 2 H). 13C NMR (75.5 MHz, CDCl3, DEPT):
d = 8.64 (CH2, cPr-C), 15.4 (CH2, cPr-C), 28.2 (CH2), 39.5 (CH2),
40.3 (C, cPr-C), 42.5 (CH2, NCH2), 48.0 (CH2, OCH2), 52.4 (CH3,
CO2CH3), 62.8 (CH), 66.8 (CH2Ph), 68.9 (CH2Ph), 127.8 (CH,
aryl-C), 128.0 (CH, aryl-C), 128.30 (CH, aryl-C), 128.34 (CH,
HRMS (ESI): calcd. for C38H41N6O9 [M + H+] 725.2935; found
725.2928.
Spiro[cyclopropane[1,9]8-(3-guanidinopropyl)-1,4,7-trioxo-octa-
hydro[2H]pyrazino[1,2-a]pyrazin-3-yl]acetic acid (cis- and trans-
11). A suspension of 10 (1.34 g, 2.63 mmol) and Pd/C (335 mg,
10% Pd/C, ~5 mol%) in MeOH (20 mL) was stirred under an
atmosphere of H2 (balloon) at 20 ◦C for 12 h and the catalyst
=
aryl-C), 128.7 (CH, aryl-C), 134.5 (C), 136.7 (C), 155.6 (C N),
R
removed by filtration over Celiteꢀ. Removal of the solvent in vacuo
160.2 (C, CO2Bn), 163.7 (C, CO2Bn), 172.6 (C, NCO), 172.8 (C,
CO). MS (70 eV, DCI); m/z (%): 552 (22) [M + H+], 342 (15), 296
(17), 279 (100), 222 (18), 208 (28), 126 (45), 104 (14). C28H33N5O7
(551.6): calcd C 60.97, H 6.03, N 12.70; found C 61.15, H 6.16, N
12.92%.
yielded 737 mg (99%) of cis- and trans-11 as a foam (mixture of
two diastereomers 1.1 : 1 [according to NMR]). IR (KBr): n =
3326 cm-1 (NH), 3187, 2929, 2850, 1718 (CO), 1668 (CO), 1626
1
(CO), 1435, 1224. H NMR (CD3OD, 300 MHz): d = 0.99–1.40
(m, 2 cPr-H, 4 H), 1.58–1.73 (m, CH2, 4 H), 2.62–3.00 (m, CH2,
2 H), 3.03–3.19 (m, 1 H), 3.34–3.40 (m, 1 H), 3.54–3.88 (m, 1 H),
3.97–4.10 (m, 1 H), 4.14–4.50 (m, 1 H), 4.66–4.88 (m, 1 H). 13C
NMR (125.7 MHz, CD3OD): d = 9.9 (CH2, cPr-C), 10.2 (CH2,
cPr-C), 13.5 (CH2, cPr-C), 14.9 (CH2, cPr-C), 26.1 (CH2), 26.4
(CH2), 26.7 (CH2), 27.0 (CH2), 29.0 (CH2), 29.1 (CH2), 30.1 (CH2),
30.3 (CH2), 39.0 (CH2), 39.9 (CH2), 40.4 (CH2), 40.5 (CH2), 46.4
(C, cPr-C), 46.9 (C, cPr-C), 52.4 (CH), 52.6 (CH), 54.1 (CH), 54.4
(CH), 158.60 (C), 158.63 (C), 167.0 (CO), 168.8 (CO), 169.8 (CO),
171.3 (CO), 172.8 (CO), 175.9 (CO). MS (ESI); m/z (%), positive
peaks: 1099 (12) [3 M + H+], 733 (22) [2 M + H+], 367 (100) [M +
H+]; negative peaks: 365 (100) [(M - H)-]. C15H22N6O5 (366.4):
calcd. C 49.17, H 6.05, N 22.94; found C 50.43, H 6.21, N 23.55%.
Benzyl spiro[cyclopropane[1,9](8-{3-[bis(benzyloxycarbonyl)-
guanidino]propyl}-1,4,7-trioxooctahydro[2H]pyrazino[1,2-a]pyra-
zin-3-yl)]acetate (10). To a solution of 8 (4.45 g, 8.07 mmol) and
Boc-(S)-Asp-(4-OBn)-OH (3.39 g, 10.5 mmol) in CH2Cl2 (30 mL)
were added at 0 ◦C 2,4,6-collidine (1.40 mL, 10.5 mmol) and DCC
(2.01 g, 9.70 mmol), and the mixture was stirred with rewarming
◦
to 20 C for 8 h. Addition of CH2Cl2 (80 mL), extraction of the
◦
organic phase with cold (0 C) 1 N HCl (30 mL), water (30 mL)
and sat. NaHCO3 solution (2 ¥ 30 mL), drying of the organic
phase over Na2SO4, removal of the solvent in vacuo and filtration
of the residue over 50 g of silica gel (3 ¥ 15 cm, CH2Cl2–MeOH
10 : 1, ninhydrin, Rf = 0.75) yielded 9 with a dipeptide fragment.
This compound was dissolved in CH2Cl2 (10 mL),◦and methanolic
HCl (50.0 mL, 75 mmol, 1.5 M) was added at 0 C, the mixture
was stirred for 8 h, and all volatiles were removed in vacuo. The
residue was taken up in CH2Cl2 (200 mL), the solution neutralized
with sat. NaHCO3 solution, the phases were separated, the aq.
phase extracted with CH2Cl2 (3 ¥ 50 mL), and the combined
organic phases were dried over Na2SO4. Removal of the solvent,
suspension of the residue in toluene (10 mL), heating at 90 ◦C
for 24 h, removal of the solvent in vacuo and chromatographic
purification of the residue on 40 g of silica gel (3 ¥ 30 cm, CH2Cl2–
MeOH 50 : 1; MOPS, Rf = 0.27) yielded 3.35 g (57%) of 10 as
a foam [unseparable mixture of two diastereomers, ratio 1 : 1
(according to NMR)]. IR (KBr): n = 3333 cm-1 (NH), 3032,
2923, 2835, 1752 (CO), 1710 (CO), 1668 (CO). 1H NMR (C2D2Cl4,
Acknowledgements
This work was supported by BayerCropScience AG as well as
Celanese AG throughgenerous gifts ofchemicals. M. L. is indebted
to the German Merit Foundation (Studienstiftung des deutschen
Volkes) for a student (2000–2002) and a doctoral fellowship (2002–
2004). V. S. K. is grateful to the Degussa-Stiftung (Degussa AG)
for a graduate student fellowship.
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66.7 (CH2), 67.8 (CH2), 68.70 (CH2), 68.72 (CH2), 127.1 (CH,
aryl-C), 127.2 (CH, aryl-C), 127.25 (CH, aryl-C), 127.27 (CH,
aryl-C), 127.7 (CH, aryl-C), 127.8 (CH, aryl-C), 127.97 (CH, aryl-
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C), 128.3 (CH, aryl-C), 128.4 (CH, aryl-C), 128.6 (CH, aryl-C),
129.18 (CH, aryl-C), 129.22 (CH, aryl-C), 133.8 (CH, aryl-C),
135.1 (C, aryl-C), 135.2 (C, aryl-C), 138.38 (C, aryl-C), 138.41 (C,
aryl-C), 162.9 (CO), 163.4 (CO), 164.2 (CO), 164.9 (CO), 166.9
(CO), 168.2 (CO), 169.2 (CO), 169.4 (CO). MS (EI, 70 eV); m/z
(%): 725.0 (5) [M+], 505.0 (12), 414.0 (16), 399.0 (8), 91 (100).
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3822 | Org. Biomol. Chem., 2008, 6, 3816–3822
This journal is
The Royal Society of Chemistry 2008
©