Synthesis of tetrasubstituted symmetrical pyrazines from β-keto γ-amino esters: A mild strategy for self-dimerization of peptides

Article abstract of DOI:10.1002/ejoc.201403237

A facile synthesis of highly symmetrical tetrasubstituted pyrazines through simple aerial oxidation of β-keto γ-amino esters is reported. The scope of the reaction was examined by use of various amino acid side-chain functional groups andpeptides. The mil

Full text of DOI:10.1002/ejoc.201403237

Job/Unit: O43237
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FULL PAPER
DOI: 10.1002/ejoc.201403237
Synthesis of Tetrasubstituted Symmetrical Pyrazines from β-Keto γ-Amino
Esters: A Mild Strategy for Self-Dimerization of Peptides
Mothukuri Ganesh Kumar,[a] Varsha J. Thombare,[a] Rupal D. Bhaisare,[a] Anindita Adak,[a]
and Hosahudya N. Gopi*[a]
Keywords: Amino acids / Peptides / Oxidation / Cyclization / Nitrogen heterocycles
A facile synthesis of highly symmetrical tetrasubstituted pyr-
azines through simple aerial oxidation of β-keto γ-amino es-
ters is reported. The scope of the reaction was examined by
use of various amino acid side-chain functional groups and
peptides. The mild and efficient transformation of β-keto γ-
amino esters into pyrazines may serve as an attractive strat-
egy for self-dimerization of peptides.
Introduction
Pyrazines are a class of heterocyclic aromatic compounds
found ubiquitously in nature. Many of these natural pyr-
azine derivatives and their synthetic counterparts have
shown excellent biological activities, including anticancer,[1]
antiviral,[2] and antituberculosis[3] properties etc. Further-
more, the roles of tetrasubstituted pyrazines as sex phero-
mones in female insects[4] and as alarm pheromones in
ponerine ants[5] have been documented. Some sexually de-
ceptive orchid species produce pyrazines as semiochemicals
to attract male wasps as pollinators.[4] Many bacterial
strains use various substituted pyrazines as a sole source
of carbon and energy.[6] Besides their biological activities,
pyrazines have also been extensively used in the food and
perfume industries, due to their taste and aromas.[7] In ad-
dition, pyrazine heterocycles have also been finding applica-
tions in materials science.[8] Some naturally occurring pyr-
azine derivatives – tetramethylpyrazine,[9] a representative
barrenazine,[10] and cephalostatin[1a] – are shown in Fig-
ure 1. The majority of the natural pyrazines are derived
from amino acids.[11] Recently, Kutonovas et al. delineated
the possible catabolic pathway for the degradation of pyr-
azines.[12]
Figure 1. Examples of naturally occurring pyrazine deriva-
tives.[9,10,1a]
these, various other protocols involving α-haloenol acet-
ates,[16] nitroepoxides,[17] ruthenium-catalyzed dehydro-
genative coupling thermal treatment of azirine phosphona-
tes and β-amino alcohols,[18] tosylketoximes,[19] and con-
densation of amino diketones with hydroxylamines[20] etc.
have been developed. Recently, Sperry and colleagues exam-
ined the self-condensation of various amino aldehydes to
produce substituted pyrazines; the study revealed the im-
portance of Cbz protection over Boc and Fmoc protection
for the amino aldehydes.[21] In addition, Rojas et al. re-
ported the self-condensation of α-amino aldehydes to pyr-
azines in a three-step one-pot procedure.[22] In an accidental
discovery, Pettit et al. also observed the formation of tetra-
substituted pyrazine from the methyl ester of β-keto γ-leu-
cine as a byproduct in the total synthesis of respirantin.[23]
Recently, we reported the facile synthesis of β-keto γ-
amino esters and their utility in the synthesis of β-hydroxy-
γ-amino acids (statines) as well as fluorescent coumarin
As a consequence of the growing demand for substituted
pyrazine derivatives in the pharma, food, and perfume in-
dustries, various synthetic protocols have been developed.
Generally, pyrazines are synthesized variously by condensa-
tion of 1,2-diamines with 1,2-dicarbonyl compounds,[13]
self-condensation of α-amino ketones,[11d,14] or through re-
actions between α-haloketones and ammonia.[15] Besides
[a] Department of Chemistry, Indian Institute of Science
Education and Research,
Dr. Homi Bhabha Road, Pune-411 008, India
E-mail: hn.gopi@iiserpune.ac.in
www.iiserpune.ac.in
Supporting information for this article is available on the
WWW under http://dx.doi.org/10.1002/ejoc.201403237.
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H. N. Gopi et al.
FULL PAPER
amino acids.[24] The wide applications of pyrazine deriva- further examined whether Boc-phenylalanal can undergo
tives motivated us to investigate the synthesis of tetrasubsti- similar self-condensation and subsequent aromatization af-
tuted pyrazines from β-keto γ-amino esters (Scheme 1). ter Boc group removal. Unlike the β-keto γ-amino ester, the
Here we report the mild synthesis of highly symmetrical α-amino aldehyde did not undergo cyclization and subse-
tetrasubstituted pyrazines from β-keto γ-amino esters quent aromatization.[21]
through a simple aerial oxidation. Through the utilization
of this mild method for aromatization of β-keto esters, a
new strategy for the self-dimerization of peptides has been
developed.
Figure 2. X-ray structures of 2a and 2b.
Scheme 1. Tetrasubstituted pyrazines from β-keto γ-amino esters.
Inspired by the smooth aromatization of the unprotected
β-keto ester of 1a and the highly symmetric nature of
pyrazine 2a, we subjected other hydrophobic γ-amino β-
keto esters 1b–1f to the pyrazine synthesis. Although the β-
keto esters 1e and 1f, derived from the β-branched amino
Results and Discussion
In order to establish whether β-keto γ-amino esters can acids Val and Ile, respectively, did not react, the others – 1b
undergo self-condensation and subsequent aerial oxidation to 1d – gave the corresponding pyrazine products 2b–2d in
to give pyrazines, we synthesized ethyl esters of various β- moderate to good yields. We did not isolate pyrazine deriva-
keto γ-amino acids, starting from N-Boc-protected amino tives from 1e or 1f. In addition to 2a, 2b also gave single
aldehydes and ethyl diazoacetate in the presence of an- crystals, and its X-ray structure is also shown in Figure 2.
hydrous tin chloride.[24a,25] A schematic representation of
The scope of the reaction was investigated further with γ-
the reaction is shown in Scheme 2. All ethyl esters of β-keto amino β-keto esters 1g–1j, containing side-chain functional
γ-amino esters were isolated in good yields after column groups. We observed no interference of ester, thioether, or
purification. To achieve the formation of pyrazine deriva- amide functional groups in the self-condensation and sub-
tives, we initially subjected β-keto γ-phenylalanine ethyl es- sequent aromatization. All pyrazine products 2g–2j were
ter 1a to the self-condensation reaction in the open air after isolated in moderate to good yields and are listed in Table 1.
the removal of the N-Boc group. The TFA salt of the γ-
Inspired by the mild and biocompatible self-condensa-
amino β-keto ethyl ester was dissolved in THF and neutral- tion and the aromatization of β-keto esters containing
ized with DIPEA (Scheme 3). The transformation of the amine and carboxylic acid side-chains, we sought to investi-
γ-amino β-keto ester into tetrasubstituted pyrazine 2a was gate whether this strategy can be further extended to pept-
achieved after the reaction mixture had been stirred over- ides. In addition, nature selectively utilizes α-amino ketones
night. Pure 2a was isolated in good yield (75%) after col- in the synthesis of symmetrical cephalostatins (Figure 1)
umn chromatography. Further, the pure 2a gave X-ray-qual- and ritterazines.[1c,26] We anticipated that, as in the case of
ity single crystals, and its structure is shown in Figure 2. We cephalostatins, it should also be possible to “self-staple”
Scheme 2. Synthesis of N-Boc β-keto γ-amino esters starting from amino acids.
Scheme 3. Schematic representation of the synthesis of tetrasubstituted pyrazines starting from N-Boc β-keto γ-amino esters.
2
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Self-Dimerization of Peptides
Table 1. List of pyrazines synthesized from β-keto γ-amino esters.
peptides through aerial oxidation, if the side-chain amino
groups of 1h and 1i and the carboxylic acid side-chain of 1j
were coupled with peptides.
To verify this, we designed a new strategy for the synthe-
sis of peptide β-keto esters, as shown in Scheme 4. The free
amine in the Boc-Dap side-chain was directly coupled to
the N-hydroxysuccinimide ester of N-Cbz-Val to give a di-
peptide carboxylic acid. The free carboxylic acid was con-
verted to afford the corresponding dipeptide aldehyde
through LAH reduction of the corresponding Weinreb
amide. The dipeptide aldehyde P1 was transformed into β-
keto ester P2 by treatment with ethyl diazoacetate in the
presence of tin(II) chloride. The pure dipeptide β-keto ester
P2 was isolated in good yield after the column purification.
The Boc group of the dipeptide P2 was selectively removed
with TFA, and the TFA salt P3 was neutralized with DI-
PEA as base. The free dipeptide β-keto ester was stirred
overnight in open air. Conversion of dipeptide β-keto ester
into pyrazine C1 was observed within 12 h. The pure pept-
ide pyrazine derivative C1 was isolated after column
chromatography in 55% yield.
We further extended this strategy to a tripeptide. A sche-
matic representation of the synthesis of a tripeptide β-keto
ester and the subsequent transformation of this β-keto ester
into a pyrazine is shown in Scheme 5. The tripeptide alde-
hyde P4 obtained after treatment of a Weinreb amide with
LAH was converted into the corresponding β-keto ester P5
by treatment with ethyl diazoacetate in the presence of an-
hydrous tin(II) chloride in CH2Cl2.
The tripeptide β-keto ester was subjected to the same
aerial oxidation after removal of the Boc group as described
earlier. The reaction was monitored by MALDI-TOF mass
spectrometry, and the transformation of the β-keto ester
into the corresponding pyrazine C2 was observed within
12 h. We further noticed that an additional supply of oxy-
gen from a balloon increased the rate of formation of pyr-
azine products from peptides, but it is not essential. The
tripeptide pyrazine derivative C2 was isolated in 50% yield
after reversed-phase HPLC purification on a C18 column
with a methanol/H2O gradient system. These results clearly
Scheme 4. Synthesis of a dipeptide β-keto ester and subsequent pyrazine synthesis.
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H. N. Gopi et al.
FULL PAPER
are reported in Hz. Mass spectra were recorded by MALDI TOF/
TOF and Electron Spray Ionization (ESI).
General Procedure for the Synthesis of N-Protected γ-Amino β-Keto
Esters: The appropriate N-protected amino aldehyde (2.0 mmol)
was dissolved in CH2Cl2 (15 mL) at room temperature, and then
tin(II) chloride (0.0756 g, 20 mol-%) was added, followed by ethyl
diazoacetate (0.239 g, 2.1 mmol). Immediate gas evolution was ob-
served. The reaction mixture was stirred and the progress of the
reaction was monitored by TLC. After completion of the reaction
(ca. 30 min), it was quenched with HCl (0.5 n, 10 mL) and the reac-
tion mixture was extracted with CH2Cl2 (30 mLϫ 3). The com-
bined organic layer was washed with brine (20 mL), dried with an-
hydrous sodium sulfate, and concentrated under reduced pressure
to afford a greenish oily crude product, which was purified by silica
gel column chromatography.
Ethyl (S)-4-[(tert-Butoxycarbonyl)amino]-3-oxo-5-phenylpentanoate
(1a):[24a] White crystals (0.521 g, 78%). [α]2D5 = –54.5 (c = 0.6,
MeOH). 1H NMR (400 MHz, CDCl3): δ = 12.16 (s, 1 H, enolic
17%), 7.24–7.15 (m, 5 H), 5.03–5.01 (d, J = 7.3 Hz, 1 H), 4.57–
4.52 (q, J = 6.4 Hz, 1 H), 4.18–4.12 (q, J = 7.2 Hz, 2 H), 3.51–3.40
(dd, J = 16, 11.4 Hz, 2 H), 3.15–2.95 (m, 2 H), 1.38 (s, 9 H), 1.26–
1.22 (t, J = 7.2 Hz, 3 H) ppm. 13C NMR (100 MHz, CDCl3): δ =
201.9, 166.8, 155.1, 136.0, 129.24, 128.6, 127.0, 80.2, 61.4, 60.4,
46.8,36.8, 28.2, 14.0 ppm. HRMS: calcd. for C18H25NO5 [M +
Na]+ 358.1630; found 358.1633.
Scheme 5. Transformation of a tripeptide β-keto ester into a pyr-
azine.
Ethyl (S)-4-[(tert-Butoxycarbonyl)amino]-3-oxopentanoate (1b):[24a]
Light yellow liquid (0.398 g, 76%). [α]2D5 = –35.69 (c = 1, MeOH).
1H NMR (500 MHz, CDCl3): δ = 12.13 (s, 1 H), 5.17 (s, 1 H),
4.43–4.37 (m, 1 H), 4.24–4.20 (q, J = 7 Hz, 2 H), 3.62–3.54 (dd, J
= 14.5, 10.5 Hz, 2 H), 1.46 (s, 9 H), 1.38–1.36 (d, J = 6.5 Hz, 3 H),
1.31–1.28 (t, J = 7 Hz, 3 H) ppm. 13C NMR (125 MHz, CDCl3): δ
= 202.5,166.9, 155.1, 80.1, 61.5, 55.4, 45.9, 28.3, 17.1, 14.1 ppm.
HRMS: calcd. for C12H21NO5 [M + Na]+ 282.1317; found
282.1317.
suggest that, like the γ-amino-β-keto esters, peptide β-keto
esters can also undergo self-condensation and subsequent
aromatization to give pyrazine derivatives. The strategy de-
scribed here provides a unique means for self-dimerization
of peptides under very mild conditions.
Conclusions
Ethyl (S)-4-(tert-Butoxycarbonylamino)-6-methyl-3-oxoheptanoate
(1c):[24a] Light yellowish liquid (0.476 g, 79%). [α]2D5 = –53.70 (c =
Overall, we have demonstrated the conversion of ethyl
esters of β-keto γ-amino acids into highly symmetrical
tetrasubstituted pyrazines under very mild conditions. With
the exceptions of the γ-amino β-keto esters derived from
the sterically hindered amino acids valine and isoleucine, all
other β-keto esters gave the corresponding tetrasubstituted
pyrazines in moderate to good yields. In addition to the β-
keto amino esters, we also demonstrated the synthesis of
peptide β-keto esters and their subsequent transformation
into highly symmetrical tetrasubstituted pyrazines. We are
currently extending this new peptide “self-stapling” strategy
to solid-phase synthesis. The mild and biocompatible aro-
matization of β-keto esters of amino acids and peptides re-
ported here may find applications in medicinal chemistry
and materials science.
1
1, MeOH). H NMR (500 MHz, CDCl3): δ = 12.10 (s, 1 H), 4.96–
4.94 (d, J = 9.5 Hz, 1 H), 4.40–4.36 (m,1 H), 4.24–4.20 (q, J =
7 Hz, 2 H), 3.63–3.53 (dd, J = 16.0, 18.5 Hz, 2 H), 1.74–1.67 (m,
3 H), 1.46 (s, 9 H), 1.32–1.29 (t, J = 7 Hz, 3 H), 0.97 (b, s, 6
H) ppm. 13C NMR (125 MHz, CDCl3): δ = 203.02, 167.07, 155.55,
80.15, 61.51, 58.19, 46.35, 39.90, 28.31, 24.83, 23.28, 21.59,
14.12 ppm. HRMS: calcd. for C15H27NO5 [M + Na]+ 324.1786;
found 324.1784.
Ethyl (S)-4-[(tert-Butoxycarbonyl)amino]-3-oxo-4-phenylbutanoate
(1d): Yellowish liquid (0.46 g, 72%). [α]2D5 = –4.0 (c = 1, MeOH).
1H NMR (400 MHz, CDCl3): δ = 7.38–7.29 (m, 5 H), 5.86 (d, J =
4 Hz, 1 H), 5.44 (d, J = 8 Hz), 4.09 (q, J = 8 Hz, 2 H), 3.39 (dd, J
= 16, 60 Hz, 2 H), 1.39 (s, 9 H), 1.19 (t, J = 8 Hz, 2 H) ppm. 13C
NMR (100 MHz, CDCl3): δ = 198.5, 166.3, 154.8, 135.9, 129.4,
128.9, 128.2, 80.2, 64.4, 61.6, 46.3, 28.3, 14.3 ppm. HRMS: calcd.
for C17H23NO5 [M + Na]+ 344.1468; found 344.1469.
Ethyl (S)-4-(tert-Butoxycarbonylamino)-5-methyl-3-oxohexanoate
(1e):[24a] Colorless liquid (0.48 g, 84%). [α]2D5= –32.64 (c = 1,
MeOH). 1H NMR (500 MHz, CDCl3): δ = 12.11 (s, 1 H, enolic
form, 6.5%), 5.06 (br. s, 1 H), 4.35–4.32 (m, 1 H), 4.22–4.18 (q, J
= 7 Hz, 2 H), 3.57–3.50 (dd, J = 15.5, 3 Hz, 2 H), 2.27–2.23 (m, 1
H), 1.44 (s, 9 H), 1.29–1.26 (t, J = 7 Hz, 3 H), 1.02–0.82 (m, 6
H) ppm. 13C NMR (125 MHz, CDCl3): δ = 202.23, 166.75, 155.86,
80.03, 64.38, 61.55, 47.14, 29.56, 28.31, 19.84, 16.67, 14.10 ppm.
HRMS: calcd. for C14H25NO5 [M + Na]+ 310.1630; found
310.1620.
Experimental Section
General Experimental Details: Solvent THF was dried with sodium
and immediately distilled prior to use. Dichloromethane (CH2Cl2)
was distilled prior to use. Column chromatography was performed
with silica gel (120–200 mesh). 1H NMR and 13C NMR spectra
were recorded at 400 MHz and 100 MHz respectively, with use of
residual solvent as internal standards (CDCl3). Chemical shifts (δ)
are reported in parts per million (ppm) and coupling constants (J)
4
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Self-Dimerization of Peptides
Ethyl (4S,5R)-4-[(tert-Butoxycarbonyl)amino]-5-methyl-3-oxohept- Diethyl 2,2Ј-(3,6-Dibenzylpyrazine-2,5-diyl)diacetate (2a): White
anoate (1f):[24a] Colorless liquid (0.453 g, 76%). [α]2D5 = –25.08 (c =
1, MeOH). H NMR (400 MHz, CDCl3): δ = 12.09 (s, 1 H, enolic
7.5%), 5.03–5.01 (d, J = 8.24 Hz, 1 H), 4.31–4.28 (m, 1 H), 4.19–
4.13 (q, J = 6.88 Hz, 2 H), 3.51 (s, 2 H), 1.97–1.90 (m, 1 H), 1.63–
crystals (0.32 g, 79%), m.p. 95–96 °C. 1H NMR (400 MHz,
CDCl3): δ = 7.27–7.14 (m, 10 H), 4.18 (s, 4 H), 4.07 (q, J = 7.1 Hz,
4 H), 3.81 (s, 4 H), 1.19 (t, J = 7.1 Hz, 6 H) ppm. 13C NMR
(100 MHz, CDCl3): δ = 169.9, 151.8, 147.0, 137.9, 128.9, 128.7,
1
1.57 (m, 2 H), 1.41 (s, 9 H), 1.27–1.23 (t, J = 7.2 Hz, 3 H), 0.97–0.95 126.7, 61.2, 40.9, 40.8, 28.4, 14.2 ppm. UV: λmax = 204, 281 nm.
(dd, J = 3.64, 3.24 Hz, 3 H), 0.89–0.85 (t, J = 6.9 Hz, 3 H) ppm. 13
C
Fluorescence: λex = 281 nm, λem = 445 nm. HRMS: calcd. for
NMR (100 MHz, CDCl3): δ = 202.38, 166.72, 155.74, 79.99, 64.26,
61.47, 47.28, 36.28, 28.25, 24.00, 16.02, 14.05, 11.60 ppm. MALDI
TOF/TOF-: calcd. for C15H27NO5 [M + Na]+ 324.1787; found
324.1709.
C26H28N2O4 [M + H]+ 433.2127; found 433.2194.
Diethyl 2,2Ј-(3,6-Dimethylpyrazine-2,5-diyl)diacetate (2b): Colorless
1
solid (0.174 g, 67%), m.p. 94–95 °C. H NMR (400 MHz, CDCl3):
δ = 4.17 (q, J = 7.1 Hz, 4 H), 3.82 (s, 4 H), 2.48 (s, 6 H), 1.24 (s, 6
H) ppm. 13C NMR (100 MHz, CDCl3): δ = 170.03, 149.54, 146.47,
Ethyl (S)-4-[(tert-Butoxycarbonyl)amino]-7-(methylthio)-3-oxohept-
anoate (1g): Yellow viscous liquid (0.4 g, 64%). [α]2D5 = –3.75 (c =
61.34, 41.33, 21.20, 14.31 ppm. UV (λmax
) 278 nm, 211 nm.
1
0.8, MeOH). H NMR (400 MHz, CDCl3): δ = 5.30 (d, J = 8 Hz,
Fluorescence: λex = 278 nm, λem = 435 nm. HRMS: calcd. for
C14H20N2O4 [M + Na]+ 281.1501; found 281.1503.
1 H), 4.44–4.39 (m, 1 H), 4.14 (q, J = 8 Hz, 2 H), 3.59–3.50 (m, 2
H), 2.49 (m, 2 H), 2.13 (m, 1 H), 2.04 (s, 3 H), 1.83–1.77 (m, 1 H),
1.39 (s, 9 H), 1.23 (t, J = 8 Hz, 3 H) ppm. 13C NMR (100 MHz,
CDCl3): δ = 202.0,167.0, 155.5, 80.3, 61.5, 60.4, 58.3, 46.3, 30.2,
30.0, 28.3, 15.4, 14.1 ppm. HRMS: calcd. for C15H27NO5S [M +
Na]+ 342.1350; found 342.1357.
Diethyl 2,2Ј-(3,6-Diisobutylpyrazine-2,5-diyl)diacetate (2c): Yellow
1
liquid (0.2 g, 60%). H NMR (400 MHz, CDCl3): δ = 4.16 (q, J =
7.1 Hz, 4 H), 3.85 (s, 4 H), 2.61 (d, J = 7.3 Hz, 4 H), 2.17–2.10 (m,
2 H), 1.23 (t, J = 7.2 Hz, 6 H), 0.92 (d, J = 6.7 Hz, 12 H) ppm.
13C NMR (100 MHz, CDCl3): δ = 170.2, 151.6,146.2, 61.0, 42.6,
40.8, 28.2, 22.4, 14.1 ppm. UV: λmax = 215 nm, 267 nm. Fluores-
cence: λex = 267 nm, λem = 429 nm. HRMS (ESI): calcd. for
C20H32N2O4 [M + H]+ = 365.2240; found 365.2240.
Ethyl (S)-5-{[(Benzyloxy)carbonyl]amino}-4-[(tert-butoxycarbonyl)-
amino]-3-oxopentanoate (1h): Yellow liquid (0.54 g, 66%). [α]2D5
=
1
–3.00 (c = 1, MeOH). H NMR (400 MHz, CDCl3): δ = 7.30 (s, 5
H), 5.54 (s, 1 H), 5.05 (s, 2 H), 4.41 (s, 1 H), 4.14 (q, J = 8 Hz, 2
H), 3.68 (d, J = 8 Hz, 2 H), 3.58 (s, 2 H), 3.53 (d, J = 8 Hz, 2 H),
1.40 (s, 9 H), 1.23 (t, J = 4 Hz, 3 H) ppm. 13C NMR (100 MHz,
CDCl3): δ = 201.4,172.6, 167.2, 156.4,155.7, 136.0, 128.4, 128.0,
127.8, 80.2, 66.9, 62.0, 61.0, 60.4, 45.9, 39.7, 29.5, 28.1, 19.1,
13.9 ppm. HRMS: calcd. for C20H28N2O7 [M + Na]+ 431.1794;
found 431.1790.
Diethyl 2,2Ј-[(3,6-Diphenylpyrazine)-2,5-diyl]diacetate (2d): White
1
solid (0.24 g, 62%), m.p. 94–97 °C. H NMR (400 MHz, CDCl3):
δ = 7.62 (dd, J = 7.8, 1.7 Hz, 4 H), 7.52–7.42 (m, 6 H), 4.13 (q, J
= 7.1 Hz, 4 H), 3.99 (s, 4 H), 1.21 (t, J = 7.1 Hz, 6 H) ppm. 13C
NMR (100 MHz, CDCl3): δ = 169.3, 151.0, 146.3, 61.2, 40.7, 33.7,
32.5, 15.8, 14.2 ppm. UV: λmax = 250 nm, 292 nm. Fluorescence:
λex
= 292 nm, λem = 371 nm. HRMS (ESI): calcd. for
C18H28N2O4S2 [M + H]+ 405.1814; found 405.1815.
Ethyl (S)-4,8-Bis[(tert-butoxycarbonyl)amino]-3-oxooctanoate (1i):
Light yellow (0.66 g, 74%). [α]2D5 = –2.0 (c = 1, MeOH). H NMR
1
Diethyl 2,2Ј-{3,6-Bis[2-(methylthio)ethyl]pyrazine-2,5-diyl}diacetate
(400 MHz, CDCl3): δ = 7.34–7.27 (m, 5 H), 5.12 (m, 2 H), 4.82
(br., 1 H), 4.18 (q, J = 8 Hz, 2 H), 3.50 (m, 2 H), 3.18 (m, 2 H),
1.68–1.46 (m, 8 H), 1.42 (s, 9 H), 1.26 (t, J = 8 Hz, 3 H) ppm. 13C
NMR (100 MHz, CDCl3): δ = 202.4, 167.0, 156.6, 136.6, 128.5,
128.1, 80.2, 77.4, 76.8, 66.73 61.6, 46.2, 40.4, 30.4, 28.5, 22.25
14.16 ppm. HRMS: calcd. for C23H34N2O7 [M + Na]+ 473.2264;
found 473.2264.
1
(2g): White solid (0.252 g, 67%), m.p. 83–87. H NMR (400 MHz,
CDCl3): δ = 4.18 (q, J = 7.1 Hz, 1 H), 3.89 (s, 1 H), 3.07–3.00 (m,
1 H), 2.93–2.86 (m, 1 H), 2.12 (s, 2 H), 1.27 (t, J = 7.2 Hz, 2
H) ppm. 13C NMR (100 MHz, CDCl3): δ = 169.3, 151.0, 146.3,
61.2, 40.7, 33.7, 32.5, 15.8, 14.1 ppm. UV: λmax = 210 nm, 280 nm.
Fluorescence: λex = 280 nm, λem = 332 nm. HRMS (ESI): calcd.
for C18H28N2O4S2 [M + H]+ 401.1568; found 401.1564.
7-Benzyl (S)-1-Ethyl 4-[(tert-Butoxycarbonyl)amino]-3-oxoheptane-
dioate (1j): White solid (0.46 g, 57%). [α]2D5 = –2.0 (c = 1, MeOH).
1H NMR (400 MHz, CDCl3): δ = 7.38–7.32 (m, 5 H), 5.25 (d, J =
8 Hz, 1 H), 5.12 (s, 2 H), 4.42 (m, 1 H), 4.18 (q, J = 4 Hz, 2 H),
3.57 (m, 2 H), 2.46 (m, 2 H), 2.27 (m, 1 H), 1.85 (m, 1 H), 1.43 (s,
9 H), 1.26 (t, J = 8 Hz, 3 H) ppm. 13C NMR (100 MHz, CDCl3):
δ = 201.8, 172.8, 166.9, 155.6, 135.8, 128.7, 80.4, 66.7, 61.7, 59.0,
46.3, 30.0, 29.8, 28.4, 26.0, 14.2 ppm. HRMS: calcd. for
C28H29NO7 [M + Na]+ 430.1841; found 430.1840.
Diethyl 2,2Ј-[3,6-Bis({[(benzyloxy)carbonyl]amino}methyl)pyrazine-
2,5-diyl]diacetate (2h): Light yellow oil (0.36 g, 63%). 1H NMR
(400 MHz, CDCl3): δ = 7.35 (dd, J = 12.8, 4.3 Hz, 10 H), 6.03 (s,
4 H), 5.13 (s, 4 H), 4.52 (s, 4 H), 4.17 (q, J = 7.0 Hz, 4 H), 3.92 (s,
4 H), 1.26 (t, J = 7.1 Hz, 6 H) ppm. 13C NMR (100 MHz, CDCl3):
δ = 169.5, 156.4, 148.5, 145.8, 136.4, 128.7, 128.3, 67.1, 61.7, 42.7,
40.0, 14.2 ppm. UV: λmax = 216 nm, 276 nm. Fluorescence: λex
=
267 nm, λem = 441 nm. HRMS (ESI): calcd. for C30H34N4O8 [M +
H]+ 579.2455; found 579.2455; calcd. for [M + Na]+ 601.2274;
found 601.2287.
General Procedure for the Synthesis of Pyrazine Products: The ap-
propriate Boc-γ-amino β-keto ester (1 mmol) was dissolved in
CH2Cl2 (2 mL) and the solution was cooled in an ice bath. Then,
TFA (2 mL) was added slowly to this solution. After completion
of the reaction (ca. 30 min), TFA was removed from the reaction
mixture under reduced pressure. The TFA salt of the γ-amino β-
keto ester was dissolved in THF (5 mL) and the pH was adjusted
to ca. 8 by slow addition of DIPEA (ca. 1 mL). The reaction mix-
ture was stirred overnight in an open flask. The transformation of
the γ-amino β-keto ester into the pyrazine was monitored by TLC.
After completion of the reaction, the solvent was evaporated under
reduced pressure and the crude product was directly purified by
column chromatography with an EtOAc/pet. ether (5%) solvent
system.
Diethyl 2,2Ј-(3,6-Bis{4-[(tert-butoxycarbonyl)amino]butyl}pyrazine-
2,5-diyl)diacetate (2i): Pale yellow (0.52 g, 78%), m.p. 94–96 °C. 1H
NMR (400 MHz, CDCl3): δ = 7.37–7.27 (m, 10 H), 5.07 (s, 4 H),
4.98 (s, 2 H), 4.13 (q, J = 7.1 Hz, 4 H), 3.81 (s, 4 H), 3.20 (q, J =
6.5 Hz, 4 H), 2.74 (t, J = 7.4 Hz, 4 H), 1.74 (p, J = 7.6 Hz, 4 H),
1.55 (p, J = 7.0 Hz, 4 H), 1.21 (t, J = 7.1 Hz, 6 H) ppm. 13C NMR
(100 MHz, CDCl3): δ = 170.2, 146.1, 136.7, 128.1, 128.1, 66.6, 61.3,
40.7, 33.1, 29.3, 25.2, 14.2 ppm. UV: λmax = 278 nm. Fluorescence:
λex = 278 nm, λem = 319 nm. HRMS: calcd. for C36H46N4O8 [M +
H]+ 663.3388; found 663.3384.
Dibenzyl 3,3Ј-[3,6-Bis(2-ethoxy-2-oxoethyl)pyrazine-2,5-diyl]diprop-
anoate (2j): White solid (0.343 g, 60%), m.p. 95–97 °C. 1H NMR
Eur. J. Org. Chem. 0000, 0–0
© 0000 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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H. N. Gopi et al.
FULL PAPER
(400 MHz, CDCl3): δ = 7.55–7.12 (m, 10 H), 5.09 (s, 4 H), 4.13 (q,
J = 7.1 Hz, 1 H), 3.80 (s, 4 H), 3.04 (t, J = 7.1 Hz, 4 H), 2.87 (t, J
= 7.2 Hz, 4 H), 1.22 (t, J = 7.1 Hz, 6 H) ppm. 13C NMR (101 MHz,
DIPEA (ca. 1 mL). The reaction mixture was stirred in an open
flask. The transformation of the tripeptide γ-amino β-keto ester
into the pyrazine was monitored by MALDI-TOF/TOF. After
CDCl3): δ = 201.80, 172.79, 166.93, 135.74, 128.38, 80.41, 66.69, completion of the reaction, the solvent was evaporated under re-
61.68, 60.50, 58.99, 46.25, 30.04, 28.36, 26.00, 14.15 ppm. UV: λmax
= 208 nm, 278 nm. Fluorescence: λex = 278 nm, λem = 337 nm.
HRMS: calcd. for C32H36N2O8 [M + H]+ 577.2550; found
577.2532.
duced pressure and the crude product was then purified by re-
versed-phase HPLC to afford the desired pyrazine C2 (0.12 g) in
50% yield. Light yellow solid (0.12 g). Retention time (tR) = 22 min
(mobile phase MeOH/H2O). 1H NMR (400 MHz, CDCl3): δ =
7.75–7.22 (m, 14 H), 6.06 (s, 2 H), 5.25–5.01 (m, 6 H), 4.55 (s, 4
H), 4.21–4.16 (m, 3 H), 3.93 (s, 4 H), 1.64–1.20 (m, 16 H), 1.67 (s,
8 H), 1.39 (s, 4 H), 1.39 (s, 4 H), 1.28 (m, 6 H), 0.93 (m, 12 H) ppm.
UV: λmax = 278 nm. Fluorescence: λex = 280 nm, λem = 320 nm.
MALDI-TOF/TOF: calcd. for C48H66N8O12 [M + Na]+ 969.4689;
found 969.6588.
Synthesis of Diethyl 2,2Ј-[3,6-Bis({[(benzyloxy)amino]-3-methyl-
butanamido}methyl)pyrazine-2,5-diyl]diacetate (C1): Cbz-Val-Dap-
(Boc)-CHO was dissolved in CH2Cl2 (10 mL) at room temperature,
and then tin(II) chloride (0.059 g, 20 mol-%) was added, followed
by ethyl diazoacetate (0.152 g, 1.3 mmol). Immediate gas evolution
was observed. The reaction mixture was stirred and the progress of
the reaction was monitored by TLC. After completion of the reac-
tion (ca. 30 min), the mixture was quenched with HCl (0.5 n, 5 mL)
and then extracted with CH2Cl2 (30 mLϫ 3). The combined or-
ganic layer was washed with brine (20 mL), dried with anhydrous
sodium sulfate, and concentrated under reduced pressure to afford
a greenish oily crude product. This was purified by silica gel col-
umn chromatography to afford pure peptide P2 (0.336 g, 60%),
which was directly used for the pyrazine synthesis. Peptide P2
(0.4 g, 0.8 mmol) was dissolved in CH2Cl2 (2 mL) and the solution
was cooled in an ice bath. Then TFA (2 mL) was added slowly to
this solution. After completion of the reaction (ca. 30 min), TFA
was removed from the reaction mixture under reduced pressure.
The TFA salt P3 was dissolved in THF (5 mL), and the pH was
adjusted to ca. 8 by slow addition of DIPEA (ca. 1 mL). The reac-
tion mixture was stirred overnight in an open flask. The transfor-
mation of the γ-amino β-keto ester into the pyrazine was monitored
by TLC. After completion of the reaction, the solvent was evapo-
rated under reduced pressure, and the crude product was directly
subjected to column chromatography and purified with an EtOAc/
pet. ether (35%) solvent system to afford pure C1 (0.15 g) in 55%
Supporting Information (see footnote on the first page of this arti-
1
cle): ORTEP diagrams of compounds 2a and 2b, copies of H and
13C NMR and mass spectra for all synthesized compounds and
RP-HPLC profile of pyrazine C2 are available.
Acknowledgments
The authors thank Indian Institute of Science Education and Re-
search (IISER-Pune) for financial support. M. G. K. is grateful to
the Council of Scientific and Industrial Research (CSIR), New
Delhi for a Senior Research Fellowship.
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6
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Self-Dimerization of Peptides
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Received: September 19, 2014
Published Online: ᭿
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© 0000 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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Job/Unit: O43237
/KAP1
Date: 24-11-14 12:10:02
Pages: 8
H. N. Gopi et al.
FULL PAPER
Peptide Self-Dimerization
M. Ghanesh Kumar, V. J. Thombare,
R. D. Bhaisare, A. Adak,
H. N. Gopi* ...................................... 1–8
Synthesis of Tetrasubstituted Symmetrical
Pyrazines from β-Keto γ-Amino Esters: A
Mild Strategy for Self-Dimerization of
Peptides
A facile synthesis of highly symmetrical
tetrasubstituted pyrazines through simple
aerial oxidation of β-keto γ-esters derived
from amino acids and peptides is reported.
Keywords: Amino acids / Peptides / Oxi-
dation / Cyclization / Nitrogen heterocycles
8
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