Large-Scale Synthesis of Piperazine-2,6-dione and Its Use in the Synthesis of Dexrazoxane Analogues

Article abstract of DOI:10.1055/s-0035-1562618

An efficient, large-scale synthesis of piperazine-2,6-dione was developed. The advantages of this procedure include the use of inexpensive starting materials, satisfactory yields, and a convenient workup without the need for chromatographic techniques. Furthermore, this procedure can be easily modified for the preparation of 1-substituted piperazine-2,6-dione hydrobromides. The utility of the prepared piperazine-2,6-dione was demonstrated in the synthesis of a novel analogue of the only drug used in clinical practice to prevent anthracycline-induced cardiotoxicity, dexrazoxane.

Full text of DOI:10.1055/s-0035-1562618

SYNTHESIS
0
0
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9
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7
8
8
1
1
4
3
7
-
2
1
0
X
© Georg Thieme Verlag Stuttgart · New York
2016, 48, A–I
paper
A
J. Roh et al.
Paper
Synthesis
Large-Scale Synthesis of Piperazine-2,6-dione and Its Use in the
Synthesis of Dexrazoxane Analogues
Jaroslav Roh*
1. CbzCl, H2O
2. Ac2O
3. aq NH3
4. Ac2O
Galina Karabanovich
Veronika Novakova
Tomáš Šimůnek
Kateřina Vávrová
O
HN
O
O
O
O
COOH
COOH
5. HBr, AcOH
N
BrCH2COBr
+H2N
Br–
NH
O
HN
N
NH
O
39% overall yield
43% yield
over 2 steps
26.6 g (0.2 mol)
15.3 g
Charles University in Prague, Faculty of Pharmacy in
Hradec Králové, Heyrovského 1203, 50005 Hradec
Králové, Czech Republic
-no chromatography
-scalable
Dexrazoxane analogue
jaroslav.roh@faf.cuni.cz
Received: 07.06.2016
O
Accepted after revision: 22.07.2016
Published online: 31.08.2016
DOI: 10.1055/s-0035-1562618; Art ID: ss-2016-z0415-op
HOOC
O
N
COOH
N
HN
O
N
H2NOC
N
NH
O
CONH2
Abstract An efficient, large-scale synthesis of piperazine-2,6-dione
was developed. The advantages of this procedure include the use of in-
expensive starting materials, satisfactory yields, and a convenient work-
up without the need for chromatographic techniques. Furthermore,
this procedure can be easily modified for the preparation of 1-substitut-
ed piperazine-2,6-dione hydrobromides. The utility of the prepared
piperazine-2,6-dione was demonstrated in the synthesis of a novel ana-
logue of the only drug used in clinical practice to prevent anthracycline-
induced cardiotoxicity, dexrazoxane.
DEX
ADR-925
Scheme 1 Structures of the cardioprotective drug dexrazoxane (DEX)
and its metabolite ADR-925
To date, few structure–cardioprotective activity rela-
tionship studies of DEX analogues have been published,
mostly with negative results.5,8–10 Such studies have been
hampered by the unmet need for large amounts of DEX an-
alogues because no in vitro model of chronic ANT cardio-
toxicity has been established to date. Thus, each DEX ana-
logue must be repeatedly administered to experimental an-
imals together with selected ANT for several weeks or
months; that is, a period of time necessary for the develop-
ment of chronic ANT cardiotoxicity in an appropriate in
vivo model.
DEX is prepared on a large scale through the cyclization
of (S)-1,2-diaminopropane-N,N,N′,N′-tetraacetic acid in
formamide by heating at 100–110 °C under reduced pres-
sure for 1–2 h, followed by heating at 150–160 °C for 4–5 h.
The majority of DEX analogues that have previously been
described were prepared by using this or a similar method-
ology starting from diaminoalkane-N,N,N′,N′-tetraacetic ac-
ids.11,12 We successfully applied this method for the synthe-
sis of racemic razoxane, 4,4′-(ethylene)bis(piperazine-2,6-
dione) or 4,4′-(propane-1,3-diyl)bis(piperazine-2,6-dione).
However, only DEX analogues with linkers that are stable
under the harsh cyclization conditions, for example 4,4′-
(alkanediyl)bis(piperazine-2,6-diones), can be prepared by
using this method. To overcome the limitations connected
with the cyclization in the last step of the DEX analogue
Key words piperazine-2,6-dione, large-scale synthesis, dexrazoxane,
cyclization, synthesis, formamide
Dexrazoxane (DEX, ICRF-187, ADR-529, (+)-(S)-4,4′-
(propane-1,2-diyl)bis(piperazine-2,6-dione), Scheme 1) is
the only drug used in clinical practice to prevent anthracy-
cline (ANT)-induced cardiotoxicity. DEX can be used to ef-
fectively protect the heart against the chronic type of ANT
cardiotoxicity, which represents the most serious limitation
of the use of ANT in anticancer treatment.1 However, the
mechanism(s) of ANT cardiotoxicity and DEX-induced car-
dioprotection are still not fully understood.2–4 There are
two main hypotheses. The first (traditional) hypothesis fo-
cuses on the iron chelation properties of the DEX metabo-
lite ADR-925, which may prevent ANT-induced oxidative
stress.5 The second (and more recent) hypothesis involves
the specific interaction of DEX with topoisomerase II, an
enzyme that regulates the topology of DNA and manages its
tangles and supercoils.6,7
Hence, new analogues of DEX are needed as potential
novel cardioprotective agents and as tools to dissect the
mechanism(s) of ANT-induced cardiotoxicity as well as to
provide effective and cardio-specific protection.
© Georg Thieme Verlag Stuttgart · New York — Synthesis 2016, 48, A–I

B
J. Roh et al.
Paper
Synthesis
synthesis, piperazine-2,6-dione would be a valuable inter-
mediate for the preparations of DEX analogues with less
stable linkers between the two rings.
Hence, the aim of this study was to develop a reliable,
scalable, and inexpensive synthesis of piperazine-2,6-dione
and to demonstrate its use in the synthesis of DEX ana-
logues.
O
O
HCOOHNH3
+
HOOC
N
COO–
HOOC
N
H
COOH
N
NH
O
+
DMF, toluene
150–170 °C
4 h
NH4
O
4
5
6
Scheme 3 Synthesis of 4-formylpiperazine-2,6-dione (5) from imino-
diacetic acid (4)
We first attempted to prepare piperazine-2,6-dione by
using known methods.13–17 Piperazine-2,6-dione can be
prepared by hydrolysis of 2,6-bishydroxyiminopiperazine.
However, the reported preparation of 2,6-bishydroxyimino-
piperazine starting from iminodiacetonitrile and hydroxyl-
amine suffers from very low yield (14%).13 The reaction be-
tween methyl glycinate hydrochloride (1) with haloacet-
amide (2) in boiling acetonitrile was reported to give
reasonable yields of piperazine-2,6-dione (40 and 33%)
without the need for column chromatography.14,15 Unfortu-
nately, we did not obtain any piperazine-2,6-dione by using
either of those two methods; only methyl N-(carbamoyl-
methyl)glycinate (3) was isolated from the reaction mix-
tures (Scheme 2).
Melting acid 4 with urea gave 4-carbamoylpiperazine-
2,6-dione (7) in 44% yield (Scheme 4). However, the isola-
tion was complicated by the residues of urea and its decom-
position products; thus, column chromatography was
needed, which is a serious drawback in large-scale proce-
dures.
O
O
CO(NH2)2
HOOC
N
H
COOH
N
NH
O
1.5 h, 155 °C
vacuum
H2N
44%
4
7
Scheme 4 Synthesis of 4-carbamoylpiperazine-2,6-dione (7) through
melting of iminodiacetic acid (4) with urea
O
H2NOC
MeOOC
X
KHCO3
H2N
NH
+
MeCN
reflux, 8 h
MeO
NH2·HCl
O
Therefore, we decided to protect iminodiacetic acid (4)
with a Cbz group before the cyclization reactions. The Cbz-
protected acid 8 was cyclized to Cbz-protected piperazine-
2,6-dione (9) by using three different approaches involving
formamide,12 ammonium formate in N,N-dimethylforma-
mide (DMF),16 or urea.18 The only successful procedure was
the latter: melting protected acid 8 with urea under vacu-
um gave 32–54% yield of product 9 at a gram scale, but only
29% yield on a 50-gram scale. The crucial step of this pro-
cess, the urea-assisted cyclization, was hampered by the in-
convenient work-up that resulted in variable yields of Cbz-
protected piperazine-2,6-dione 9. Upon cooling the melt, a
hard, glass-like material was obtained that had to be me-
chanically removed from the reaction vessel, ground and
further purified. This method is problematic especially at
larger scales. Final deprotection with HBr in acetic acid gave
the final piperazine-2,6-dione hydrobromide (10; Scheme
5).
X = Br or Cl
1
2
3
Scheme 2 Unsuccessful preparation of piperazine-2,6-dione through
the reaction of methyl glycinate hydrochloride with bromo/chloroacet-
amide according to previous procedures14,15
Another potentially scalable procedure is the prepara-
tion of 4-formylpiperazine-2,6-dione (5) from iminodia-
cetic acid (4), followed by hydrolysis of the formyl group.16
Although the reaction using a Dean–Stark trap proceeded
well, we were not able to isolate the product 5 by using the
described procedure (by evaporation of the reaction mix-
ture and dilution with methanol). No solids were formed,
even after one month of crystallization at 4 °C. The addition
of acetone to the methanol solution enabled us to obtain a
beige solid after one week of crystallization at 4 °C. Howev-
er, NMR and HRMS experiments showed that the product
was N-formyliminodiacetic acid (6). When the reaction was
carried out without a Dean–Stark trap and the water and
volatiles were azeotropically distilled off during the reac-
tion, crude 5 was obtained in 8% yield (Scheme 3). However,
the low yield meant that this method was not suitable for
the large-scale synthesis of piperazine-2,6-dione.
O
CO(NH2)2
HOOC
N
R
COOH
R
N
NH
3 h, 150–165 °C
vacuum
32–54%
29% at a 50-g scale
O
HBr
AcOH
rt, 2 h
98%
CbzCl
aq NaOH
rt, 3 h
9, R = Cbz
4, R = H
8, R = Cbz
The reaction of 4 with formamide according to the cy-
clization protocol used in the DEX synthesis12 expectedly
gave no product; only black tar was obtained after evapora-
tion of the reaction mixture, from which no solids crystal-
lized after dilution with various organic solvents.
10, R = H·HBr
97%
Scheme 5 Large-scale synthesis of piperazine-2,6-dione hydrobro-
mide 10 through urea-assisted cyclization
© Georg Thieme Verlag Stuttgart · New York — Synthesis 2016, 48, A–I

C
J. Roh et al.
Paper
Synthesis
O
O
O
O
1. aq. NH3
2. HCl
COOH
Ac2O
Ac2O
HOOC
N
R
COOH
Cbz
N
Cbz
N
R
N
NH
rt, 3 days
91–95%
0 °C to rt, 2 h
88–93%
140 °C, 2 h
50–56%
CONH2
CbzCl
aq NaOH
rt, 3 h
O
4, R = H
11
12
HBr
9, R = Cbz
AcOH
rt, 2 h
98%
8, R = Cbz
95–97%
10, R = H·HBr
Scheme 6 Large-scale synthesis of piperazine-2,6-dione hydrobromide (9) through acetic anhydride assisted cyclization
To optimize the above method for additional conve-
nience, we attempted to replace the urea-assisted cycliza-
tion with a three-step imide synthesis.19,20 In the first step,
the Cbz-protected acid 8 was dehydrated to Cbz-protected
morpholine-2,6-dione 11 in acetic anhydride.21 Simple
evaporation of the reaction mixture afforded product 11,
which was pure enough to be used in the next step. The ob-
tained Cbz-protected morpholine-2,6-dione 11 was added
to ice-cold aqueous ammonia. After complete dissolution,
the reaction mixture was acidified to obtain crystalline N-
Cbz-N-(carbamoylmethyl)glycine (12).
Product 12 was then heated in acetic anhydride to
140 °C for 2 h. Cooling of the reaction mixture and addition
of diethyl ether provided Cbz-protected piperazine-2,6-di-
one (9) in good yields and with high purity. Final deprotec-
tion was accomplished by using HBr in acetic acid. The
overall yield of this method was 39% at the 0.2-mol scale
(15.3 g of 10 was obtained from 26.6 g of 4). This method
repeatedly gave similar yields and was scalable, robust, and
convenient; the whole reaction sequence involved only
evaporations, crystallizations, and filtrations, with no chro-
matographic methods being employed (Scheme 6).
sis12 starting from glycinamide-N,N,N′,N′-tetraacetic acid
(16), failed. Only 4-(carbamoylmethyl)piperazine-2,6-dione
(17) was isolated in the second reaction (Scheme 7).
Therefore, 10 was used in the synthesis of DEX analogue
13. First, we attempted a one-step reaction between two
equivalents of 10 and one equivalent of bromoacetyl bro-
mide in acetonitrile or DMF by using various bases. Howev-
er, these methods gave very low yields (less than 10%).
In an effort to increase the yields of DEX analogue 13,
we attempted to use an imide protection/deprotection
strategy, which would increase the solubility of the pipera-
zine-2,6-dione moiety and also protect the imide nitrogen
against unwanted side reactions. Thus, we modified the
large-scale procedure described in Scheme 6 to be generally
applicable for the preparation of 1-substituted piperazine-
2,6-diones and used it for the preparation of 1-(4-methoxy-
phenyl)piperazine-2,6-dione (20a) and 1-(4-methoxyben-
zyl)piperazine-2,6-dione hydrobromides (20b, Scheme 8).
4-Methoxyphenyl (PMP) and 4-methoxybenzyl (PMB) sub-
stituents were chosen because of their use as protective
groups in similar compounds – hydantoins23 and phthalim-
ides.24
To demonstrate the usefulness of 10 in the synthesis of
DEX analogues, we report here the synthesis of a new DEX
analogue 4,4′-(1-oxoethane-1,2-diyl)bis(piperazine-2,6-di-
one) (13). It should be noted that attempts to prepare 13 by
using the method described previously,22 starting from te-
tramethyl glycinamide-N,N,N′,N′-tetraacetate (15) and
through urea-assisted cyclization or standard DEX synthe-
O
COOH
Ac2O
RNH2, Et3N
THF
Cbz
N
R'
N
N
R
11
140 °C, 2 h
CONHR
O
18a, R = PMP, 81%
18b, R = PMB, 95–98%
19a, R' = Cbz, R = PMP
81%
19b, R' = Cbz, R = PMB
72–75%
HBr
AcOH
rt, 1 h
BrCH2COBr
DIPEA
R'OOC
R'OOC
O
20a, R' = H.HBr, R = PMP
86–94%
20b, R' = H.HBr, R = PMB
77–87%
N
COOR'
COOR'
R'OOC
N
R
COOR'
MeCN, rt, 48 h
63%
N
1. aq NaOH
rt, 3.5 h
SOCl2
15, R' = Me
16, R' = H
4, R = R' = H
Scheme 8 Synthesis of 1-PMP- and 1-PMB-protected piperazine-2,6-
dione hydrobromides 20a and 20b
MeOH
–20 °C to rt
81%
2. H+
83%
14, R = H·HCl, R' = Me
O
HN
O
O
O
O
O
Compounds 20a and 20b were used in the synthesis of
PMP- and PMB-protected DEX analogues 21a and 21b, re-
spectively (Scheme 9). Cleavage of the PMP and PMB groups
with ceric ammonium nitrate23,25–27 and cleavage of PMB
through palladium-catalyzed hydrogenolysis,28 or with tri-
H2N
HCONH2
1. 110 °C, 1 h,
vacuum
2. 150–160 °C, 4 h
N
16
N
NH
O
N
NH
O
17
13%
13
not detected
31
fluoroacetic acid29,30 or AlCl3 failed; the target DEX ana-
Scheme 7 Unsuccessful preparation of 13 according to standard DEX
synthesis
logue 13 was not detected in any of these reactions. Unfor-
© Georg Thieme Verlag Stuttgart · New York — Synthesis 2016, 48, A–I

D
J. Roh et al.
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Synthesis
and used as received. TLC was performed on Merck aluminum plates
with silica gel 60 F254. Merck silica gel 60 (0.040–0.063 mm) was used
for column chromatography. Melting points were recorded with a Bü-
chi B-545 apparatus and are uncorrected. 1H and 13C NMR spectra
were recorded with Varian Mercury Vx BB 300 or VNMR S500 NMR
spectrometers (Varian Inc., Palo Alto, CA). Chemical shifts are report-
ed as δ values in parts per million (ppm) and are indirectly referenced
to tetramethylsilane (TMS) via the solvent signal. The elemental anal-
ysis was carried out with an Automatic Microanalyser EA1110CE
(Fisons Instruments S.p.A., Milano, Italy). Atmospheric pressure
chemical ionization (APCI) was performed with an Agilent 500 Ion
Trap LC/MS (Agilent Technologies, Santa Clara, California, USA) by di-
rect infusion into the detector of the sample dissolved in methanol.
HRMS (ESI+, ESI–) experiments were performed with a Q-Exactive
Plus Mass Spectrometer (Thermo Scientific, Bremen, Germany) by di-
rect infusion into the detector of the sample dissolved in acetonitrile–
water (1:1). IR spectra were measured with a Nicolet 6700 Spectro-
photometer in the ATR mode (Thermo Scientific, Waltham, MA, USA).
O
O
O
O
N
O
BrCH2COBr
Br–
NH2
DIPEA
R
N
O
N
+
R
N
N
R
MeCN
0 °C to rt
O
20a, R = PMP
20b, R = PMB
21a, R = PMP, 63%
21b, R = PMB, 77%
Scheme 9 Synthesis of PMP- and PMB-protected DEX analogues 21a
and 21b
tunately, this imide protection/deprotection strategy was
not successful in this application.
Hence, we proceeded with unprotected piperazine-2,6-
dione hydrobromide (10) and developed a two-step proce-
dure that consists of the synthesis and isolation of 4-(2-bro-
moacetyl)piperazine-2,6-dione (22) and its conversion into
the target DEX analogue 13. Gratifyingly, this procedure led
to 43% overall yield of the product (Scheme 10).
Synthesis of 4-Formylpiperazine-2,6-dione (5)16
Method A: Iminodiacetic acid (13.1 g, 0.1 mol) and ammonium for-
mate (18.9 g, 0.3 mol) were heated to reflux in a mixture of DMF (100
mL) and toluene (40 mL) for 4 h. A reaction vessel equipped with a
Dean–Stark trap was used to remove the water formed as an azeo-
tropic mixture. The reaction mixture was then evaporated under vac-
uum, and the residue was dissolved in MeOH (20 mL). Acetone (20
mL) was added, and the mixture was cooled to 4 °C. After one week,
the crystalline solid was collected by filtration.
O
HN
O
O
HN
O
BrCH2COBr
DIPEA
Br–
NH2
O
+
N
MeCN
0 °C to rt
41–62%
Br
10
22
O
HN
O
O
O
DIPEA
N
Yield: 6.9 g (39%) of crude N-formyliminodiacetic acid monoammoni-
um salt (6) as a beige solid.
1H NMR (500 MHz, DMSO-d6): δ = 8.01 (s, 1 H), 3.91 (s, 2 H), 3.76 (s,
22 + 10
DMF
N
NH
O
rt, 24 h
69%
13
2 H).
Scheme 10 Piperazine-2,6-dione-based synthesis of DEX analogue
4,4′-(1-oxoethane-1,2-diyl)bis(piperazine-2,6-dione) (13)
1H NMR (500 MHz, D2O): δ = 8.10 (s, 1 H), 4.15 (s, 2 H), 4.06 (s, 2 H).
13C NMR (126 MHz, DMSO-d6): δ = 173.05, 171.63, 164.21, 53.08,
50.16.
13C NMR (126 MHz, D2O): δ = 175.70, 174.30, 167.02, 51.93, 47.49.
HRMS (ESI–): m/z [M – H]– calcd for C5H6NO5: 160.02515; found:
To conclude, a novel large-scale synthesis of piperazine-
2,6-dione hydrobromide was developed and optimized. The
advantage of this procedure is the use of inexpensive mate-
rials, satisfactory yields, and a convenient workup without
the need for column chromatography. We also demonstrat-
ed that 1-substituted piperazine-2,6-dione hydrobromides
can be easily prepared by using this method after small
modifications.
The synthetic value of the obtained piperazine-2,6-di-
one hydrobromide was demonstrated by the synthesis of a
new DEX analogue 4,4′-(1-oxoethane-1,2-diyl)bis(pipera-
zine-2,6-dione) (13), which was not possible to prepare by
using standard DEX synthetic approaches. This robust and
convenient synthetic methodology opens up possibilities to
further vary the DEX structure to explore the importance of
the individual structural fragments in the cardioprotective
activity of DEX and its mechanism of action.
160.02271.
Anal. Calcd for C5H10N2O5: C, 33.71; H, 5.66; N, 15.73. Found: C, 33.61;
H, 5.65; N, 16.39.
Method B: Iminodiacetic acid (13.1 g, 0.1 mol) and ammonium for-
mate (18.9 g, 0.3 mol) were heated in DMF (100 mL) in distillation ap-
paratus at a bath temperature of 150–160 °C for 1 h. Toluene (40 mL)
was added and the mixture was heated for 3 h at a bath temperature
of 160–170 °C, distilling off the toluene with water together with a
part of other volatiles. The reaction mixture was then evaporated un-
der vacuum, the residue was dissolved in MeOH (30 mL), and the
mixture was cooled to 4 °C overnight. The crystalline solid was col-
lected by filtration and washed with MeOH (20 mL).
Yield: 1.2 g (8%) of crude 4-formylpiperazine-2,6-dione (5);16 beige
solid.
1H NMR (500 MHz, DMSO-d6): δ = 11.33 (s, 1 H), 8.08 (s, 1 H), 4.29 (s,
2 H), 4.20 (s, 2 H).
13C NMR (126 MHz, DMSO-d6): δ = 169.54, 169.25, 161.98, 47.32,
42.42.
The structural identities of the prepared compounds were confirmed
by 1H NMR and 13C NMR spectroscopic analysis. All chemicals used for
synthesis were obtained from Sigma–Aldrich (Schnelldorf, Germany)
HRMS (ESI+): m/z [M + H]+ calcd for C5H7N2O3: 143.04512; found:
143.04457.
© Georg Thieme Verlag Stuttgart · New York — Synthesis 2016, 48, A–I

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Synthesis
4-Carbamoylpiperazine-2,6-dione (7)
Anal. Calcd for C12H14N2O5: C, 54.13; H, 5.30; N, 10.52. Found: C,
54.01; H, 5.14; N, 10.33.
Iminodiacetic acid (1 g, 0.0075 mol) and urea (0.9 g, 0.015 mol) were
melted together at 155 °C under vacuum (30 mbar) for 1.5 h. The melt
was cooled to r.t. and stirred with MeOH (10 mL) at r.t. overnight. The
resulting suspension was evaporated, and the product was purified
by column chromatography (CHCl3–MeOH, 3:1).
4-(Benzyloxycarbonyl)piperazine-2,6-dione (9)
Through urea-catalyzed cyclization of 8: N-(Benzyloxycarbonyl)imino-
diacetic acid (8; 1.6 g, 0.006 mol) and urea (0.8 g, 0.013 mol) were
melted together at 150–165 °C under vacuum (30 mbar) for 3 h.
MeOH (15 mL) was added to the cooled melt, and the mixture was
stirred for 2 days until none of the melt remained in the mixture.
MeOH was then evaporated and the residue was partitioned between
CHCl3 (30 mL) and sat. aq NaHCO3 (30 mL). The aqueous phase was
then extracted with CHCl3 (2 × 30 mL) and the combined chloroform
extracts were dried over Na2SO4 and evaporated.
Yield: 0.52 g (44%); white solid; mp 187–188 °C.
IR (ATR): 3350, 3234, 3182, 1726, 1681, 1657, 1469, 1397, 1329, 1293,
1251, 1185, 1119, 1104, 995, 963, 910, 759 cm–1
.
1H NMR (300 MHz, DMSO-d6): δ = 10.67 (s, 1 H), 7.53 (s, 1 H), 7.17 (s,
1 H), 3.90 (s, 2 H), 3.79 (s, 2 H).
13C NMR (75 MHz, DMSO-d6): δ = 172.15, 169.97, 157.71, 51.91,
44.53.
Yield: 0.8 g (54%; 29% when started from 67 g of 8); white solid.
Anal. Calcd for C5H7N3O3: C, 38.22; H, 4.49; N, 26.74. Found: C, 37.82;
H, 4.4; N, 26.75.
Through cyclization of 12: N-Benzyloxycarbonyl-N-(carbamoylmeth-
yl)glycine (12; 42.7 g, 0.160 mol) and acetic anhydride (160 mL) were
heated with stirring at 140 °C for 2 h. The reaction mixture was then
cooled and Et2O (160 mL) was added. After 3 days at 4 °C, the white
crystals were filtered off, washed with Et2O, and dried.
N-(Benzyloxycarbonyl)iminodiacetic Acid (8)32
Iminodiacetic acid (26.6 g, 0.2 mol) was dissolved in 2 M NaOH (200
mL) and the resulting solution was cooled in an ice bath. Benzyloxy-
carbonyl chloride (37.5 g, 31.4 mL, 0.22 mol) and 2 M NaOH (120 mL)
were subsequently added to the stirred solution, the reaction mixture
was stirred at r.t. for 6 h and then washed with Et2O (2 × 100 mL). The
aqueous phase was acidified with conc. HCl to pH 1–2 and extracted
with Et2O (3 × 200 mL). The organic extract was dried over Na2SO4
and evaporated. The product was used in the subsequent reactions
without further purification.
Yield: 19.9 g (50%; 54% and 56% when started from 12 g and 9 g of 12,
respectively); white crystalline solid; mp 170–172 °C.
IR (ATR): 3193, 3089, 2884, 1732, 1711, 1459, 1445, 1428, 1389, 1363,
1344, 1300, 1239, 1199, 1118, 977, 862, 761, 741 cm–1
.
1H NMR (300 MHz, DMSO-d6): δ = 11.41 (s, 1 H), 7.47–7.27 (m, 5 H),
5.11 (s, 2 H), 4.22 (s, 4 H).
13C NMR (75 MHz, DMSO-d6): δ = 169.58, 154.10, 136.41, 128.65,
128.27, 127.95, 67.32, 46.58.
Yield: 51 g (95%); colorless viscous oil.
1H NMR (300 MHz, CDCl3): δ = 10.37 (s, 1 H), 8.53 (s, 1 H), 7.38–7.23
(m, 5 H), 5.14 (s, 2 H), 4.16 (s, 2 H), 4.11 (s, 2 H).
Anal. Calcd for C12H12N2O4: C, 58.06; H, 4.87; N, 11.29. Found: C,
57.79; H, 4.73; N, 11.14.
13C NMR (75 MHz, CDCl3): δ = 174.10, 173.39, 156.09, 135.45, 128.55,
128.31, 127.86, 68.50, 50.06, 49.94.
Piperazine-2,6-dione Hydrobromide (10)
4-(Benzyloxycarbonyl)piperazine-2,6-dione (9; 19.9 g, 0.08 mol) was
added in several portions to a stirring solution of HBr (33%) in acetic
acid (60 mL). The resulting suspension was stirred vigorously for 2 h
at r.t., then Et2O (100 mL) was added and the solid was filtered off,
washed with Et2O and left under vacuum over NaOH in a dessicator
for 24 h.
N-Benzyloxycarbonyl-N-(carbamoylmethyl)glycine (12)
N-(Benzyloxycarbonyl)iminodiacetic acid (8; 51 g, 0.19 mol) was dis-
solved in acetic anhydride (100 mL) and stirred at r.t. for 3 days. The
reaction mixture was then evaporated under high vacuum and the re-
sulting oil crystallized in one day at r.t. under an Ar atmosphere to
give N-(benzyloxycarbonyl)morpholine-2,6-dione (11)21 as a white
solid, which was used immediately without further purification.
Yield: 15.3 g (98%); white solid; mp 290–300 °C (decomp.).
IR (ATR): 3068, 2946, 2783, 1709, 1674, 1541, 1525, 1427, 1398, 1272,
1224, 1180, 1058, 976, 923, 801, 766 cm–1
.
N-(Benzyloxycarbonyl)morpholine-2,6-dione (11; 44 g, 0.177 mol)
was added in several portions into ice cold 25% aqueous ammonia
(240 mL) under vigorous stirring. The reaction mixture was stirred for
2 h in an ice bath. During this period, the entire solid dissolved. The
reaction mixture was then acidified to pH 1 with 36% HCl, and the
formed precipitate was filtered, washed with water and dried.
1H NMR (300 MHz, DMSO-d6): δ = 11.83 (s, 1 H), 9.84 (br s, 2 H), 4.01
(s, 4 H).
13C NMR (75 MHz, DMSO-d6): δ = 166.49, 44.21.
Anal. Calcd for C4H7BrN2O2: C, 24.64; H, 3.62; N, 14.36. Found: C,
24.91; H, 3.69; N, 13.99.
Yield: 42.7 g (91%; 93% and 88% when started from 5 g and 15 g of 11,
respectively); white crystalline solid; mp 173–174 °C.
Dimethyl Iminodiacetate Hydrochloride (14)14
IR (ATR): 3365, 3235, 1706, 1646, 1558, 1541, 1472, 1450, 1428, 1400,
Methanol (500 mL) was cooled to –20 °C, and SOCl2 (36 mL, 0.5 mol)
was added dropwise under vigorous stirring while maintaining the
temperature of the reaction mixture below –10 °C. Iminodiacetic acid
(13.3 g, 0.1 mol) was then added in one portion. The reaction mixture
was stirred for 24 h at r.t. and then left to stand at r.t. for two days.
The volatiles were evaporated, and the residue were crystallized from
methanol.
1363, 1338, 1258, 1200, 1142, 1017, 974, 916, 773 cm–1
.
1H NMR (500 MHz, DMSO-d6): δ = 13.02 (s, 1 H), 7.62 (d, J = 5.5 Hz,
1 H), 7.39–7.26 (m, 5 H), 7.21 (d, J = 7.2 Hz, 1 H), 5.07 (d, J = 4.1 Hz,
2 H), 4.01 (d, J = 15.4 Hz, 2 H), 3.90 (d, J = 13.1 Hz, 2 H).
13C NMR (126 MHz, DMSO-d6): δ = 171.44 (d, J = 10.2 Hz), 171.30,
155.68 (d, J = 10.8 Hz), 136.73 (d, J = 4.0 Hz), 128.46, 127.88 (d, J =
1.6 Hz), 127.24 (d, J = 2.2 Hz), 66.74 (d, J = 6.8 Hz), 51.30 (d, J =
45.4 Hz), 50.60 (d, J = 60.1 Hz).
Yield: 16 g (81%); white crystalline solid.
1H NMR (300 MHz, D2O): δ = 4.13 (s, 4 H), 3.85 (s, 6 H).
13C NMR (75 MHz, D2O): δ = 168.08, 54.12, 47.66.
MS (APCI+): m/z (%) = 223.1 (100) [M – CO2 + H]+, 267.0 (30) [M + H]+.
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J. Roh et al.
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Synthesis
Anal. Calcd for C6H12ClNO4: C, 36.47; H, 6.12; N, 7.09. Found: C, 36.52;
H, 6.01; N, 6.98.
13C NMR (75 MHz, DMSO-d6): δ = 171.35, 170.77, 57.85, 55.16.
Anal. Calcd for C6H9N3O3: C, 42.11; H, 5.30; N, 24.55. Found: C, 41.83;
H, 5.32; N, 24.52.
Tetramethyl Glycinamide-N,N,N′,N′-tetraacetate (15)
Dimethyl iminodiacetate hydrochloride (14; 16 g, 0.081 mol) and
DIPEA (20.93 g, 28.2 mL, 0.162 mol) were added to acetonitrile (150
mL), and then bromoacetyl bromide (8.17 g, 3.55 mL, 0.04 mol) was
added dropwise. The reaction mixture was stirred at r.t. for 48 h, then
the acetonitrile was evaporated and the residue was dissolved in
EtOAc (200 mL). The organic solution was washed with H2O (3 × 150
mL), dried over Na2SO4 and evaporated. The pure product was ob-
tained by column chromatography (CHCl3–EtOAc, 5:1).
N-Benzyloxycarbonyl-N-{2-[(4-methoxyphenyl)amino]-2-oxoeth-
yl}glycine (18a)
p-Toluidine (1 g, 0.0081 mol) and triethylamine (1.1 mL, 0.008 mol)
were dissolved in anhydrous THF (25 mL), then N-(benzyloxycarbon-
yl)morpholine-2,6-dione (11; 2 g, 0.008 mol) was added to the stirred
solution, and the mixture was stirred at r.t. for 1 h. The THF was evap-
orated and the residue was dissolved in 2.5% aq NaOH (40 mL). The
aqueous solution was washed with EtOAc (2 × 20 mL) and acidified to
pH 1 with HCl. The resulting solid was filtered off and dried.
Yield: 9.2 g (63%); colorless solid; mp 57–59 °C.
IR (ATR): 2956, 1754, 1741, 1647, 1475, 1433, 1410, 1373, 1304, 1205,
Yield: 2.4 g (81%); white solid; mp 137–138 °C.
1183, 1146, 1002, 892, 728 cm–1
.
IR (ATR): 3260, 3099, 2834, 1705, 1607, 1562, 1510, 1445, 1401, 1365,
1H NMR (300 MHz, CDCl3): δ = 4.60 (s, 2 H), 4.13 (s, 2 H), 3.74 (s, 3 H),
1345, 1323, 1244, 1134, 1031, 982, 833, 769, 748, 733, 716 cm–1
.
3.70 (s, 3 H), 3.68 (s, 6 H), 3.65 (s, 2 H), 3.53 (s, 4 H).
13C NMR (75 MHz, CDCl3): δ = 170.86, 170.04, 169.97, 169.40, 56.52,
54.41, 52.26, 52.12, 51.59, 49.83, 48.19.
MS (APCI+): m/z (%) = 363.3 (100) [M + H]+.
1H NMR (500 MHz, DMSO-d6): δ = 12.99 (s, 1 H), 10.02 (d, J = 12.5 Hz,
1 H), 7.52–7.44 (m, 2 H), 7.38–7.30 (m, 2 H), 7.30–7.21 (m, 2 H), 6.92–
6.86 (m, 2 H), 5.09 (d, J = 4.3 Hz, 2 H), 4.11 (s, 2 H), 4.09 (d, J = 2.9 Hz,
2 H), 3.72 (d, J = 2.5 Hz, 3 H).
13C NMR (126 MHz, DMSO-d6): δ = 171.90 (d, J = 3.3 Hz), 167.13 (d, J =
15.5 Hz), 155.78 (d, J = 3.8 Hz), 155.47 (d, J = 5.1 Hz), 136.73 (d, J =
5.7 Hz), 131.99 (d, J = 7.3 Hz), 128.46 (d, J = 12.9 Hz), 127.90 (d, J =
10.9 Hz), 127.24 (d, J = 13.3 Hz), 120.82 (d, J = 10.2 Hz), 114.06, 66.81
(d, J = 10.4 Hz), 55.34, 52.06 (d, J = 31.1 Hz), 50.53 (d, J = 65.9 Hz).
Anal. Calcd for C14H22N2O9: C, 46.41; H, 6.12; N, 7.73. Found: C, 46.16;
H, 6.03; N, 7.55.
Glycinamide-N,N,N′,N′-tetraacetic Acid (16)
Tetramethyl glycinamide-N,N,N′,N′-tetraacetate (15; 2.84 g, 0.0078
mol) was dissolved in 1 M NaOH (31.3 mL), and the mixture was
stirred at r.t. for 3.5 h. The reaction mixture was then acidified to pH 3
by using Amberlyst 15. The reaction mixture was filtered and evapo-
rated to dryness and the residue was further dried over P2O5. Product
16 was contaminated with inorganic residues, as detected by elemen-
tal analysis. Product 16 was used in the next step without further pu-
rification.
Anal. Calcd for C19H20N2O6: C, 61.28; H, 5.41; N, 7.52. Found: C, 61.62;
H, 5.75; N, 7.16.
N-Benzyloxycarbonyl-N-{2-[(4-methoxybenzyl)amino]-2-oxoeth-
yl}glycine (18b)
4-Methoxybenzylamine (1.23 g, 0.009 mol) and triethylamine (1.1
mL, 0.008 mol) were dissolved in anhydrous THF (20 mL), then N-
(benzyloxycarbonyl)morpholine-2,6-dione (11; 2 g, 0.008 mol) was
added to the stirred solution and the mixture was stirred at r.t. for 2
h. The THF was then evaporated and the residue was dissolved in 2.5%
aq NaOH (40 mL). The aqueous solution was washed with Et2O (2 × 20
mL) and acidified to pH 1 with HCl. The aqueous layer was extracted
with EtOAc (3 × 40 mL), and the combined organic extract was dried
over Na2SO4 and evaporated.
Yield: 2 g (83%); white solid.
1H NMR (300 MHz, D2O): δ = 4.35 (s, 2 H), 4.15 (s, 2 H), 4.06 (s, 2 H),
3.88 (s, 4 H).
13C NMR (75 MHz, D2O): δ = 173.84, 173.67, 169.80, 166.45, 57.27,
55.06, 51.19, 50.55.
MS (APCI+): m/z (%) = 307.0 (100), 308.0 (10) [M + H]+.
HRMS (ESI+): m/z [M + H]+ calcd for C10H15N2O9: 307.07721; found:
307.07673.
Yield: 3.04 g (98%; 95% when started from 10 g of 11); colorless oil
that crystallized after several weeks; mp 149–150 °C.
IR (ATR): 3303, 1717, 1655, 1543, 1515, 1469, 1446, 1410, 1398, 1364,
1316, 1243, 1230, 1183, 1143, 1036, 970, 958, 915, 819, 774, 761, 695
Anal. Calcd for C10H14N2O9: C, 39.22; H, 4.61; N, 9.15. Found: C, 34.65;
H, 4.46; N, 7.76.
cm–1
.
1H NMR (500 MHz, DMSO-d6): δ = 12.97 (s, 1 H), 8.66–8.50 (m, 1 H),
7.42–7.26 (m, 5 H), 7.15 (dd, J = 29.6, 8.6 Hz, 2 H), 6.84 (dd, J = 34.9,
8.6 Hz, 2 H), 5.08 (d, J = 7.1 Hz, 2 H), 4.23 (dd, J = 12.6, 5.8 Hz, 2 H),
4.04 (d, J = 12.5 Hz, 2 H), 3.99 (d, J = 9.3 Hz, 2 H), 3.72 (d, J = 9.1 Hz,
3 H).
13C NMR (126 MHz, DMSO-d6): δ = 171.57 (d, J = 4.2 Hz), 168.92 (d, J =
11.9 Hz), 158.41 (d, J = 4.1 Hz), 155.75 (d, J = 5.2 Hz), 136.72 (d, J =
7.2 Hz), 131.13 (d, J = 3.4 Hz), 128.70 (d, J = 2.8 Hz), 128.52, 127.94,
127.32 (d, J = 5.4 Hz), 113.85 (d, J = 5.9 Hz), 66.84, 55.24, 51.59 (d, J =
37.0 Hz), 50.61 (d, J = 68.2 Hz), 41.77 (d, J = 3.0 Hz).
4-(Carbamoylmethyl)piperazine-2,6-dione (17)33
Crude glycinamide-N,N,N′,N′-tetraacetic acid (16; 2 g, 0.0065 mol)
was heated in formamide (12 mL) under vacuum (30 mbar) at 110 °C
for 1 h and then under an argon atmosphere at 155 °C for 3 h. The for-
mamide was then distilled off, and the residue was purified by col-
umn chromatography (EtOAc–acetone, 6:1).
Yield: 145 mg (13%); beige solid; mp 210–215 °C (Lit.33 212–215 °C).
IR (ATR): 3420, 3286, 3144, 3065, 2837, 1689, 1670, 1615, 1592, 1435,
1397, 1362, 1330, 1296, 1277, 1205, 1163, 1148, 1028, 933, 874, 852
cm–1
.
Anal. Calcd for C20H22N2O6: C, 62.17; H, 5.74; N, 7.25. Found: C, 61.8;
H, 5.34; N, 7.37.
1H NMR (300 MHz, DMSO-d6): δ = 11.12 (s, 1 H), 7.39 (s, 1 H), 7.11 (s,
1 H), 3.37 (s, 4 H), 3.06 (s, 2 H).
© Georg Thieme Verlag Stuttgart · New York — Synthesis 2016, 48, A–I

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J. Roh et al.
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Synthesis
4-Benzyloxycarbonyl-1-(4-methoxyphenyl)piperazine-2,6-dione
(19a)
13C NMR (126 MHz, DMSO-d6): δ = 165.84, 159.41, 129.55, 125.91,
114.53, 55.56 (d, J = 4.0 Hz), 45.49.
N-Benzyloxycarbonyl-N-{2-[(4-methoxyphenyl)amino]-2-oxoethyl}gly-
cine (18a; 2 g, 0.0056 mol) and acetic anhydride (8 mL) were heated
to 140 °C under an Ar atmosphere with stirring for 2 h. The reaction
mixture was then evaporated under vacuum, the residue was dis-
solved in EtOAc (50 mL), and the organic solution was washed with
sat. aq NaHCO3 (2 × 25 mL) and H2O (30 mL). The organic fraction was
dried over Na2SO4 and evaporated. The pure product was obtained by
column chromatography (hexane–EtOAc, 2:1).
Anal. Calcd for C11H13BrN2O3: C, 43.87; H, 4.35; N, 9.30. Found: C,
43.72; H, 4.28; N, 8.96.
1-(4-Methoxybenzyl)piperazine-2,6-dione Hydrobromide (20b)
HBr (33% in acetic acid, 3 mL) was added to 4-benzyloxycarbonyl-1-
(4-methoxybenzyl)piperazine-2,6-dione (19b; 0.9 g, 0.0024 mol), and
the resulting mixture was stirred vigorously at r.t. for 1 h. Et2O (20
mL) was added and the solid was filtered off, washed with Et2O and
left under vacuum over NaOH in a dessicator for 24 h.
Yield: 1.53 g (81%); colorless oil that solidified into an amorphous col-
orless solid; mp 118–119 °C.
Yield: 0.67 g (87%; 77% when started from 7 g of 19b); white solid;
mp 185–188 °C (with decomp.).
IR (ATR): 1746, 1693, 1609, 1511, 1456, 1363, 1289, 1232, 1199, 1108,
1030, 970, 831, 759, 723 cm–1
.
IR (ATR): 3007, 2942, 2837, 1741, 1693, 1613, 1514, 1432, 1393, 1343,
1H NMR (500 MHz, DMSO-d6): δ = 7.45–7.32 (m, 5 H), 7.10 (d, J =
8.9 Hz, 2 H), 6.99 (d, J = 8.9 Hz, 2 H), 5.17 (s, 2 H), 4.48 (s, 4 H), 3.78 (s,
3 H).
13C NMR (126 MHz, DMSO-d6): δ = 168.59, 159.15, 153.94, 136.38,
129.83, 128.64, 128.26, 127.97, 126.70, 114.23, 67.34, 55.48 (d, J =
3.8 Hz), 47.44.
1303, 1250, 1217, 1175, 1114, 1030, 938, 893, 769 cm–1
.
1H NMR (500 MHz, DMSO-d6): δ = 7.24 (d, J = 8.7 Hz, 2 H), 6.86 (d, J =
8.7 Hz, 2 H), 4.78 (s, 2 H), 4.21 (s, 4 H), 3.71 (s, 3 H).
13C NMR (126 MHz, DMSO-d6): δ = 165.67, 158.75, 129.78 (d, J =
7.8 Hz), 128.16, 113.81, 55.32 (d, J = 12.6 Hz), 44.87, 41.78.
Anal. Calcd for C12H15BrN2O3: C, 45.73; H, 4.80; N, 8.89. Found: C,
46.10; H, 4.72; N, 8.66.
Anal. Calcd for C19H18N2O5: C, 64.40; H, 5.12; N, 7.91. Found: C, 64.05;
H, 5.32; N, 7.55.
4,4′-(1-Oxoethane-1,2-diyl)bis[1-(4-methoxyphenyl)piperazine-
2,6-dione] (21a)
4-Benzyloxycarbonyl-1-(4-methoxybenzyl)piperazine-2,6-dione
(19b)
Bromoacetyl bromide (0.34 g, 0.15 mL, 0.00168 mol) was slowly add-
ed to a solution of 1-(4-methoxyphenyl)piperazine-2,6-dione hydro-
bromide (20a; 1 g, 0.0033 mmol) and DIPEA (0.87 g, 1.17 mL, 0.0067
mmol) in MeCN (30 mL) under an inert atmosphere at 0 °C. The reac-
tion mixture was stirred at 0 °C for 15 min and then at r.t. for 7 days.
The solvent was evaporated under reduced pressure, and the residue
was dissolved in EtOAc (35 mL) and washed with 0.5% HCl (35 mL).
The organic layer was separated, dried over anhydrous sodium sul-
fate, and evaporated under reduced pressure. The product was puri-
fied by column chromatography (EtOAc).
N-Benzyloxycarbonyl-N-{2-[(4-methoxybenzyl)amino]-2-oxoethyl}gly-
cine (18b; 5 g, 0.0129 mol) and acetic anhydride (20 mL) were heated
to 140 °C under an Ar atmosphere with stirring for 2 h. The reaction
mixture was then evaporated under vacuum, the residue was dis-
solved in EtOAc (70 mL), and the organic solution was washed with
sat. aq NaHCO3 (2 × 40 mL) and H2O (100 mL). The organic fraction
was dried over Na2SO4 and evaporated. The pure product was ob-
tained by column chromatography (hexane–EtOAc, 2:1).
Yield: 3.43 g (72%; 75% when started from 15 g of 18b); colorless oil.
IR (ATR): 2957, 1684, 1611, 1513, 1442, 1385, 1344, 1310, 1237, 1214,
Yield: 0.5 g (63%); white solid; mp 262–265 °C.
1178, 1115, 1031, 970, 921, 892, 852, 822, 759 cm–1
.
IR (ATR): 3014, 2838, 1737, 1687, 1642, 1612, 1512, 1381, 1359, 1301,
1H NMR (500 MHz, DMSO-d6): δ = 7.40–7.30 (m, 5 H), 7.17 (d, J =
8.7 Hz, 2 H), 6.84 (d, J = 8.7 Hz, 2 H), 5.12 (s, 2 H), 4.75 (s, 2 H), 4.41 (s,
4 H), 3.71 (s, 3 H).
13C NMR (126 MHz, DMSO-d6): δ = 168.49, 158.62, 153.96, 136.31,
129.42, 128.82, 128.62, 128.24, 127.91, 113.87, 67.35, 55.22 (d, J =
7.1 Hz), 47.22, 41.54.
1250, 1185, 1170, 1030, 979, 834, 724 cm–1
.
1H NMR (500 MHz, DMSO-d6): δ = 7.07 (d, J = 8.9 Hz, 2 H), 7.01 (d, J =
8.9 Hz, 2 H), 6.98–6.93 (m, 4 H), 4.61 (s, 2 H), 4.53 (s, 2 H), 3.77 (s,
6 H), 3.75 (s, 4 H), 3.69 (s, 2 H).
13C NMR (126 MHz, DMSO-d6): δ = 170.29, 167.93, 159.15, 159.05,
129.88, 129.80, 126.76, 126.65, 114.23, 114.18, 55.72, 55.48, 55.46,
48.86, 45.57.
MS (ESI+): m/z (%) = 391.12 (100) [M + Na]+.
MS (APCI+): m/z (%) = 481.1 (100) [M + H]+.
1-(4-Methoxyphenyl)piperazine-2,6-dione Hydrobromide (20a)
Anal. Calcd for C24H24N4O7: C, 60.00; H, 5.03; N, 11.66. Found: C,
59.98; H, 5.06; N, 11.26.
4-Benzyloxycarbonyl-1-(4-methoxyphenyl)piperazine-2,6-dione (19a;
1.4 g, 0.004 mol) was added at r.t. in several portions to stirring 33%
HBr in acetic acid (7 mL). The resulting suspension was stirred vigor-
ously at r.t. for 1 h, then Et2O (30 mL) was added and the solid was
filtered off, washed with Et2O, and left under vacuum over NaOH in a
dessicator for 24 h.
4,4′-(1-Oxoethane-1,2-diyl)bis[1-(4-methoxybenzyl)piperazine-
2,6-dione] (21b)
Bromoacetyl bromide (0.32 g, 0.14 mL, 0.00159 mol) was slowly add-
ed to a solution of 1-(4-methoxybenzyl)piperazine-2,6-dione hydro-
bromide (20b; 1 g, 0.0032 mol) and DIPEA (0.82 g, 1.08 mL, 0.00635
mol) in MeCN (30 mL) under an inert atmosphere at 0 °C. The reac-
tion mixture was stirred at 0 °C for 15 min and then at r.t. for 24 h.
The solvent was evaporated under reduced pressure and the residue
was dissolved in EtOAc (35 mL) and washed with 0.5% HCl (35 mL).
The organic layer was separated, dried over anhydrous sodium sul-
Yield: 1.12 g (94%; 86% when started from 3 g of 19a); white solid;
mp 253–256 °C.
IR (ATR): 3057, 2748, 2617, 1757, 1688, 1610, 1575, 1513, 1469, 1391,
1359, 1307, 1275, 1259, 1241, 1182, 1068, 1027, 937, 862, 839, 792
cm–1
.
1H NMR (500 MHz, DMSO-d6): δ = 7.09 (d, J = 9.0 Hz, 2 H), 7.03 (d, J =
9.0 Hz, 2 H), 4.25 (s, 4 H), 3.78 (s, 3 H).
© Georg Thieme Verlag Stuttgart · New York — Synthesis 2016, 48, A–I

H
J. Roh et al.
Paper
Synthesis
fate, and evaporated. The product was purified by column chromatog-
raphy (EtOAc–hexane, 7:1).
Acknowledgment
This work was supported by the Czech Science Foundation (project
no. 13-15008S).
Yield: 0.62 g (77%); white solid; mp 77–80 °C.
IR (ATR): 2838, 1737, 1680, 1612, 1514, 1430, 1389, 1344, 1302, 1247,
1178, 1112, 1030, 973, 894, 823, 764 cm–1
.
1H NMR (500 MHz, CDCl3): δ = 7.32 (d, J = 8.6 Hz, 4 H), 6.82 (d, J =
8.8 Hz, 4 H), 4.87 (s, 2 H), 4.86 (s, 2 H), 4.46 (s, 2 H), 4.30 (s, 2 H), 3.78
(s, 3 H), 3.77 (s, 3 H), 3.56 (s, 4 H), 3.38 (s, 2 H).
13C NMR (126 MHz, CDCl3): δ = 168.84, 166.29, 159.28, 159.05,
130.79, 130.43, 128.58, 127.99, 113.84, 113.76, 56.78, 55.79, 55.20,
48.48, 45.26, 42.46, 41.83.
Supporting Information
Supporting information for this article is available online at
http://dx.doi.org/10.1055/s-0035-1562618.
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References
MS (APCI+): m/z (%) = 509.2 (100) [M + H]+.
(1) Gianni, L.; Herman, E. H.; Lipshultz, S. E.; Minotti, G.; Sarvazyan,
N.; Sawyer, D. B. J. Clin. Oncol. 2008, 26, 3777.
(2) Jirkovsky, E.; Lencova-Popelova, O.; Hroch, M.; Adamcova, M.;
Mazurova, Y.; Vavrova, J.; Micuda, S.; Simunek, T.; Gersl, V.;
Sterba, M. Toxicology 2013, 311, 191.
Anal. Calcd for C26H28N4O7: C, 61.41; H, 5.55; N, 11.02. Found: C,
61.46; H, 5.63; N, 10.68.
4-(2-Bromoacetyl)piperazine-2,6-dione (22)
Bromoacetyl bromide (5.17 g, 2.24 mL, 0.0256 mol) was slowly added
to a suspension of piperazine-2,6-dione hydrobromide (10; 5 g,
0.0256 mol) and DIPEA (6.63 g, 8.77 mL, 0.051 mol) in MeCN (50 mL)
at 0 °C in an ice bath. The reaction mixture was stirred for 15 min at
0 °C and then at r.t. for 1 h. The solvent was then evaporated, and the
product was purified by column chromatography (CHCl3–EtOAc, 1:1).
(3) Simunek, T.; Sterba, M.; Popelova, O.; Adamcova, M.; Hrdina, R.;
Gersl, V. Pharmacol. Rep. 2009, 61, 154.
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P.; Gersl, V.; Simunek, T. Antioxid. Redox Signal. 2013, 18, 899.
(5) Hasinoff, B. B.; Hellmann, K.; Herman, E. H.; Ferrans, V. J. Curr.
Med. Chem. 1998, 5, 1.
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2003, 100, 10629.
(7) Zhang, S.; Liu, X.; Bawa-Khalfe, T.; Lu, L.-S.; Lyu, Y. L.; Liu, L. F.;
Yeh, E. T. H. Nat. Med. 2012, 18, 1639.
Yield: 3.73 g (62%); white solid; mp 157–159 °C (with decomp.).
IR (ATR): 3092, 2831, 1713, 1698, 1628, 1472, 1376, 1310, 1276, 1240,
1187, 1118, 1046, 976, 857 cm–1
.
1H NMR (500 MHz, acetone-d6): δ = 10.20 (s, 1 H), 4.50 (s, 2 H), 4.42
(s, 2 H), 4.22 (s, 2 H).
13C NMR (126 MHz, acetone-d6): δ = 169.13, 168.73, 166.16, 49.63,
45.53, 27.15.
(8) Herman, E. H.; Elhage, A. N.; Creighton, A. M.; Witiak, D. T.;
Ferrans, V. J. Res. Commun. Mol. Pathol. Pharmacol. 1985, 48, 39.
(9) Herman, E. H.; Zhang, J.; Hasinoff, B. B.; Chadwick, D. P.; Clark, J.
R.; Ferrans, V. J. Cancer Chemother. Pharmacol. 1997, 40, 400.
(10) Jirkovska-Vavrova, A.; Roh, J.; Lencova-Popelova, O.; Jirkovsky,
E.; Hruskova, K.; Potuckova-Mackova, E.; Jansova, H.; Haskova,
P.; Martinkova, P.; Eisner, T.; Kratochvil, M.; Sus, J.; Machacek,
M.; Vostatkova-Tichotova, L.; Gersl, V.; Kalinowski, D. S.; Muller,
M. T.; Richardson, D. R.; Vavrova, K.; Sterba, M.; Simunek, T. Tox-
icol. Res. (Cambridge, U. K.) 2015, 4, 1098.
(11) Prevention and treatment of cardiac conditions: Cross, M. R.;
Dilly, S. G.; Scaife, M. US2008/275036, 2008.
(12) Bis diketopiperazines: Creighton, A. M. US3941790, 1976.
(13) Barot, N. R.; Elvidge, J. A. J. Chem. Soc., Perkin Trans. 1 1972,
1009.
(14) Mancilla, T.; Carrillo, L.; Zamudio-Rivera, L. S.; Beltran, H. I.;
Farfan, N. Org. Prep. Proced. Int. 2002, 34, 87.
(15) Hou, X.; Ge, Z.; Wang, T.; Guo, W.; Wu, J.; Cui, J.; Lai, C.; Li, R.
Arch. Pharm. 2011, 344, 320.
(16) Shvedaite, I. P. Chem. Heterocycl. Compd. 1995, 31, 63.
(17) Dinsmore, C. J.; Beshore, D. C. Tetrahedron 2002, 58, 3297.
(18) Čapek, K.; Čapková, J.; Jarý, J.; Trška, P. Collect. Czech. Chem.
Commun. 1984, 49, 1914.
(19) Cignarella, G.; Nathanson, G. J. Org. Chem. 1961, 26, 1500.
(20) Henry, D. W. J. Heterocycl. Chem. 1966, 3, 503.
(21) Gregory, H.; Morley, J. S.; Smith, J. M.; Smithers, M. J. J. Chem.
Soc. C 1968, 715.
Anal. Calcd for C6H7BrN2O3: C, 30.66; H, 3.00; N, 11.92. Found: C,
30.31; H, 2.96; N, 11.62.
4,4′-(1-Oxoethane-1,2-diyl)bis(piperazine-2,6-dione) (13)
DIPEA (1.1 g, 1.48 mL, 0.0085 mol) was added to a suspension of pip-
erazine-2,6-dione hydrobromide (10; 1.66 g, 0.0085 mol) in DMF (13
mL) under an argon atmosphere. The suspension became a solution in
several seconds. Then, 4-(2-bromoacetyl)piperazine-2,6-dione (22; 2
g, 0.0085 mol) in DMF (7 mL) was added, followed by the addition of
further DIPEA (1.1 g, 1.48 mL, 0.00851 mol, 1 equiv). The reaction
mixture was stirred at r.t. for 24 h. Upon completion, as determined
by TLC (EtOAc–acetone, 1:1), the solvent was evaporated under re-
duced pressure. The crude product was washed with warm MeCN (30
mL) and with H2O (2 × 15 mL) to give the pure product (57%). An addi-
tional 12% of the pure product crystallized from MeCN upon cooling.
The product was filtered, washed with water and dried.
Yield: 1.57 g (69%); white solid; mp 247–250 °C.
IR (ATR): 3016, 2834, 1715, 1617, 1488, 1410, 1325, 1299, 1248, 1224,
1181, 1147, 1027, 887, 837, 741, 722 cm–1
.
1H NMR (500 MHz, DMSO-d6): δ = 11.36 (s, 1 H), 11.10 (s, 1 H), 4.30 (s,
2 H), 4.24 (s, 2 H), 3.50 (s, 2 H), 3.42 (s, 4 H).
13C NMR (126 MHz, DMSO-d6): δ = 171.24, 169.56, 169.35, 167.80,
55.94, 54.58, 47.67, 44.40.
(22) Snapka, R. M.; Woo, S. H.; Blokhin, A. V.; Witiak, D. T. Biochem.
Pharmacol. 1996, 52, 543.
(23) Pham, T. Q.; Pyne, S. G.; Skelton, B. W.; White, A. H. Tetrahedron
Lett. 2002, 43, 5953.
(24) Hennessy, E. J.; Buchwald, S. L. J. Org. Chem. 2005, 70, 7371.
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2765.
MS (APCI+): m/z (%) = 269.0 (100) [M + H]+.
Anal. Calcd for C10H12N4O5: C, 44.78; H, 4.51; N, 20.89. Found: C,
44.39; H, 4.53; N, 20.55.
© Georg Thieme Verlag Stuttgart · New York — Synthesis 2016, 48, A–I

I
J. Roh et al.
Paper
Synthesis
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© Georg Thieme Verlag Stuttgart · New York — Synthesis 2016, 48, A–I