Synthesis and reactions of paramagnetic aromatic aldehydes as useful synthetic building blocks

Article abstract of DOI:10.1055/s-2004-829172

Starting from paramagnetic benzaldehyde, a benzophenone-type photoactivable spin label, paramagnetic warfarin and phenindione were synthesized. Nitration of protected benzylic alcohol led to 2-nitrobenzylmethanethiosulfonate, a thiol specific spin label,

Full text of DOI:10.1055/s-2004-829172

PAPER
2115
Synthesis and Reactions of Paramagnetic Aromatic Aldehydes as Useful
Synthetic Building Blocks
Paramagnetic
A
a
romatic
A
l
m
dehydes as
U
seful
S
á
ynthetic Bu
s
ilding Blocks Kálai,a Győző Kulcsár,a József Jekő,b Erzsébet Ősz,c Kálmán Hideg*a
a
Institute of Organic and Medicinal Chemistry, University of Pécs, P. O. Box 99, 7602 Pécs, Hungary
Fax +36(72)536219; E-mail: kalman.hideg@aok.pte.hu
b
c
ICN Hungary, P. O. Box 1, 4440 Tiszavasvári, Hungary
Department of Biochemistry and Medical Chemistry, University of Pécs, P. O. Box 99, 7602 Pécs, Hungary
Received 6 May 2004
Dedicated to Prof. Károly Lempert on the occasion of his 80th birthday
tions of this compound to spin labels and biologically ac-
tive paramagnetic compounds.
Abstract: Starting from paramagnetic benzaldehyde, a benzophe-
none-type photoactivable spin label, paramagnetic warfarin and
phenindione were synthesized. Nitration of protected benzylic alco-
hol led to 2-nitrobenzylmethanethiosulfonate, a thiol specific spin
label, and 2-nitroaldehyde, which was a key compound for para-
magnetic indigo, salicylic acid and nifedipine.
Treatment of 1 with phenylmagnesium bromide and oxi-
dation of the intermediate secondary alcohol (not isolated)
with MnO2 gave the spin label 2 with the benzophenone
photoactivable group.13 Reduction of 1 with NaBH4 yield-
ed the benzyl alcohol 3. The nitration of 3 with a mixture
of nitric acid–sulfuric acid unfortunately oxidized the al-
cohol 3 to aldehyde 1 without nitration. The problem
could be solved by protecting of benzylic hydroxyl group
as its acetate.14 To achieve this, compound 3 was treated
with acetyl chloride in the presence of Et3N to afford ester
4. Nitration of 4 at low temperature gave the desired 2-ni-
tro derivative without hydrolysis of protecting group and
the crude product was deacetylated by Zemplen’s method
to give 2-nitroalcohol 5. The alcohol was converted to the
corresponding mesylate, with methanesulfonyl chloride
in the presence of Et3N and this mesylate was substituted
with bromine by treatment with LiBr in acetone15 to give
compound 6. The methanethiosulfonate reagent 7 was ob-
tained by treatment of bromide 6 with NaSSO2CH3 in
aqueous acetone. This reagent is capable of making re-
versible disulfide bond with SH group of cysteine in pro-
teins,16 while the nitro group is capable of making further
secondary interactions, such as H-bonds.
Key words: anticoagulant, free radicals, drugs, indole, pyridines
The past 40 years have witnessed the more extended utili-
zation of nitroxides on many fields in chemistry such as
co-oxidants,1 stoichiometric and catalytic antioxidants,2
spin labels,3 MRI agents4 and spin traps.5 The first and the
most wide-spread utilization from the above mentioned
applications is the spin label, which is used to evaluate
function and structure of a protein or an enzyme. This
means that a paramagnetic reagent modifies the protein
side chain to get information on structure and function by
EPR spectroscopy. The other approach for this kind of bi-
ological study is modification of a small molecule, actual-
ly a substrate, without significant change in its structure.
In practice this requires that the characteristic functional
groups must remain intact, so a carbon–carbon bond for-
mation is needed to form in the presence of nitroxide free
radical moiety. In our laboratory we have synthesized sev-
eral biologically active molecules modified with stable ni-
troxide free radical such as flavones,6 galactose,7 ebselen,8
omeprazole,9 and menadione.9 These paramagnetically
modified biologically active compounds can be obtained
through different methods. One of the most important ap-
proach is the Grignard reaction of nitrones, followed by
oxidation of the N-hydroxylamine product.10–12 Treatment
of TMPO (2,2,5-trimethyl-3,4-dihydro-2H-pyrrol-1-ox-
ide) with 4-dimethoxymethylphenylmagnesium bromide
followed by removal of protecting group gave paramag-
netic benzaldehyde 1,8 which could be oxidized or re-
duced for synthesis of useful substrates. This readily
available spin labeled benzaldehyde was published from
our laboratory earlier,8 however, further utilization was
not outlined. In this paper we report possible transforma-
Aldol condensation of 1 with excess acetone gave the
paramagnetic benzalacetone 8 in the presence of base, the
Michael addition of this product with 4-hydroxycouma-
rine in water gave the 9 paramagnetic derivative of war-
farin, the most widely prescribed anticoagulant drug.17
We hope nitroxide moiety will synergize the effect of
warfarin, because nitroxides were reported to exhibit also
anticoagulant effect.18 An other paramagnetic anticoagu-
lant compound was achieved with condensation of com-
pound 1 with phthalide in ethanolic NaOEt solution
followed by rearrangement to give indan-1,3-dione 10,
the paramagnetic phenindione.19 Treatment of compound
10 with hydrazine hydrate followed by oxidation of hy-
droxylamine in pyrrolidine ring with MnO2 afforded com-
pound 1120 as a paramagnetic derivative of phthalazinone
type PARP inhibitors21 (Scheme 1).
Oxidation of compound 5 with activated MnO2 in
chloroform22 yielded nitroaldehyde 12, a key compound
for further synthesis. To prove the substitution pattern of
SYNTHESIS 2004, No. 13, pp 2115–2120
Advanced online publication: 27.07.2004
DOI: 10.1055/s-2004-829172; Art ID: P04204SS
© Georg Thieme Verlag Stuttgart · New York
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x.
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x
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T. Kálai et al.
PAPER
Scheme 1 Reagents and conditions: (a) phenylmagnesium bromide (1.1 equiv), THF, reflux 1 h, r.t., 8 h, then aq NH4Cl, activated MnO2
(5.0 equiv), CHCl3, reflux, 1 h, 47%; (b) NaBH4 (1.1 equiv), EtOH, r.t., 72%; (c) H2SO4–HNO3 (2:1), 0 °C → r.t., 2 h, then K2CO3 to pH 8,
66%; (d) AcCl (1.1 equiv), Et3N (1.1 equiv), CH2Cl2, 0 °C → r.t., 1 h, 75%; (e) for 5 H2SO4–HNO3 (2:1), 0 °C → r.t., 3 h, then K2CO3 to pH 8,
extraction with CHCl3, then NaOMe (0.1 equiv), MeOH, r.t., 30 min, 42%; (f) MsCl (1.1 equiv), Et3N (1.1 equiv), CH2Cl2, 0 °C → r.t., 1 h,
then LiBr (2.0 equiv), acetone, reflux, 1 h, 54%; (g) NaSSO2CH3 (2.0 equiv), acetone–H2O, 1 h, reflux, 43%; (h) acetone, aq NaOH (1.0 equiv),
r.t., 12 h, 61%; (i) 4-hydroxycoumarine (1 equiv), H2O, reflux, 12 h, 30%; (j) phthalide (1.0 equiv), MeOH, NaOMe (4 equiv), 0 °C → r.t., then
reflux 5 h, 36%; (k) N2H4·H2O, reflux, 3 h, then H2O, extraction with CHCl3, activated MnO2 (1 equiv), O2, 15 min, r.t., 42%.
the benzene ring by NMR spectroscopy compound 12 was oxidized pyridine compound 18 as a side product
reduced by ascorbic acid followed by treatment with (Scheme 2).27–29
acetyl chloride in the presence of Et3N to give the diamag-
In conclusion, paramagnetic benzaldehyde and 2-ni-
trobenzaldehyde are important starting materials for the
netic O-acetyl compound.23 The substitution pattern of the
aromatic region could be unequivocally established by the
synthesis for both spin labels and paramagnetically modi-
multiplicity of the aromatic protons as well as the cou-
fied bioactive molecules. Further application of these
pling constant values determined from the 1H NMR spec-
molecules is under progress.
trum of diamagnetic O-acetyl compound.
The reaction of 12 with acetone and NaOH gave a para-
Melting points were determined with a Boetius micro melting point
magnetic indigo biradical 13 in a typical Baeyer–Drewson
reaction featuring the 2-nitrobenzaldehyde.24 Oxidation of
12 with Ag2O in aqueous alkali solution12 gave the 2-ni-
trobenzoic acid 14, which was reduced with ammonium
formate in the presence of Pd/C catalyst25 to the paramag-
netic anthranilic acid 15. To obtain the paramagnetic sali-
cylic acid this compound was diazotized with NaNO2 in
formic acid and the diazonium compound was hydrolyzed
in aqueous formic acid to give paramagnetic salicylic acid
16.26 Reaction of 12 with ammonium acetate and methyl
acetoacetate in a trimethylsilyl iodide mediated reaction
under mild condition gave the dihydropyridine methyl es-
ter 17 as a paramagnetic derivative of the nifedipine drug,
which is a well known calcium channel blocker and the
apparatus and are uncorrected. Elemental analyses (C, H, N, S)
were performed on Fisons EA 1110 CHNS elemental analyzer. The
IR (Specord 85) spectra were in each case consistent with the as-
signed structure. Mass spectra were recorded on a VG TRIO-2 in-
strument in the EI mode. NMR spectra were recorded with Varian
1
UNITYINOVA 400 WB (400/100 MHz for H/13C) spectrometer.
Chemical shifts are referenced to Me4Si (1H) or to the residual sol-
vent signal (13C). Measurements were run at 298 K probe tempera-
ture in CDCl3 solution. Structure elucidation was based on gradient
enhanced 13C-1H HSQC and 13C-1H HMBC experiments executed
using standard Varian software.
ESR spectra were taken on Miniscope MS 200 in 10–4 M CHCl3 so-
lution and all monoradicals gave triplett line aN = 14.0–14.4 G, and
13 biradical gave quintet line aN1 = 14.0 G, aN2 = 7.3 G. Flash col-
umn chromatography was performed on Merck Kieselgel 60
Synthesis 2004, No. 13, 2115–2120 © Thieme Stuttgart · New York

PAPER
Paramagnetic Aromatic Aldehydes as Useful Synthetic Building Blocks
2117
2-(4-Acetoxymethylphenyl)-2,5,5-tetramethylpyrrolidin-1-
yloxy Radical (4)
To a stirred solution of compound 3 (4.68 g, 20.0 mmol) and Et3N
(2.22 g, 22.0 mmol) in CH2Cl2 (40 mL) was added AcCl (1.72 g,
22.0 mmol) dropwise at 0 °C. The mixture was allowed to warm to
r.t. and the stirring was continued for further 1 h. The mixture was
washed with brine (20 mL), the organic phase was dried (MgSO4),
filtered and evaporated. The residue was purified by flash column
chromatography (hexane–Et2O, 2:1) to give compound 4 as an or-
ange oil; yield: 4.14 g (75%); Rf = 0.41 (hexane–EtOAc, 2:1).
2-(4-Hydroxymethyl-3-nitrophenyl)-2,5,5-tetramethylpyrroli-
din-1-yloxy Radical (5)
To a vigorously stirred solution of 65% HNO3 (5 mL) and 98%
H2SO4 (10 mL) was added compound 4 (4.17 g, 15.0 mmol) drop-
wise at 0 °C. The mixture was stirred for further 3 h at 0 °C, then it
was allowed to warm to r.t. It was poured in portions into crushed
ice (100 g) in a 500 mL beaker with stirring and basified with solid
K2CO3 to pH 8 (extensive foaming !). The solution was extracted
with CHCl3 (3 × 40 mL), the combined organic phases were dried
(MgSO4), filtered and evaporated. The residue was dissolved in
MeOH (20 mL) and NaOMe [prepared freshly by dissolving Na
metal (35 mg, 1.5 mmol) in MeOH (10 mL)] was added and allowed
to stand for 30 min. at r.t. The MeOH was evaporated off and the
residue was dissolved in CHCl3 (40 mL). The CHCl3 layer was
washed with 5% aq H2SO4 solution (15 mL), brine (15 mL), dried
(MgSO4), filtered and evaporated and the residue was purified by
flash column chromatography (hexane–EtOAc) to give compound
5 as a red oil; yield: 1.75 g, (42%); Rf = 0.39 (CHCl3–Et2O, 2:1).
2-(4-Bromomethyl-3-nitrophenyl)-2,5,5-tetramethylpyrrolidin-
1-yloxy Radical (6)
Scheme 2 Reagents and conditions: (a) MnO2 (5.0 equiv), CHCl3,
reflux, 1 h, 59%; (b) acetone, aq 10% NaOH (1.0 equiv), r.t., 30 min,
33%; (c) for 14 Ag2O (2.0 equiv), 10% aq NaOH–THF, 60 °C, 2 h,
then H+, 44%; (d) HCO2NH4 (8.0 equiv), MeOH, 10% Pd/C (cat.), N2,
40 °C, 2 h, then activated MnO2, O2, CHCl3, 10 min, 32%; (e) 80% aq
HCO2H, NaNO2 (1.0 equiv), 0 °C, 30 min, then H2O, 0 °C → 40 °C,
25%; (f) methyl acetoacetate (2 equiv), NH4OAc (2 equiv), TMSCl
(1.0 equiv), NaI (1.0 equiv), MeCN, r.t., 3d, 17% for 17 and 10% for
18.
To a stirred solution of alcohol 5 (2.79 g, 10.0 mmol) and Et3N (1.1
g, 11.0 mmol) in CH2Cl2 (20 mL) was added MeSO2Cl (1.26 g, 11.0
mmol) dropwise at 0 °C. The mixture was allowed the warm to r.t.
and stirred for further 1 h. The organic phase was washed with H2O
(2 × 10 mL), dried (MgSO4), filtered and evaporated. The residue
was dissolved in anhyd acetone (20 mL), LiBr (1.74 g, 20.0 mmol)
was added and the mixture was stirred and refluxed for 1 h. The sol-
vent was evaporated, the residue was dissolved in Et2O (20 mL) and
was washed with H2O (2 × 10 mL). The organic phase was separat-
ed, dried (MgSO4), filtered, evaporated and the residue was purified
by flash column chromatography (hexane–Et2O) to give compound
6 as a red solid; yield: 1.84 g (54%); mp 58–60 °C; Rf = 0.42 (hex-
ane–EtOAc, 2:1).
(0.040–0.063 mm). Qualitative TLC was carried out on commer-
cially prepared plates (20 × 20 × 0.02 cm) coated with Merck Kie-
selgel GF254. Compounds 18 and 38 were prepared according to
published procedures. The physical and spectral data of all new
compounds are listed in Table 1.
2-(3-Nitro-4-methanethiosulfonylmethylphenyl)-2,5,5-tetra-
methylpyrrolidin-1-yloxy Radical (7)
4-(1-Oxyl-2,5,5-trimethylpyrrolidin-2-yl)benzophenone Radi-
cal (2)
To a solution of compound 6 (1.71 g, 5.0 mmol) in acetone (10 mL)
and H2O (5 mL) was added NaSSO2Me (1.34 g, 10.0 mmol) and the
mixture was refluxed for 1 h. The solvent was evaporated, the resi-
due dissolved in CHCl3 (20 mL) and washed with brine (10 mL).
The organic phase was separated, dried (MgSO4), filtered, evaporat-
ed and the residue was purified by flash column chromatography
(hexane–EtOAc) to give the title compound 7 as an amorphous ma-
terial; yield: 803 mg (43%); Rf = 0.58 (CHCl3–Et2O, 2:1).
To a solution of phenylmagnesium bromide [prepared from bro-
mobenzene (942 mg, 6.0 mmol) and Mg turnings (144 mg, 6.0
mmol) in THF (10 mL)], was added a solution of compound 1 (1.16
g, 5.0 mmol) in THF (10 mL) dropwise and the mixture stirred and
refluxed for 1 h and allowed to stand overnight at r.t. After quench-
ing with sat. aq NH4Cl solution, Et2O (10 mL) was added, the or-
ganic layer was separated, dried (MgSO4), filtered and evaporated.
The residue was dissolved in CHCl3 (30 mL), activated MnO2 (2.18
g, 25.0 mmol) was added and the mixture was stirred and refluxed
for 1 h. After cooling, the MnO2 was filtered off, the filtrate evapo-
rated and the residue was purified by flash column chromatography
(hexane–Et2O, 2:1) to give compound 2 as a yellow solid; yield: 693
mg (47%); mp 87–88 °C; Rf = 0.47 (hexane–EtOAc, 2:1).
4-[4-(1-Oxyl-2,5,5-trimethylpyrrolidin-2-yl)phenyl]but-3-en-2-
one Radical (8)
To a solution of compound 1 (1.39 g, 6.0 mmol) in acetone (15 mL)
was added NaOH (240 mg, 6.0 mmol) in H2O (6 mL) and the mix-
ture was stirred for 5 h at r.t. and allowed to stand overnight at r.t.
The acetone was evaporated off, the aqueous phase was extracted
with CHCl3 (2 × 30 mL). The combined organic phases were dried
(MgSO4), filtered, evaporated and after purification by flash col-
umn chromatography (hexane–EtOAc) compound 8 was obtained
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2118
T. Kálai et al.
PAPER
Table 1 Compounds 2–18 Prepared
Compounda Yield
(%)
mp
ºC
IR (Neat or Nujol)
cm–1
MS
m/z (%)
2
4
5
6
47
75
42
54
87–88
oil
1660 (C=O), 1590, 1550 (C=C)
1720 (C=O), 1600, 1550, 1500 (C=C)
3400 (OH), 1620 (C=C), 1530 (NO2)
1650, 1600 (C=C), 1525 (NO2)
308 (M+, 8), 294 (26), 278 (66), 105 (100)
276 (M+, 10), 262 (11), 246 (100), 217 (13)
279 (M+, 9), 265 (13), 234 (20), 148 (100)
oil
58–60
341/343 (M+, 3/3), 327/329 (9/9), 311/313 (2/2),
176 (100)
7
8
43
61
30
36
42
59
semisolid
60–62
1620, 1550 (C=C), 1540 (NO2)
1660 (C=O), 1600, 1560 (C=C)
1710(C=O), 1650, 1620, 1570 (C=C)
1700 (C=O), 1550 (C=C)
373 (M+, 6), 343 (5), 293 (30), 159 (100)
272 (M+, 13), 258 (41), 242 (49), 43(100)
434 (M+, 6), 404 (26), 346 (13), 242 (100)
348 (M+, 8), 334 (24), 318 (100), 262 (40)
362 (M+, 2), 332 (100), 318 (9), 276 (8)
277 (M+, 8), 263 (17), 232 (15), 146 (100)
9
115–118
86–87
10
11
12
201–204
51–53
3180 (NH), 1650 (C=O), 1560 (C=C)
1680 (C=O), 1620, 1590 (C=C),
1520 (NO2)
13
14
15
33
44
32
192–193
188–190
197–200
1640 (C=O), 1610 (C=C)
514 (M+, 11), 499 (20), 484 (31), 469 (100)
293 (M+, 11), 263 (22), 177 (76), 159 (100)
263 (M+, 15), 233 (42), 177 (80), 159 (100)
1660 (C=O), 1600 (C=C), 1525 (NO2)
3480, 3310 (NH2), 1650 (C=O), 1620,
1590, 1550 (C=C)
16
17
25
17
232–234
115–117
1660 (C=O), 1590, 1560 (C=C)
264 (M+, 4), 250 (35), 234 (100), 216 (69)
TSP: 473 [M + H] +
3300 (NH), 1670 (C=O), 1600 (C=C),
1530 (NO2)
18
10
90–92
1700 (C=O), 1650 (C=C), 1540 (NO2)
TSP: 471 [M + H] +
a Satisfactory microanalyses obtained: C 0.18; H 0.18; N 0.17. Exception: 18, H –0.46.
as a yellow solid; yield: 995 mg (61%); mp 60–62 °C; Rf = 0.26
(hexane–EtOAc, 2:1).
tography (hexane–EtOAc) to give compound 10 as a pink solid;
yield: 627 mg (36%); mp 86–88 °C; Rf = 0.51 (CHCl3–Et2O, 2:1).
4-Hydroxy-3-{1-[4-(1-Oxyl-2,5,5-trimethylpyrrolidin-2-yl)phe-
nyl]-3-oxobutyl}chromen-2-one Radical (9)
4-[4-(1-Oxyl-2,5,5-trimethylpyrrolidin-2-yl)benzyl]-2H-
phthalazin-1-one Radical (11)
A mixture of paramagnetic benzalacetone 8 (1.08 g, 4.0 mmol) and
4-hydroxycoumarine (648 mg, 4.0 mmol) was added to H2O (10
mL), and the suspension was refluxed for 12 h. After cooling, the
mixture was extracted with CHCl3 (20 mL). The organic layer was
separated, dried (MgSO4), filtered, evaporated and after purification
by flash column chromatography (hexane–EtOAc) compound 9
was obtained as a light-yellow solid; yield: 520 mg (30%); mp 115–
118 °C; Rf = 0.32 (CHCl3–Et2O, 2:1).
A solution of compound 10 (348 mg, 1.0 mmol) in hydrazine hy-
drate (7 mL) was refluxed for 3 h upon which the red solution be-
came pale yellow. After cooling, H2O (20 mL) was added, the
mixture was saturated with solid NaCl and extracted with CHCl3
(2 × 20 mL). The combined organic phases were dried (MgSO4),
activated MnO2 (87 mg, 1.0 mmol) was added and O2 was bubbled
through for 15 min. The mixture was filtered, evaporated and the
residue was purified by flash column chromatography (CHCl3–
Et2O) to give the title compound as a pale yellow solid; yield: 152
mg (42%); mp 201–204 °C; Rf = 0.27 (CHCl3–Et2O, 2:1).
2-[4-(1-Oxyl-2,5,5-trimethylpyrrolidin-2-yl)phenyl]indan-1,3-
dione Radical (10)
To a stirred solution of aldehyde 1 (1.16 g, 5.0 mmol) and phthalide
(670 mg, 5.0 mmol) in MeOH (20 mL) was added methanolic
NaOMe solution [freshly prepared by dissolving Na metal (460 mg,
20.0 mmol) in MeOH (20 mL)] dropwise at 0 °C. After warming up
to r.t., the mixture was refluxed for 5 h and cooled. It was then
poured into ice water (100 mL). The mixture was extracted with
Et2O (30 mL), the organic phase was discarded, and the aqueous
phase was acidified with aq 2.0 M HCl and extracted with EtOAc
(2 × 20 mL). The combined organic phases were dried (MgSO4),
filtered, evaporated and after purification by flash column chroma-
2-Nitro-4-(1-oxyl-2,5,5-trimethylpyrrolidin-2-yl)benzaldehyde
Radical (12)
To a stirred solution of alcohol 5 (5.58 g, 20.0 mmol) in CHCl3 (80
mL) was added activated MnO2 (8.70 g, 0.1 mol) and the mixture
was refluxed for 1 h. The MnO2 was filtered out the solvent was
evaporated and after purification of the residue by flash column
chromatography (hexane–Et2O) compound 12 was obtained as a
wine-red solid; yield: 3.26 g (59%); mp 51– 53 °C; Rf = 0.30 (hex-
ane–EtOAc, 2:1).
Synthesis 2004, No. 13, 2115–2120 © Thieme Stuttgart · New York

PAPER
Paramagnetic Aromatic Aldehydes as Useful Synthetic Building Blocks
2119
2-Nitro-4-(1-acetoxy-2,5,5-trimethylpyrrolidin-2-yl)benzalde-
hyde for NMR Study
2-Hydroxy-4-(1-oxyl-2,5,5-trimethylpyrrolidin-2-yl)benzoic
Acid Radical (16)
To a solution of radical 12 (277 mg, 1.0 mmol) in dioxane (10 mL),
was added a solution of ascorbic acid (880 mg, 5.0 mmol) in H2O (5
mL) and the mixture was stirred at 40 °C for 15 min under N2. The
pale yellow or colorless solution was extracted with CHCl3 (2 × 20
mL), dried (MgSO4) under N2. First acetyl chloride (86 mg, 1.1
mmol) and then slowly Et3N (111 mg, 1.1 mmol) were added, the
temperature being kept below 10 °C. The stirring was continued for
1 h, after which the mixture was filtered and the filtrate was evapo-
rated in vacuo to dryness. The residue was taken up in brine (10 mL)
and extracted with EtOAc (2 × 15 mL). The combined organic lay-
ers were dried (MgSO4), filtered, evaporated and after flash chro-
matography (hexane–Et2O) the O-acetyl derivative was obtained as
a yellow oil; yield: 112 mg (35%); Rf = 0.48 (hexane–EtOAc, 2:1).
1H NMR (400 MHz, CDCl3): d = 1.22 (s, 3 H, CH3), 1.23 (s, 3 H,
CH3), 1.56 (s, 3 H, CH3), 1.68 (m, 1 H, CH2), 1.83 (m, 1 H, CH2),
1.97 (s, 3 H, COCH3), 1.98 (m, 1 H, CH2), 2.17 (m, 1 H, CH2), 7.82
(d, J = 8.1 Hz, 1 Harom), 8.13 (dd, J = 8.1, 1.5 Hz, 1 Harom), 8.43 (d,
J = 1.5 Hz, 1 Harom), 10.28 (s, 1 H, CHO).
To a stirred solution of compound 15 (526 mg, 2.0 mmol) in 80%
aq HCO2H (8 mL) was added a solution of NaNO2 (138 mg, 2.0
mmol) in H2O (2 mL) dropwise and the mixture was stirred at 0 °C
for 30 min. The mixture was diluted with H2O (10 mL) and slowly
warmed to 40 °C and after the solvents were evaporated off to dry-
ness, the residue was dissolved in brine (10 mL), extracted with
CHCl3 (2 × 15 mL). The combined organic phases were dried
(MgSO4), filtered, evaporated and after flash chromatography
(CHCl3–Et2O, 3:1) compound 16 was obtained as a yellowish-
brown solid; yield: 132 mg (25%); mp 232–234 °C; Rf = 0.26
(CHCl3–MeOH, 9:1).
4-[4-(1-Oxyl-2,5,5-trimethylpyrrolidin-2-yl)-2-nitrophenyl]-
2,6-dimethyl-1,4-dihydropyridine-3,5-dicarboxylic Acid Di-
methyl Ester Radical (17) and 4-[4-(1-Oxyl-2,5,5-trimethylpyr-
rolidin-2-yl)-2-nitrophenyl]-2,6-dimethylpyridine-3,5-
dicarboxylic Acid Dimethyl Ester Radical (18)
To a stirred solution of compound 12 (1.38g, 5.0 mmol) in MeCN
(10 mL), methyl acetoacetate (1.16 g, 10.0 mmol), and NH4OAc
(770 mg, 10.0 mmol), was added Me3SiCl (543 mg 5.0 mmol) drop-
wise and then NaI (745 mg, 5.0 mmol) was added in one portion and
the mixture was stirred for further 3 d at r.t. The mixture was poured
into iced-water (100 mL) and extracted with EtOAc (3 × 20 mL).
The combined organic phases were dried (MgSO4), filtered, evapo-
rated and after purification by flash column chromatography (hex-
ane–EtOAc) compounds 17 as the fifth and 18 as the fourth band
were obtained following aminocrotonate (Rf 0.51, hexane–EtOAc,
2:1), unreacted starting aldehyde 12 and probably a condensation
product of aldehyde 12 and methyl acetoacetate (m/z = 375, M+)
(Rf = 0.19, hexane–EtOAc, 2:1).
13C NMR (100 MHz, CDCl3): d = 18.9 (COCH3), 23.3 (CH3), 24.1
(CH3), 28.9 (CH3), 35.5 (CH2), 38.8 (CH2), 66.0 (quat C), 69.8 (quat
C), 122.3 (CHarom), 129.0 (arom C–CHO), 129.3 (CHarom), 131.6
(CHarom), 149.6 (arom C–NO2), 156.8 (quat arom C), 169.7
(COCH3), 188.1 (CHO).
(E)-6,6¢-Bis-(1-oxyl-2,5,5-trimethylpyrrolidin-2-yl)-1H,1¢H-
[2,2¢]biindolylidene-3,3¢-dione Biradical (13)
To a solution of compound 12 (277 mg, 1.0 mmol) in acetone (4
mL) was added aq 10% NaOH (0.4 mL, 1.0 mmol). The mixture
was allowed to stand at r.t. for 30 min., then it was diluted with H2O
(70 mL) and the precipitated blue solid was filtered to give com-
pound 13; yield: 87 mg (33%); mp 192–193 °C; Rf = 0.34 (CHCl3–
Et2O, 2:1).
17
Yield: 401 mg (17%); mp 115–117 °C; Rf = 0.11 (CHCl3–MeOH,
9:1).
2-Nitro-4-(1-oxyl-2,5,5-trimethylpyrrolidin-2-yl)benzoic Acid
Radical (14)
18
To a stirred suspension of freshly precipitated Ag2O (4.63 g 20.0
mmol) in 10% aq NaOH solution (20 mL) was added a solution of
aldehyde 12 (2.77 g, 10.0 mmol) in THF (5 mL) at 60 °C and the
mixture was stirred for 2 h. After cooling, the solution was filtered
through a Celite pad, and washed with MeOH (20 mL). The organic
solvents were evaporated off, the residue aqueous phase was acidi-
fied with 5% aq H2SO4 to pH 2 and extracted with CHCl3 (3 × 10
mL). The organic phase was dried (MgSO4), filtered, evaporated to
afford compound 14 as a yellow solid; yield: 1.29 g (44%); mp 188–
190 °C; Rf = 0.12 (CHCl3–MeOH, 9:1).
Yield: 235 mg (10%); mp 90–92 °C; Rf = 0.21 (CHCl3–Et2O, 2:1).
Acknowledgment
This work was supported by a grant from the Hungarian National
Research Foundations (OTKA T34307, T042951 and M 045190)
and Hungarian Ministry of Health, Social and Family Affairs (ETT
512/2003). Bolyai fellowships for T. K. and E. Ő. from the Hunga-
rian Academy of Sciences are gratefully acknowledged. The au-
thors thank Noémi Lazsányi for elemental analysis and Mária Szabó
for mass spectral measurements (ICN, Hungary).
2-Amino-4-(1-oxyl-2,5,5-trimethylpyrrolidin-2-yl)benzoic Acid
Radical (15)
To a stirred solution of compound 14 (1.76 g, 6.0 mmol) in MeOH
(40 mL) was added ammonium formate (3.0 g, 48.0 mmol) at 40 °C.
After dissolution of ammonium formate, 10% Pd/C catalyst (100
mg) was added and the mixture stirred for 2 h at this temperature un-
der N2. After consumption of the nitro acid 14, the mixture was fil-
tered through a Celite pad and washed with MeOH (30 mL) and the
filtrate was evaporated. The residue was dissolved in a mixture of
CHCl3 (30 mL) and MeOH (3 mL), and the organic solution was
washed with brine, and dried (MgSO4). Activated MnO2 (869 mg,
10.0 mmol) was added and O2 was bubbled through the solution for
10 min. The mixture was filtered, evaporated and after purification
by flash column chromatography (CHCl3–Et2O, 2:1) compound 15
was obtained, as a beige solid; yield: 505 mg (32%); mp 197–200
°C; Rf = 0.26 (CHCl3–MeOH, 9:1).
References
(1) (a) Formaggio, F.; Bonchio, M.; Crisma, M.; Peggion, C.;
Mezzato, S.; Polese, A.; Barazza, A.; Antonello, S.; Maran,
F.; Broxterman, Q. B.; Kaptein, B.; Kamphuis, J.; Vitale, R.
M.; Saviano, M.; Benedetti, E.; Toniolo, C. Chem.–Eur. J.
2002, 8, 84. (b) Nerbouh, N.; Bobbit, J. M.; Brückner, C.
Org. Prep. Proced. Int. 2004, 36, 1.
(2) Krishna, M. C.; DeGraff, W.; Hankovszky, H. O.; Sár, P. C.;
Kálai, T.; Jekő, J.; Russo, A.; Mitchell, J. B.; Hideg, K. J.
Med. Chem. 1998, 41, 3477.
(3) Hubbell, W. L.; Altenbach, C.; Hubbell, C. M.; Khorana, H.
G. Adv. Protein Chem. 2003, 63, 243.
Synthesis 2004, No. 13, 2115–2120 © Thieme Stuttgart · New York

2120
T. Kálai et al.
PAPER
(4) (a) Jászberényi, Z.; Brücher, E.; Jekő, J.; Hideg, K.; Kálai,
T.; Király, R. Eur. J. Inorg. Chem. 2003, 3601. (b) Malikal,
A. J.; Turro, N. J.; Bosman, A. W.; Cornel, J.; Meijer, E. W.
J. Phys. Chem. A 2003, 107, 8467.
(17) (a) Ikawa, M.; Stahmann, M. A.; Link, P. K. J. Am. Chem.
Soc. 1944, 66, 902. (b) Sadler, E. J. Nature 2004, 427, 493.
(c) Li, T.; Chang, C.-Y.; Jin, D.-J.; Lin, P.-J.; Khvorova, A.;
Stafford, D. W. Nature 2004, 427, 541.
(5) Li, B. B.; Blough, N. V.; Gutierrez, P. L. Free Radicals Biol.
Med. 2000, 29, 548.
(6) Müller, É.; Kálai, T.; Jekő, J.; Hideg, K. Synthesis 2000,
1415.
(7) Zhao, M.; Kálai, T.; Hideg, K.; Altenbach, C.; Hubbell, W.;
Kaback, R. H. Biochemistry 2000, 39, 11381.
(8) Gadányi, Sz.; Kálai, T.; Jekő, J.; Berente, Z.; Hideg, K.
Synthesis 2000, 2039.
(18) Zhdanov, R. I.; Kubatiev, A. A. In Bioactive Spin Labels;
Zhdanov, R. I., Ed.; Springer Verlag: Berlin, 1992, 461.
(19) Dieckmann, W. Ber. Dtsch. Chem. Ges. 1914, 47, 1435.
(20) Martin, N. M. B.; Smith, G. C. M.; White, C. R.; Newton, R.
F.; Douglas, D. G.; Eversley, P. J.; Vile, J. PCT Int. Appl.
WO 0236576, 2002; Chem. Abstr. 2002, 136, 355242.
(21) Cosi, C. Expert Opin. Ther. Pat. 2002, 12, 1047; Chem.
Abstr. 2003, 138, 100226.
(9) Kálai, T.; Jekő, J.; Hideg, K. Synthesis 2000, 831.
(10) Hideg, K.; Lex, L. J. Chem. Soc., Perkin Trans. 1 1986,
1431.
(11) Hankovszky, H. O.; Hideg, K.; Lovas, M. J.; Jerkovich, Gy.;
Rockenbauer, A.; Győr, M.; Sohár, P. Can. J. Chem. 1989,
67, 1392.
(22) Hideg, K.; Hankovszky, H. O.; Lex, L.; Kulcsár, Gy.
Synthesis 1980, 911.
(23) Hideg, K.; Sár, P. C.; Hankovszky, H. O.; Tamás, T.;
Jerkovich, Gy. Synthesis 1993, 390.
(24) Baeyer, A.; Drevson, B. Ber. Dtsch. Chem. Ges. 1882, 15,
2856.
(12) Kálai, T.; Jekő, J.; Hubbell, W. L.; Hideg, K. Synthesis 2003,
2084.
(13) Hermanson, G. T. Bioconjugate Techniques; Academic
Press: San Diego, 1995.
(14) (a) Green, W. T.; Wuts, P. G. M. Protective Groups in
Organic Synthesis, 2nd ed.; Wiley: New York, 1991.
(b) Kocienski, P. Protecting Groups, 2nd ed.; Georg Thieme
Verlag: Stuttgart, 2000.
(15) Hankovszky, H. O.; Hideg, K.; Lex, L. Synthesis 1980, 914.
(16) Berliner, L. J.; Grünwald, J.; Hankovszky, H. O.; Hideg, K.
Anal. Biochem. 1982, 119, 450.
(25) Ram, S.; Ehrenkaufer, R. E. Synthesis 1988, 91.
(26) Atkinson, A. R.; Holm-Hansen, D.; Nevers, A. D.; Marino,
A. S. J. Am. Chem. Soc. 1943, 65, 476.
(27) (a) Hantzsch, A. Liebigs Ann. Chem. 1882, 215, 1.
(b) Marciniak, G.; Delgado, A.; Leclerc, G.; Velly, J.;
Decker, N.; Schwartz, J. J. Med. Chem. 1989, 32, 1402.
(28) (a) Hankovszky, H. O.; Sár, P. C.; Hideg, K.; Jerkovich, Gy.
Synthesis 1991, 91. (b) Santander-G, I. P.; Nunez-Vergara,
L. J.; Squella, J. A.; Navarrete-Encina, P. A. Synthesis 2003,
2781.
(29) Sabitha, G.; Reddy, K. K. S. G.; Reddy, S. Ch.; Yadav, J. S.
Tetrahedron Lett. 2003, 44, 4129.
Synthesis 2004, No. 13, 2115–2120 © Thieme Stuttgart · New York