The Journal of Organic Chemistry
Article
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1180, 1142, 853, 754, 716, 650, 605; H NMR (500 MHz, CDCl3 +
15% CF3COOH, a mixture of two isomeric cations, slow exchange in
NMR time scale) major isomer (60%) 1.38 (d, J = 1.6 Hz, 3H), 1.50
(d, J = 1.6 Hz, 3H), 1.54 (m, 3H), 1.67 (d, J = 2.3 Hz, 3H), 2.27 (ddd,
J1 = 7.7 Hz, J2 = 13.7 Hz, J3 = 2.6 Hz, 1H), 2.50 (dd, J1 = 12.1 Hz, J2 =
13.7 Hz, 1H), 3.38 (dd, J1 = 7.7 Hz, J2 = 12.1 Hz, 1H), 10.64 (d, 0.5H
(one-half of the multiplet)), minor isomer (40%) 1.45 (d, J = 1.3 Hz,
3H), 1.55 (m, 3H), 1.60 (m, 6H), 2.35 (ddd, J1 = 14.4 Hz, J2 = 8.8 Hz,
1H) 2.39 (dd, J1 = 14.4 Hz, J2 = 7.3 Hz, J3 = 4.7 Hz, 1H) 3.27 (br t, J =
8.2 Hz, 1H), 10.46 (d, J = 80.5 Hz, 1H); 13C NMR (125.77 MHz;
CDCl3 + 10% CF3COOH) major isomer 16.1 (CH3), 23.7 (CH3),
25.0 (CH3), 27.1 (CH3), 37.4 (d, J =1.9 Hz, CH2), 47.3 (d, J = 4.7,
CH), 72.2 (d, J = 1.6 Hz, C), 74.4 (d, J = 3.2 Hz, C), 174.3 (br, C),
minor isomer 21.9 (CH3), 22.3 (CH3), 23.9 (CH3), 25.3 (CH3), 36.6
(d, J = 3.1 Hz, CH2), 49.8 (d, J = 3.1 Hz, CH), 72.4 (d, J = 2.1 Hz, C),
74.7(d, J = 2.9 Hz, C), 175.5 (C).
of nitroxide and/or destabilization of hydroxylamine by bulky
substituents.
EXPERIMENTAL SECTION
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Synthesis. Nitroxides In1,30 In2, In3, In5, In7−In10, Im2, Im3,31
In4,5 In11,32 Im1,33 Im5,14 PCA,34 In4D, and Im4D23 were prepared
according to the procedures described in the corresponding literature;
CPH-15N hydrochloride was prepared via hydrogenation of PCA-15N
using the procedure developed for the 14N-containing compound32
(see below); PCA-15N was prepared from triacetonamine-15N
(TAA-15N) according to a literature procedure;35 TAA-15N was
prepared from 15NH4Cl (99.9% isotope enrichment) according to the
method by Pirrwitz and Schwarz36 with minor modifications (Scheme
3).37
Scheme 3. Synthetic Scheme of CPH-15N Preparation
Method B (analogous to that in ref 34). Hydrochloric acid (36%, 1
mL, 11.6 mmol) was added to a solution of PCA (210 mg, 1.13 mmol)
in ethanol (2 mL). The mixture was heated to slow boiling and stirred
for 20 min to form a nearly colorless solution, which was allowed to
cool to ambient temperature and evaporate to dryness under reduced
pressure. Hydrochloric acid (36%, 2 mL) was added to the residue,
and the mixture was heated to slow boiling and stirred for 2 h,
evaporating to approximately one half of the initial volume. The
solution was allowed to cool to ambient temperature and placed in a
refrigerator (−18 °C) for 12 h. The crystalline precipitate of CPH was
filtered off and washed with cold hydrochloric acid to yield 206 mg
(80%).
Rate Constants of Nitroxide Reduction with Ascorbate. The
solutions of nitroxide (0.1 mM) were mixed under anaerobic
conditions in a glovebox at an oxygen level of <1 ppm with various
concentrations of ascorbate (1−100 mM) in 0.1 M Na-phosphate
buffer, pH 7.6. The mixture was transferred to the 50 μL glass capillary
tube, and the EPR spectrum was recorded using an X-band
spectrometer within 1−2 min after mixing. For the kinetics studies,
the double integral of the low-field component of the EPR spectrum
was monitored.
Cyclic Voltammetry. Cyclic voltammograms (CVs) of nitroxides
(2.0 × 10−3−10−2 M solutions in dry MeCN, 0.1 M Et4NClO4) were
performed at 295 K under an argon atmosphere with an SVA-1BM
electrochemical system (Bulgaria) equipped with a digital interface. A
stationary working cylindrical Pt electrode (S = 0.08 cm2) was used. A
Pt spiral was used as an auxiliary electrode. The peak potentials were
quoted with reference to saturated aq Hg/Hg2Cl2. Measurements were
carried out in a mode of triangular pulse potential with the sweep rate,
v, varied from 0.05 to 10 V/s. Measurements were performed in the
voltage range from −2.3 to +2.5 V.
1-Hydroxy-2,2,5,5-tetramethylpyrrolidine-3-carboxylic acid hy-
drochloride (CPH). This compound was incorrectly described
previously;34 for the characteristics of the zwitterion, see ref 35. The
samples of CPH were prepared using two different methods, both
giving equal yields, and the samples have identical spectral character-
istics.
Method A (analogous to that in ref 32). A solution of PCA (1.87 g,
10 mmol) in MeOH (50 mL) was placed in a 200 mL Erlenmeyer
flask equipped with an adapter for the gas supply with a valve at the
outlet, and then the wet catalyst (Pd/C, 1%, 200 mg) was added. A
magnetic stirring bar was placed in the flask. Then, the flask was
purged with nitrogen followed by hydrogen and connected to a
gasometer. The reaction mixture was vigorously stirred at ambient
temperature until gas absorption stopped (usually the amount of the
gas slightly exceeded the calculated value of 112 mL). The flask was
opened, and the reaction mixture was acidified with HCl to pH 2−3.
The catalyst was filtered off; the solvent was distilled off under reduced
pressure, and the residue was crystallized from concentrated
hydrochloric acid to give CPH (1.8 g, 80%): mp 190−192 °C dec
(from aq concentrated HCl); IR (KBr) νmax (cm−1) 1724, 1500, 1466,
1417, 1406, 1391, 1379, 1309, 1292, 1274, 1242, 1202, 1180, 1142,
Equilibrium in CPH-15N−Nitroxide Mixtures. Fresh stock
solutions (0.5 mM) were prepared by dissolving the nitroxides and
CPH-15N in a 15 mM solution of KOH in methanol containing 100
μM DTPA under anaerobic conditions in a glovebox filled with argon
(oxygen concentration below 5 ppm). The aliquots of each nitroxide
solution were mixed with an aliquot of CPH-15N, sealed, and left for
24 h or more in the glovebox at 25 °C. The solutions were then
transferred into 50 μL glass capillary tubes and sealed. The EPR
spectra were recorded on a spectrometer using a modulation
amplitude of 0.5 G and microwave power of 6.36 mW.
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1049, 986, 853, 764, 721, 650, 601; H NMR (400 MHz, (CD3)2SO)
1.24 (br s, 3H), 1.38 (br s, 3H), 1.41 (br s, 3H), 1.53 (br s, 3H), 2.08
(dd, J1 = 7.7 Hz, J2 = 13.4 Hz, 1H), 2.19 (br m, 1H), 3.13 (br m, 1H),
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11.6 (br s, 1H); H NMR (300 MHz, CDCl3 + 10% CF3COOH, a
mixture of two isomeric cations, slow exchange in NMR time scale)
major isomer (60%) 1.42 (s, 3H), 1.53 (s, 3H), 1.60 (s, 3H), 1.72 (s,
3H), 2.30 (dd, J1 = 7.7 Hz, J2 = 13.7 Hz, 1H), 2.54 (dd, J1 =12.1 Hz, J2
= 13.7 Hz, 1H), 3.41 (dd, J1 = 7.7 Hz, J2 = 12.1 Hz, 1H), 10.49 (s,
1H), minor isomer (40%) 1.49 (s, 3H), 1.59 (s, 3H), 1.63 (s, 3H),
1.64 (s, 3H), 2.38 and 2.42 (ABd, JAB = 14.5 Hz, Jd = 8.5 Hz, 2H), 3.30
(t, J = 8.5 Hz, 1H), 10.41 (s, 1H); 13C NMR (75 MHz; CDCl3 + 10%
CF3COOH) major isomer 16.0 (CH3), 23.6 (CH3), 25.0 (CH3), 27.1
(CH3), 37.4 (CH2), 47.3 (CH), 72.2 (C), 74.4 (C), 174.3 (C), minor
isomer 21.9 (CH3), 22.2 (CH3), 23.8 (CH3), 25.3 (CH3), 36.5 (CH2),
49.7 (CH), 72.4 (C), 74.7(C), 175.5 (C); Anal. Calcd for
C9H18NClO3 (%) C 48.32, H 8.11, N 6.26, Cl 15.85; found (%) C
48.17, H 8.12, N 6.26, Cl 15.78.
ASSOCIATED CONTENT
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S
* Supporting Information
The Supporting Information is available free of charge on the
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Copies of the H and 13C NMR spectra of CPH and
CPH-15N, EPR spectrum of PCA-15N, and calculation of
parameters Es,n and σI,n for multiple regression (PDF)
CPH-15N: mp 189−192 °C dec (from aq concentrated HCl); IR
(KBr) νmax (cm−1) 1724, 1495, 1466, 1408, 1391, 1381, 1292, 1202,
G
J. Org. Chem. XXXX, XXX, XXX−XXX