Asymmetric synthesis of L-proline regio- and stereoselectively labelled with deuterium

Article abstract of DOI:10.1016/S0957-4166(99)00083-X

A synthesis of L-proline where all of the ring methylenes are stereoselectively labelled with deuterium is described. A catalytic deuteration of protected 3,4-dehydro-L-proline using transition metal catalyst followed by RuO₄-oxidation gave a [3,4-D₂]pyroglutamic acid derivative. A syn-selective deuteration of the aminal derived from the pyroglutamate with Et₃SiD-BF₃ · OEt₂ furnished (2S,3S,4R,5S)-[3,4,5- D₃]proline. The present procedure is also applied to the synthesis of the corresponding (2S,3S,4R,5R)-isomer.

Full text of DOI:10.1016/S0957-4166(99)00083-X

TETRAHEDRON:
ASYMMETRY
Pergamon
Tetrahedron: Asymmetry 10 (1999) 937–945
Asymmetric synthesis of L-proline regio- and stereoselectively
labelled with deuterium
Makoto Oba, Tsutomu Terauchi, Akiko Miyakawa and Kozaburo Nishiyama ∗
Department of Material Science and Technology, Tokai University, 317, Nishino, Numazu, Shizuoka 410-0395, Japan
Received 5 February 1999; accepted 1 March 1999
Abstract
A synthesis of L-proline where all of the ring methylenes are stereoselectively labelled with deuterium is
described. A catalytic deuteration of protected 3,4-dehydro-L-proline using transition metal catalyst followed by
RuO4-oxidation gave a [3,4-D2]pyroglutamic acid derivative. A syn-selective deuteration of the aminal derived
from the pyroglutamate with Et3SiD–BF3·OEt2 furnished (2S,3S,4R,5S)-[3,4,5-D3]proline. The present procedure
is also applied to the synthesis of the corresponding (2S,3S,4R,5R)-isomer. © 1999 Elsevier Science Ltd. All rights
reserved.
1. Introduction
The availability of amino acids regio- and stereoselectively labelled with stable isotopes, such as D,
13C and 15N, is valuable both for stereochemical studies on the enzymatic transformations1 and for the
determination of the three-dimensional structure of the proteins by NMR spectroscopy.2 In particular,
stereoselective deuterium-labelling of the diastereotopic hydrogen and the methyl group is the most
important and essential technique for these investigations. In the light of the importance of these amino
acids, we have recently developed methods for regio- and stereoselective deuterium-labelling of amino
acid side chains.3
Proline is the sole cyclic amino acid in mammalian proteins and is usually located in bend and loop
structures where it provides important conformational constraints in globular proteins.4 Furthermore,
proline and its metabolites, mainly hydroxyproline, have also figured as components of biologically-
active compounds such as antibiotics.5 Therefore, specimens of L-proline stereospecifically labelled
with deuterium(s) in the ring methylene(s) are required for understanding its unusual conformational
preference and the stereochemical course of its metabolic reactions.
∗
Corresponding author. Fax: +81-559-68-1155; e-mail: nishiyam@wing.ncc.u-tokai.ac.jp
0957-4166/99/$ - see front matter © 1999 Elsevier Science Ltd. All rights reserved.
PII: S0957-4166(99)00083-X

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In connection with stereochemical studies on proline hydroxylase, stereoselective labelling at the γ-
6
carbon was achieved by reduction of tosylates derived from 4-hydroxyproline with LiAlD4 or LiAlT4
with an improved protocol being recently developed by Baldwin et al.7 The samples of [3,4-T2]proline
prepared by catalytic tritiation of the dehydroproline were also used in the above study.8 For stereoselec-
tive labelling at the β-9 or δ-position,10 methods based on a combination of chemical and enzymatic
reactions were explored. In this paper we wish to report on the first synthesis of [3,4,5-D3]proline in
which all of the prochiral methylenes are stereoselectively labelled with deuterium.11
2. Results and discussion
In order to incorporate deuterium atoms stereoselectively into the 3- and 4-positions of L-proline,
a catalytic deuteration of the corresponding dehydro-L-proline seems to be the most efficient and
convenient procedure. We therefore chose both ethyl and t-butyl 3,4-dehydro-L-prolinate 1a12 and 1b
derived from trans-4-hydroxy-L-proline as starting materials (Scheme 1).
Scheme 1.
Although a few papers concerning a catalytic deuteration13 or tritiation8 of racemic 3,4-dehydroproline
itself have been published, no papers concerning the protected derivatives were found in the literature. A
catalytic deuteration of compound 1a was carried out using 10% palladium on carbon in MeOD for 24 h
under medium pressure of deuterium gas (5 kgf/cm2). The regio- and stereoselectivities of the deuteration
1
1
were checked by H NMR spectroscopy after being deprotected because the H NMR spectrum of the
deuterated ethyl prolinate 2a was complicated by the s-cis/s-trans isomerism often found in the N-
acylproline derivatives.14 The 1H NMR spectrum of the deuterated proline 3 so obtained revealed that a
considerable extent of H–D scrambling occurred. Screening for a transition metal catalyst such as PdO,
PtO2, Rh/C, Ru/C, RhCl(PPh3)3 and RuCl2(PPh3)3 was carried out and the selected examples which
resulted in relatively clean formation of the [3,4-D2]proline 3 are listed in Table 1.
When RuCl2(Ph3)3 was used as a catalyst, the selectivity of deuteration was the best, giving the
[3,4-D2]proline 3 in 77% yield after deprotection of the ethyl [3,4-D2]prolinate 2a with 1 M HCl
followed by Dowex 50W-X8 (entry 2). The enantiomeric excess was checked by HPLC analysis using
a chiral stationary phase column, and was found to be 98% ee (L-form). The 1H NMR spectrum of the
[3,4-D2]proline 3 was shown in Fig. 1A. Although the deuterium content of the 4R-proton cannot be
estimated due to the signal overlapping, the signal intensity of the 3S-proton corresponds only to the
0.03 proton, indicating a stereoselective formation of (2S,3S,4R)-isomer. This stereochemical outcome

M. Oba et al. / Tetrahedron: Asymmetry 10 (1999) 937–945
939
Table 1
Stereoselective deuteration of protected dehydroprolinate 1a and 1b
can be attributed to a predominant deuterium addition to the olefin 1a anti to the resident ester group.
The corresponding t-butyl ester 1b was also subjected to the above deuteration and Ru on carbon proved
to be the most efficient catalyst in this case (entry 6, vide infra).
Figure 1. 400 MHz 1H NMR spectra of (A) (2S,3S,4R)-[3,4-D2]proline 3, (B) (2S,5S)-[5-D]proline 10, (C)
(2S,3S,4R,5S)-[3,4,5-D3]proline 6, (D) (2S,3S,4R,5R)-[3,4,5-D3]proline 12, and (E) unlabelled proline in D2O

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M. Oba et al. / Tetrahedron: Asymmetry 10 (1999) 937–945
We next examined the selective deuterium-labelling of the diastereotopic hydrogen at the δ-position.
Since a stereoselective reduction of the amido carbonyl moiety was considered to be the most convenient
approach for this purpose, a preliminary examination was performed using unlabelled ethyl and t-butyl
pyroglutamate 7a and 7b as starting materials. Among several methods for conversion of the pyrogluta-
mic acid into proline,15 we adopted a procedure of Pedregal et al.16 which involves a stepwise process for
reducing the carbonyl moiety. An additional advantage of this method for our purpose was the availability
of the corresponding deuterated-reducing agents. As shown in Scheme 2, a reduction of the protected
pyroglutamate 7 with lithium triethylborohydride was carried out at −78°C to give 5-hydroxyprolinate 8
and the resulting amino alcohol 8 was directly, or after conversion to the corresponding methyl ether
9, subjected to a reductive deuteration using the deuteriosilane–Lewis acid system. The results are
summarised in Table 2. For example, t-butyl 5-methoxyprolinate 9b was treated with Et3SiD in the
presence of BF3·OEt2 at −78°C for 2 h and subsequent deprotection procedure gave [5-D]proline 10 in
78% yield based on the 5-methoxyprolinate 9b (entry 9). The enantiomeric excess at the C-2 position
was checked by HPLC analysis to be 99% ee (L-form) and the stereoselectivity of the deuteration was
1
determined by H NMR integration of the δ-proton signals by comparison with the reported data.10,13
As shown in Fig. 1B, formation of (2S,5S)-[5-D]proline 10, the syn-isomer, was predominant over the
(2S,5R)-isomer in the ratio of 90:10. Similar syn-selective addition in the acyliminium intermediate which
seemed to be inconsistent with the steric interaction was also observed in a few Lewis acid catalysed
amido alkylation reactions.17 According to Danishefsky’s hypothesis,18 favorable orbital interaction over
steric interaction, so-called Cieplak-type stereoelectronic effect,19 was responsible for the observed syn-
selectivity. In our case the emerging σ* orbital at the δ carbon may be more stabilised by interacting with
the σ C–H bond rather than σ C-ester bond at the α carbon.
Scheme 2.
Other silanes such as tris(trimethylsilyl)deuteriosilane (TMS3SiD), Ph3SiD or Ph2SiD2, were also
tested as a reducing agent and a significant decrease in the selectivity was observed, probably due to
Table 2
Transformation of prolinate 8 or 9 into [5-D]proline 10

M. Oba et al. / Tetrahedron: Asymmetry 10 (1999) 937–945
941
an additional steric repulsion between the bulky silane and the ester moiety (entries 3–5). We previously
reported that the deuteriogermane and deuteriostannane were slightly more effective than deuteriosilane
for the syn-selective deuteration;11 however, here we employed only deuteriosilanes from ecological
and economical perspectives. Utilisation of trimethylsilyl trifluoromethanesulfonate (TMSOTf) and trityl
perchlorate (Ph3CClO4) as a Lewis acid resulted in the same degree of selectivity (entries 6 and 7).
Focusing on the difference in the ester moiety, the t-butyl ester proved to be slightly advantageous (entries
8 and 9).
Since a preparation of [3,4,5-D3]proline from ethyl [3,4-D2]prolinate 2a was minutely described in
our previous communication, we here report a synthesis of (2S,3S,4R,5S)-[3,4,5-D3]proline from t-butyl
[3,4-D2]prolinate 2b obtained using the reaction conditions shown in entry 6 (Table 1). An oxidation of
the prolinate 2b was carried out using RuO4, prepared in situ from RuO2 and NaIO4,20 to afford t-butyl
[3,4-D2]pyroglutamate 4b in a quantitative yield. After conversion of the pyroglutamate 4b into t-butyl
5-methoxy[3,4-D2]prolinate 5b in 75% yield, the 5-methoxyprolinate 5b was subjected to the above
deuteration procedure (Table 2, entry 9) to afford [3,4,5-D3]proline 6 in 94% yield. The enantiomeric
excess at the α-position was determined by HPLC analysis to be 99% ee (L-form) and the relative
1
configuration was confirmed by H NMR spectrum. As shown in Fig. 1C, the signal intensity of the
residual 3S-proton of compound 6 corresponded to 0.08 proton and the ratio of 5S- and 5R-proton signals
was 10:90, indicating a selective formation of (2S,3S,4R,5S)-isomer.
Using the present protocol, it was logically possible to obtain the corresponding (5R)-isomer simply
by changing the deuterium source. However, attempts to reduce the amido carbonyl moiety of the
ethyl [3,4-D2]pyroglutamate 4a with lithium triethylborodeuteride resulted in failure due to insufficient
deuterium incorporation. The origin of the hydrogen atom incorporated in the δ-position is still not clear.
Therefore, we attempted to prepare the (5R)-isomer from the t-butyl [3,4-D2]pyroglutamate 4b as shown
in Scheme 3. Thus, the pyroglutamate 4b was initially reduced with lithium triethylborodeuteride and
the deuterated amino alcohol so formed was converted to methyl ether 11 in 55% yield. Finally, the
5-methoxy[3,4-D2]prolinate 11 was subjected to Et3SiH–BF3·OEt2 system followed by deprotection to
give (2S,3S,4R,5R)-[3,4,5-D3]proline 12 in 85% yield with 99% ee (L-form). The relative configuration
1
was also confirmed by H NMR spectrum (Fig. 1D) and it was found that the ratio of (5S)- and (5R)-
isomer was 13:87.
Scheme 3.
In conclusion, we have completed a stereoselective synthesis of (2S,3S,4S,5S)-[3,4,5-D3]proline 6
based on the catalytic deuteration of the dehydro-L-prolinate 2 and the syn-selective reduction of aminal
5 derived from the [3,4-D2]pyroglutamate 4 with Et3SiD–BF3·OEt2 system. During the course of
these investigations, [3,4-D2]proline 3 and [5-D]proline 10 were also obtained. In addition, the present
procedure was applied to the synthesis of the corresponding (2S,3S,4R,5R)-isomer 12 simply by changing
the deuterium source. These amino acids are considered to be useful for unambiguous assignments of 1H
NMR signals of the ring methylenes, especially the (2S,3S,4R,5S)-isomer which is a promising probe for
monitoring the ring dynamics through the vicinal spin–spin coupling constants.

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3. Experimental
1H and 13C NMR spectra were recorded in CDCl3 or D2O on a Varian UNITY-400 spectrometer. All
chemical shifts are reported as δ values (ppm) relative to residual chloroform (7.26 ppm) or sodium
3-(trimethylsilyl)[2,2,3,3-D4]propionate (0 ppm). High resolution mass spectra (EI) were obtained on
a JEOL JMS-AX-500 spectrometer with DA7000 data system using perfluorokerosene as an internal
standard. For ion-exchange chromatography, Dowex 50W-X8 activated with 1 M HCl was used. Enan-
tiomeric excess was determined on a Senshu SSC-3100 high-pressure liquid chromatography system
equipped with chiral MCIGEL CRS10W column from Mitsubishi Kasei Co. and 2 mM CuSO4 solution
as an eluent. Catalytic hydrogenation was performed in an Ishii CHA-S medium-pressure catalytic
hydrogenator. All other reagents were of commercial grade and used as supplied.
3.1. t-Butyl (2S)-N-t-butoxycarbonyl-3,4-dehydroprolinate 1b
The t-butyl 3,4-dehydroprolinate 1b was obtained as a colorless oil in 51% yield (5 steps) from
trans-4-hydroxy-L-proline according to the reported procedure12 described for the preparation of the
corresponding ethyl ester 1a. 1H NMR (CDCl3) δ 1.440, 1.443, 1.45 and 1.47 (4 s, 18H), 4.10 and 4.27
(2 m, 2H), 4.81 and 4.88 (2 m, 1H), 5.69 and 5.73 (2 m, 1H), 5.90 and 5.94 (2 m, 1H). HRMS m/z
269.1586 (M+, calcd for C14H23NO4: 269.1627).
3.2. (2S,3S,4R)-[3,4-D2]Proline 3
To a solution of ethyl (2S)-N-t-butoxycarbonyl-3,4-dehydroprolinate (1a, 1.21 g, 5.00 mmol) in MeOD
(25 ml) was added a solution of RuCl2(PPh3)3 (23.9 mg, 0.025 mmol) in benzene (2 ml), and the resultant
solution was stirred under medium pressure (5 kgf/cm2) using deuterium gas at room temperature
overnight. After removal of the solvent, the residue was submitted to a short chromatography on silica
gel to remove the catalyst, giving ethyl N-t-butoxycarbonyl[3,4-D2]prolinate (2a, 1.23 g) as an oil in a
1
quantitative yield. H NMR (CDCl3) δ 1.24 and 1.26 (2 t, J=7 Hz, 3H), 1.40, 1.45 (2 s, 9H), 1.92 (m,
2H), 3.33–3.56 (m, 2H), 4.10–4.28 (m, 3H). HRMS m/z 246.1698 [(M +H)+, calcd for C12H20D2NO4:
246.1674].
Deprotection of the [3,4-D2]prolinate 2a (0.363 g, 1.50 mmol) was carried out in 1 M HCl (15 ml) at
110°C for 3 h followed by a treatment with Dowex 50W-X8 to give [3,4-D2]proline (3, 135 mg, 77%)
as a colorless solid, mp 214–217°C (dec.). 1H NMR (D2O) δ 1.97 (ddd, J=7, 7 and 7 Hz, 1H), 2.05 (dd,
J=7 and 7 Hz, 1H), 2.33 (dd, J=7 and 7 Hz, 0.03H), 3.33 (dd, J=12 and 7 Hz, 1H), 3.41 (dd, J=12 and 7
Hz, 1H), 4.13 (d, J=7 Hz, 1H). HRMS m/z 117.0762 (M+, calcd for C5H7D2NO2: 117.0759).
3.3. t-Butyl (2S,3S,4R)-N-t-butoxycarbonyl[3,4-D2]prolinate 2b
A mixture of t-butyl-N-t-butoxycarbonyl-3,4-dehydroprolinate (1b, 1.03 g, 4.00 mmol) and Ru/C (0.1
g) in MeOD (40 ml) was stirred at room temperature overnight under the medium pressure (5 kgf/cm2)
of deuterium gas. After removal of the catalyst using a Celite pad, evaporation of the solvent gave [3,4-
1
D2]prolinate 2b (1.63 g, 97%) as an oil. H NMR (CDCl3) δ 1.43, 1.44 and 1.45 (3 s, 18H), 1.89 (m,
1.8H), 2.00–2.18 (m, 0.2H), 3.32–3.54 (m, 2H), 4.09 and 4.17 (2 br s, 1H). HRMS m/z 273.1874 (M+,
calcd for C14H23D2NO4: 273.1909).

M. Oba et al. / Tetrahedron: Asymmetry 10 (1999) 937–945
943
3.4. t-Butyl (2S,3S,4R)-N-t-butoxycarbonyl[3,4-D2]pyroglutamate 4b
To an aqueous solution of 10% sodium metaperiodate (120 ml) was added RuO2 (219 mg, 1.65 mmol),
and the black suspension turned into a yellow solution. Then to this solution was added a solution of
[3,4-D2]prolinate 2b (3.00 g, 11.1 mmol) in ethyl acetate (45 ml). The resulting two-phase mixture was
vigorously stirred at room temperature and monitored to completion by TLC. The layers were separated
and to the organic phase was added 2-propanol (2 ml) and stirred for 1 h. After removal of the precipitated
RuO2 using a Celite pad, the filtrate was dried over MgSO4, concentrated, and chromatographed on silica
gel to afford [3,4-D2]pyroglutamate 4b (3.18 g, 100%) as an oil. 1H NMR (CDCl3) δ 1.47 (s, 9H), 1.49
(s, 9H), 1.96 (dd, J=10 and 2 Hz, 0.9H), 2.25 (dd, J=10 and 10 Hz, 0.1H), 2.43 (d, J=10 Hz, 0.1H),
2.57 (d, J=10 Hz, 0.9H), 4.46 (d, J=2 Hz, 1H). 13C NMR (CDCl3) δ 21.19 (t, J=21 Hz), 27.85 (6 C),
30.64 (t, J=21 Hz), 59.47, 82.10, 83.09, 149.37, 170.32, 173.11. HRMS m/z 288.1778 [(M+H)+, calcd
for C14H22D2NO5: 288.1780).
3.5. (2S,3S,4R,5S)-[3,4,5-D3]Proline 6
To a solution of t-butyl [3,4-D2]pyroglutamate 4b (578 mg, 2.00 mmol) in THF (20 ml) was added a
solution of 1 M LiEt3BH in THF (2.4 ml, 2.40 mmol) at −78°C under Ar atmosphere and the reaction
mixture was stirred for 0.5 h. Then the reaction was quenched with saturated aqueous NaHCO3 (6 ml)
at −78°C and the mixture was warmed up to 0°C. After addition of 30% H2O2 (1 ml), the mixture was
stirred for a further 20 min and concentrated. The residue was extracted with ether, washed with H2O
and dried over MgSO4. After removal of the solvent, the crude t-butyl 5-hydroxyprolinate was directly
treated with MeOH (13 ml) in the presence of p-toluenesulfonic acid (30 mg, 0.174 mmol) for 16 h.
The mixture was diluted with CHCl3, washed with saturated aqueous NaHCO3 and dried over MgSO4.
After removal of the solvent, the residue was chromatographed on silica gel. Elution with a mixture of
hexane and ethyl acetate (7:3) afforded oily t-butyl (2S,3S,4R,5RS)-N-t-butoxycarbonyl-5-methoxy[3,4-
D2]prolinate (5, 457 mg, 75%) as unseparable diastereoisomers. 1H NMR (CDCl3) δ 1.437, 1.440, 1.45,
1.46, 1.48 and 1.49 (6 s, 18H), 1.84–2.11 (m, 2H), 3.34, 3.39, 3.40 and 3.43 (4 s, 3H), 4.14–4.20 (m,
1H), 5.12–5.29 (m, 1H). HRMS m/z 303.2043 (M+, calcd for C15H25D2NO5: 303.2015).
To a solution of t-butyl 5-methoxy[3,4-D2]prolinate 5 (457 mg, 1.50 mmol) in CH2Cl2 (15 ml) was
added Et3SiD (352 mg, 3.00 mmol) and trifluoroborane etherate (468 mg, 3.30 mmol) in two portions at
intervals of 0.5 h at −78°C under argon atmosphere and the resulting solution was stirred for 2 h. Then the
reaction mixture was quenched with saturated aqueous Na2CO3 (3 ml) and the organic layer was dried
over MgSO4 and evaporated. Deprotection of the resultant crude [3,4,5-D3]prolinate was carried out in 1
M HCl (25 ml) at 110°C for 3 h, followed by a treatment with Dowex 50W-X8 to give [3,4,5-D3]proline
(6, 166 mg, 94%) as a colorless solid, mp 217–220°C (dec.). 1H NMR (D2O) δ 1.97 (dd, J=8 and 7 Hz,
1H), 2.05 (dd, J=8 and 7 Hz, 1H), 2.34 (m, 0.08H), 3.32 (d, J=7 Hz, 0.9H), 3.41 (d, J=7 Hz, 0.1H), 4.13
(d, J=7 Hz, 1H). HRMS m/z 118.0813 (M+, calcd for C5H6D3NO2: 118.0822).
3.6. (2S,5S)-[5-D]Proline 10
Using the procedure described for the synthesis of the [3,4-D2]prolinate 5, t-butyl (2S,5RS)-N-t-
butoxycarbonyl-5-methoxyprolinate 9b was obtained as an oil from the pyroglutamate 7b in 74% yield.
1H NMR (CDCl3) δ 1.43, 1.44, 1.446, 1.454, 1.47 and 1.48 (6 s, 18H), 1.71–2.46 (m, 4H), 3.33, 3.385,
3.393 and 3.42 (4 s, 3H), 4.11–4.24 (m, 1H), 5.11–5.29 (m, 1H). HRMS m/z 301.1936 (M+, calcd for
C15H27NO5: 301.1936).

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M. Oba et al. / Tetrahedron: Asymmetry 10 (1999) 937–945
To a solution of 5-methoxyprolinate 9b (300 mg, 1.00 mmol) in CH2Cl2 (10 ml) was added Et3SiD
(234 mg, 2.00 mmol) and trifluoroborane etherate (312 mg, 2.20 mmol) in two portions at intervals of
0.5 h at −78°C under argon atmosphere and the resulting solution was stirred for 2 h. Then the reaction
mixture was quenched with saturated aqueous Na2CO3 and the organic layer was dried over MgSO4 and
evaporated. Deprotection of the resultant crude [5-D]prolinate was carried out in 1 M HCl (15 ml) at
110°C for 3 h followed by treatment with Dowex 50W-X8 to give (2S,5S)-[5-D]proline (10, 91.0 mg,
1
78%) as a colorless solid, mp 212–216°C (dec.). H NMR (CDCl3) δ 2.01 (m, 2H), 2.08 (m, 1H), 2.35
(m, 1H), 3.33 (dd, J=7 and 7 Hz, 0.9H), 3.41 (dd, J=7 and 7 Hz, 0.1H), 4.13 (dd, J=9 and 6 Hz). HRMS
m/z 116.0692 (M+, calcd for C5H8DNO2: 116.0696).
3.7. (2S,3S,4R,5R)-[3,4,5-D3]Proline 12
t-Butyl (2S,3S,4R,5RS)-N-t-butoxycarbonyl-5-methoxy[3,4,5-D3]prolinate 11 was obtained as an oil
in 55% yield following the procedure described for the preparation of the [3,4-D2]prolinate 5 except that
LiEt3BD was used instead of LiEt3BH. 1H NMR (CDCl3) δ 1.44, 1.45, 1.46, 1.47, 1.486 and 1.491 (6 s,
18H), 1.85–2.11 (m, 2H), 3.35, 3.40, 3.41 and 3.44 (4 s, 3H), 4.14–4.21 (m, 1H). HRMS m/z 304.2116
(M+, calcd for C15H24D3NO5: 304.2078).
To a solution of 5-methoxy[3,4,5-D3]prolinate 11 (318 mg, 1.04 mmol) in CH2Cl2 (10 ml) was added
Et3SiH (242 mg, 2.08 mmol) and trifluoroborane etherate (324 mg, 2.28 mmol) in two portions at
intervals of 0.5 h at −78°C under argon atmosphere and the resulting solution was stirred for 2 h. Then the
reaction mixture was quenched with saturated aqueous Na2CO3 (2 ml) and the organic layer was dried
over MgSO4 and evaporated. Deprotection of the resultant crude [3,4,5-D3]prolinate was carried out in 1
M HCl (17 ml) at 110°C for 3 h followed by a treatment with Dowex 50W-X8 to give [3,4,5-D3]proline
(12, 104 mg, 85%) as a colorless solid, mp 207–213°C (dec.). 1H NMR (D2O) δ 1.97 (dd, J=8 and 7 Hz,
1H), 2.05 (dd, J=8 and 7 Hz, 1H), 2.34 (m, 0.08H), 3.32 (d, J=7 Hz, 0.13H), 3.41 (d, J=7 Hz, 0.87H),
4.13 (d, J=7 Hz, 1H). HRMS m/z 118.0786 (M+, calcd for C5H6D3NO2: 118.0822).
Acknowledgements
This work has been supported by CREST (Core Research for Evolutional Science and Technology)
of Japan Science and Technology Corporation (JST) and a Grant-in-Aid for Scientific Research (No.
09740480) from the Ministry of Education, Science, Sports and Culture of Japan. We also thank Ms H.
Oikawa for MS measurements.
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