Chirally deuterated benzyl chlorides from benzyl alcohols via hexachloroacetone/polymer-supported triphenylphosphine: Synthesis of protected (2S, 3S)-[3-2H, 15N]-tyrosine

Article abstract of DOI:10.1002/jlcr.3004

Chirally deuterated benzyl chlorides were prepared using novel, general hexachloroacetone/polymer-supported triphenylphosphine treatment of chirally deuterated benzyl alcohols. Doubly labeled protected tyrosine was obtained in 62% yield with 86% de at the α-carbon and 82% de at the β-carbon. Key in the synthesis was the alkylation of ¹⁵N-labeled (-)-8-phenylmenthylhippurate with R-(-)-4-triisopropylsilyloxybenzyl-α-d chloride. Chirally deuterated benzyl chlorides were prepared in high yield with inversion from benzyl alcohols using solid-phase methods. One of the benzyl chlorides obtained was used in the synthesis of protected tyrosine labeled with ¹⁵N and chirally deuterated on the β-position. Copyright

Full text of DOI:10.1002/jlcr.3004

Research Article
Received 4 October 2012,
Revised 2 November 2012,
Accepted 9 November 2012
Published online 21 December 2012 in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/jlcr.3004
Chirally deuterated benzyl chlorides from benzyl
alcohols via hexachloroacetone/polymer-
supported triphenylphosphine: synthesis
15
of protected (2S, 3S)-[3-2H, N]-tyrosine
Derek W. Barnett,a Maryanne S. Refaei,b and Robert W. Curley, Jr.a,b
*
Chirally deuterated benzyl chlorides were prepared using novel, general hexachloroacetone/polymer-supported triphenyl-
phosphine treatment of chirally deuterated benzyl alcohols. Doubly labeled protected tyrosine was obtained in 62% yield
with 86% de at the a-carbon and 82% de at the b-carbon. Key in the synthesis was the alkylation of 15N-labeled (À)-8-
phenylmenthylhippurate with R-(À)-4-triisopropylsilyloxybenzyl-a-d chloride.
Keywords: chiral; benzylic; deuteration; chlorides; alcohols; tyrosine
aldehyde 2 was treated with morpholine perchlorate and KCN
Introduction
to give the morpholinonitrile derivative, which was then
deprotonated and treated with D2O to effect 1H–2H exchange.5
Stable isotope labeled amino acids are valuable research tools that
have many uses in the fields of mechanistic enzymology and
Deuteration proceeded with high efficiency as determined by
1H NMR spectroscopy (relative integration of the residual methine
biomolecular nuclear magnetic resonance (NMR) spectroscopy.
These compounds are often employed to study biosynthesis, the
proton vs. the morpholine methylene protons; reconfirmed by
stereochemical course of enzyme-mediated processes,1 and
integration of proton resonances of aldehydes 3 and 4), and acid
protein structure by NMR methods.2 Alkylation of glycine equiva-
hydrolysis of the morpholinonitrile returned 4-benzyloxybenzalde-
lents is a well precedented route to a-amino acid derivatives.3
hyde-a-d (3). Benzyl group removal (trimethylsilyl iodide) and
In earlier work,4 we used (S)-benzyl-a-d mesylate to alkylate
reprotection as the triisopropylsilyl ether gave 4-triisopropylsilylox-
15N-labeled (À)-8-phenylmenthyl hippurate3t to give a (2S,3R)-
ybenzaldehyde-a-d (4). Finally, Alpine BoraneW reduction of 4 to
phenylalanine derivative. It was also shown that (R)-benzyl-a-d
give 1b introduced asymmetry with approximately 99% ee as
determined by NMR analysis of the (À)-camphanate ester.6
chloride could be used as the electrophilic species to generate
the (2S,3S)-epimer. This method allows synthetic access to all
stereoisomers at C2 and C3 by varying the chirality of the ester-
protecting group and the electrophile. Availability of labeled
glycine in multiple isotopomeric forms also allows the incorpora-
tion of different labeling patterns into the final product.
The preparation of tyrosine is a logical extension of this
To demonstrate the versatility of the strategy, preparation of
the b-position epimer of tyrosine, relative to the phenylalanine
synthesis,4 was planned. This would require the use of the chloride
derived from 1b as the electrophile.4 Several halogenation condi-
tions using alkoxyphosphonium intermediates to ensure inversion
of the benzylic center during alcohol to halide conversion
were investigated.7 Unfortunately, these reactions also produced
work. However, a key consideration is the impact of the
para-hydroxy group on the alkylation and deprotection reac-
tions. We now report novel hexachloroacetone (HCA)/polymer-
supported triphenylphosphine (psTPP) chlorination conditions to
produce an enantiotopically deuterated benzylic electrophile and
use it in the stereoselective synthesis of a protected doubly labeled
tyrosine.
by-products, which interfered with or prohibited isolation of the
labile halides, as others have also documented.8,9
The chlorination of alcohols using HCA (5) and soluble
triphenylphosphine (TPP) was first reported by Magid and
co-workers.10 They detailed the regioselective conversion of
aDivision of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The
Ohio State University, Columbus, OH 43210, USA
Results and discussion
Shown in Scheme 1 is the synthesis of (S)-4-triisopropylsilyloxybenzyl-
a-d alcohol (1b) that served as the precursor for the electrophilic bOhio State Biochemistry Program, The Ohio State University, Columbus,
OH 43210, USA
species in the tyrosine synthesis. Alternative phenol-protecting
groups investigated included the methyl, benzyl, and tert-
butyldimethylsilyl ethers, but triisopropylsilyl displayed the best
*Correspondence to: Robert W. Curley, Ohio State Biochemistry Program, The
Ohio State University, Columbus, OH 43210, USA.
combination of stability and reactivity as the electrophile. Thus, E-mail: curley.1@osu.edu
J. Label Compd. Radiopharm 2013, 56 6–11
Copyright © 2012 John Wiley & Sons, Ltd.

D. W. Barnett et al.
O
Ph
O
i
O
D
H
15
O
N
BnO
BnO
H
O
3
2
ii
1) 2 eq. LDA, 2 eq. TMEDA,
-78 oC
2) 1.1 eq. 6b, -78 oC to 0 oC;
O
H
D
iii
D
OH
(62%)
TIPSO
TIPSO
1b
4
Scheme 1. Synthesis of 1b. Reagents and conditions: (a) i, morpholine, perchloric
acid, KCN, 90 ꢀC (quant.); ii, NaH, D2O, THF (86%, 99% 2H); iii, 2 N HCl (92%; 99%
2H); (b) i, trimethylsilyl iodide, CH2Cl2 (59%); ii, triisopropylsilyl chloride, NEt3, THF;
(c) (R)-Alpine BoraneW (79% from unprotected phenol; 99% ee).
Ph
TIPSO
H15
D
N
O
O
O
allylic alcohols into chlorides with inversion of stereochemistry
under very mild conditions with excellent yields. Villeneuve and
Chan also reported these conditions to be an acid-free procedure
to prepare acid chlorides.11
7
(86% de at C2, 82% de at C3)
Scheme 3. Synthesis of doubly labeled protected tyrosine 7.
In our work, model chlorinations with HCA and soluble TPP still
suffered from purification difficulties as mentioned previously.
Therefore, a source of TPP allowing unreacted phosphine and by-
products to be removed by filtration was employed. Shown
in Scheme 2 are the conditions used for the synthesis of
(R)-4-triisopropylsilyloxybenzyl-a-d chloride (6b). The stoichiometry
is based on earlier findings.11 Chloride 6b was obtained with
excellent apparent stereoselectivity and in high yield from
treatment of 1b with 2.5 eq. of psTPP and 0.5 eq. of HCA in
THF at 0 ꢀC. Because optically pure 6b is not known, the optical
purity estimate is based on analysis of the alkylation product of
6b in the labeled protected tyrosine synthesis (Scheme 3).
Although this is an indirect stereochemical analysis of the chloride,
it establishes the lower limit of its optical purity at 82% ee. The
filterable source of phosphine greatly facilitated product isolation,
and to our knowledge, this represents the first use of psTPP for
conversion of benzyl alcohols to chlorides.12
chlorides were then prepared as in Scheme 2. Optical rotations of
the alcohols and chlorides were recorded and the optical purities
of the alcohols were determined using NMR analysis of the
(À)-camphanate esters.6 As shown in Table 1, yields of the chlorina-
tion reaction are essentially quantitative except in the case of the
unsubstituted benzyl alcohol (1a). The specific rotation observed
for 6a here is comparable to that reported for optically pure
material.13 Optical rotation data for the remaining chlorides 6 (as
well as 1c–e) have not been reported. If desired, the optical purity
of these chlorides could be indirectly determined using the
alkylation strategy similar to 6b. It should be noted that the
chlorination of the nitro compound (1e) proceeded noticeably
faster than with the other benzyl alcohols.
With 6b in hand, alkylation of 15N-labeled (À)-8-phenyl-
menthylhippurate to provide protected tyrosine 7 proceeded
as for phenylalanine (Scheme 3).4 These conditions afforded
doubly labeled 7 in 62% yield with 86% de at the a-carbon and
82% at the b-carbon as determined by 1H NMR spectroscopy
and HPLC. We saw an even larger decrease in b-position de in
our phenylalanine synthesis from chirally deuterated benzyl
mesylate4 further suggesting 6b has high ee. Figure 1 shows
Because we were interested in the production of chiral benzylic
chlorides, we investigated the effect of the para-substituent on the
stereoselectivity of the reaction. A small number of enantiotopi-
cally deuterated alcohols were prepared as in Scheme 1 and are
shown in Table 1. In the case of the 4-nitro analog, the propensity
1
of the deuterated morpholinonitrile to undergo H–2H exchange
during hydrolysis required an alternative synthesis. Thus, we
reduced methyl 4-nitrobenzoate with NaBD4 and oxidized to the
dueterio aldehyde analogous to 4 and then converted to the
benzyl alcohol as in Scheme 1. The enantioselectively deuterated
1
the amide proton region of the 400-MHz H NMR spectrum of
crude 7. The pair of doublets (~90 Hz 15N–H coupling and
7.5 Hz three bond coupling to the b-proton) centered at
6.27 ppm is the amide proton resonance of the (2S)-product,
whereas the doublet centered at 5.90 ppm is the (2R)-product
(the triplet pair centered at 6.03 ppm is unreacted starting
material showing coupling to the additional a-proton). As for
protected phenylalanine,4 the HPLC chromatogram (Figure 2)
shows the (2S)-diastereomer (tR = 21 min) elutes after the (2R)-
diastereomer (tR = 16 min) with starting material at tR = 8 min.
Integration of these bands is consistent with the NMR data
and supports the assigned a-position de. Integration of the
D
H
0.5 (Cl3C)2CO (5)
Cl
1b
2.5 psTPP, THF,
0 oC (96%)
TIPSO
6b
>82% ee
98% ee
1
benzylic proton region of the H NMR spectrum of 7 (Figure 3)
Scheme 2. Synthesis of 6b using hexachloroacetone/polymer-supported
triphenylphosphine conditions.
was used to estimate the de at the b-position.
J. Label Compd. Radiopharm 2013, 56 6–11
Copyright © 2012 John Wiley & Sons, Ltd.
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D. W. Barnett et al.
Table 1. Chlorination studies using hexachloroacetone/polymer-supported triphenylphosphine conditions
Entry
X
ee of (S) – 1a
[a]2D1of (S) – 1
Chloride yield
[a]2D1of (R) – 6
a
b
c
d
e
H
96%
98%
92%
95%
95%
+1.34ꢀ (neat)
64%
96%
97%
quant.
quant.
À1.52ꢀ (c 6.9, THF)b
À0.39ꢀ (c 15.4, THF)
À0.18ꢀ (c 16.6, THF)
À0.37ꢀ (c 13.6, THF)
À1.31ꢀ (c 6.9, THF)
OTIPS
OCH3
OBn
NO2
+0.30ꢀ (neat)
+0.91ꢀ (neat)
+0.59ꢀ (c 5.1, THF)
+2.46ꢀ (c 6.1, THF)
1
aestimated by H NMR analysis of (À)-camphanate derivative.
b[a]2D5 = À1.53ꢀ reported for specific rotation of optically pure material.13
2.95
2.90
2.85
2.80
2.75
2.70
2.65
2.60
ppm
6.45 6.40 6.35 6.30 6.25 6.20 6.15 6.10 6.05 6.00 5.95 5.90 5.85 5.80 ppm
Figure 3. Benzylic proton region of 1H NMR spectrum (400 MHz) of 7 showing an
approximate 82% de at C3.
Figure 1. Amide proton region of 1H NMR spectrum (400 MHz) of 7 showing an
approximate 86% de at the a-carbon.
stereoselectivity of a-position alkylation,3r–t perhaps a more
readily removed benzamide-protecting group would improve
stereochemical outcome. To this end, we modeled hydrolysis
rates by colorimetric (ninhydrin) assay of liberated glycine
from 6M HCl treatment of ethyl hippurate and its para-nitro and
3,5-dinitro analogs. As expected, the rate of hydrolysis increases
with degree of nitration, and the 3,5-dinitro analog hydrolyzes at
least 3.5–5 times faster than the unsubstituted benzamide, which
our studies show may prove useful for the current synthesis.
Desilylation of 7 gave 8 in high yield (Scheme 4) with no
impact on stereochemistry as evidenced by H NMR and HPLC
1
analysis. However, final deprotection of 8 in conc. HCl at 70 ꢀC
gave tyrosineÁHCl (9) in 59% yield but with an approximate
73% de at the a-carbon. This slight epimerization was seen in
our phenylalanine synthesis4 and has been seen by others.14–16
Inspection of the benzylic region of the 1H NMR spectrum of
9 showed that the product now also has about 44% de at the
b-position. Although this decrease will not likely hinder use of
9, it is surprising and may result via formation of a quinone
methide species known to form in many processes17 and
implicated in epimerization of chiral benzylic centers in natural
Conclusions
products under similar conditions.18 Regardless of mechanism, In summary, our previous method for preparation of 15N-labeled
1H NMR studies of intermediates during the hydrolysis of 8 L-phenylalanine with chiral b-deuteration has been successfully
showed that the ester is cleaved quickly with no b-position extended to protected tyrosine. Some degradation of b-position
epimerization, but the longer reaction times needed for benza- stereochemical integrity occurred during benzamide group
mide hydrolysis permit decrease of stereochemical integrity. hydrolysis to give tyrosine, and we offer a solution for this
Because the benzamide group participates in ensuring the high problem. The labeled tyrosine should still be useful for a variety
Figure 2. Chromatogram of crude 7 showing retention times of starting material, (2R)-, and (2S)-diastereomer, left to right.
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Copyright © 2012 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2013, 56 6–11

D. W. Barnett et al.
temperatures, as well as the method of purification, varied slightly
depending on the starting aldehyde.
A flask was charged with morpholine (20 mL) and cooled to 0 ꢀC. Perchloric
acid (70%, 9.5 mL; 0.11 mol) was added dropwise to the stirred solution
followed by careful addition of benzaldehyde (0.11 mol). The mixture became
a yellow solid, which dissolved on warming to 70 ꢀC. After 4 h at 70 ꢀC, the
mixture was cooled to room temperature and an aqueous solution of KCN
(0.12 mol, minimal water) was added. The mixture was warmed to 90 ꢀC for
1 h and then cooled to room temperature and poured onto ice with stirring.
The resulting precipitate was washed with water, recrystallized from absolute
ethanol, and dried under vacuum at 40 ꢀC for 12 h to give 18.8 g (91%) of a-
phenyl-4-morpholineacetonitrile: mp 67–68 ꢀC (lit. 68–70 ꢀC); 19 1H NMR
(CDCl3) d 2.27–2.38 (m, 4H), 3.45–3.57 (m, 4H), 4.65 (s, 1H), 7.15–7.22
(m, 3H), 7.34–7.36 (m, 2H); 13C NMR (CDCl3) d 49.79, 62.31, 66.55,
115.03, 127.80, 128.61, 128.79, 132.55; HRMS-ES (m/z): [M + Na]+ calcu-
lated for 225.1004, found 225.0991.
Ph
RO
H15
D
N
O
O
O
7
8
R = TIPS
R = H
TBAF
9:1 THF-H2O
(98%)
conc. HCl, 70 o
(59%)
C
A dry, argon flushed flask was charged with a-phenyl-4-morpholineace-
tonitrile (50mmol) and THF (50 mL). Sodium hydride (95%; 99mmol) was
carefully added, and the pink mixture was heated to 40ꢀC for 1h. The
mixture was cooled to 0ꢀC, and deuterium oxide (1.0mol) was carefully
added dropwise as the pink color faded over 30min of stirring. The solution
was acidified (pH 1–2) by addition of freshly distilled thionyl chloride and
poured onto ice with stirring. The resulting precipitate was washed with
water and dried to give white solid a-phenyl-4-morpholinoeacetonitrile-
a-d (9.91 g, 99%; 99% 2H incorporation as determined by 1H NMR):
HO
15NH3Cl
OH
D
O
mp 67–68 ꢀC (lit. 69–70 ꢀC)19 1H NMR (CDCl3) d 2.42 (t, 4H, J = 4.4Hz),
;
9
3.51–3.60 (m, 4H), 7.15–7.28 (m, 3H), 7.38–7.40 (m, 2H); 13C NMR (CDCl3)
d 49.75, 61.85 (t, J= 22.1 Hz), 66.60, 115.00, 127.82, 128.74, 132.46; HRMS-
ES (m/z): [M+ Na]+ calculated for 226.1082, found 226.1070.
Scheme 4. Deprotection of doubly labeled tyrosine 7.
A
flask was charged with a-phenyl-4-morpholineacetonitrile-a-d
of purposes and methods to enhance hydrolysis rate should
diminish this problem. More importantly, in accomplishing
this synthesis, a new solid phase-assisted method for the
preparation of chirally deuterated benzyl chlorides has been
(25 mmol) and 2M HCl (60 mL). The suspension was refluxed for 12 h,
and the resulting two-phase mixture was cooled to room temperature
and extracted with ether. The combined organic phases were washed
with saturated NaHCO3, water and brine, dried over MgSO4, filtered,
developed. The process is high yielding, proceeds with apparent and concentrated to the oil benzaldehyde-a-d (2.5 g, 93%; 98% 2H incor-
poration as determined by 1H NMR), which was used as obtained: 1H NMR
excellent stereochemical fidelity, and allows convenient work-up
(CDCl3) d 7.39–7.68 (m, 3H), 7.87 (d, 2H, J= 4.9Hz), 10.01 (s, residual CHO).
and purification.
(S)-(+)-benzyl-a-d alcohol (1a)
Experimental section
A THF solution of 0.5M R-Alpine-BoraneW (70 mL, 35 mmol) was added to
All reactions were carried out under a dry argon atmosphere unless
benzaldehyde-a-d (2.3 g, 21.5 mmol). The mixture was stirred at room
otherwise noted and in dry glassware when necessary. 15N-Glycine and
temperature for 12 h then refluxed for 1.5 h. After cooling to room
D2O were from Cambridge Isotope Laboratories (Cambridge Isotope
temperature, acetaldehyde (5 mL) was added and the mixture was stirred
Laboratories Inc., Andover, MA, USA). All other reagents and solvents
for 30 min, and then solvent was removed. Some of the pinene by-
were from Sigma-Aldrich (Sigma-Aldrich Corp., St. Louis, MO, USA) and
products were then removed under high vacuum at 50 ꢀC for 5 h. The
were used as obtained unless otherwise indicated. Analytical thin-layer
resulting orange oil was dissolved in ether (75 mL and, cooled to 0 ꢀC,
chromatography (TLC) and column chromatography materials were from
and ethanolamine (2.1 g, 35 mmol) was added and the mixture was
EM Separations (EM Separations, Gibbstown, NJ, USA), whereas preparative
stirred for 30min at 0ꢀC. The resulting white precipitate was filtered and
TLC plates were from Analtech (Analtech, Inc., Newark, DE, USA). Optical
washed with ether. The combined ether filtrate was concentrated, and
rotations were obtained using a Perkin-Elmer 241 polarimeter (Perkin-Elmer
the resulting oil was dissolved in 10% aqueous methanol and washed
Inc., Waltham, MA, USA). Analytical HPLC analysis was performed on a
several times with heptane. Isopropanol was added to the methanolic
Beckman Ultrasphere ODS column (Beckman Coulter Inc., Brea, CA, United
fraction, and the solution was concentrated under reduced pressure. The
States) (4.6 Â 250mm) using a Beckman System Gold 127 solvent module
resulting yellow oil was chromatographed (silica gel; 5% ethyl acetate/
and 166 UV detection module. NMR spectra were recorded on a Bruker
hexane) to give 2.0g (87%; >96% ee) of 1a as a colorless oil: [a]2D1 + 1.34o
DRX400 instrument (Bruker Corp., Billerica, MA, USA) operating at
(neat)20 1H NMR (CDCl3) d 1.90 (s, 1H), 4.64 (t, 1H, J = 1.8Hz), 7.26–7.37
;
400 MHz for 1H and 100MHz for 13C measurements, respectively. A Bruker
DRX800 instrument in The Ohio State University Campus Chemical Instru-
ment Center was used for measuring 800-MHz (1H) spectra. Mass spectra
were recorded at The Ohio State University Campus Chemical Instrument
Center using a Micromass QTOF spectrometer.
(m, 5H); 13C NMR (CDCl3) d 64.20 (t, J= 22.1Hz), 126.79, 126.23, 128.21,
140.64; HRMS-EI (70 eV) m/z: calculated 109.0637, found 109.0569.
(S)-(+)-4-triisopropylsilyloxybenzyl-a-d alcohol (1b)
[a]2D1 + 0.30o (neat); 1H NMR (CDCl3) d 1.08 (d, 18H, J = 7.04 Hz), 1.23
(m, 3H), 4.57 (br t, 1H), 6.85 (d, 2H, J = 8.6 Hz), 7.20 (d, 2H, J = 8.6 Hz);
HRMS-ES (m/z): [M + Na]+ calculated for 304.1835, found 304.1837.
General procedure for synthesis of deuterated benzyl alcohols
The procedure used for the synthesis of the deuterated aldehydes and
their resulting stereoselectively deuterated benzyl alcohols consisted of
four steps. The procedure that follows was used for the preparation of
(S)-(+)-4-methoxybenzyl-a-d alcohol (1c)
benzaldehyde-a-d/(S)-(+)-benzyl-a-d alcohol (1a) and is representative [a]2D1 + 0.91o (neat); 1H NMR (CDCl3) d 3.79 (s, 3H), 4.58 (br t, 1H), 6.87
of the other deuterated electrophile precursors. The reaction times and
(d, 2H, J = 8.6 Hz), 7.27 (d, 2H, J = 8.6 Hz); 13C NMR (CDCl3) d 55.26, 64.50
J. Label Compd. Radiopharm 2013, 56 6–11
Copyright © 2012 John Wiley & Sons, Ltd.
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D. W. Barnett et al.
(t, J = 22.0 Hz), 113.89, 128.63, 133.09, 159.12; HRMS-ES (m/z): [M + Na]+ 62.71, 66.15, 69.47, 75.46, 114.78, 127.39, 127.66. 128.29, 130.10, 137.13,
calculated for 162.0641, found 162.0645.
158.81; HRMS-ES (m/z): [M+ Na]+ calculated for 256.0615, found 256.0617.
(S)-(+)-4-benzyloxybenzyl-a-d alcohol (1d)
(R)-(À)-4-nitrobenzyl-a-d chloride (6e)
[a]2D1 + 0.59o (c 5.1, THF); 1H NMR (CDCl3) d 2.07 (br s, 1H), 4.55 (br s, 1H),
5.06 (s, 2H), 6.96 (d, 2H, J = 8.8 Hz), 7.27 (d, 2H, J = 8.8 Hz), 7.31–7.45 (m,
5H); 13C NMR (CDCl3) d 22.1 (t, J = 22.1 Hz), 69.90, 114.79, 127.32,
127.84, 128.46, 128.52, 133.26, 136.84, 158.23; HRMS-ES (m/z): [M + Na]+
calculated for 238.0954, found 238.0946.
[a]2D1 À 1.31o (c 6.9, THF); 1H NMR (acetone-d6) d 4.81 (s, 1H), 7.71 (d, 2H,
J= 8.6 Hz), 8.21 (d, 2H, J=8.6Hz); 13C NMR (acetone-d6) d 43.98 (t, J=23Hz),
123.54, 129.58, 144.82, 147.57; HRMS-EI (70 eV) m/z: calculated 172.01488,
found 172.01506.
15N-(À)-8-phenylmenthylhippurate
(S)-(+)-4-nitrobenzyl-a-d alcohol (1e)
A mixture of 15N-hippuric acid (prepared from 15N-glycine and benzoyl
[a]2D1 + 2.46o (c 6.1, THF); 1H NMR (CDCl3) d 4.81 (s, 1H), 7.52 (d, 2H,
J = 8.8 Hz), 8.21 (d, 2H, J = 8.8 Hz); 13C NMR (CDCl3/acetone-d6) d 62.24
(t, J = 23 Hz), 123.03, 126.79, 146.78, 150.19; HRMS-ES (m/z): [M + Na]+
calculated for 177.0386, found 177.0378.
chloride; 2.0 g, 11.2 mmol), (À)-8-phenylmenthol (prepared by the
procedure of Ort21
; 0.75 g, 3.2 mmol), and p-toluenesulfonic acid
(100 mg) was refluxed under Dean–Stark conditions. Upon disappear-
ance of the alcohol by TLC, the reaction was cooled to room temperature,
saturated NaHCO3 was added (15 mL) and the mixture was stirred for
General procedure for stereochemical analysis of deuterated 15 min. The phases were separated, and the organic phase was washed
with water and brine, concentrated, and purified by silica gel flash
column chromatography to yield a white solid (1.2 g, 93%): 1H NMR
(CDCl3) d 0.87 (d, 3H, J = 6.5 Hz), 0.89–1.02 (m), 1.52–2.11 (m), 3.41
(dd, 1H, J = 18.4, 5.7 Hz), 3.58 (dd, 1H, J = 18.4, 4.6 Hz), 4.91 (dd, 1H,
J = 10.7, 4.6 Hz), 6.05 (dt, 1H, J = 91.4, 5.0 Hz), 7.12–7.73 (m, 10H).
benzyl alcohols
The procedure used for the preparation of diastereomeric derivatives of
labeled benzyl alcohols was adapted from Williams and co-workers.6 The
procedure that follows was used for the preparation of the (À)-camphanate
ester of (S)-(+)-benzyl-a-d alcohol (1a) and is representative of the other
deuterated alcohols. A solution of the alcohol in a small amount
of toluene/pyridine was added to a solution of (À)-camphanic chloride
(2 equivalents) in toluene at 0 ꢀC. The reaction mixture was stirred at 0ꢀC
for 30min then 5h at 25 ꢀC. The reaction mixture was concentrated under
reduced pressure, purified by preparative TLC, and analyzed by NMR
spectroscopy. For the (À)-camphanate ester of 1a: 1H NMR (800 MHz; CDCl3)
d 5.35 (br s, major), 5.39 (br s, minor), major/minor >98:2.
(2S, 3S)-N-benzoyl-O-triisopropylsilyl-[3-2H, 15N]-tyrosine-(À)-8-
phenylmenthyl ester (7)
Under dry conditions, a 0.5M solution of LDA was prepared by the
addition of a solution of n-BuLi (1 equivalent) in hexanes to a solution of dii-
sopropylamine (1 equivalent) in dry THF at À78 ꢀC. The solution was stirred
for 10min, and then tetramethylethylenediamine (1 equivalent) was added.
15N-(À)-8-Phenylmenthylhippurate (0.5 equivalent) was dissolved in dry
THF (0.25M solution) and added slowly. The mixture was stirred at
À78 ꢀC for 20min; after which, it was warmed to 0ꢀC, and electrophile 1b
(slight excess) was added slowly with further stirring for 2 h. The reaction
was quenched with 1M HCl, warmed to room temperature, diluted with
ethyl acetate, and the phases were separated. The ethyl acetate layer was
dried (MgSO4) and evaporated, and the residue was purified by silica gel
preparative TLC (15% ethyl acetate/hexane) to give 32mg (62%) of 7 as a
white solid: [a]2D1 + 11.2o (c 13.5, THF); 1H NMR (CDCl3) d 0.85–1.30 (m),
0.88 (d, 3H, J = 6.6Hz), 1.04 (s), 1.06 (s), 1.65–1.86 (m, 5H), 2.06 (dt, 1H,
J = 11.8, 3.5 Hz), 2.65 (br t, 1H, J= 3.7Hz), 4.17 (dt, 1H, J = 2.4, 1.3Hz), 4.78
(dt, 1H J= 10.7, 4.2Hz), 6.27 (dd, 1H, J = 91.4, 7.5 Hz), 6.71 (d, 2H, J = 8.3Hz),
6.85 (d, 2H, J = 8.3Hz), 7.24–7.26 (m, 4H), 7.66 (d, 2H, J = 7.2Hz); 13C NMR
(CDCl3) d 12.28, 14.12, 17.83, 20.93, 21.62, 23.82, 26.37, 28.81, 31.24, 34.41,
35.99 (t, J= 18.9 Hz), 39.40, 41.54, 50.33, 53.15, 53.27, 60.28, 76.06, 119.80,
125.19, 125.34, 126.92, 128.02, 128.43, 130.37, 131.42, 134.29, 134.37,
151.23, 154.80, 166.31, 166.47, 170.91; HPLC (95% CH3OH/H2O): tR = 20.2
min (major), 15.6min (minor).
General procedure for synthesis of deuterated benzyl chlorides
Polymer-supported triphenylphosphine was washed with THF and dried
under high vacuum at 100 ꢀC for 12 h prior to use. To a dry flask under
argon was added TPP resin (~3 mmol/g, 1.89 mmol) and dry THF
(25 mL). The suspension was cooled to 0 ꢀC, and HCA (0.5 equivalents)
was added. After it was stirred at 0 ꢀC for 20 min, a solution of the
benzyl-a-d alcohol (1.0 equivalent) in dry THF was added dropwise. The
reaction mixture was stirred for 1 h at 0 ꢀC then warmed to room
temprerature. When TLC of the mixture showed no remaining starting
material, the resin was removed by filtration and washed with CH2Cl2,
and the combined solvents were removed under reduced pressure. The
product chlorides were then purified using silica gel chromatography
with yields shown in Table 1.
(R)-(À)-benzyl-a-d chloride (6a)
[a]2D1 À 1.52o (c 6.9, THF); 1H NMR (CDCl3) d 4.53 (s, 1H), 7.28–7.36 (m, 5H);
13C NMR (CDCl3) d 45.76 (t, J = 23 Hz), 128.18, 128.28, 137.29; HRMS-EI
(70 eV) m/z: calculated 127.02980, found 127.03019.
(2S, 3S)-N-benzoyl-[3-2H, 15N]-tyrosine-(À)-8-phenylmenthyl ester (8)
To a stirred solution of 14 mg (0.024 mmol) of 7 in THF-H2O (9:1) was
added 13 mg (2 equivalent) of TBAFÁH2O. After 3 h of stirring, TLC showed
consumption of starting material, and the solution was diluted with
water and extracted with ethyl acetate. The ethyl acetate layer was
washed with brine, dried (MgSO4), concentrated, and purified by
preparative silica gel TLC (40% ethyl acetate/hexane) to give 10 mg
(R)-(À)-4-triisopropylsilyloxybenzyl-a-d chloride (6b)
[a]2D1 À 0.39o (c 15.4, THF); 1H NMR (CDCl3) d 1.09 (d, 18H, J = 7.4 Hz), 1.25
(m, 3H), 4.63 (br t, 1H), 6.89 (d, 2H, J = 8.5 Hz), 7.31 (d, 2H, J = 8.5 Hz); 13C
NMR (CDCl3) d 13.27, 18.15, 46.45 (t, J= 23.2 Hz), 120.65, 131.07, 131.50,
156.93; HRMS-ES (m/z): [M+ Na]+ calculated for 322.1480, found 322.1470.
1
(98%) of 8 as a white solid: H NMR (CDCl3) d 0.89–1.29 (m), 0.90 (d, 3H,
J = 6.4 Hz), 1.02 (s), 1.20 (s), 1.47 (br s, 1H), 1.70 (d, 1H, J = 12.7 Hz), 1.84
(dt, 2H, J = 13.8, 3.1 Hz), 2.12 (dt, 1H, J = 11.6, 3.1 Hz), 2.65 (m, 1H), 4.20
(t, 1H, J = 6.1 Hz), 4.86 (dt, 1H, J = 11.0, 4.2 Hz), 6.44 (dd, 1H, J = 91.2,
7.9 Hz), 6.70 (d, 2H, J = 8.8 Hz), 6.88 (d, 2H, J = 8.8 Hz), 7.17–7.30 (m, 4H),
7.37–7.51 (m, 4H), 7.68 (d, 2H, J = 7.2 Hz); 13C NMR (CDCl3) d 12.24,
14.07, 17.62, 20.95, 21.69, 23.37, 26.28, 29.12, 31.19, 34.35, 36.10 (m),
39.34, 41.48, 50.23, 53.36, 53.49, 60.45, 76.19, 115.37, 125.16, 125.35,
126.94, 128.01, 128.53, 130.35, 131.66, 133.83, 133.91, 151.51, 155.49,
(R)-(À)-4-methoxybenzyl-a-d chloride (6c)
[a]2D1 À 0.18o (c 16.6, THF); 1H NMR (CDCl3) d 3.79 (s, 3H), 4.53 (br t, 1H,
J = 1.6 Hz), 6.87 (d, 2H, J = 8.8 Hz), 7.30 (d, 2H, J = 8.8 Hz); 13C NMR
(acetone-d6) d 45.89 (t, J = 23.2 Hz), 55.15, 113.99, 129.91, 159.52,
175.42; HRMS-EI (70 eV) m/z: calculated 157.04037, found 157.04137.
(R)-(À)-4-benzyloxybenzyl-a-d chloride (6d)
[a]D21 À 0.37o (c 13.6, THF); 1H NMR (CDCl3) d 4.53 (br s, 1H), 5.50 (s, 2H), 6.93 167.03, 167.19, 170.97; HPLC (85% CH3OH/H2O): tR = 8.4 min (major),
(d, 2H, J = 8.6Hz), 7.28–7.42 (m, 7H); 13C NMR (CDCl3) d 45.64 (t, J = 23.0 Hz), 7.4 min (minor).
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J. Label Compd. Radiopharm 2013, 56 6–11

D. W. Barnett et al.
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A suspension of 40mg (0.08 mmol) of 8 in concentrated HCl (10 mL) was
warmed to 70ꢀC for 12 h. The cooled solution was then washed with
CH2Cl2, and the aqueous phase was evaporated to give 10.3mg (59%)
1
of 9 as a white solid: H NMR (D2O) d 2.2 (br dd, 1H, J= 7.5, 2.2Hz), 3.10
1
(residual H, br t, J= 3.4Hz), 4.08 (d, 1H, J= 7.5Hz), 6.75 (d, 2H, J = 8.3Hz),
7.05 (d, 2H, J = 8.3Hz); 13C NMR (D2O/CD3OD) d 35.25 (t, J= 19.6Hz), 55.13,
116.56, 126.40, 131.47, 155.87, 172.43; HRMS-ES (m/z): [M]+ calculated for
184.0958, found 184.0951.
Acknowledgements
We gratefully acknowledge financial support of this work by the
National Science Foundation (NSF MCB9723642).
[5] R. W. Curley, Jr., M. J. Panigot, A. P. Hansen, S. W. Fesik, J. Biomol. NMR
1994, 4, 335–340.
Conflict of Interest
[6] R. M. Williams, D. Zhai, P. J. Sinclair, J. Org. Chem. 1986, 51, 5021–5022.
[7] O. Mitsunobu, Synthesis 1981, 1–28.
The authors did not report any conflict of interest.
[8] Pollastri, M. P.; Sagal, J. F.; Chang, G. Tetrahedron Lett. 2001, 42,
2459–2460.
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J. Label Compd. Radiopharm 2013, 56 6–11
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