Preparation of optically active allothreonine via optical resolution by replacing crystallization.

Article abstract of DOI:10.1248/cpb.50.287

An attempt was made to use a simple procedure to obtain D- and L-allothreonine (D- and L-aThr), which are non-proteinogenic alpha-amino acids and are useful as chiral reagents in asymmetric syntheses. DL-aThr that exists as a conglomerate was optically resolved by replacing crystallization with L-alanine (L-Ala) as an optically active co-solute. D-aThr was preferentially crystallized from an aqueous solution of DL-aThr in the presence of L.-Ala, as was L-aThr in the presence of D-Ala. Furthermore, a diasteroisomeric mixture of D-aThr and L-threonine (L-Thr) and one of L-aThr and D-Thr were prepared, respectively, by epimerization of L- and D-Thr using salicylaldehyde as the catalyst in acetic acid. Based on the result of the replacing crystallization, D- and L-aThr were separated from aqueous solutions of the diastereoisomeric mixtures in the presence of L- and D-Ala. The partially resolved D- and L-aThr were recrystallized from water to yield the corresponding enantiomers in optically pure forms.

Full text of DOI:10.1248/cpb.50.287

February 2002
Notes
Chem. Pharm. Bull. 50(2) 287—291 (2002)
287
Preparation of Optically Active Allothreonine via Optical Resolution by
Replacing Crystallization
Tadashi SHIRAIWA,* Keiji FUKUDA, and Motoki KUBO
Unit of Chemistry, Faculty of Engineering, Kansai University, Yamate-cho, Suita, Osaka 564–8680, Japan.
Received September 10, 2001; accepted October 16, 2001
An attempt was made to use a simple procedure to obtain D- and L-allothreonine (D- and L-aThr), which are
non-proteinogenic a-amino acids and are useful as chiral reagents in asymmetric syntheses. DL-aThr that exists
as a conglomerate was optically resolved by replacing crystallization with L-alanine (L-Ala) as an optically active
co-solute. D-aThr was preferentially crystallized from an aqueous solution of DL-aThr in the presence of L-Ala, as
was L-aThr in the presence of D-Ala. Furthermore, a diasteroisomeric mixture of D-aThr and L-threonine (L-Thr)
and one of L-aThr and D-Thr were prepared, respectively, by epimerization of L- and D-Thr using salicylaldehyde
as the catalyst in acetic acid. Based on the result of the replacing crystallization, D- and L-aThr were separated
from aqueous solutions of the diastereoisomeric mixtures in the presence of L- and D-Ala. The partially resolved
D- and L-aThr were recrystallized from water to yield the corresponding enantiomers in optically pure forms.
Key words allothreonine; optical resolution; alanine; threonine; epimerization
7
)
D- and L-Allothreonine [D- and L-aThr; (2R,3R)- and aThr. Although L-aThr will be preferentially crystallized
2S,3S)-2-amino-3-hydroxybutanoic acid] are useful as chiral from the racemic solution in the presence of D-Hyp, L-Hyp is
(
1
)
reagents in asymmetric syntheses. However, D- and L-aThr, expensive and D-Hyp is not commercially available. There-
non-proteinogenic a-amino acids, are difficult to produce fore, we selected D- and L-Ala, which are commercially avail-
commercially in large quantities. Therefore, synthetic DL- able and are the most inexpensive optically active a-amino
2
—4)
aThr
ing the diastereoisomeric salts of DL-aThr derivatives to ob- by replacing crystallization of DL-aThr (Chart 1).
has been subjected to optical resolution by separat- acids, as the optically active cosolutes in optical resolution
5
,6)
7)
tain the enantiomers. In our previous paper, DL-aThr was
Optical resolution by replacing crystallization is based on
found to exist as a conglomerate and to be optically resolved different interactions between enantiomers and the optically
by preferential crystallization and replacing crystallization of active co-solute. Therefore, the solubilities (mole fractions)
DL-aThr. The optical resolution by replacing crystallization is of D-, L-, and DL-aThr were first measured in the presence of
3
a procedure for obtaining an enantiomer from a conglomer- L-Ala (16.0 mmol) in 100 cm of water at 10 °C; these values
7
)
ate, and is achieved by allowing an optically active co-solute are summarized in Table 1, together with the solubilities in
8
)
to coexist in a racemic supersaturated solution. DL-aThr was the absence of L-Ala.
optically resolved using 4-hydroxy-L-proline (L-Hyp) as the
When L-Ala was present in aqueous solutions of D-, L-, and
optically active cosolute, and D-aThr was allowed to prefer- DL-aThr, D-aThr was less soluble than L-aThr. In addition, D-,
entially crystallize from a supersaturated solution of DL- L-, and DL-aThr were each less soluble in the presence of L-
Chart 1. Preparation of Optically Active Allothreonine (aThr)
Reagents: (a) i) N-Bromosuccinimide, H O, ii) concentrated aqueous ammonia; (b) L-Ala as an optically active co-solute; (c) D-Ala as an optically active co-solute; (d) salicyl-
2
aldehyde, acetic acid, 80 °C.
∗
To whom correspondence should be addressed. e-mail: shiraiwa@ipcku.kansai-u.ac.jp
© 2002 Pharmaceutical Society of Japan

2
88
Vol. 50, No. 2
a)
Table 1. Solubilities of DL-, D-, and L-Allothreonine
1.00—4.00 mmol of L-Ala, the yield of L-aThr rapidly de-
creased with increasing amounts of L-Ala. When a larger
amount (4.00—6.00 mmol) of L-Ala was present in the
racemic solution, the yield of L-aThr was approximately con-
stant, and D-aThr of optical purity of 80—85% was obtained
in yields of 0.54—0.58 g. Therefore, to optimize the condi-
tions, the optical resolution was conducted by stirring mix-
tures containing 4.00 mmol of L-Ala with varying degrees of
supersaturation from 174—214% for 80 min.
When 176—200% supersaturated solutions were used, L-
aThr seemed to be hardly crystallized because the optical
resolution afforded D-aThr of optical purities of 80—86% in
yields of 0.37—0.59 g; the yield tended to increase with in-
creasing supersaturation. When a 204% supersaturated solu-
tion was used, L-aThr began to rapidly crystallize, hence D-
aThr of optical purity of 71% was obtained. In addition, the
optical resolution for the 196% supersaturated solution was
carried out by stirring for 70—180 min. Although almost no
aThr was crystallized by stirring for 70 min, D-aThr of an op-
tical purity of 81% was obtained in a yield of 0.580 g by stir-
ring for 80 min. However, D-aThr, obtained by stirring for
L-Ala as an optically
Solubility
(g (100 cm of water)
[mole fraction]
3
Ϫ1
Allothreonine
active co-solute
)
(
mmol)
D-aThr 5.15 [0.0077]
L-aThr 5.15 [0.0077]
5.28 [0.0079]
D-aThr 4.51 [0.0067]
L-aThr 4.81 [0.0072]
4.93 [0.0074]
5.06 [0.0076]
b)
c)
DL-aThr
—
10.30
{
b)
c)
L-aThr
—
DL-aThr
16.0
9.32
{
D-aThr
L-aThr
16.0
16.0
3
a) Conditions: Water, 100 cm ; temperature, 10 °C. b) See ref. 7. c) None.
9
0 min, showed a low optical purity (44%), because of the
onset of rapid crystallization of L-aThr.
On the other hand, when D-Ala was used as the co-solute,
L-aThr of an optical purity of 85% was obtained in a yield of
0
.574 g from the solution containing DL-aThr (4.384 g) and D-
3
Ala (4.00 mmol) in 25 cm of water by stirring for 80 min.
The obtained D- and L-aThr were recrystallized from water
to give optically pure D- and L-aThr. For example, optically
pure D-aThr (3.90 g) was obtained from 5.00 g of D-aThr of
an optical purity of 85%, and optically pure L-aThr (3.05 g)
was obtained from 4.00 g of L-aThr of an optical purity of
Fig. 1. Relationship between Yield of Enantiomer and L-Alanine in Opti-
cal Resolution of DL-Allothreonine
3
Conditions: DL-aThr, 4.384 g (36.8 mmol); solvent, 25 cm of water; L-Ala, 0.0891—
0
.535 g (1.00—6.00 mmol); temperature, 10 °C; resolution time, 80 min. Yield of enan-
tiomer: ᭺, D-aThr; ᭹, L-aThr.
8
1%, as described in the Experimental section.
Based on the above results, successive optical resolution
Ala than in its absence. Therefore, a repulsive interaction is was attempted by stirring the 196% supersaturated solution
thought to occur between D-aThr and L-Ala, and between L- of DL-aThr with 4.00 mmol of L-Ala for 80 min. As described
aThr and L-Ala, and the interaction between D-aThr and L- above for solubility, repulsive interactions are estimated to
1
1)
Ala may be stronger than that between L-aThr and L-Ala.
occur not only between D-aThr and L-Ala, but also between
These findings suggested that when L-Ala was present in the L-aThr and L-Ala. These interactions promote the crystalliza-
supersaturated aqueous solution of DL-aThr, D-aThr was pref- tion of D- or L-aThr during the preferential crystallization of
erentially crystallized from the solution.
their enantiomers. Therefore, after optical resolution of the
Based on the above findings, the optical resolution of initial solution, a small amount of D- or L-aThr was added as
DL-aThr was attempted by stirring a mixture containing seed crystals to the solution in subsequent optical resolu-
3
6.8 mmol (4.384 g) of DL-aThr and 1.00—6.00 mmol of L- tions. These results are summarized in Table 2. The degrees
3
Ala, as the optically active co-solute, in 25 cm of water for of resolution of D- and L-aThr [DR(%)] in Table 2 were cal-
0 min at 10 °C. The yields of enantiomers [YE (g) and culated:
8
L
YE (g)] of L- and D-aThr were calculated from
D
DR(%)ϭ[(Yield(g)ϫOP(%)/100ϪSC)/
YE (g)ϭ(1/2)[Yield(g)ϫ(100ϪOP(%))/100]
(1)
(2)
(operation amount of D- or L-aThϪS or S )]ϫ100
(3)
L
D
L
and
where S (1.232 g) is the solubility of L-aThr in DL-aThr in
L
3
2
5 cm of water at 10 °C, and S (0.050 g) is the amount of
C
YE (g)ϭYield(g)ϪYE (g),
D
L
seed crystals. The operation amounts (g) are the amounts of
where OP (%) is the optical purity of obtained D-aThr, calcu- D- and L-aThr in the solution, and the values in runs 2—6 are
2
0
lated based on the specific rotation of authentic D-aThr: [a]
calculated based on the results from runs 1—5, respectively.
A successive optical resolution was achieved to afford D-
D
Ϫ3
7)
Ϫ32.8° (cϭ1.00, 1 mol dm HCl). The results are shown
in Fig. 1. The purity of the crystallized aThr was confirmed aThr of optical purities of 81—90%, and L-aThr with purities
1
by its H-NMR spectrum to be free of L-Ala.
of 96 and 100% at 48—60% degrees of resolution. The par-
When L-Ala was present as an optically active co-solute, tially resolved D-aThr was recrystallized from water to give
D-aThr was preferentially crystallized from the racemic solu- optically pure D-aThr.
tion. The yield of D-aThr slightly decreased with increasing
Next, we attempted to more simply obtain optically active
amounts of L-Ala. On the other hand, in the presence of aThr, based on the above result. In general, optically active

February 2002
289
a)
Table 2. Successive Optical Resolution by Replacing Crystallization of DL-Allothreonine
Operation amounts of
D- and LaThr (g)
aThr obtained
b)
Added amount of
DL-aThr (g)
Resolution time
Run
1
c)
d)
(min)
Yield
Optical purity
Degree of resolution
D-aThr
L-aThr
(g)
(%)
(%)
4.384
0
0.500
0
0.380
0
2.192
1.666
1.916
1.485
1.675
1.429
2.192
2.138
1.886
1.860
1.672
1.653
80
90
60
80
70
90
D 0.580
L 0.552
D 0.507
L 0.428
D 0.315
L 0.296
81.4
47.7
55.4
56.6
60.2
48.2
55.7
e)
2
3
4
5
100
89.6
100
88.2
96.1
f)
e)
f)
e)
6
3
a) Conditions: optically active co-solute, 0.356 g (4.00 mmol) of L-Ala; solvent, 25 cm of water; temperature, 10 °C. b) The operation amounts in runs 2—6 were calcu-
lated from the results in 1—5, respectively. c) The Yield is the sum of the amounts of the crystallized aThr and seed crystals. d) The optical purities of D- and L-aThr obtained
7)
26
Ϫ3
were calculated on the basis of the specific rotation of authentic D-aThr; lit. [a]D Ϫ32.8° (cϭ1.00, 1 mol dm HCl). e) Seed crystals, 0.050 g of L-aThr. f) Seed crystals,
0
.050 g of D-aThr.
Fig. 2. Relationship between Yield of Diastereoisomer and L-Alanine in
Separation of D-Allothreonine and L-Threonine
Fig. 3. Relationship between Yield of Diastereoisomer and Stirring Time
in Separation of D-Allothreonine and L-Threonine
Conditions: Diastereoisomeric mixture (L-Thr in 18%de), 5.965 g (50.0 mmol); sol-
vent, 25 cm of water; L-Ala, 0.178—0.802 g (2.00—9.00 mmol); temperature, 10 °C;
stirring time, 3 h. Yield of diastereoisomer: ᭺, D-aThr; ᭹, L-Thr.
Conditions: Diastereoisomeric mixture (L-Thr in 18%de), 5.965 g (50.0 mmol); sol-
vent, 25 cm of water; L-Ala, 0.535 g (6.00 mmol); temperature, 10 °C; stirring time,
1—5 h. Yield of diastereoisomer: ᭺, D-aThr; ᭹, L-Thr.
3
3
3
a-amino acids undergo racemization with carbonyl com- 17.5%de) in 25 cm of water for 3 h at 10 °C in the presence
pounds, such as salicylaldehyde, which acts as a catalyst in of 2.00—9.00 mmol of L-Ala, as shown in Fig. 2.
9
)
acetic acid. Therefore, epimerization at the C-2 position of
The yield of D-aThr tended to gradually increase with in-
threonine (Thr) will yield a diastereoisomeric mixture of Thr creasing amounts of L-Ala. On the other hand, when L-Ala
and aThr. However, Thr and aThr are difficult to separate (2.00—6.00 mmol) was present, the yield of L-Thr rapidly
from the mixture without transformation into their deriva- decreased with increasing amounts of L-Ala. However, in the
1
0)
tives, such as O-methyl-N-acyl derivatives. In our previous presence of 6.00—9.00 mmol of L-Ala, the yield of L-Thr
1
1)
paper, L-Thr was prevented from crystallizing from an was approximately constant. Therefore, D-aThr was crystal-
aqueous solution of DL-Thr in the presence of L-Ala. There- lized in 74—78%de from a solution containing 6.00—
fore, we attempted to separate D- and L-aThr from mixtures 9.00 mmol of L-Ala. When the solution of the mixture was
of Thr and aThr (Chart 1).
stirred for 1—5 h in the presence of 6.00 mmol of L-Ala, the
L-Thr was subjected to epimerization using salicylalde- yield of D-aThr was approximately constant, as shown in Fig.
hyde as the catalyst, and was then stirred for 1—5 h in acetic 3. On the other hand, L-Thr was hardly crystallized during
acid at 80 °C. The epimerization gave a mixture of L-Thr and the first 1—2 h, and then rapidly crystallized with prolonged
D-aThr in a molar ratio of 1 : 0.7, regardless of the reaction stirring. Therefore, D-aThr was obtained in 98%de and in a
time after 1 h; the mixture was L-Thr with a diastereoiso- yield of more than 1 g by stirring for 1 and 2 h. In addition,
meric excess (de) of about 17%. The molar ratio of L-Thr and the aqueous solution of the diastereoisomeric mixture pre-
D-aThr in the mixture was determined by the intensity ratios pared from D-Thr was stirred for 1 h in the presence of
1
of the methine proton signals at the C-2 positions in the H- 6.00 mmol of D-Ala to give L-aThr in 97%de. D- and L-aThr
NMR spectrum of the mixture. The yield of the diastereoiso- obtained in low de were recrystallized from water to give D-
meric mixture was approximately constant (1.1 g) during the and L-aThr as single diastereoisomers. For example, D-aThr
first 1—3 h, then rapidly decreased with increasing reaction (1.54 g) with 100%de and L-aThr (1.65 g) with 100%de were
time, due to decomposition of L-Thr and D-aThr. Separation obtained from 2.00 g of D-aThr of 78%de and L-aThr of
of D-aThr from the diastereoisomeric mixture was attempted 84.5%de, respectively.
by stirring 5.965 g (50.0 mmol) of the mixture (L-Thr of
We reported optical resolution by preferential crystalliza-

2
90
Vol. 50, No. 2
2
0
Ϫ3
tion of DL-aThr, which is another procedure for obtaining following manner: D-aThr (5.00 g) ([a] Ϫ27.9° (cϭ1.00, 1 mol dm
D
2
0
Ϫ3
HCl)) or L-aThr (4.00 g) ([a] ϩ26.6° (cϭ1.00, 1 mol dm HCl)) was
added to water (7.5 cm ). The mixture was vigorously stirred for 3 h at 10 °C
before the purified D- or L-aThr was collected by filtration and dried.
D-aThr: yield, 3.90 g; [a] Ϫ32.8° (cϭ1.00, 1 mol dm HCl). L-aThr:
yield, 3.05 g; [a]D ϩ32.8° (cϭ1.00, 1 mol dm HCl). The H-NMR spectra
enantiomers from a conglomerate, and D- and L-aThr were
obtained in yields of about 10% based on the amounts of D-
and L-aThr in the supersaturated solution. On the other
hand, optical rersolution by replacing crystallization gave D-
D
3
7
)
20
D
Ϫ3
2
0
Ϫ3
1
and L-aThr in yields of more than 20%. However, successive of D- and L-aThr were virtually identical to that of DL-aThr.
Successive Optical Resolution DL-aThr (4.384 g, 36.8 mmol) and L-Ala
optical resolution required a small amount of L-aThr as seed
crystals to promote the preferential crystallization of L-aThr.
Separation of optically active aThr from the diastereoiso-
3
(
0.356 g, 4.00 mmol) were dissolved in 25 cm of water at 50 °C. After cool-
ing the solution to 10 °C, followed by stirring for 80 min at 10 °C, precipi-
tated D-aThr (0.580 g) was collected by filtration and dried (run 1 in Table
meric mixture was more simply achieved using D- and L-Ala 2). After adding 0.050 g of L-aThr to the filtrate at 10 °C, followed by stirring
as the co-solutes, and afforded D- amd L-aThr in high de and for 90 min, precipitated L-aThr (0.552 g) was collected by filtration (run 2 in
Table 2). DL-aThr (0.500 g) was dissolved in the filtrate at 50 °C. After
adding 0.050 g of D-aThr to the filtrate at 10 °C, followed by stirring for
yields of more than 40%, based on the amounts of D- and L-
aThr in the mixtues.
6
0 min, precipitated D-Thr (0.507 g) was collected by filtration (run 3 in
Table 2). Optical resolution was carried out at 10 °C in a manner similar to
that just described; the detailed conditions are given for runs 4—6 in Table
Experimental
General Specific rotation values were measured at 589 nm with a 2.
Horiba Seisakusho SEPA-300 auto-polarimeter equipped with a quartz cell
Epimerization of Optically Active Threonine L-Thr (2.38 g, 20.0
1
3
with a 5.00 cm path length. H-NMR spectra were recorded with a JNM-
mmol) was dissolved in 100 cm of acetic acid at 80 °C. After adding salicy-
FX270 FT NMR system using sodium 3-(trimethylsilyl)propane-1-sulfonate laldehyde (0.244 g, 2.00 mmol) to the solution, the mixture was stirred for
(
DSS) as an internal standard. Chemical shift values were reported in d units 1—5 h at 80 °C. The mixture was concentrated in vacuo at 60 °C to give a
downfield from DSS. Melting points were measured with a Yanaco MP-500
D micro melting point apparatus.
mixture of L-Thr and D-aThr as the diastereoisomeric residue. After adding
3
50 cm of methanol to the residue, followed by stirring for 0.5 h at 40 °C, the
D- and L-Thr and D- and L-Ala were purchased from Wako Pure Chemical
Ind. DL-aThr was synthesized starting from (E)-2-butenoic acid, purchased
mixture was collected by filtration, washed thoroughly with methanol, and
dried. The molar ratio of L-Thr and D-aThr in the mixture was determined by
7
)
from Wako Pure Chemical Ind.; mp 241—243 °C (decomp) (lit, mp 242— the intensity ratios of the methine proton signals at the C-2 positions in the
2
)
3)
7)
1
2
43 °C (decomp); mp 260 °C (decomp); mp 240—242 °C (decomp)).
H-NMR spectrum of the mixture.
The mixture obtained at 1 h: yield, 1.13 g; [a]D Ϫ21.9° (cϭ1.00,
1
20
H-NMR (270 MHz, D O, DSS) d: 4.36 (1H, qd, Jϭ6.8, 4.1 Hz, 3-CH), 3.83
1H, d, Jϭ4.1 Hz, 2-CH), 1.20 (3H, d, Jϭ6.8 Hz, 4-CH ).
Optical Resolution by Replacing Crystallization DL-aThr (4.384 g,
6.8 mmol) and L-Ala (0.0891—0.535 g, 1.00—6.00 mmol) were dissolved
2
Ϫ3
(
1 mol dm HCl); the mixture was composed of L-Thr and D-aThr in the
3
20
molar ratio of 1 : 0.69. The mixture obtained at 1.5 h: yield, 1.12 g; [a]
D
Ϫ3
3
Ϫ22.0° (cϭ1.00, 1 mol dm HCl); the mixture was composed of L-Thr and
3
in 25 cm of water at 50 °C. After cooling the solution to 10 °C over 30 min, D-aThr at a molar ratio of 1 : 0.70. The mixture obtained at 2 h: yield, 1.11 g;
2
0
Ϫ3
followed by stirring for 80 min with a blade (0.80 cm width; 2.5 cm length) [a] Ϫ22.1° (cϭ1.00, 1 mol dm HCl); the mixture was composed of L-
at 100 rpm and 10 °C, the precipitated D-aThr was collected by filtration,
washed with a small amount of methanol, and dried.
D
Thr and D-aThr in the molar ratio of 1 : 0.71. The mixture obtained at 2.5 h:
yield, 1.11 g; [a]D Ϫ22.2° (cϭ1.00, 1 mol dm HCl); the mixture was com-
posed of L-Thr and D-aThr in the molar ratio of 1 : 0.72. The mixture ob-
tained at 3 h: yield, 1.10 g; [a]D Ϫ22.2° (cϭ1.00, 1 mol dm HCl); the
mixture was composed of L-Thr and D-aThr in the molar ratio of 1 : 0.72.
2
0
Ϫ3
2
0
D-aThr obtained using 1.00 mmol of L-Ala: yield, 0.909 g; [a] Ϫ7.83°
D
Ϫ3
20
Ϫ3
(
0
3
cϭ1.00, 1 mol dm HCl). D-aThr obtained using 2.50 mmol of L-Ala: yield,
2
0
Ϫ3
.671 g; [a] Ϫ20.1° (cϭ1.00, 1 mol dm HCl). D-aThr obtained using
D
2
0
Ϫ3
20
.00 mmol of L-Ala: yield, 0.636 g; [a]D Ϫ22.0° (cϭ1.00, 1 mol dm HCl). The mixture obtained at 4 h: yield, 0.884 g; [a] Ϫ22.0° (cϭ1.00,
D
2
0
Ϫ3
D-aThr obtained using 4.00 mmol of L-Ala: yield, 0.580 g; [a] Ϫ26.7° (cϭ 1 mol dm HCl); the mixture was composed of L-Thr and D-aThr in the
D
Ϫ3
20
1
0
6
.00, 1 mol dm HCl). D-aThr obtained using 5.00 mmol of L-Ala: yield,
.544 g; [a] Ϫ26.4° (cϭ1.00, 1 mol dm HCl). D-aThr obtained using
.00 mmol of L-Ala: yield, 0.542 g; [a]D Ϫ27.7° (cϭ1.00, 1 mol dm HCl).
molar ratio of 1 : 0.70. The mixture obtained at 5 h: yield, 0.515 g; [a]D
Ϫ21.9° (cϭ1.00, 1 mol dm HCl); the mixture was composed of L-Thr and
D-aThr in the molar ratio of 1 : 0.69. H-NMR of the mixture obtained at
2
0
Ϫ3
Ϫ3
D
2
0
Ϫ3
1
Optical resolution was carried out for the 176—214% supersaturated so- 1.5 h (270 MHz, D O, DSS) d: 4.36 (0.7H, qd, Jϭ6.8, 4.1 Hz, 3-CH (D-
2
lutions of DL-aThr (3.931—4.765 g, 33.0—40.0 mmol) in the presence of L-
Ala (0.356 g, 4.00 mmol) by stirring for 80 min at 10 °C in a manner similar
to that described above.
aThr)), 4.25 (1H, qd, Jϭ6.5, 4.9 Hz, 3-CH (L-Thr)), 3.83 (0.7H, d, Jϭ4.1 Hz,
2-CH (D-aTr)), 3.58 (1H, d, Jϭ4.9 Hz, 2-CH (L-Thr)), 1.32 (3H, d,
Jϭ6.6 Hz, 4-CH (L-Thr)), 1.20 (2.1H, d, Jϭ6.8 Hz, 4-CH (D-aThr)). The
H-NMR spectra of the other mixtures were similar to that obtained at 1.5 h.
Epimerization of D-Thr (2.38 g, 20.0 mmol) was carried out at a reaction
3
3
2
0
1
D-aThr obtained from 176% supersaturated solution: yield, 0.371 g; [a]
D
Ϫ3
Ϫ26.4° (cϭ1.00, 1 mol dm HCl). D-aThr obtained from 182% supersatu-
2
0
Ϫ3
rated solution: yield, 0.449 g; [a]D Ϫ26.7° (cϭ1.00, 1 mol dm HCl). D-
time of 1.5 h in a manner similar to L-Thr.
2
0
20
aThr obtained from 187% supersaturated solution: yield, 0.516 g; [a]
The mixture of D-The and L-aThr: yield, 1.14 g; [a]D ϩ20.1° (cϭ1.00,
D
Ϫ3
Ϫ3
Ϫ28.3° (cϭ1.00, 1 mol dm HCl). D-aThr obtained from 192% supersatu-
1 mol dm HCl); the mixture was composed of D-Thr and L-aThr at a molar
2
0
Ϫ3
1
rated solution: yield, 0.552 g; [a]D Ϫ26.9° (cϭ1.00, 1 mol dm HCl). D-
ratio of 1 : 0.70. The H-NMR spectrum was virtually identical to that of the
2
0
aThr obtained from 200% supersaturated solution: yield, 0.588 g; [a]
mixture of L-Thr and D-aThr.
D
Ϫ3
Ϫ26.1° (cϭ1.00, 1 mol dm HCl). D-aThr obtained from 204% supersatu-
Separation of Optically Active Allothreonine from the Diastereoiso-
meric Mixture The diastereoisomeric mixture, composed of L-Thr and D-
aThr at a molar ratio of 1 : 0.7, (5.955 g, 50.0 mmol) and L-Ala (0.178—
2
0
Ϫ3
rated solution: yield, 0.631 g; [a]D Ϫ23.3° (cϭ1.00, 1 mol dm HCl). D-
2
0
aThr obtained from 209% supersaturated solution: yield, 0.654 g; [a]
D
Ϫ3
3
Ϫ22.5° (cϭ1.00, 1 mol dm HCl). D-aThr obtained from 214% supersatu-
0.802 g, 2.00—9.00 mmol) were dissolved in 25 cm of water at 50 °C. After
2
0
Ϫ3
rated solution: yield, 0.728 g; [a]D Ϫ18.6° (cϭ1.00, 1 mol dm HCl).
cooling the solution to 10 °C over 30 min, followed by stirring for 3 h with a
blade (0.80 cm width; 2.5 cm length) at 100 rpm and 10 °C, the precipitated
D-aThr was collected by filtration, washed with a small amount of methanol,
and dried. The diastereoisomeric excess (%de) of the obtained D-aThr was
Optical resolution was carried out for a solution of DL-aThr (4.384 g,
3
9
6.8 mmol) in the presence of L-Ala (0.356 g, 4.00 mmol) by stirring for
0—180 min at 10 °C in a manner similar to that described above.
2
0
D-aThr obtained at 90 min: yield, 0.830 g; [a] Ϫ14.5° (cϭ1.00, determined based on the intensity ratios of the methine proton signals at the
D
Ϫ3
20
1
1
mol dm HCl). D-aThr obtained at 120 min: yield, 1.079 g; [a]D Ϫ6.41°
C-2 positions of L-Thr and D-aThr in the H-NMR spectrum.
Ϫ3
20
D
20
(
cϭ1.00, 1 mol dm HCl). D-aThr obtained at 150 min: yield, 1.215 g; [a]
D-aThr obtained in 7.3%de using 2.00 mmol of L-Ala: yield, 1.86 g; [a]D
Ϫ3
Ϫ3
Ϫ3.32° (cϭ1.00, 1 mol dm HCl). D-aThr obtained at 180 min: yield,
Ϫ24.3° (cϭ1.00, 1 mol dm HCl). D-aThr obtained in 23.3%de using
2
0
Ϫ3
20
Ϫ3
1
.279 g; [a] Ϫ3.10° (cϭ1.00, 1 mol dm HCl).
3.00 mmol of L-Ala: yield, 1.62 g; [a]D Ϫ25.8° (cϭ1.00, 1 mol dm HCl).
D
20
Optical resolution was carried out for a solution of DL-aThr (4.384 g,
6.8 mmol) in the presence of D-Ala (0.356 g, 4.00 mmol) by stirring for
0 min at 10 °C in a manner similar to that described above.
L-aThr: yield, 0.574 g; [a]D ϩ27.9° (cϭ1.00, 1 mol dm HCl).
The partially resolved D- and L-aThr were recrystallized from water in the
D-aThr obtained in 44.3%de using 4.00 mmol of L-Ala: yield, 1.45 g; [a]
D
Ϫ3
3
8
Ϫ27.7° (cϭ1.00, 1 mol dm HCl). D-aThr obtained in 66.2%de using 5.00
2
0
Ϫ3
mmol of L-Ala: yield, 1.35 g; [a]D Ϫ29.7° (cϭ1.00, 1 mol dm HCl). D-
2
0
Ϫ3
20
aThr obtained in 74.4%de using 6.00 mmol of L-Ala: yield, 1.23 g; [a]D
Ϫ3
Ϫ30.5° (cϭ1.00, 1 mol dm HCl). D-aThr obtained in 77.3%de using

February 2002
291
2
0
Ϫ3
3
7
.00 mmol of L-Ala: yield, 1.24 g; [a]D Ϫ30.7° (cϭ1.00, 1 mol dm HCl).
lution containing 0.356 g (4.00 mmol) of L-Ala in 25 cm of water at 60 °C.
After vigorously stirring the solution for 10 h at 10 °C, the precipitated aThr
was rapidly collected by filtration and thoroughly dried. The solubility at
2
0
D-aThr obtained in 78.2%de using 9.00 mmol of L-Ala: yield, 1.29 g; [a]
D
Ϫ3
Ϫ30.8° (cϭ1.00, 1 mol dm HCl).
Separation of D-aThr from the diastereoisomeric mixture was carried out 10 °C was calculated on the basis of the weight of aThr. For the dissolution
in a solution of the mixture (4.384 g, 36.8 mmol) in the presence of L-Ala
of DL-aThr, the solubility of D- and L-aThr was estimated based on the opti-
0.535 g, 6.00 mmol) by stirring for 1—5 h in a manner similar to that de- cal purity of aThr obtained by filtration, and by its weight.
scribed above.
(
2
0
D-aThr obtained in 98.2%de at 1 h: yield, 1.03 g; [a] Ϫ32.6° (cϭ1.00, References
D
Ϫ3
20
D
1
mol dm HCl). D-aThr obtained in 97.3%de at 2 h: yield, 1.06 g; [a]
1) Coppola G. M., Schuster H. F., “Asymmetric Syntheses, Construction
of Chiral Molecules Using Amino Acids,” John Wiley & Sons, 1987,
pp. 163—166.
2) Carter H. E., Zirkle C. L., J. Biol. Chem., 178, 709—714 (1949).
3) Liwschitz Y., Rabinsohn Y., Perera D., J. Chem. Soc., 1962, 1116—
1119.
4) Harada K., Oh-hashi J., Bull. Chem. Soc. Jpn., 39, 2311—2312 (1966).
5) West H. D., Carter H. E., J. Biol. Chem., 122, 611—617 (1938).
6) Arold H., J. Prakt. Chem., 24, 23—26 (1964).
7) Miyazaki H., Morita H., Shiraiwa T., Kurokawa H., Bull. Chem. Soc.
Jpn., 67, 1899—1903 (1994).
Ϫ3
Ϫ32.6° (cϭ1.00, 1 mol dm HCl). D-aThr obtained in 48.7%de at 4 h: yield,
20
Ϫ3
1
3
.43 g; [a] Ϫ28.1° (cϭ1.00, 1 mol dm
HCl). D-aThr obtained in
D
2
0
Ϫ3
6.2%de at 5 h: yield, 1.59 g; [a]D Ϫ27.0° (cϭ1.00, 1 mol dm HCl).
Separation of L-aThr from the mixture composed of D-Thr and L-aThr at a
molar ratio of 1 : 0.7 was carried out in a solution of the mixture (4.384 g,
6.8 mmol) in the presence of D-Ala (0.535 g, 6.00 mmol) by stirring for 1 h
in a manner similar to that described above.
3
2
0
L-aThr was obtained in 97.2%de at 1 h: yield, 1.07 g; [a] ϩ32.5°
D
Ϫ3
(
cϭ1.00, 1 mol dm HCl).
The partially separated D- and L-aThr were recrystallized from water in
the following manner: mixture of D-aThr (2.00 g) of 78%de or L-aThr
8) Addadi L., Weinstrin S., Gati E., Weissbuch I., Lahav M., J. Am.
Chem. Soc., 104, 4610—4617 (1982).
3
(
2.00 g) of 85%de in 10 cm of water was vigorously stirred for 4 h at 10 °C,
then the purified D- or L-aThr was collected by filtration and dried.
9) Yamada S., Hongo C., Yoshioka R., Chibata I., J. Org. Chem., 48,
843—846 (1983).
mol dm HCl). L-aThr: yield, 1.65 g; mp 269—272 °C (decomp) (lit, mp 10) Carter H. E., West H. D., J. Biol. Chem., 119, 109—119 (1937).
2
0
D-aThr: yield, 1.52 g; mp 270—272 °C (decomp); [a] Ϫ32.8° (cϭ1.00,
D
Ϫ3
7)
1
2
2
0
Ϫ3
1
69—270 °C (decomp)); [a]D ϩ32.8° (cϭ1.00, 1 mol dm HCl). The H- 11) Shiraiwa T., Kubo M., Fukuda K., Kurokawa H., Biosci. Biotech.
NMR spectra of D- and L-aThr were virtually identical to that of DL-aThr.
Biochem., 63, 2212—2215 (1999).
Solubility DL-, D-, or L-aThr (4.765 g, 40.0 mmol) was dissolved in a so-