Optical resolution of DL-cysteine

Disclosed is a method of optical resolution of DL-cysteine utilizing optically active mandelic acid. The method facilitates commercial production of optically active cysteine, particularly L-cysteine which is in great demand.

BACKGROUND OF THE INVENTION 
1. Filed of the Invention 
The present invention concerns optical resolution of DL-cysteine, or, in 
other words, a method of producing optically active cysteine or cystine. 
2. State of the Art 
L-cysteine is produced nowadays mainly by extracting it from natural raw 
materials. Because this way of production cannot meet rapidly increasing 
demand, other ways of industrial production such as chemical synthesis or 
fermentation methods have been sought. In cases according to the 
synthesis, the product will be in DL-form, and therefore, it should be 
optically resolved to obtain L-cysteine. 
General methods of optically resolving DL-amino acids known to date are 
preferential crystallization, resolution with enzymes, and diastereomer 
resolution. As far as DL-cysteine is concerned, because it is chemically 
rather unstable and racemization reaction is difficult to occur, there has 
been established no method of commercially practicable optical resolution 
by the preferential crystalization or diastereomer resolution. 
With respect to the method of resolution using the enzyme, Japanese Patent 
Disclosure No. 29293/1976 describes a method of effecting acylase to 
S-alkyl- or aryl-mercapto-N-acyl-DL-cysteine to obtain optically active 
S-alkyl- or aryl-mercapto-cysteine, and reducing the latter to form 
optically active cysteine. The method cannot be said advantageous for 
commercial practice, because the method not only uses the enzyme which is 
an organism substance, but also requires multiple steps of N-acylation, 
S-alkyl- or aryl-mercaptization, and further, dealkyl- or 
dearyl-mercaptization. 
On the other hand, as an example of optically resolving certain DL-amino 
acids with optically active resolving agent, a method using mandelic acid 
was disclosed in Japanese Patent Disclosure No. 119844/1978. Also, use of 
L-phenyl alanine for optical resolution of mandelic acid was reported in 
J. Chem. Soc. Jpn. 92 (11) 999 (1971). 
SUMMARY OF THE INVENTION 
The object of the present invention is to provide a commercially 
advantageous method of optical resolution of DL-cysteine using optically 
active mandelic acid, not taking the steps of substituting the amino group 
and the thiol group but directly from DL-form to the optically active 
forms of a high purity with a high yield. 
The method of the present invention is characterized in that an optically 
active mandelic acid is reacted with DL-cysteine to form a complex, and 
that the complex substance is resolved by utilizing difference in 
solubilities thereof. 
PREFERRED EMBODIMENTS OF THE INVENTION 
The first step of carrying out the present method is to dissolve free 
DL-cysteine or cysteine containing excess amount of one of the enantiomers 
thereof in a liquid medium. Also, optically active mandelic acid is 
dissolved in the same kind of liquid medium. The latter solution (mandelic 
acid solution) is gradually added to the former solution (cysteine 
solution). Then, the less soluble complex will preferentially precipitates 
or crystallizes out in the mixed solution. Separation of the solid from 
the liquid gives L-cysteine L-mandelic acid or D-cysteine D-mandelic acid. 
From these complexes substances, optically active cysteine can be obtained 
by decomposing the complex with a mineral acid, concentrating, and 
extracting the mandelic acid with an organic solvent, and optionally, 
recrystallization. 
Preferable materials for the liquid medium are, usually, water, methanol, 
ethanol, 1-propanol, 2-propanol, and mixtures thereof. 
At the crystallization, seeding is not always necessary but, in order to 
facilitate the crystallization, it is useful to add a small amount of the 
less soluble complex to the solution. 
There is no particular limit in the addition ratio of cysteine and mandelic 
acid. It is the most preferable to use mandelic acid in the amount 
equivalent to the content of the optically active cysteine from the view 
point of the resolution efficiency. 
The starting cysteine may be not only the above mentioned free cysteine, 
but also of the other forms such as hydrochloride and sulfate. In the 
latter case, the resolution can be facilitated by adding a substance, 
e.g., sodium hydroxide, which release cysteine at the time of the 
reaction. Even if the hydrochloride, sulfate or the like of cysteine 
remaines in the reaction solution, there will be no trouble in the 
resolution. 
The reaction temperature and pressure may be chosen from wide ranges. 
However, it is preferable that, in view of the relatively low stability of 
cysteine, the reaction is carried out at a room temperature or, even under 
cooling with cold water or ice water.

The present invention will now be illustrated with Examples below: 
EXAMPLE I 
17.6 g (0.10 mole) of DL-cysteine hydrochloride was dissolved in 10 ml of 
water. Also, 3.2 g (0.08 mole) of sodium hydroxide was dissolved in 20 ml 
of water, and, in the resulting solution, 7.6 g (0.05 mole) of 
L-(+)-mandelic acid was dissolved. These two solutions were mixed, and 5 
mg of crystal of less soluble L-cysteine.L-mandelic acid was seeded to the 
mixed solution, which was allowed to stand at a room temperature for 1.5 
hours. Precipitated crystal was separated by filtration, washed with 3.0 
ml of water, and then dried to give 10.01 g of crystal of 
L-cysteine.L-mandelic acid. [.alpha.].sub.435.sup.20 +195.0.degree. 
(C=1.00, N-HCl). Through recrystallization of 9.91 g of the crude crystal 
with 15 ml of water, 6.81 g of refined crystal was obtained. 
[.alpha.].sub.435.sup.21.5 +202.3.degree. (C=1.00, 2N-HCl). 
18.6 ml of 2N-HCl was added to 6.76 g of the refined crystal to completely 
dissolve the crystal by heating to 40.degree. to 50.degree. C., and then, 
the solution was cooled to 15.degree. C. for 30 minutes. Precipitated 
crystal was filtered out, and the filtrate solution was evaporated to dry. 
20 ml of isopropyl ether was added to the residue, and the admixture was 
stirred for 30 minutes. Isopropyl ether phase was separated by 
decantation, and further 10 ml of isopropyl ether was added for washing. 
Through recrystallization of the residual solid from 5 ml of 95 % ethanol, 
3.31 g of L-cysteine hydrochloride was obtained. [.alpha.].sub.D.sup.21 
+5.6.degree. (C=1.00, 2N-HCl) m.p. 179.degree. to 181.degree. C. (dec.). 
EXAMPLE II 
35.2 g (0.20 mole) of DL-cysteine hydrochloride was dissolved in 20 ml of 
water. Also, 6.4 g (0.16 mole) of sodium hydroxide was dissolved in 40 ml 
of water, and, in the resulting solution 15.2 g (0.10 mole) of 
D-(-)-mandelic acid was dissolved. These two solutions were mixed, and 
allowed to stand at a room temperature for 3.5 hours. Precipitated crystal 
was taken out by filtration, washed with 6.0 ml of water, and then dried 
to give 19.78 g of D-cysteine.D-mandelic acid. [.alpha.].sub.435.sup.21 
-197.2.degree. (C =1.00, 2N-HCl) Through recrystallization of 19.73 g of 
the crude crystal with 30 ml of water, 13.86 g of refined crystal was 
obtained. [.alpha.].sub.435.sup.18- 203.2.degree. (C=1.00, 2N-HCl). 
19.8 ml of 2.5 N-HCl was added to 13.71 g of the refined crystal to 
completely dissolve the crystal by heating to 40.degree. to 50.degree. C., 
and then, the solution was cooled to 12.degree. C. for 30 minutes. 
Precipitated crystal was filtered out, and the filtrate solution was 
evaporated to dry. 30 ml of isopropyl ether was added to the residue, and 
the admixture was heated to 40.degree. to 50.degree. C. and stirred for 30 
minutes. Isopropyl ether phase was separted by decantation, and further, 
each 15 ml of isopropyl ether was used twice for the same procedure. 
Through recrystallization of the residual solid from 8 ml of 95% ethanol, 
4.93 g of D-cysteine hydrochloride was obtained. [.alpha.].sub.D.sup.21 
-5.7.degree. (C=1.00, 2N-HCl) m.p. 179.degree. to 181.degree. C. (dec.). 
EXAMPLE III 
17.6 g (0.10 mole) of DL-cysteine hydrochloride was dissolved in 20 ml of 
water. Also, 2.8 g (0.07 mole) of sodium hydroxide was dissolved in 40 ml 
of water, and, in the resulting solution 9.12 g (0.06 mole) of 
L-(+)-mandelic acid was dissolved. These two solutions were mixed, and 5 
mg of crystal of less soluble L-cysteine.L-mandelic acid was seeded to the 
mixed solution, which was allowed to stand at a room temperature for 1.5 
hours, and then cooled with ice water for 1.5 hours. Precipitated solution 
was separated by filtration, washed with 6.0 ml of water, and then dried 
to give 6.69 g of crude crystal of L-cysteine.L-mandelic acid. 
[.alpha.].sub.435.sup.20 +202.0.degree. (C=1.00, 2N-HCl). 
2.73 g of the crude crystal was dissolved in 20 ml of 0.5 N-NaOH, and, 
after addition of 0.16 g of ferric chloride to the resulting solution, pH 
of the solution was adjusted to 7 to 8 with 1N-NaOH. Air was blown into 
thus prepared solution for 1.5 hours, and formed crystal was filtered out. 
Washing of the crystal with 5 ml of water and drying gave 0.58 g of 
L-cystine. [.alpha.].sub.D.sup.23 -135.9.degree. (C=1.00, 1N-HCl). Through 
recrystallization of 0.53 g of the crude crystal from 60 ml of water, 0.20 
g of the refined crystal was obtained. [.alpha.].sub.D.sup.22 
-212.7.degree. (C=1.00, 1N-HCl). 
EXAMPLE IV 
175.6 g (1.00 mole) of DL-cysteine hydrochloride was dissolved in 100 ml of 
water. Also, 32.0 g (0.80 mole) of sodium hydroxide was dissolved in 170 
ml of water, and, in the resulting solution 76.0 g (0.50 mole) of 
L-(+)-mandelic acid was dissolved. These two solutions were mixed, and 50 
mg of crystal of less soluble L-cysteine L-mandelic acid was seeded to the 
mixed solution, which was cooled to 17.degree. to 20.degree. C. for 1.5 
hours, and then, 7.degree. to 10.degree. C. for 2 hours. Precipitated 
crystal was separated by filtration, washed with 27 ml of water, and then 
dried to give 121.7 g of crystal of L-cysteine.L-mandelic acid. 
[.alpha.].sub.435.sup.22.5 +189.7.degree. (C=1.00, 2N-HCl). Through 
recrystallization of the crude crystal with 150 ml of water, 91.5 g of 
refined crystal was obtained. [.alpha.].sub.435.sup.22.5 +202.0.degree. 
(C=1.00, 2 N-HCl). 
86 ml of 3.8 N-HCl was added to 91.4 g of the refined crystal to completely 
dissolve the crystal by heating to about 60.degree. C., and then, the 
solution was cooled to 7.degree. to 10.degree. C. for 30 minutes. 
Precipitated crystal was filtered out, and the filtrate solution was 
subjected to extraction with isopropyl ether in a liquid-liquid extractor 
for 2 hours. Water phase was concentrated to water content of 10%. 66 ml 
of isopropyl alcohol was added to the concentrated water solution for 
recrystallization, by which 33.8 g of L-cysteine hydrochloride was 
obtained. [.alpha.].sub.D.sup.25 +5.5.degree. (C=1.00, 2N-HCl). 
Then, 29.5 ml of concentrated hydrochloric acid was added to the mother 
liquor given by separation of the crude L-cysteine.L-mandelic acid, and 
the liquor was concentrated to the volume of about 200 ml. Precipitated 
sodium chloride was filtered out, and the filtrate solution was extracted 
with isopropyl ether in the liquid-liquid extractor for 2 hours. Water 
phase was concentrated to the volume of about 100 ml, and the concentrated 
solution received 98 ml of 20% sodium hydroxide aqueous solution for 
adjusting pH of the solution to 5 to 6. Water was so added to the solution 
that the volume increased to 300 ml, and 68.4 g (0.45 mole) of 
D-(-)-mandelic acid was added to the diluted solution, which was heated to 
about 70.degree. C. for dissolution. 50 mg of less soluble 
D-cysteine.D-mandelic acid was seeded to the solution, and the solution 
was cooled to 17.degree. to 20.degree. C. for 1.5 hours, and then, 7 to 
10.degree. C. for 2 hours. Precipitated crystal was filtered out, washed 
with 25 ml of water, and then dried to give 124.3 g of crude crystal of 
D-cysteine.D-mandelic acid. [.alpha.].sub.435.sup.23.5 -162.9.degree. 
(C=1.00, 2N-HCl). 
Through adding 100 ml of water to the crude crystal, heating to 70.degree. 
C., subsequent cooling and filtration, 80.5 g of refined crystal of 
D-cysteine.D-mandelic acid was obtained. [.alpha.].sub.435 -198.9.degree. 
(C=1.00, 2N-HCl). 
The same procedures as described in connection with L-cysteine.L-mandelic 
acid were carried out using the refined crystal, and 36.0 g of D-cysteine 
hydrochloride was obtained. [.alpha.].sub.D.sup.21.5 -5.3.degree. (C=1.00, 
2N-HCl).