JPH083131A - Penicillamine derivative, chirality discriminating agent and separating agent - Google Patents

Abstract

PURPOSE:To obtain the subject new compound useful as a chirality discriminating agent, especially a separating agent for carrying out the resolution of various optical isomer mixtures such as amino acids in liquid chromatography. CONSTITUTION:This S- and N-alkyl derivative of penicillamine is represented by the formula [R1 and R2 are each an alkyl (which may contain an unsaturated bond and an aromatic group as a substituent group) or R1 is a group after removing R1 from the formula and R2 has the meaning described above or R1 and R2 together denote an alkanediyl; R3 is H or an alkyl (same as the described above); the total number of carbon atoms in R1 to R3 is >=8], e.g. N,S-dioctyl-D-penicillamine. The compound represented by the formula is obtained by introducing the two (R1 and R2) or three alkyl groups (R1, R2 and R3) into penicillamine stepwise or at a stroke, etc.

Description

Detailed Description of the Invention

[0001]

This invention relates to penicillamine S-
And N-alkyl derivatives and their uses. In the above applications, the use as a chirality discriminating agent, in particular, as a separating agent for performing resolution of an enantiomer mixture (for example, racemate) of a compound having chirality such as an amino acid in, for example, liquid chromatography. included.

[0002]

2. Description of the Related Art Separation agents for optical resolution in which a silica gel carrier for reversed phase is coated with an amino acid derivative are already known.
[(1) Chromatographia, Vol. 13, page 667 (1980), (2) JP-A-58-96062)
No. 3, (3) Japanese Patent Laid-Open No. 64-26523]. Above (1) and
The separating agent described in (2) uses an N-alkyl derivative of proline or hydroxyproline as an amino acid to be supported, and the separating agent described in (3) uses serine as an amino acid to be supported. , N, N-dialkyl derivatives of alanine or norparin are used.

[0003]

The fillers using the amino acid derivatives of the above (1) and (2) show excellent resolving ability for specific amino acids, but serine, histidine, lysine, glutamic acid, etc. It has a drawback that it shows almost no optical resolution for the hydrophilic amino acid. Also
The filler using the amino acid derivative of (3) shows optical resolution with respect to the hydrophilic amino acids described above, but the optical resolution of asparagine, cysteine, 3,4-dihydroxyphenylalanine (hereinafter abbreviated as DOPA), etc. Has the drawback of being difficult. The present invention seeks to provide a separating agent which does not have the above-mentioned drawbacks.

[0004]

That is, the present invention provides (1) a general formula

Embedded image [Wherein R ₁ and R ₂ are alkyl groups (which may include an unsaturated bond and an aromatic group as a substituent), or R ₁ is the same as the above formula (I) except R ₁ R _{2 in the} resulting group
Is as defined above, or R ₁ and R ₂ together represent an alkanediyl group, R ₃ is hydrogen or an alkyl group (including an unsaturated bond and an aromatic group as a substituent) Also means). However, the total number of carbon atoms of R ₁ , R ₂ and R ₃ is 8 or more]. (2) A chirality identifying agent comprising the compound according to (1) above. (3) To provide a separating agent obtained by coating a reverse phase carrier of the compound according to (1) above.

In the above general formula (I), the alkyl group represented by R ₁ preferably has 5 to 20 carbon atoms, and
-18 is more preferable, and 8-15 is most preferable. Further, as the alkyl group of R ₂ and R ₃ ,
Those having 1 to 20 carbon atoms are preferable, those having 5 to 19 or 7 to 18 are more preferable, and those having 8 to 15 are most preferable. These alkyl groups may be linear or may have one or more, for example 1-2, branches. Further, these alkyl groups may have one or more, for example, 1-3 unsaturated bonds (double bond or triple bond) in the main chain or side chain. Further, these alkyl groups have one or more, for example, 1-2 aromatic groups as substituents (preferably phenyl, tolyl, xylyl,
A monocyclic aromatic group such as cumenyl).

Representative examples of alkyl groups are: for R ₁ ,
Heptyl, isoheptyl, 2-ethylhexyl, octyl, isooctyl, nonyl, 2,4,5-trimethylhexyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, 3-methyl-4-ethylundecyl, gentadecyl, hexadecyl, heptadecyl, octadecyl. , Nonadecyl, icosanyl, and unsaturated forms thereof, such as 7-octenyl, 3,5-decadienyl, 6
-Dodecynyl, as well as aromatic substituents such as benzyl,
Phenyl, styryl, 2-phenylpropyl, etc., and in the case of R ₂ and R ₃ , in addition to the above, methyl, ethyl, propyl, isopropyl, butyl, pentyl, hexyl, allyl, butenyl and the like are included. The alkanediyl group formed by combining R ₁ and R ₂ preferably has ₁ to 10 carbon atoms, more preferably 2 to 5 carbon atoms, and has a side chain. Examples of representative alkanediyl groups are lower alkylene and lower alkylidene, such as methylene, ethylene, trimethylene, methylethylene, ethylidene, propylidene, isopropylidene, butylidene, pentylidene, hexylidene and the like. The total number of carbon atoms of R ₁ and R ₂ or R ₁ , R ₂ and R ₃ (when other than hydrogen) in the above compound needs to be 8 or more in order to obtain appropriate hydrophobicity. Is preferably 8 to 25, more preferably 10 to 20, and most preferably 13 to 18. When R ₁ is a group obtained by removing R ₁ from the formula (I), the compound (I)
Is in the form of a dimer via a —S—S— bond.

Examples of compounds of this invention are N, S-dioctyl-D-penicillamine, N, N-dimethyl-S-dodecyl-D-penicillamine, N, N, S-tripentyl-D.
-Penicillamine, N, N-dioctyl-di-D-penicillamine, 2,2,5,5-tetramethyl-3-octylthiazolidine-4-carboxylic acid. The above compound (I) is to be used in the form of a pure optical isomer or an isomer mixture in which one optical isomer is predominant. The above-mentioned compound (I) is, for example, penicillamine 2
(R ₁ and R ₂ ) or 3 (R ₁ , R ₂ and R ₃ ) alkyl groups are introduced stepwise (2 or 3 steps) or all at once, or R ₁ and R ₂ are alkanes. It is prepared by reacting an aldehyde or a ketone to obtain a compound forming a diyl group, or by oxidizing a corresponding -SH compound to form a dimer via an -SS- bond.

The reaction for introducing an alkyl group is carried out by a conventional method used for alkylating S and N atoms. An exemplary method is for penicillamine, preferably in the presence of a base, of the formula: R—X, where R is R ₁ , R ₂ or R _2.
3 (other than hydrogen) alkyl group, and X is a reactive ester group of an inorganic acid or an organic acid). In the above formula, the reactive ester group includes halides (eg chloride, bromide), sulfonic acid (eg methanesulfonic acid, toluenesulfonic acid) ester, tartaric acid ester and the like. As the base, inorganic bases such as sodium hydroxide and potassium hydroxide and organic bases such as trialkylamine and dimethylaniline are used. This reaction is preferably carried out in the presence of a promoter such as iodide (eg potassium iodide).
The reaction is accelerated by heating. Alternatively, the aldehyde R'- may be added under reducing conditions (for example, a metal catalyst for catalytic reduction such as platinum and palladium and hydrogen gas, a metal such as zinc and an acid).
A method of reacting CHO (wherein R ′ is an alkyl group or hydrogen atom having one less carbon atom than R above) is also possible. The introduction reaction of the alkanediyl group is carried out in the same manner as the cyclic acetalization of glycol. A typical method is a method of reacting an aldehyde or a ketone in the presence of an acid catalyst (hydrochloric acid, toluenesulfonic acid, zinc chloride, etc.). The reaction of forming an —S—S— bond is performed by a method generally used for producing a disulfide. A typical method is air oxidation using a metal ion catalyst such as iron or copper, or oxidation with an oxidizing agent (eg hydrogen peroxide, halogen, hypohalous acid, manganese oxide, etc.). The reaction product can be separated and purified by a commonly used separation means such as solvent extraction, recrystallization, chromatography and the like.

[Usage] When the compound represented by the formula (I) is brought into contact with a compound having chirality such as an amino acid, the physicochemically different qualitatively or qualitatively with respect to the two optical isomers. Since it interacts, it can be used as a chirality discrimination agent. The simplest application example in this application is separation of two optical isomers or removal of one optical isomer using a separating agent obtained by binding compound (I) to a solid. As the separating agent, a solid carrier whose surface is coated with the compound (I) is convenient. As the solid carrier, reverse phase carrier granules are preferable. As such a carrier, an inorganic type or an organic type can be used as long as it has a hydrophobic surface. Preferred are silica gel surface-treated with an alkylsilane having 3 to 18 carbon atoms, porous glass surface-treated with an alkylsilane having an alkyl group having 3 to 18 carbon atoms, and styrene-based and acrylic. Examples include hydrophobic polymer gels such as a system. The shape of the carrier may be crushed or spherical, and may be porous or non-porous.
It is preferably a spherical porous carrier having a particle size of μm. In this case, by using a porous carrier having a large surface area, the coating amount of compound (I) can be increased,
Further, it is possible to adjust the coating amount by controlling the surface area of the carrier. The pore diameter of the porous carrier is 10
~ 1000 Å, surface area 1-1000 m ²
It is preferably in the range of. As a method for coating the solid carrier with the compound (I), the method described in the literature [Chromatography (Chr
omatographia) Vol. 13, p. 667 (1980)], a method of directly introducing the alcohol aqueous solution in which the compound (I) is dissolved into a column for liquid chromatography packed with a reversed phase carrier and coating the column is convenient. Alternatively, a method of mixing a reverse phase carrier and a solution of compound (I) and distilling off the solvent under reduced pressure may be used. The support coated with compound (I) is then treated with a suitable metal ion such as Cu ²⁺ ,
It is in the form of a complex salt of Ni ²⁺ , Zn ²⁺ , Cd ²⁺ , Hg ²⁺ or Co ³⁺ and is used for optical resolution of a racemate by a liquid chromatography method. This can be done, for example, as follows. A column made of glass, stainless steel, titanium or the like having an appropriate length, for example, 1 to 100 cm, is closely packed with a separating agent, and a liquid feed pump and a sample injector are provided at the inlet side of the column, and On the outlet side, connect a detector such as an ultraviolet detector or an optical rotation detector and a fraction collector if necessary. Separation can be performed by a usual method. That is, after injecting a racemate, an eluent is sent by a liquid feed pump to develop the injected racemate in a column and divide it. Suitable eluents include aqueous solutions of metal salts such as 0.1-10 mM copper sulfate or copper acetate. In this case, an organic solvent that is miscible with water, such as methanol or acetonitrile,
When it is added to the 30% by volume eluent, favorable results are easily obtained. Generally, the retention time of the racemate tends to decrease as the amount of organic solvent added increases. In addition, the eluent
Keeping the pH constant is extremely preferable in order to obtain a chromatogram with good reproducibility. The eluate eluted from the column is introduced into a detector such as an ultraviolet detector, each divided enantiomer is detected by the absorbance at 254 nm, and the chromatogram is recorded by a recorder.

[0010]

INDUSTRIAL APPLICABILITY The separating agent of the present invention can separate various optical isomer mixtures, particularly amino acid optical isomer mixtures, by a chromatographic method. Regarding amino acids, not only proline and arosreonine, which were conventionally separable, but difficult asparagine, cysteine, DOPA
In addition to a wide range of amino acids including hydrophilic amino acids such as serine, histidine, lysine and glutamic acid, and the above metal ions such as α-oxycarboxylic acid, 2-aminoalcohol, β-aminocarboxylic acid and 1,2-diamine. Optical resolution of various racemates with coordinate bonds is possible.
Moreover, the separating agent of the present invention has the advantages that it is easy to manufacture and has excellent durability.

[0011]

EXAMPLES The present invention will be described below with reference to examples.
The examples do not limit the invention. Example 1 (A) D-Penicillamine 3.0 g (20.1 mmol), potassium iodide 3.3 g (20.1 mmol) and triethylamine 10.2 g (100.5 mmol) in chloroform 2
After adding a mixed solution of 55 ml and 45 ml of methanol, the mixture was stirred at room temperature for 12 hours to be dissolved. After adding 23.3 g (120.6 mmol) of 1-bromooctane to this,
The mixture was heated at a bath temperature of 70 to 80 ° C and refluxed for 8 hours. At this time, the temperature of the reaction solution was 55 ° C. After the reaction, cool to room temperature,
The deposited precipitate was filtered off. After concentrating the filtrate under reduced pressure,
400 ml of ethyl acetate and 300 ml of 1N hydrochloric acid were added to the residue.
Was added, and the mixture was shaken well, allowed to stand, and separated. 50 g of anhydrous magnesium sulfate was added to the ethyl acetate layer for dehydration, filtration was performed, and the filtrate was concentrated under reduced pressure. Ethyl acetate 50 for residue
The solution was added with ml to dissolve it, cooled to 5 ° C., and allowed to stand overnight. The precipitated crystals were collected by filtration and dried under reduced pressure to give 1.7 g of slightly yellow crystals of N, S-di-n-octyl-D-penicillamine. Melting point: 123-124 ° C In the infrared absorption spectrum of this slightly yellow crystal (potassium bromide mixing method and liquid paraffin mixing method), 3400 cm
Absorption near ^-1 based on NH stretching vibration of secondary amino group,
2850～2950Cm ^-1 strong absorption based on C-H stretching vibration of linear alkyl groups, and 1590 cm ^-1 absorption due to carboxylic acid was observed at, The mass number is based on the molecular ion +1 in FD mass spectrum 374 The spectrum of was observed. Considering these and the elemental analysis results, it is confirmed that the pale yellow crystal is N, S-di-n-octyl-D-penicillamine.

(B) The obtained N, S-di-n-octyl-D
-Penicillamine was coated on a reverse phase carrier by the following method. First, a stainless steel column (inner diameter 6 mm, length 150 mm) packed with 2.5 g of porous silica gel having an average particle diameter of 5 μm surface-treated with octadecylsilane,
A mixture of 50% by volume methanol and 50% by volume water was run. Then, the N, S-di-n-octyl-D-
0.06% methanol of pale yellow crystals of penicillamine 50
1 liter of a mixed solution containing 50% by volume of water and 50% by volume of water was passed through the column at a rate of 0.8 ml / min, and then 20 ml of water was passed through the column at a rate of 0.8 ml / min. Subsequently, 500 ml of a 1 mmol / l copper sulfate aqueous solution was passed at a rate of 1.0 ml per minute.

(C) The column coated as described above,
5 μl of 0.02 wt% amino acid racemic aqueous solution was injected, and 0.1-10 mmol / l of copper sulfate aqueous solution was added to 0.2
The solution was sent at a flow rate of 2.0 ml / min to develop a racemate. Table 1 shows an example of the results obtained by the resolution of racemates of various amino acids in this manner. The measurement conditions were as follows. Temperature: room temperature or 50 ° C. Eluent flow rate: 0.2-1.0 ml / min Detector: UV absorber (wavelength: 254 nm) In this table, retention time is the racemate after injection into the column. It means the time from the separation into the enantiomers until elution from the column, that is, the time that each enantiomer holds the column. The separation coefficient α, which represents the optical resolution of the separating agent, is calculated by the following equation. α = t ₂ −t ₀ / t ₁ −t ₀ t ₁ : Retention time of weaker adsorbed enantiomer t ₂ : Retention time of stronger adsorbed enantiomer t ₀ : Retention corresponding to dead volume of column The time separation coefficient shows that when α = 1, there is no optical resolution, and the larger the difference from 1, the higher the optical resolution.

[Table 1] An example of separating DL-asparagine is shown in FIG. In FIG. 1, peak number (1) is D-asparagine and peak number (2) is L-asparagine. An example of DL-lysine separation is shown in FIG. In Figure 2, peak number
(3) is L-lysine, and peak number (4) is D-lysine. Although DL-asparagine and DL-lysine are amino acids that are difficult to be resolved by conventional commercially available amino acid resolution columns, it can be seen that the columns packed with the separating agent of the present invention can be efficiently and completely resolved.

Example 2 Average particle size 5 surface-treated with octadecylsilane
First, a mixed solution of 50% by volume of methanol and 50% by volume of water was poured into a stainless steel column (inner diameter 4.6 mm, length 50 mm) packed with 0.5 g of porous silica gel having a size of μm, and then, 100 ml of a mixed solution of 0.04% methanol (50% by volume) and water (50% by volume) containing slightly yellow crystals of N, S-dioctyl-D-penicillamine obtained in Example 1 was passed through the column at a rate of 0.8 ml / min. After that, the following Example 1
The racemates of various amino acids were resolved in the same manner as in.
Table 2 shows an example of the results.

[Table 2]

Example 3 Using the columns obtained in Example 1 and Example 2, racemates of various oxyacids were separated. Table 3 shows an example of the results.

[Table 3]

Example 4 (A) 1.0 g (2.7 mmol) of N, S-dioctyl-D-penicillamine obtained in Example 1 was dissolved in a mixed solution of 30 ml of acetic acid and 5 ml of water, and 3 ml of 35% formalin aqueous solution. And 100 mg of platinum oxide were added. Hydrogen gas was blown into the system to replace the inside of the system with hydrogen gas, which was then sealed and stirred at 40 ° C. for 24 hours. This was filtered to remove platinum, the filtrate was concentrated under reduced pressure, and the residue was dried under reduced pressure to obtain 1.11 g of a slightly yellow oily substance of N-methyl-N, S-dioctyl-D-penicillamine. . This product had an infrared absorption spectrum of N,
340 observed with S-dioctyl-D-penicillamine
The absorption due to the N—H stretching vibration of the secondary amino group around 0 cm ⁻¹ almost disappears, and the strong absorption due to the C—H stretching vibration of the linear alkyl group at 2850 to 2950 cm ⁻¹ , and 15
Absorption due to carboxylic acid was observed at 90 cm ^-1 , and
In the FD mass spectrum, a spectrum with a mass number of 388 based on molecular ion + 1 was observed. Considering these and the elemental analysis results, the slightly yellow oily substance is N-methyl-
It is confirmed to be N, S-di-n-octyl-D-penicillamine. (B) Obtained N-methyl-N, S-di-n-octyl-
D-Penicillamine was coated on the reverse phase carrier by the following method. To a stainless steel column (inner diameter 4.6 mm, length 150 mm) packed with 1.5 g of porous silica gel having an average particle size of 5 μm, which was surface-treated with octadecylsilane,
First, a mixed solution of 50% by volume of methanol and 50% by volume of water was flowed. Then, 0.0 of the slightly yellow oily substance of N-methyl-N, S-dioctyl-D-penicillamine was obtained.
0.6 liter of a mixed solution of 50% by volume of 6% methanol and 50% by volume of water was passed through the column at a rate of 0.8 ml / min, and then the same procedure as in Example 1 was carried out. Split. Table 4 shows an example of the results.

[Table 4]

Example 5 Using the column obtained in Example 4, racemates of various oxyacids were separated. Table 5 shows an example of the results.

[Table 5]

[Brief description of drawings]

FIG. 1 is a graph showing an elution curve when DL-asparagine was separated using the separating agent of the present invention.

FIG. 2 is a graph showing an elution curve when DL-lysine was separated using the separating agent of the present invention.

[Explanation of Codes] 1 D-asparagine 2 L-asparagine 3 L-lysine 4 D-lysine

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Internal reference number FI Technical indication C07D 277/06 // C07C 227/34 7537-4H G01N 30/48 W C07M 7:00

Claims (3)

[Claims] 1. A compound of the general formula [Wherein R ₁ and R ₂ are alkyl groups (which may include an unsaturated bond and an aromatic group as a substituent), or R ₁ is the same as the above formula (I) except R ₁ R _{2 in the} resulting group
Is as defined above, or R ₁ and R ₂ together represent an alkanediyl group, R ₃ is hydrogen or an alkyl group (including an unsaturated bond and an aromatic group as a substituent) Also means). However, the total number of carbon atoms of R ₁ , R ₂ and R ₃ is 8 or more]. 2. A chirality identifying agent comprising the compound according to claim 1. 3. A separating agent obtained by coating the reverse phase carrier of the compound according to claim 1.