WO2013160926A1

WO2013160926A1 - Direct crystallization of enantiomers by heterogeneous stereoselective nucleaton on a membrane

Info

Publication number: WO2013160926A1
Application number: PCT/IT2013/000123
Authority: WO
Inventors: Efrem CURCIO; Enrico Drioli; Gianluca Dl PROFIO; Enrica FONTANANOVA
Original assignee: Consiglio Nazionale Delle Ricerche; Università Della Calabria
Priority date: 2012-04-26
Filing date: 2013-04-24
Publication date: 2013-10-31
Also published as: ITRM20120179A1

Abstract

The invention concerns a procedure for the separation of optical isomers or for the production of an enantiomeric excess from a racemic mixture or from a mixture having any composition of D and L enantiomers, through direct crystallization by stereoselective nucleation on membranes functionalized with chiral detectors, or prepared starting from materials having stereoselective properties, or made by means of molecular imprinting techniques, or anyhow endowed with the ability of recognizing a specific enantiomer. The membrane carries out the double function of: i) controlling the level of supersaturation of the enantiomeric solution by means of the selective transport of solvent (or, possibly, of antisolvent), taking into account that the trans-membrane transport rate depends both on the structural parameters of the membrane and on the operation conditions; and H) promoting, on the surface of the membrane itself, the heterogeneous selective nucleation of the specific enantiomer, D or L, that preferentially interacts with the same membrane.

Description

DIRECT CRYSTALLIZATION OF ENANTIOMERS BY HETEROGENEOUS STEREOSELECTIVE NUCLEATON ON A MEMBRANE

DESCRIPTION

Field of the invention

The present invention concerns a process for direct crystallization of enantiomers by heterogeneous selective crystallization on a membrane. More specifically, the invention concerns an innovative technique for separating optical isomers or for the production of an enantiomeric excess from a racemic mixture or having any composition of D- and L-enantiomers.

Background of the invention

Chemical compounds having the same structural formula are called isomers. The isomers are divided into two main classes: constitutional isomers, which differ in the way the atoms are bonded together, and stereoisomers, which differ only in their three-dimensional arrangement in space. The latter class includes the enantiomers (or optical isomers), i.e. molecules whose mirror images are not superimposable (chiral molecules). Two enanti- omers have substantially identical chemical and physical properties, but different interactions with other chiral molecules; in addition, they possess an optical activity, as they can rotate the plane of polarized light to an equal extent but in the opposite direction. The enantiomer that rotates the plane of polarized light counter-clockwise is said counterclockwise or levorotatory (in- dicated by the letter "L" or the "-" sign) and, conversely the enantiomer that rotates the plane of polarized light in a clockwise direction is called clockwise or dextrorotatory (indicated by the letter "D" or by the sign "+"). A mixture consisting of a pair of enantiomers present in equimolar amounts is called racemic or racemate.

Although two enantiomers exhibit chemical and physical properties substantially identical (and, therefore, they are difficult to separate), it is shown that organisms react differently to the administration of chiral drugs. Since the discovery of stereoisomerism in biochemical systems, made by Pasteur in 1860, the purification of enantiomeric mixtures has played a primary role in the pharmaceutical industry. In general only one of the two enantiomers is active (for example, the levorotatory enantiomer of tetramisole - also called levamisole - is an anthelmintic having broad spectrum of action, while its dextrorotatory form is inactive). Therefore, the excess of one enantiomeric form compared to the racemate increases the effectiveness of the drug (for example, escitalopram, dextrorotatory enantiomer used as antidepressant as being a selective serotonin reuptake inhibitor, shows an activity twice as high compared to the racemic form citalopram). In other cases, even the mere presence of one of the two enantiomers may result in serious side effects (as in the case of thalidomide, a sedative that - marketed in the fifties and sixties in the racemic form - turned out to be highly teratogenic when taken during pregnancy).

Asymmetric synthesis and separation of enantiomers from racemic mixtures represent the methods used for the preparation of pure enantiomers. The processes for the resolution of racemic mixtures are various; among them, the most common ones are cited below:

1. chromatographic separation techniques, such as traditional high perfor- mance liquid chromatography (HPLC) and gas chromatography (GC) or the more recent capillary electro-chromatography, which, while guaranteeing high chiral discrimination, are expensive and not currently applicable on a large scale;

2. preferential crystallization of conglomerates (obtained by seeding an en- antiomer which acts as seed for nucleation), which is effective but limited to conglomerates only - the latter being mixtures of crystals of pure enantiomers - which occur with a frequency of less than 10% of the cases;

3. the traditional conversion of enantiomers in diastereoisomeric salts and subsequent separation by means of physical methods: in exchange for an easier separation, diastereoisomeric salts often have a lower pharmacological activity;

4. enzyme-mediated stereoselective kinetic resolution in the event that the enantiomers present a different reaction rate. The method is expensive, complex, and applicable in a limited number of cases.

The scientific and patent literature also includes applications of membrane processes for the purification of enantiomers. The U.S. patent No. 6485650 (A.S. Bhown) describes a method for the separation of enantiomers using supported liquid membranes (SLM): the membrane as such has not enantioselective properties, but its porous structure is used to immobilize in the liquid phase a chiral carrier capable of selectively complexing a specific enantiomer and transport it through the membrane itself. The enantiomer is then recovered from a sweep fluid.

The U.S. patent No. 5080795 (W.H. Pirkle et al.) differs from the previously described method in a technical variant, which allows to control the temperature of the two phases placed in contact with the membrane. A number of possible carriers are identified in the literature: dialkyl tartrates, diben- zoyl tartaric acid, polylactic acid etc.. Such methods, however, have found little application at industrial level because of the insufficient stability of the SLMs.

The publication by J.H. Kim et al. (JH et al., Optical resolution of alpha- amino acids through enantioselective polymeric membranes based on poly- saccharides, Journal of Membrane Science, (2003) 213, 273-283) describes the preparation of enantioselective membranes from sodium alginate and chitosan and their application to the optical resolution of alpha-amino acids. The filtration tests on a solution of racemic tryptophan allowed to obtain an enantiomeric excess of approximately 98%.

The publication by M. Yoshikawa et al. (Yoshikawa M. et al., Molecular- ly imprinted polymeric membranes for optical resolution, Journal of Membrane Science, (1995) 108, 171-175) describes the preparation of molecularly imprinted polymer membranes starting from polystyrene resin and using tetrapeptide derivatives as mold molecules. Tests on racemic tryptophan solu- tions showed a higher permeation rate of the L- enantiomer through the membrane.

It is noteworthy that in all the cases previously discussed, presented for illustrative purposes only of a vast literature, the use of the membrane is aimed exclusively at preferentially transporting in liquid phase one of the two enantiomers through the membrane itself, and not to crystallization.

The scientific article by A. Swang-Ariyaksul et al. (Swang-Ariyaksul A. et al., Chiral separation using a novel combination of cooling crystallization and a membrane barrier: Resolution of DL-glutamic acid, Chemical Engineering Science, (2009) 64, 1980-1984) refers to a chiral separation process that combines crystallization by cooling and seeding, and membrane separation. Specifically, the membrane is used to prevent the crystals from passing from one compartment to the other; said membrane, however, is permeable to both enantiomers, which are transported under a concentration gradient from one compartment to the other and in the opposite direction. For example, if in the right compartment crystals of the L-enantiomer are produced by seeding and in the left one crystals of the D-enantiomer are produced, the L-enantiomer that is in solution passes through the membrane from the left (higher concentration of L) to the right (lower concentration of L) compartment, while the enantiomer D moves in the opposite direction. Ultimately, in this case, the membrane is used to transport the enantiomers but has no role in the enanti- oselective crystallization, which is instead promoted by seeding and cooling.

Summary of the invention

In the light of this prior art, the present invention has been aimed at using membranes having stereoselective properties (intrinsic or induced by the functionalization of the membrane with chiral detectors) capable of recognizing a specific enantiomer with the purpose of promoting the stereo- specific heterogeneous nucleation on the surface of the membrane itself.

Unlike the solutions described in the prior art cited above, the proposed technical solution according to the present invention: i) is aimed at achieving the direct crystallization of a specific enantiomer; ii) does not implement a selective transport of the enantiomers through the membrane (the selective transport concerns the solvent or, where appropriate, the anti-solvent); iii) uses the membrane both to control the degree of supersaturation of the enan- tiomeric solution and to promote heterogeneous stereoselective nucleation.

It has indeed been observed, in studies conducted in relation to the present invention, that it is possible to produce an enantiomeric excess using an innovative direct crystallization process via stereoselective heterogeneous nucleation on a membrane which has stereoselective properties, in that it is capable of recognizing and preferentially interacting with a given enantiomer.

According to the process proposed there is obtained, therefore, a direct crystallization by stereoselective nucleation on membranes functionalized with chiral selectors, or prepared from materials having stereoselective properties, or made with molecular imprinting techniques, and which exhibit the ability to recognize a specific enantiomer. In the process according to the invention the membrane is used for the following purposes:

i. controlling the degree of supersaturation of the enantiomeric solution through selective transport of solvent (or, possibly, of anti-solvent), bearing in mind that the rate of trans-membrane transport depends both on the structural parameters of the membrane (porosity, tortuosity, pore diameter, thickness etc..) and on the operating conditions;

ii. promoting, on the membrane surface, the heterogeneous stereoselective nucleation of the specific enantiomer that interacts preferen- tially with the membrane itself.

Detailed description of the invention

The present invention specifically provides a process for separating enantiomers or producing an enatiomeric excess by direct crystallization on a membrane, comprising the following operations:

(a) placing a solution of enantiomers containing at least a D enantiomer and an L-enantiomer (a racemic mixture or a mixture having any composition) of an optically active compound in one or more solvents in contact with a membrane, on one of the two sides of said membrane;

(b) placing an extracting solution consisting of a pure liquid, or of a mixture of liquids or of a solution of one or more solids in one or more liquids, in contact with said membrane on the opposed side with respect to said solution of enantiomers;

(c) obtaining a selective transport of solvent from the solution of enantiomers to the extracting solution; and

(d) at the same time, obtaining the formation of crystalline nuclei of a specific one of said D- and L-enantiomers on the surface or in the pores or in any other point of the structure of said membrane, according to a heterogeneous stereoselective nucleation mechanism, and recovering the crystals formed from the membrane;

wherein said membrane:

i) is such as to allow a selective transport of solvent through it; and

ii) has stereoselective properties , i.e. it is capable of recognizing and interacting preferentially with one of said D- and L-enantiomers.

For the purposes of the present description, the term "membrane" refers to any conventional membrane or selective barrier that, interposed between two phases, allows the specific transport of one or more components from one phase to the other ("trans-membrane transport"). The above steps can occur in any state of aggregation: solid, liquid or gas, or in any combination thereof. The membrane may be porous, with an average size of the pores ranging between 0.1 nm and 50 μητι, or dense, i.e. with pores smaller than 0.1 nm.

The membrane may be composed of organic or inorganic material, pol- ymeric material or a combination thereof in hybrid or composite structures. It may be produced in any possible form; typical shapes are flat membranes, tubular membranes, hollow fiber and capillary fiber membranes, membranes in the form of spiral wound sheets, etc.. The pores of the membrane can have any shape, such as - for example - round shape, slit shape, hexagonal shape, irregular shape, spongy shape or other. The membrane may be hydrophilic or hydrophobic, or it may show hydrophilic areas alternate with hydrophobic areas. These classes of membranes can be applied in the process according to the invention in dependence on the nature of the enantiomeric solution treated.

The term "functionalized" refers to a membrane in the structure of which functional groups or molecules have been grafted, by suitable chemical or physical processes, in order to promote specific interactions between the membrane and one or more components of the solution.

In the case of the present invention the following are of specific interest:

A. functionalized membranes with specific molecular recognizers as chi- ral macrocycles, linear or cyclic oligosaccharides, any other com- pound suitable to give rise to host-guest inclusion complexes of a natural or synthetic origin, or suitable to preferentially recognize a specific enantiomer;

B. membranes obtained starting from chiral, pro-chiral, syndiotactic, iso- tactic, atactic, or eutactic polymers having stereoselective properties; C. membranes obtained with molecular imprinting techniques, which result in a preferential interaction with a specific enantiomer.

In the process dealt with by the present invention, the membrane can be inserted in a "module", where this term refers to a unit able to contain one or more membranes positioned between one or more inlet fittings and one or more outlet fittings. The module can be reinforced with a material such as ceramics, metals, polymers, etc.. Optionally, the module may contain a sequential succession of multiple membranes arranged in a concentric configuration.

For the purposes of the present description, the term "enantiomeric solution" (or "crystallization solution"), is generally intended to mean a mixture in the liquid phase of at least one pair of enantiomers and a solvent, the latter being a fluid or a combination of multiple fluids in which the enantiomers have a solubility different from zero.

Further, the term "antisolvent" refers to a fluid or a combination of more fluids that differ in chemical composition from the solvent and that, when mixed with the crystallization solution in any proportion, reduces the solubility of the substance of interest. The term antisolvent may refer to a liquid, a su- percritical fluid, a liquefied and compressed gas, a dense vapor or a mixture of two or more of them. Preferred antisolvents include alcohols, ketones, carbox- ylic acids, esters, ethers, alkanes, water, amines, quaternary ammonium salts, ionic liquids, gaseous carbon dioxide and supercritical liquids. Examples of particularly preferred antisolvents include water, methanol, ethanol, hexane, pentane, polyethylene glycol, ionic liquids that comprise EDTA (metal) complexes.

The transport of matter through the membrane is promoted by a driving force which is represented, according to the most general meaning, by an electrochemical potential gradient, or, according to more specific meanings, by a difference in activity, or in temperature, or in partial pressure, or in hydrostatic pressure, or in electric potential between the two sides of the membrane, or by any combination of these parameters.

The rate of the trans-membrane transport of matter depends on both the structural parameters of the membrane (porosity, tortuosity, pore diameter, thickness, etc..) and the operating conditions: this allows to control the degree of supersaturation of the enantiomeric solution

The term "supersaturated" or "in oversaturation" refers to the solution in which the component that is intended to crystallize is present, in the operating conditions of temperature and pressure, in a concentration higher than its solubility value. The "degree of supersaturation" is defined as the ratio of the concentration of the component that it is intended to crystallize to its solubility value of at the temperature and at the pressure of the system.

The term "heterogeneous nucleation" refers, for the purposes of the present description, to the meaning assigned such term by the Classical Nucleation Theory. It is also referred to as "stereoselective" any chemical or physical interaction, or any process (including heterogeneous nucleation) that preferentially involves a specific enantiomer. The nucleation, i.e., the process of formation of crystal nuclei, is termed stereoselective (or stereospecific) if it is directed towards the formation of a specific enantiomer; it is also termed "heterogeneous" because it takes place on the surface, in the pores or in any other point of the membrane, or else because it is promoted by interactions, of any type, between the membrane and one of the enantiomers present in the crystallization solution.

According to the present invention, the direct crystallization of a specific enantiomer is meant to be promoted by the membrane since:

1. the membrane promotes the selective transport of solvent (or, possibly, of antisolvent) between the enantiomeric solution and the extracting solution - as noted above, the rate of the trans-membrane transport of matter, which depends on both the structural parameters of the membrane (porosity, tortuosity, pore diameter, thickness etc..) and the oper- ating conditions, allows to control the degree of supersaturation of the enantiomeric solution;

2. the membrane, thanks to its stereoselective properties, favors the formation of crystal nuclei of a specific enantiomer on its surface, according to a mechanism of heterogeneous stereoselective nucleation.

According to the proposed process, a membrane having the characteristics mentioned above is placed in contact with the enantiomeric solution from one side. In a typical application the membrane is preferably hydrophobic (i. e., it prevents the passage in liquid phase of both the solvent and the enantiomers dissolved in it) and is functionalized with specific molecular detectors such as chiral macrocycles, or linear or cyclic oligosaccharides, or any other compound that can give rise to host-guest inclusion complexes, or that has the ability to recognize a given enantiomer preferentially. The membrane is placed in contact, on the opposite side with respect to the enantiomeric solution, with a solution called "extractant" preferentially formed by an aqueous salt solution. To this purpose solutions of a polyelectrolyte may also be used (such as, for example, polyvinyl alcohol, polyethylene glycol, etc.), or solutions of any compound that confers on the solution an activity lower than that of the enantiomeric solution (being the activity defined, in agreement with the usual notions of thermodynamics, as the multiplication product of the concen- tration by the activity coefficient).

The activity gradient drives the (volatile) solvent to evaporate through the membrane and condense on the side of the extracting solution; the enan- tiomers, having practically no volatility, do not cross the membrane. The rate of the trans-membrane transport of matter depends both on the structural parameters of the membrane (porosity, tortuosity, pore diameter, thickness, etc.), and on the operating conditions: this fact allows to control the degree of supersaturation of the enantiomeric solution.

The presence on the membrane surface of detectors having specific molecular selectivity towards one of the two enantiomers (dextro- or levo- rotatory) determines in the supersaturated solution of enantiomers the preferential formation of nuclei of the enantiomer recognized (heterogeneous ste- reoselective nucleation). The stage of growth of the crystalline phase leads, with all experimental evidence, to the formation of a solid product (and, obviously, to a mother liquor) which has an enantiomeric excess.

Possible variations with respect to the above-described method include the application of vacuum, a gas or a gas mixture on the extractant side (par- tial pressure gradient), or the application of a temperature difference between the two sides of the membrane, or the use of an antisolvent which diffuses from the extractant side towards the enantiomeric solution.

In particular, according to a specific embodiment of the invention, said extracting solution is constituted by one or more antisolvents, whose selective transport through the membrane determines the degree of supersaturation of the enantiomeric solution. According to a further specific embodiment of the invention, also the enantiomeric solution can contain one or more antisolvents, the concentration of which increases with the progressive diffusion of the solvent through the membrane to the extracting solution, thus determining the degree of supersaturation of the enantiomeric solution. According to other specific embodiments of the proposed method, the extracting solution is instead constituted by a gas, or a gas mixture of any composition, or by vacuum.

More generally, according to the process of the invention, the removal of solvent or the addition of antisolvent can be promoted by a difference in activity between the two sides of said membrane, or by a temperature difference, by a partial pressure difference, by a hydrostatic pressure difference, an electric potential difference or by any combination of these parameters.

Other possible variants with respect to the method illustrated above include the use of hydrophilic membranes, in which the transmembrane transport of solvent or antisolvent in the liquid phase is promoted by a hydro- static pressure gradient or by an electric potential difference acting on the solutions on the two sides of the membrane.

In the process according to the invention, the contact between said enantiomeric solution and the membrane can be achieved in a static mode, or by flowing one or both said solutions in contact with the membrane tangential- ly to the surface of the membrane itself, in laminar or turbulent regimen.

Advantageously, the processes proposed according the invention may be realized in more stages, reaching the progressive purification of a given enantiomer. In this case the operations (a)-(d) are repeated one or more times, in stages, to obtain a progressive purification, the process comprising one or both the following series of operations:

I. repeating one or more times the crystals recovery, by dissolving them again in said one or more solvents and obtaining a recrys- tallization of the same through the repetition of said operations (a)-(d);

II. repeating one or more times said operations (a)-(d) on the enantiomeric solution after separation of the crystals obtained.

Finally, according to other specific embodiments of the procedure of the invention, the crystalline product enriched in the desired enantiomer can also be recovered in a compartment different from the module containing the membrane.

Considering the proposed process according to the invention, it is noteworthy that the transmembrane flow of solvent or antisolvent depends on the structural and morphological parameters of the membrane, on the physi- cochemical characteristics of the solvent/extractant/antisolvent employed, on the difference in chemical potential between the sides of the membrane. The ability to modulate these parameters gives, in an exclusive manner with respect to conventional enantioselective crystallization techniques, a greater operational flexibility and the opportunity to be able to accurately control the degree and rate of generation of the supersaturation.

A further advantage is determined by the fact that the membrane promotes, on its surface, the stereoselective heterogeneous nucleation of the specific enantiomer that preferentially interacts with it. The kinetics of nucleation depends on certain physicochemical parameters of the membrane, such as, in particular, porosity, tortuosity, surface energy; therefore, nucleation kinetics can be controlled in an exclusive manner with respect to conventional enantioselective crystallization techniques, by modulating the properties of the membrane in the manufacturing step.

In a comparison of the present invention with the techniques of membrane filtration of racemic mixtures, it is to be noted that - while the efficiency of the latter decreases over time due to the progressive occupation of the chiral sites - the efficiency of the present crystallization technique is stable over time, as the enantiopure nuclei formed to the membrane surface can act as stereoselective nucleation seeds for the molecules in solution.

Further, the modularity feature and the wide range of types of membranes (flat, tubular, hollow fiber, etc.) allow a much greater operational flexibility than conventional techniques currently in use.

Some embodiments of the method according to the invention, as well as some experimental data concerning the characteristics of the products obtained and the comparison with the prior art, are given by way of example in the following.

EXAMPLE 1

Enantioselective direct crystallization of DL-mandelic acid on polymeric membranes functionalized with con O-octyloxycarbonyl- -cyclodextrin

The enantiomeric solution, having racemic in composition, is obtained by dissolving 2.5 g of DL-mandelic acid in 10 mL of ultrapure water; the resulting solution (hereinafter referred to as "racemic solution") is stirred for 30 minutes at 20°C.

47.6 g of anhydrous magnesium sulphate are dissolved in ultrapure water added to reach a volume of 100 mL; the resulting solution (hereinafter referred to as "the extracting solution"), stirred for 30 minutes at 20°C, has a 0.5 M concentration.

Crystallization tests were carried out using the following polymer membranes:

1. dense polydimethylsiloxane membrane loaded with O-octyloxycarbonyl- β-cyclodextrin in a percentage of 16.97% with respect to the total mass of the membrane itself. The membrane, obtained by phase inversion technique by evaporation of the solvent (dichloromethane), is hereinafter referred to as "M1";

2. low porosity membrane (11.2%) of poly(vinylidene fluoride-co-hexafluo- ropropylene) loaded with O-octyloxycarbonyl-p-cyclodextrin in a percentage of 12.8% with respect of to the total mass of the membrane itself. The membrane, obtained by phase inversion technique (solvent: Ν,Ν-dimethylacetamide; non-solvent: water), is hereinafter referred to as "M2";

3. high porosity membrane (77%) of polyvinylidene fluoride loaded with O- octyloxycarbonyl-p-cyclodextrin in a percentage of 20.0% with respect to the total mass of the membrane itself. The membrane, obtained by phase inversion technique induced by a non-solvent (solvent: N,N- dimethylacetamide; non-solvent: water), is hereinafter referred to as "M3";

4. High porosity membrane (80.5%) poly(vinylidene fluoride-co-hexafluo- ropropylene) loaded with O-octyloxycarbonyl-p-cyclodextrin in a percentage of 12.8% with respect to the total mass of the membrane itself. The membrane, obtained by phase inversion technique induced by non-solvent (solvent: N, N-dimethylacetamide; non-solvent: water), is hereinafter referred to as "M4";

The crystallization experiments were carried out in plates with wells of 20 mm diameter. On each membrane 400 L of racemic solution have been placed, while the other side of the membrane was placed in contact with 5 mL of extracting solution. The plates were kept for 24 hours in a refrigerated incu- bator at 12°C. At the end of the test, as a preliminary step the solvent has been removed by vacuum evaporation from the crystals adhering to the surface of each membrane, and then the crystals were dissolved in 1 mL of water.

The solution thus obtained was analyzed by liquid chromatography

HPLC [Agilent 1100 series HPLC System - USA equipped with Astec CHIROBIOTIC™ T-USA 250 mm x 4.6 mm (ID) column, eluent: 1% TEAA/ methanol (80/20 v/v ) pH = 4.1 mL/min]. The results showed that the process of heterogeneous nucleation on membranes functionalized with molecular stereoselective detectors gave rise to the formation of an excess of crystals in L-form (eel_, %). The results obtained are reported in the following Table 1.

Table 1

Percentage of enantiomeric excess (ee of the L-form of mandelic acid from experiments of direct crystallization of racemic solution on polymeric membranes functionalized with O-octyloxycarbonyl-p-cyclodextrin

EXAMPLE 2

Enantioselective direct crystallization of DL-glutamic acid on polymeric membranes functionalized with O-octyloxycarbonyl-P-cyclodextrin The enantiomeric solution, in racemic composition, is obtained by dissolving 0.22 g of DL-glutamic acid in 10 mL of ultrapure water, the resulting solution (hereinafter referred to as "racemic solution") is put under stirring for 30 minutes at 20°C .

47.6 g of anhydrous magnesium sulphate are dissolved in ultrapure water added to reach a volume of 100 mL, the resulting solution (hereinafter referred to as "the extracting solution"), stirred for 30 minutes at 20°C, has a concentration of 0.5 M.

The crystallization tests were carried out using of the polymer membranes M1 , M2, M3 and M4 described in Example 1.

The crystallization experiments were carried out in plates with wells of 20 mm diameter. On each membrane 400 L of racemic solution have been placed, while the other side of the membrane was placed in contact with 5 mL of extracting solution. The plates were kept for 24 hours in a refrigerated incubator at 12°C. At the end of the test, as a preliminary step, the solvent has been removed by vacuum evaporation from the crystals adhering to the sur- face of each membrane, and then the crystals were dissolved in 1 mL of water.

The solution thus obtained was analyzed by liquid chromatography HPLC [Agilent 1100 series HPLC System - USA, equipped with Astec CHIROBIOTIC™ T-USA 250 mm x 4.6 mm (ID) column, eluent: metha- no l/water/formic acid (80/20/0.02 v/v), 1 ml_/min]. The results showed that the process of heterogeneous nucleation on membranes functionalized with molecular stereoselective detectors gave rise to the formation of an excess of crystals of the L-form (ee_L,%). The results obtained are reported in the following Table 2.

Table 2

Percentage of enantiomeric excess (ee of the L-form of glutamic acid from experiments of direct crystallization of racemic solution on polymeric membranes functionalized with O-octyloxycarbonyl-p-cyclodextrin

The present invention has been disclosed with particular reference to some specific embodiments thereof, but it should be understood that modifica- tions and changes may be made by the persons skilled in the art without departing from the scope of the invention as defined in the appended claims.

Claims

1. A process for separating enantiomers or producing an entantiomeric excess by direct crystallization on a membrane, comprising the following op- erations:

(a) placing a solution of enantiomers containing at least a D- enantio- mer and an L-enantiomer of an optically active compound in one or more solvents in contact with a membrane, on one of the two sides of said membrane;

(c) obtaining a selective transport of solvent from the solution of enanti- omers to the extracting solution; and

(d) at the same time, obtaining the formation of crystalline nuclei of a specific one of said D- and L-enantiomers on the surface or in the pores of said membrane, or in any other point of the structure of said membrane, according to a heterogeneous stereoselective nu- cleation mechanism, and recovering the crystals formed from the membrane;

wherein said membrane:

i) is such as to allow a selective transport of solvent through it; and

ii) has stereoselective properties , i.e. it is capable of recognizing and interacting preferentially with one of said D and L enantiomers.

2. A process according to claim 1 , wherein said membrane is a membrane functionalized with specific molecular recognizers selected from the group consisting of: chiral macrocycles, linear or cyclic oligosaccharides, compounds suitable to give rise to host-guest inclusion complexes of a natural or synthetic origin, compounds suitable to preferentially recognize a specific one of said D- or L-enantiomers.

3. A process according to claim 1 , wherein said membrane is a membrane obtained starting from chiral, pro-chiral, or syndiotactic, isotactic, atactic or eutactic polymers having stereoselective properties.

4. A process according to claim 1 , wherein said membrane is a membrane obtained with molecular imprinting techniques, which result in a preferential interaction with a specific one of said D- or L-enantiomers.

5. A process according to any one of claims 2-4, wherein in the extracting solution and/or in the enantiomers solution one or more antisolvents are present, the selective transport of which through said membrane controls the oversaturation degree of the enantiomers solution.

6. A process according to any one of claims 2-4, wherein said extracting solution consists of a gas, or a gas mixture of any composition, or is replaced by vacuum.

7. A process according to any one of claims 2-4, wherein the removal of solvent or the addition of antisolvent is promoted by a difference of activity between the two sides of said membrane, or by a temperature difference, a difference in partial pressure, in hydrostatic pressure, in electric potential or by any combination of said parameters.

8. A process according to claim 7, wherein the contact between said solution of enantiomers and the membrane is obtained statically, or by having one or both solutions tangentially flow in contact with the membrane surface, in laminar or turbulent flow.

9. A process according to claim 7, wherein said operations (a)-(d) are repeated more than once, in stages, to obtain a progressive purification of a given enantiomer, the process comprising one or both the following series of operations:

I. repeating one or more times the recovery of the crystals, by dissolving them again in said one or more solvents and obtaining a recrystallization of the same through repetition of said operations

(a)-(d);

II. repeating one or more times the said operations (a)-(d) on the enantiomers solution after separation from it of the crystals obtained.

10. A process according to claims 8 or 9, wherein the crystalline product is recovered in a compartment different from the module containing said membrane.