Resolution Separation of Enantiomers

Resolution can be thought of as the converse of racemization (Section 2.4). One starts with a 50 50 mixture of both enantiomers and separates this mixture into the individual enantiomers. Of course, for some purposes one may only want one enantiomer, and recovery of the second enantiomer can be painstaking. Since enantiomers have identical properties, including solubility, separation of enantiomers by recrystallization is quite rare. It was, however, such a crystallization by Pasteur in 1848 that opened up the field of resolution. Pasteur s key observation was that two distinct but related types of crystal were obtained from an aqueous solution of the sodium ammonium salt of racemic tartaric acid. The two types of crystal were related as object and non-superimposable mirror image, and one type was identical to the dextrorotatory crystals of sodium ammonium tartrate obtained from (+)-tartaric acid, itself obtained as a by-product of wine-making. 

Typically, resolution depends on the conversion of enantiomers, which possess identical physical properties, into diastereoisomers, which do not, and then exploiting the difference in physical properties in order to separate the diastereoisomers. Finally, the diastereoisomers are reconverted to the component, and now separated, enantiomers. 

Resolution by recrystallization depends on conversion of enantiomers into diastereoisomers, usually salts, and then exploiting solubility differences. 

Resolution of a carboxylic acid can be achieved directly by formation of diastereomeric salts with an amine. Naturally occurring bases such as brucine (34) or ephedrine (35) are often used, though it should be mentioned that choice of a suitable amine is usually a matter of trial and 

Resolution of alcohols usually involves synthesis of a phthaiate half-ester, then formation of a cfiastereomeric salt, 

So far we have left unanswered an important question about optically active compounds and racemic forms How are enantiomers separated Enantiomers have identical solubilities in ordinary solvents, and they have identical boiling points. Consequently, the conventional methods for separating organic compounds, such as crystallization and distillation, fail when applied to a racemic form. 

It was, in fact, Louis Pasteur s separation of a racemic form of a salt of tartaric acid in 1848 that led to the discovery of the phenomenon called enantiomerism. Pasteur, consequently, is often considered to be the founder of the field of stereochemistry. 

Pasteur s discovery of enantiomerism and his demonstration that the optical activity of the two forms of tartaric acid was a property of the molecules themselves led, in 1874, to the proposal of the tetrahedral structure of carbon by van t Hoff and Le Bel. 

Unfortunately, few organic compounds give chiral crystals as do the (+)- and (—)-tartaric acid salts. Few organic compounds crystallize into separate crystals (containing separate enantiomers) that are visibly chiral like the crystals of the sodium ammonium salt of tartaric acid. Pasteur s method, therefore, is not generally apphcable to the separation of enantiomers. 

One of the most useful procedures for separating enantiomers is based on the following  

The bases generally employed in such resolutions have been natural alkaloids, such as strychnine, brucine, and ephedrine. These alkaloids are more complex than the general case shown in the figure, in that they contain several chiral centres (ephedrine is shown in Section 3.4.4). Tartaric acid (see Section 3.4.5) has been used as an optically active acid to separate racemic bases. Of course, not all materials contain acidic or basic groups that would lend themselves to this type of resolution. There are ways of introducing such groups, however, and a rather neat one is shown here. 

A racemic alcohol may be converted into a racemic acid by reaction with one molar equivalent of phthalic anhydride the product is a half ester of a dicarboxylic acid (see Section 7.9.1). This can now be subjected to the resolution process for acids and, in due course, the alcohols can be regenerated by hydrolysis of the ester. 

A significant improvement on the fractional crystallization process came with the introduction of chiral 

With the appropriate choice of enzyme, it has been found that only one enantiomer of the racemic mixture is hydrolysed, whilst the other remains unreacted. It is then a simple matter to separate the unreacted ester from the alcohol. The unreacted ester may then be hydrolysed chemically, thus achieving resolution of the enantiomeric alcohols. 

---

The potential for use of chiral natural materials such as cellulose for separation of enantiomers has long been recognized, but development of efficient materials occurred relatively recently. Several acylated derivatives of cellulose are effective chiral stationary phases. Benzoate esters and aryl carbamates are particularly useful. These materials are commercially available on a silica support and imder the trademark Chiralcel. Figure 2.4 shows the resolution of y-phenyl-y-butyrolactone with the use of acetylated cellulose as the adsorbent material. 

Another means of resolution depends on the difference in rates of reaction of two enantiomers with a chiral reagent. The transition-state energies for reaction of each enantiomer with one enantiomer of a chiral reagent will be different. This is because the transition states and intermediates (f -substrate... f -reactant) and (5-substrate... R-reactant) are diastereomeric. Kinetic resolution is the term used to describe the separation of enantiomers based on different reaction rates with an enantiomerically pure reagent. 

Since the first separation of enantiomers by SMB chromatography, described in 1992 [95], the technique has been shown to be a perfect alternative for preparative chiral resolutions [10, 21, 96, 97]. Although the initial investment in the instrumentation is quite high - and often prohibitive for small companies - the savings in solvent consumption and human power, as well as the increase in productivity, result in reduced production costs [21, 94, 98]. Therefore, the technique would be specially suitable when large-scale productions (>100 g) of pure enantiomers are needed. Despite the fact that SMB can produce enantiomers at very high enantiomeric excesses, it is sometimes convenient to couple it with another separation... 

Recently, two examples of the separation of enantiomers using CCC have been published (Fig. 1-2). The complete enantiomeric separation of commercial d,l-kynurenine (2) with bovine serum albumin (BSA) as a chiral selector in an aqueous-aqueous polymer phase system was achieved within 3.5 h [128]. Moreover, the chiral resolution of 100 mg of an estrogen receptor partial agonist (7-DMO, 3) was performed using a sulfated (3-cyclodextrin [129, 130], while previous attempts with unsubstituted cyclodextrin were not successful [124]. The same authors described the partial resolution of a glucose-6-phosphatase inhibitor (4) with a Whelk-0 derivative as chiral selector (5) [129]. 

The chiral recognition ability of a CSP is quantitatively evaluated from the results of chromatographic separation of enantiomers. Figure 3.4 shows a chromatogram of the resolution of benzoin (19) on cellulose tris(3,5-dimethylphenylcarbamate). The (+)-isomer elutes first followed by the (—)-isomer complete baseline separation is achieved. The results of the separation can be expressed by three parameters—capacity factors (k1), separation factor (a), and resolution factor (Rs)—defined as follows ... 

A lot of published data on the separation of enantiomers of flavors and fragrances by GC is reviewed by Chirbase/Flavor database. Table 1. summarizes the enantiomer separation of oxygenated monoterpenes on chiral stationary phases of cyclodextrin derivatives by high resolution gas chromatography. 

Chankvetadze et al. prepared a 20-cm-long silica capillary column modified by in situ coating with amylase tris(3,5-dimethylphenylcarbamate) [199] for the fast separation of enantiomers. They showed the separation of 10 pairs of enantiomers. The monolith columns exhibit a slightly lower resolution but significant faster separation compared with a conventional 25 cm packed HPLC column. 

Optical. - This adjective exclusively refers to measurements of optical rotation. It must not be used other than in the combinations optical rotation and optical purity. In particular, the terms optical resolution (better separation of enantiomers) and optical yield must be avoided. 

Several types of CSPs are commercially available. However, many of these columns provide only adequate separation of enantiomers at best and often do not provide baseline resolution. In addition, many CSPs are expensive, costing as much as 18,000 per column. CSPs using mesoporous silica may be able to enhance the separation of enantiomers and are also produced more cheaply than commercial materials... 

Biopolymers in Chiral Chromatography. Biopolymers have had a tremendous impact on the separation of nonsupernnposable. mirror-image isomers known as enantiomers. Enantiomers have identical physical and chemical properties in an achiral environment except that they rotate the plane of polarized light in opposite directions. Thus separation of enantiomers by chromatographic techniques presents special problems. Direct chiral resolution by liquid chromatography (lc) involves diastereomenc interactions between the chiral solute and the chiral stationary phase. Because biopolymers are chiral molecules and can form diastereomeric... 

---