S -Cyanohydrins

For the selective generation of (S)-cyanohydrins the enzymes from S. bicolor, M. esculenta and H. brasiliensis are available for use. However the enzyme from S. bicolor (from the seedlings), which was first isolated by Bove and Conn in 1960 [66],has two restrictions. First, after a six-step purification from 10 kg of S. bicolor plants only a limited quantity (160 mg, with a specific activity of approximately 260 U/mg) of pure enzyme in three isomeric forms is obtained [13,14,30, 31, 67] which represents a time-consuming procedure. Second, the substrate specificity of this enzyme is high, as it accepts only aromatic aldehydes as sub- 

Thus the recent developments made for the (S)-Hnls for M. esculenta and H. brasiliensis are of significant value and have widened the scope of applications in the area of (S)-cyanohydrin production. 

Following the initial isolation of the Hnl from M. esculenta [33] in which the peptide sequence was established, an overexpressed version of this enzyme (in E. coli) was prepared [41]. This system is not limited for enzyme quantity (as outlined in Sect. 2.3), and can accept a wide range of aromatic, heterocyclic and aliphatic aldehydes, as well as ketones, as substrates. In practical terms, a measure of the degree of enzyme inhibition by substrates is of significant value and for this system this has been quantified for a range of aldehydes, ketones and alcohols [70]. It was deduced that ketones and alcohols are competitive inhibitors, whilst aldehydes are noncompetitive inhibitors. 

Similar progress has also been made for the (S)-Hnl from H. brasiliensis. The purified protein has been overexpressed in a number of systems [39,44] (see Sect. 2.3), which adequately fulfills the practical demands required for transformations using this system. 

The substrate range of this Hnl is also broad and to date a wide range of aldehydes of varying structural nature [71] and chain length [72,73] can be easily transformed. 

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Recently, the enantioselective addition of hydrocyanic acid to aldehydes, analogous to the synthesis of (/ )-cyanohydrins, yielding (.S)-cyanohydrins in very high optical purity, with (S )-oxynitrilase as catalyst, was reported20,21. 

The enantiomeric excess values of the (S)-cyanohydrins are obtained from the ( + )-(R)-Mosher ester derivatives [a-methoxy-a-(trifluoromethyl)phenylacetates], whereas the corresponding benzeneacetic acids are first converted into their isopropyl carboxylates which then yield the ( + )-(ft)-Mosher ester derivatives. 

The enzyme-catalyzed stereoselective synthesis of (/ )- and (S )-cyanohydrins allows a simple access to compounds which can be easily transformed into the corresponding a-hydroxy-car-boxylic acids (see Table 2)20,21,23, a-hydroxyaldehydes26 or acyloins27, without racemization. 

Table 2. (S)-Cyanohydrins by Enzymatic Formation from Aromatic Aldehydes and Hydrocyanic Acid as well as (.S )-a-1 lydroxycarboxylic Acids by Hydrolysis20...

R)- and (S )-cyanohydrins are, respectively, hydrolyzed in hydrochloric acid to give the corresponding (R)- and (5 )-2-hydroxy acids in excellent yields and with complete retention of configuration. Under milder reaction conditions (lower temperature, shorter reaction times), the corresponding 2-hydroxy acid amides can be obtained selectively (Scheme A) ... 

Forster, S., Roos, J., Effenberger, F. et al. (1996) The first recombinant hydroxynitrile lyase and its application in the synthesis of (S)-cyanohydrins. Angewandte Chemie (International Edition in English), 35, 457 459. 

Rhodococcus erythropolis NCIMB 11540 has been employed as biocatalyst for the conversion of (R)- or (.S )-cyanohydrins to the corresponding (R)- or (S)-a-hydroxycarboxylic acids with an optical purity of up to >99% enatiomeric excess (ee) [27-29] the chiral cyanohydrins can separately be produced using hydroxynitrile lyase from Hevea braziliensis or from Prunus anygdalis [30]. Using the combined NHase-amidase enzyme system of the Rhodococcus erythropolis NCIMB 11 540, the chiral cyanohydrins were first hydrolyzed to the... 

It is of some historical interest that Kiliani s cyanohydrin synthesis (24) enabled Emil Fischer (25) to carry out the first asymmetric synthesis. Lapworth (26) used this base-catalyzed nucleophilic 1,2-addition reaction in one of the first studies of a reaction mechanism. Bredig (27,28) appears to have been the first to use quinine (29) in this reaction as the chiral basic catalyst. More recently, others (20) have used basic polymers to catalyze the addition of cyanide to aldehydes. The structure of quinine has been known since 1908 (30). Yet it is of critical importance that Prelog s seminal work on the mechanism of this asymmetric transformation (eq. [4]) could not have begun (16) until the configuration of quinine was established in 1944 (31,32). 

At present, these latter two Hnls represent the most superior catalysts for the production of (S)-cyanohydrins and it is expected that their development will rapidly continue to yield processes of industrial and technical relevance. 

One of the earliest examples in this field [94] was reported by Mao and Anderson who showed that the SbHnl was specific for the dehydrocyanation of p-hydroxybenzaldehyde cyanohydrin. In a recent patent [95] the process of dehydrocyanation was successfully demonstrated using the PaHnl as the catalyst and a gas-membrane extraction method to remove the undesired aldehydes and HCN, which yielded a wide range of (S)-cyanohydrins, often with excellent enantiomeric excess. In further patent literature, Niedermeyer [96] also generated (S)-cyanohydrins via decomposition of a racemic mixture, but the problems of workup resulted in decreased yields. 

Scheme 5. Selective (S)-cyanohydrin formation by enantioselective decomposition of a racemic mixture...

Undoubtedly, the cyanohydrin formation reactions catalysed by the Me- and HbHnls remain the preferred method for (S)-cyanohydrin formation. 

Recently the Merck research laboratories demonstrated the synthesis of both (R) and (S)-cyanohydrins (59, 60) with >93% through hydrocyanation of 3-pyridinecarboxyaldehyde (58) in >65% yield using dichloromethane (Scheme 7.23)... 

S)-Cyanohydrins Paa 99 71c Pal-Ini adsorbed on Auricel, disopropyl ether/acetate buffer (pH 3.3), HCN, room temperature... 

F. Effenberger, B. Hoersch, S. Foerster, andT. Ziegler, Enzyme-catalyzed reactions. 5. Enzyme-catalyzed synthesis of (S)-cyanohydrins and subsequent hydrolysis to (S)-a-hydroxy-carboxylic adds, Tetrahedron Lett. 1990, 31, 1249-1252. 

F. Effenberger, and K. Pfizenmaier, Enantioselective synthesis of aliphatic (S)-cyanohydrins in organic solvents using hydroxynitrile lyase from Manihot esculenta, Ann. N. Y. Acad. Sci. 1996, 799, 771-776. 

In an earlier investigation by the author, (S)-cyanohydrins were prepared using (S)-oxynitrilase obtained freeze-dried manioc leaves from Columbia (4). 

Scheme 2.6 HbHNL encapsulated in an aqua gel catalyzes the enantioselective formation of S-cyanohydrins.

Recently, it has been demonstrated that the enzymatic synthesis of (S)-cyanohydrins was possible using an oxynitrilase isolated from Sorghum biocolor (95,96), These optically active cyanohydrins can be subsequently converted chemically to chiral ot-hydroxyadds, aminoalcohols, and acyloins. 

Asymmetric synthesis by means of a cyandiydrin is an imprvtant process in organic synthesis, because the cyanohydrin can be easily converted into a variety of valuable synthetic intermediates, such as a-hy-droxy ketones, a-hydroxy acids, y-diketones, p-amino alcohols, 4-oxocarboxylic esters, 4 xonitriles, a-amino acids and acyl cyanides. More specifically, the (S)-cyanohydrin of m-phenoxybenzaldehyde is a building block for the synthesis of the insecticide deltamethrin, or (IR)-cis-pyrethroids. ... 

The non-equivalence of enantiomers through the spontaneous breaking of mirror-symmetry in nature is amplified by asymmetric autocatalytic reaction [34], e.g. Frank s spontaneous asymmetric synthesis [35, 36] (Fig. 7-8). Alberts and Wyn-berg have reported in enantioselective autoinduction that chiral lithium alkoxide products may be involved in the reaction to increase the enantioselectivity (Eq. (7.9)) [37]. The product % ee however does not exceed the level of catalyst % ee. In asymmetric hydrocyanation catalyzed by cyclic dipeptides, the (Si-cyanohydrin product complexes with the cyclic peptide to increase the enantioselectivity in the (S)-cyanohydrin product, the reaction going up to 95.8% ee (Eq. (7.10)) [38]. In the presence of achiral amine, (/ )-l-phenylpropan-l-ol catalyzed carbonyl-addition reaction of diethylzinc has been reported to show lower % ee than that of the catalyst employed [39]. 

Progress has also been made in the overexpression of the oxynitrilase from Manihot esculenta (cassava) [67,70] in Escherichia coli. As mentioned previously, this oxynitrilase is very similar to the Hevea enzyme [16,32], because both plants belong to the same plant family, the Euphorbiaceae. A fermentation on the 40 1 scale gave, after simple purification, a total amount of about 40,000 lU of oxynitrilase activity and allowed the exploration of its ability to catalyse the formation of (S)-cyanohydrins [67]. 

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