Acylation enzymatic

Enzymatic acylation reactions offer considerable promise in the synthesis of specific ester derivatives of sucrose. For example, reaction of sucrose with an activated alkyl ester in /V, /V- dim ethyl form am i de in the presence of subtilisin gave 1 -0-butyrylsucrose, which on further treatment with an activated fatty acid ester in acetone in the presence of Hpase C. viscosum produced the 1, 6-diester derivative (71,72). 

Mono cylDiols. Enzymatic synthesis of chiral monoacyl diols can be carried out either by direct enzymatic acylation of prochiral diols or by hydrolysis of chemically synthesized dicarboxylates. 

In contrast to the hydrolysis of prochiral esters performed in aqueous solutions, the enzymatic acylation of prochiral diols is usually carried out in an inert organic solvent such as hexane, ether, toluene, or ethyl acetate. In order to increase the reaction rate and the degree of conversion, activated esters such as vinyl carboxylates are often used as acylating agents. The vinyl alcohol formed as a result of transesterification tautomerizes to acetaldehyde, making the reaction practically irreversible. The presence of a bulky substituent in the 2-position helps the enzyme to discriminate between enantiotopic faces as a result the enzymatic acylation of prochiral 2-benzoxy-l,3-propanediol (34) proceeds with excellent selectivity (ee > 96%) (49). In the case of the 2-methyl substituted diol (33) the selectivity is only moderate (50). 

Cychc alcohols are excellent targets for enantioselective enzymatic acylations. For example, acylation of (65) with vinyl acetate catalyzed by Hpase SAM-II gives the (R),(3)-ester with 95% ee (81). Similarly (66), which is a precursor for seratonin uptake inhibitor, is resolved in a high yield and selectivity with Amano Hpase P (82). The prostaglandin synthon (67) is resolved by the same method into the optically pure alcohol in 35% yield (83). 

ACOCH2CF3, porcine pancreatic lipase, THE, 60 h, 77% yield. This enzymatic method was used to acetylate selectively the primary hydroxyl group of a variety of carbohydrates. The selective enzymatic acylation of carbohydrates has been partially reviewed. ... 

Another example of an enzymatic one-pot multiple Diels-Alder reaction is illustrated in Table 4.20 [83]. Racemic furfuryl alcohols 130 in the presence of ethoxy vinyl methyl fumarate 131 and enzyme TOYOBO-LIP undergo enzymatic acylation followed by kinetic enzymatic resolution to give the acyl derivatives 132 which then affords the adducts 133 and 134 by intramolecular Diels-Alder reaction 3-methyl-furfuryl alcohol 130 (R = Me) in acetone gives the best results. 

Over the past years, interest in the preparation of chiral amines and amides by enzymatic ammonolysis or aminolysis reactions [4] has greatly increased for academic and industrial sectors. The role that the enzymatic acylation of amines or ammonia plays for the preparation of some pharmaceuticals is noteworthy [5]. 

Scheme 7.4 Enzymatic acylation of amines with carboxylic acids...

Here, we have selected a few representative examples of the enzymatic resolution of esters by aminolysis or ammonolysis reactions. On the other hand, the enzymatic acylation of racemic amines is also of great utility for the preparation of optically pure... 

Gotor-Fernandez, V., ReboDedo, F. and Gotor, V. (2007) Preparation of chiral pharmaceuticals through enzymatic acylation of alcohols and amines, in Biocatalysis in the Pharmaceutical and Biotechnology Industry, (ed. R.M. Patel), Dekker, Taylor and Francis, New York, Chapter 7, pp. 203-248. 

The DKR of secondary alcohols can be efficiently performed via enzymatic acylation coupled with simultaneous racemization of the substrates. This method was first used by BackvaU for the resolution of 1-phenylethanol and 1-indanol [38]. Racemization of substrate 18 by a mthenium catalyst (Scheme 5.11) was combined with transesterification using various acyl donors and catalyzed by C.antarctica B Hpase. From aU the acyl donors studied, 4-chlorophenyl acetate was found to be the best. The desired product 19 was obtained in 80% yield and over 99% ee. 

The method is not restricted to secondary aryl alcohols and very good results were also obtained for secondary diols [39], a- and S-hydroxyalkylphosphonates [40], 2-hydroxyalkyl sulfones [41], allylic alcohols [42], S-halo alcohols [43], aromatic chlorohydrins [44], functionalized y-hydroxy amides [45], 1,2-diarylethanols [46], and primary amines [47]. Recently, the synthetic potential of this method was expanded by application of an air-stable and recyclable racemization catalyst that is applicable to alcohol DKR at room temperature [48]. The catalyst type is not limited to organometallic ruthenium compounds. Recent report indicates that the in situ racemization of amines with thiyl radicals can also be combined with enzymatic acylation of amines [49]. It is clear that, in the future, other types of catalytic racemization processes will be used together with enzymatic processes. 

Bellemin-Laponnaz S, Twedel J, Ruble JC, Breitling FM, Fu GC (2000) The kinetic resolution of allylic alcohols by a non-enzymatic acylation catalyst application to natural product synthesis. Chem Conunun 1009-1010... 

Fig. 9. Enzymatic acylation reaction of 18 and 19. Structure 21 is proposed as the reaction intermediate...

The aminoalkanephosphonic acids which bear an additional hydroxy group in the molecule were usually resolved via enzymatic acylation of this hydroxy group. For example, resolution of //-Cbz-phosphoserine dimethyl ester 60 using various lipases gave poor results. However, lipase PS-promoted acetylation of //-Chz-phosphoisoserine diethyl ester 61 gave both the unreacted substrate 61 and the 0-acetylated product 62 with almost 100% enantiomeric excess (E = 1000). ... 

As mentioned above, Met(0) must be converted to Met before it can be incorporated into proteins. There are a wide variety of organisms that have been shown to be capable of enzymatically reducing Met(O) residues. The enzymatic reduction of free Met(O) to Met has been observed in yeast , E. cofi - , Pseudomonas , plants and animal tissues . The enzyme from E. coli has been purified about 1100-fold using a newly developed very sensitive assay . The assay involves first the conversion of [ S]Met(0) to [ S]Met by the Met(O) reductase followed by the measurement of [ S]Met-tRNA after enzymatic acylation of tRNA. Since Met(O) is not a substrate for the acylation reaction , the amount of [ S]Met-tRNA formed is proportional to the amount of [ S]Met(0) converted to [ S]Met. The assay is sensitive to Met levels of less than 1 pmol. 

Sphinganine is acylated, then dehydrogenated to form ceramide. Free sphingosine, also termed sphingenine, salvaged from sphingolipid breakdown can be enzymatically acylated with acyl-CoA to form ceramide. 

Scheme 4 6 -0-Acyl-lactose (acetyl, propionyl, butyryl) derivatives prepared via the selective enzymatic acylation of lactose by the protease subtilisin were used as acceptors for enzymatic transglycosylations catalyzed by a-D-galactosidase from Talaromyces flavus, forming iso-globotriose a-Gal(l -> 3)-p-Gal-(l -> 4).92 (/) Protease N, pyridine, 37 °C (ii) a-galactosidase from Talaromyces flavus.

Analogous to the reactions of chiral alcohols, enantiomerically pure amines can be prepared by (D)KR of the racemate via enzymatic acylation. In the case of alcohols the subsequent hydrolysis of the ester product to the enantiomerically pure alcohol is trivial and is generally not even mentioned. In contrast, the product of enzymatic acylation of an amine is an amide and hydrolysis of an amide is by no means trivial, often requiring forcing conditions. 

P-Adrenergic-blocking agents, such as propranolol, have been synthesized by different chemoenzymatic methods where the key step to introduce the chirality is an enzymatic acylation or a hydrolysis process. The main reason to prepare these amino alcohols in optically pure form is due to the fact that the activity of these pharmaceuticals resides in the (S)-enantiomer. In Scheme 10.1 we have represented a chemoenzymatic approach that has been carried out for the preparation of this dmg where the key step is the resolution of the key intermediate 1-chloro-... 

The resolution of primary or secondary alcohols is easy by enzymatic acylation however, few examples have been described for the resolution of tertiary alcohols [26]. 

Although in recent years transesterification processes of racemic alcohols have received major attention, enzymatic acylation of amines for synthetic purposes is also being employed as a conventional tool for the synthesis of chiral amines and amides [31], using CALB as the biocatalyst in the majority of these reactions [31a]. The main difference between enzymatic acylation of alcohols and amines is the use of the corresponding acyl donor, because activated esters which are of utility... 

Kinetic optical resolution of racemic alcohols and carboxylic acids by enzymatic acyl transfer reactions has received enormous attention in recent years56. The enzymes generally employed are commercially available lipases and esterases, preferentially porcine liver esterase (PLE) or porcine pancreatic lipase (PPL). Lipases from microorganisms, such as Candida cylindracea, Rhizopus arrhizus or Chromobacterium viscosum, are also fairly common. A list of suitable enzymes is found in reference 57. Standard procedures are described in reference 58. Some examples of the resolution of racemic alcohols are given39. 

In favorable cases, enzymatic acylation and deacylation are stereochemically complementary and provide access to both enantiomers. 

NPIL). The chiral triol was produced in a whole cell reduction process and was then selectively protected at the primary alcohol by enzymatic acylation using Novozyme 435 (CAL-B). Finally the key intermediate was produced by protecting the two secondary alcohols as they cleanly reacted with dimethoxypropane to give a crystalline product (>99 % ee, >99 % de, 96 % purity) which was subsequently used for Lipitor production (Figure 1.44). 

Regioselective enzymatic acylation of large, insoluble polysaccharides is still a quite difficult task and therefore it is not surprising that only scant data have been reported up to now, most of them describing reaction outcomes which met with limited success. Nevertheless, enzymatic derivatization of polysaccharides has been performed in nonpolar organic solvents using insoluble polysaccharides with soluble [51] or suspended enzymes [52]. Chemically modified celluloses with either enhanced solubility or more readily accessible hydroxyl groups, like cellulose acetate or hydroxypropyl cellulose, were acylated by CalB, as reported by Sereti and coworkers [53]. However, the same authors failed to modify crystalline cellulose under the same reaction conditions. 

The application of enzymatic acylation for the resolution of racemic alcohols in organic solvent has shown to be an effective method to rapidly synthesize chiral alcohols. The racemic alcohols are treated with the lipase and acylating agent one enantiomer remains unconverted whereas the second enantiomer is esterified and easily separated by distillation (Scheme 7.2). Vinyl acetate or isopropenyl acetate are typical acylating agents, as the generated vinyl alcohol tautomerizes rapidly... 

Scheme 7.2 Resolution of racemic alcohols through enzymatic acylation.

Scheme 7.3 The synthesis of styrene oxides through the enzymatic acylation of chloroalcohols.

Scheme 7.4 Resolution of racemic alcohol enzymatic acylation.

Scheme 7.8 Dynamic kinetic resolution of racemic alcohols by the combination of transition metal catalysis with enzymatic acylation.

Scheme 7.17 Enzymatic acylation of 2-amino-9- 3-D-arabinofuranosyl-6-methoxy-9H-purine (Glaxo Wellcome).

In addition, at very low water contents, ampicillin accumulation curves do not exhibit a clear-cut maximum, inherent in the enzymatic acyl transfer reactions in aqueous medium (including quite concentrated heterogeneous aqueous solution-precipitate systems), because of the secondary hydrolysis of the target product by penicillin acylase (Figure 12.6) [84]. 

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