Lipase-catalysed reactions

Lipases catalyse reactions at interfaces, and to obtain a high rate of interesterification the reaction systems should have a large area of interface between the water immiscible reactant phase and the more hydrophilic phase which contains the lipase. This can be achieved by supporting the lipase on the surface of macroporous particles. 

Fig. 21 Short chemo-enzymatic approach to enantiomerically pure (+)-pilocarpine from an easy available precursor using two lipase-catalysed reactions (PS (from AmanoEnzyme Inc.) PLE porcine liver esterase)...

Microemulsions are used as reaction media for a variety of chemical reactions. The aqueous droplets of water-in-oil micro emulsions can be regarded as minireactors for the preparation of nanoparticles of metals and metal salts and particles of the same size as the starting microemulsion droplets can be obtained [1-3]. Polymerisation in micro emulsions is an efficient way to prepare nanolatexes and also to make polymers of very high molecular weight. Both discontinuous and bicontinuous micro emulsions have been used for the purpose [4]. Microemulsions are also of interest as media for enzymatic reactions. Much work has been done with lipase-catalysed reactions and water-in-oil microemulsions have been found suitable for ester synthesis and hydrolysis, as well as for transesterification [5,6]. 

Lif and Holmberg have demonstrated the efficiency of microemulsions as a medium for both organic and bioorganic hydrolysis of a 4-nitrophenyl ester see Scheme 3 of Fig. 3 [7]. The reactions were performed in a Winsor I type microemulsion and took place in the lower phase oil-in-water microemulsion. After the reaction was complete a Winsor I—>111 transition was induced by a rise in temperature. The products formed, 4-nitrophenol and decanoic acid, partitioned into the upper oil phase and could easily be isolated by separation of this phase and evaporation of the solvent. The principle is outlined in Fig. 4. The surfactant and the enzyme (in the case of the lipase-catalysed reaction) resided in the middle-phase microemulsion and could be reused. 

Hundreds of impressive examples of enantioselective lipase-catalysed reactions are known, including industrial processes as in the case of the BASF method of chiral amine production (Collins et al. 1997 Breuer et al. 2004 Schmid and Verger 1998). However, the classical problem of substrate acceptance or lack of enantioselectivity (or both) persists. We were able to meet this challenge in model studies regarding the hydrolytic kinetic resolution of the ester rac-1 with formation of carboxylic acid 2, catalysed by the lipase from Pseudomonas aeruginosa. The wild-type (WT) lipase is only slightly (S )-selective, the selectivity factor amounting to a mere E = 1.1 (Scheme 1). 

For the enantiopure production of human rhinovirus protease inhibitors scientists from Pfizer developed a kinetic resolution and recycling sequence (Scheme 6.14 A). The undesired enantiomer of the ester is hydrolysed and can be racemised under mild conditions with DBU. This enzymatic kinetic resolution plus racemisation replaced a significantly more expensive chemical approach [52]. An enzymatic kinetic resolution, in combination with an efficient chemically catalysed racemisation, is the basis for a chiral building block for the synthesis of Talsaclidine and Revatropate, neuromodulators acting on cholinergic muscarinic receptors (Scheme 6.14B). In this case a protease was the key to success [53]. Recently a kinetic resolution based on a Burkholderia cepacia lipase-catalysed reaction leading to the fungicide Mefenoxam was described [54]. Immobilisation of the enzyme ensured >20 cycles of use without loss of activity (Scheme 6.14 C). 

L-Proline is an efficient catalyst for the reaction between 2 -hydroxyacetophenones and aryl and heteroaryl carboxaldehydes which yields a mixture of chalcone and chroman-4-one cyclic ketones afford only the 2-spiro-linked chromanone <05TL6991> and a lipase-catalysed reaction introduces asymmetry in a multistep synthesis of 3-benzylchromanones <05H(65)761>. 

Subsequently in 1989, we prepared (5)-(-)-paraconic acid by lipase-catalysed reaction,6 and synthesized other microbial hormones with structural similarity to A-factor.7 The biological significance of A-factor in the life cycle of Streptomyces griseus was studied in detail by Horinouchi and Beppu.8... 

Figure 6.10 illustrates the preparation of two key building blocks E and I. The former was synthesized by employing lipase-catalysed reaction, while the latter was derived from L-(+)-tartaric acid (F). The known ( )-4-hydroxy-2-cyclopentenone (A) was converted to C (as a mixture of four stereoisomers) via B. Treatment of C with pig-pancreatic lipase (PPL) afforded (—)-D in 25% yield. By this enzymatic hydrolysis of the acetate C, only the acetate corresponding to (—)-D was hydrolysed even in the presence of chlorine and silicone atoms in the molecule to give the desired (—)-D. The experimental simplicity of... 

It has been shown that lipase-catalysed reactions can be used for fire large-scale production of modified tri ycerides. At present the technology is being targeted to the production of comparatively hig -value products such as confectionary fats. Wider application of file reactions to lower-value, higher-tonnage products will be dependent on the development of cheaper processes using more-productive and/or cheaper catalysts. 

The lipase-catalysed reactions of glyceride hydrolysis and synthesis are reversible. Thus, if the water content of the reaction system is limited, the non-specific enzyme results in random interesterification but the 1,3-specific enzyme and the fatty acid-specific enzyme can be used to introduce desirable fatty acids into the triglyceride molecule and to produce preferred positional fatty acid arrangements in the triglyceride molecule. 

The uses cited by Coleman and Macrae (1980) for the lipase catalysed reaction are examples of acidolysis and transesterification. Acidolysis is illustrated by the increase in l(3)-palmitoyl-3(l)-stearoyl-2-olein (POSt) resulting from the reaction of palm oil mid-fraction with stearic acid in the presence of a 1,3-specific enzyme, and by the increase in the content of linoleic acid in olive oil when that oil is reacted with linoleic acid in the presence of an enzyme with fatty acid specificity. An example of transesterification is shown by the increase in POSt triglycerides when a 1,3-specific enzyme is used to catalyse the interesterification of shea butter and palm oil mid-fraction. 

The progress of any given lipase-catalysed reaction is monitored using two main indicators. The enantiomeric excess of either the starting material or product gives a measure of the enantioselectivity of the reaction. High enantioselectivity... 

Mukherjee, K.D. and Kiewitt, I. (1991) Enrichment of y-linolenic acid from fungal oil by lipase-catalysed reactions. Appl. Microbiol. Biotechnol. 35, 579-584. 

Ohara and co-workers reported that the CALB-catalysed polycondensation of alkyl esters of lactic acid as the monomer produces oligoLAs (X = alkyl, n=2-7 in Scheme 12.1) [13].The reaction is perfectly enantioselective only the alkyl D-lactate monomer produced the oligomers. These results provide the first direct evidence that in the lipase-catalysed reaction mechanism the enantioselection is governed by the deacylation step of lipase . 

STAG. Strong research efforts in the past decade have led to the development of numerous processes, using a broad range of natural starting materials, to obtain value-added products with important and highly desirable nutritional and medical properties. It is to be expected that these achievements will lead to further commercialization of lipase-catalysed reactions for the production of STAG. 

Alkanolamide synthesis has also been achieved using lipase. Alkanolamines are susceptible to acylation both at the amine and hydroxyl group [47, 48]. However, the main product during a lipase-catalysed reaction is the amide, owing to spontaneous acyl migration from alcohol to the amine, the thermodynamically more favourable position (Figure 8.4). 

Skargelind, P. and Jasson, M. (1992) Surfactant interference on lipase catalysed reactions in microemulsions. /. Chem. Technol. Biotechnol., 54, 277-282. 

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