Lipases and

One approach called enzymatic resolution, involves treating a racemic mixture with an enzyme that catalyzes the reaction of only one of the enantiomers Some of the most commonly used ones are lipases and esterases enzymes that catalyze the hydrol ysis of esters In a typical procedure one enantiomer of the acetate ester of a racemic alcohol undergoes hydrolysis and the other is left unchanged when hydrolyzed m the presence of an esterase from hog liver... 

In 1989, two enzymes based on genetic engineering techniques were introduced, ie, a cloned alkaline protease (IBIS) and a protein engineered Subtihsin Novo (Genencor, California). Lipase and ceUulase types of detergent enzymes have also begun to appear. 

Both saturated (50) and unsaturated derivatives (51) are easily accepted by lipases and esterases. Lipase P from Amano resolves azide (52) or naphthyl (53) derivatives with good yields and excellent selectivity. PPL-catalyzed resolution of glycidyl esters (54) is of great synthetic utiUty because it provides an alternative to the Sharpless epoxidation route for the synthesis of P-blockers. The optical purity of glycidyl esters strongly depends on the stmcture of the acyl moiety the hydrolysis of propyl and butyl derivatives of epoxy alcohols results ia esters with ee > 95% (30). 

The advantages of the methoxyethyl ester over some of the other water-solubilizing esters are that many of the amino acid esters are crystalline and thus easily purified, are cleaved with a number of readily available lipases, and are useful for the synthesis of A-linked glycopeptides. ... 

Many substrates currently produced in the chemical industry are immiscible with water, but are readily miscible with organic solvents. Most enzymes, however, will not operate efficiently, or not operate at all, in non-aqueous media. Some exceptions do exist, such as lipases and esterases, which can operate in non-aqueous environments. Currently, there is considerable interest in extending the range of enzymes that do work in organic solvents. 

Table 9.9 The triglyceride content of palm oil pre- and post-incubation with lipase and stearic acid as described in the text.

By enzymatic means, chitosan can be easily depolymerized by a variety of hydrolases including lysozyme, pectinase, cellulases, hemicellulases, lipases and amylases, among others, thus showing a peculiar vulnerability to enzymes other than chitosanases [71-76]. While pectinase is of particular... 

An example that refers to the third method additives can be employed is described below. Markedly enhanced enantioselectivity was reported for P. cepacia lipase and subtilisin Carlsberg with chiral substrates converted to salts by treatment with numerous Bronsted-Lowry adds or bases [63]. This effect was observed in various organic solvents but not in water, where the salts apparently dissociate to regenerate... 

Figure 4.8 DKR of seoalcohols catalyzed by a lipase and Ru complexes at ambient temperature.

Figure 4.9 DKR of sec-alcohols using a lipase and inexpensive Al complexes.

Ogasawara ef al. took advantage of the easy racemization of acyloins in the presence of a weak base for the DKR of ewdo-3-hydroxytricyco[4.2.1.0 ]non-7-en-4-one (Figure 4.18) [43]. Acylation of the hydroxyl group was catalyzed by a lipase, and racemization took place via a transient meso-enediol. 

Faber et al. have reported a novel process for the overall deracemization of racemic mandelic acid derivatives using a combination of an enantioselective lipase and a mandelate racemase activity from Lactobacillus paracasei (Figure 5.19) [32]. 

It should also be mentioned that chemoenzymatic epoxidation reactions have been proposed using lipase and H2O2 on multigram scale [120]. 

Walker, C.H. (1994b). Interactions between pesticides and esterases in humans. In M.I. Mackness and M. Clerc (Eds.) Esterases, Lipases, and Phospholipases from Structure to Clinical Significance. NATO ASI Series. Series A, Life Sciences. New York Plenum Press 91-98. 

Recently, an interesting example of the enzymatic kinetic resolution of a-acetoxyamide 8 was demonstrated using native wheat germ lipase and immobilized lipase PS (AMANO) (Scheme 5.6). 

This model clearly shows that the catalytic machinery involves a dyad of histidine and aspartate together with the oxyanion hole. Hence, it does not involve serine, which is the key amino acid in the hydrolytic activity of lipases, and, together with aspartate and histidine, constitutes the active site catalytic triad. This has been confirmed by constructing a mutant in which serine was replaced with alanine (Serl05Ala), and finding that it catalyzes the Michael additions even more efficiently than the wild-type enzyme (an example of induced catalytic promiscuity ) [105]. 

Figure 25-8. Control of adipose tissue lipolysis. (TSH, thyroid-stimulating hormone FFA, free fatty acids.) Note the cascade sequence of reactions affording amplification at each step. The lipolytic stimulus is "switched off" by removal of the stimulating hormone the action of lipase phosphatase the inhibition of the lipase and adenylyl cyclase by high concentrations of FFA the inhibition of adenylyl cyclase by adenosine and the removal of cAMP by the action of phosphodiesterase. ACTFI,TSFI, and glucagon may not activate adenylyl cyclase in vivo, since the concentration of each hormone required in vitro is much higher than is found in the circulation. Positive ( ) and negative ( ) regulatory effects are represented by broken lines and substrate flow by solid lines.

Holm C et al Molecular mechanisms regulating hormone sensitive lipase and lipolysis. Annu Rev Nutr 2000 20 365. 

In adipose tissue, the effect of the decrease in insulin and increase in glucagon results in inhibition of lipo-genesis, inactivation of lipoprotein lipase, and activation of hormone-sensitive lipase (Chapter 25). This leads to release of increased amounts of glycerol (a substrate for gluconeogenesis in the liver) and free fatty acids, which are used by skeletal muscle and liver as their preferred metabolic fuels, so sparing glucose. 

Cutinase is a hydrolytic enzyme that degrades cutin, the cuticular polymer of higher plants [4], Unlike the oflier lipolytic enzymes, such lipases and esterases, cutinase does not require interfacial activation for substrate binding and activity. Cutinases have been largely exploited for esterification and transesterification in chemical synthesis [5] and have also been applied in laundry or dishwashing detergent [6]. 

Irrespective of the physical form of the carotenoid in the plant tissue it needs to be dissolved directly into the bulk lipid phase (emulsion) and then into the mixed micelles formed from the emulsion droplets by the action of lipases and bile. Alternatively it can dissolve directly into the mixed micelles. The micelles then diffuse through the unstirred water layer covering the brush border of the enterocytes and dissociate, and the components are then absorbed. Although lipid absorption at this point is essentially complete, bile salts and sterols (cholesterol) may not be fully absorbed and are not wholly recovered more distally, some being lost into the large intestine. It is not known whether carotenoids incorporated into mixed micelles are fully or only partially absorbed. 

As described above, the temperature effect is useful for enhancing the enantioselectivity however, one problem is the decrease in the reaction rate. For example, although in a lipase AK-catalyzed resolution of solketal, the E value (9 at 30°C, Table 1, entry 1) is increased up to 55 by lowering the temperature to —40°C, 10 times the amount of lipase and 8-fold the reaction time are required as compared with those at 30°C. Thus, the rate of acceleration is an important subject especially to make the low-temperature reaction practical. 

Practical resolution of azirine 1 by the low-temperature method combined with Toyowifc-immobilized lipase and optimized acylating... 

These results indicate that the low-temperature method increases the enantioselectivity, at least above inversion temperature, and the enantioselectivity and reaction rate can be optimized by the use of Toyon/te-immobilized lipase and a suitable acylating agent. 

The KR of secondary alcohols by some hydrolytic enzymes has been well known. The combinations of these hydrolytic enzymes with racemization catalysts have been explored as the catalysts for the efficient DKR of the secondary alcohols. Up to now, lipase and subtilisin have been employed, respectively, as the R- and 5-selective resolution enzymes in combination with metal catalysts (Scheme 2). 

DKR reactions were performed with lipase and Pd(PPh3)4 in the presence of dppf and 2-propanol in THF. 2-Propanol was used as an acyl acceptor. Various acyclic allyhc acetates were transformed to their corresponding allylic alcohols at room temperature in good yields and excellent optical purities (Table 16). 

The DKR of amine is more challenging compared to that of secondary alcohol since no metal catalysts have been known for the efficient racemizahon of amine. Reetz et al. reported for the first time the DKR of amine, in which 1-phenylethylamine was resolved by the combination of lipase and palladium (Scheme 4). In this procedure, CALB and Pd/C were employed as the combo catalysts. However, the DKR required a very long reaction time (8 days) at 50-55°C and provided a poor isolated yield (60%). Recently, an improved procedure using Pd on alkaline earth salts as the racemizahon catalyst was reported by Jacobs et al. " The DKR reachons were performed at 70°C for 24-72 h and 75-88% yields were obtained with 99% or greater enanhomeric excess. 

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