Cholesterol esterase, resolution

The resolution of racemic FTC butyrate (34) was required for the synthesis of the antiviral drug emtricitabine (Emtriva) (Scheme 7.15) a nucleoside reverse transcriptase inhibitor targeted for treatment of human immunodeficiency virus (HIV) and hepatitis infections [35]. The racemic FTC butyrate ester (34) was treated with immobilized cholesterol esterase, which cleaved the required isomer to the corresponding alcohol (-) 35 with 91% and 52% conversion [36]. The product was isolated as the hydrochloride salt to give 31% yield (98% ) from the 8 kg demonstration. The esterase was immobilized by precipitation onto an accurel polypropylene support using acetone followed by cross linking with glutaralde-... 

Cholesterol Esterase. This enzyme (EC 3.1.1.13 CAS 9026-00-0) physiologically catalyzes the hydrolysis of cholesterol esters, monoacylglycerols, and vitamin esters. It has also been used for several cyclie and noncyclic substrates with variable enantioselectivity. The resolution of racemic esters has been reported and an interesting example is the application to the racemic acetate of an hemiacetal (eq 14). ... 

S)- AND (R)-1,1 -BI-2-NAPHTH0L, obtained by resolution of its pentan-oate ester with cholesterol esterase. 

The best previous resolution of binaphthol uses fractional crystallization of the diastereomeric cinchonine salts of binaphthol cyclic phosphate ester. 8 jhe resolution using cholesterol esterase Involves fewer manipulations and thus is simpler and faster than the cinchonine method. Fewer manipulations also enable the resolutions using cholesterol esterase to be carried out on a larger scale. The high enantloselectivity of cholesterol esterase assures high ee for the (S)-enantiomer, while crystallization of (R)-blnaphthyl dipentanoate assures high enantiomeric purity for the (R)-enantiomer. 

A very recent resolution method is the selective hydrolysis of the di-pentanoic acid ester of racemic binaphthol by cholesterol esterase in practice, inexpensive bovine pancreas acetone powder is used, which cleaves only the (Sl-ester42. 

Cholesterol esterase (CE) and lipase from Candida rugosa (CRL) were very efficient in this kinetic resolution, even on a preparative scale. The best result was obtained with CRL 1 g of racemic 138 yielded 0.48 g of unreacted substrate (5)-138 in 90% ee and 0.39 g of deacetylated product (i )-139 in 88% ee. (iS)-138 was hydrolysed with NaOH, producing (iS)-139 in 75% yield and in almost optically pure form after recrystallisation. Both enantiomers of 139 were subjected to further chemical transformations. For example, the alcohol in (5)-139 was methylated giving 75% yield of enantiomerically pure (5)-140, which was finally reduced by trichlorosilane/triethylamine to give phosphine (5)-141 in 96% ee and 72% yield. This last step occurs with inversion of configuration but some erosion of enantiomeric purity was observed. 

During a study on the resolution of the sterically demanding bicyclic acetate shown in Scheme 2.56 [397], which represents an important chiral building block for the synthesis of leukotrienes [398], it was found that crude steapsin is a highly selective catalyst for its resolution. On the other hand, pure PPL and a-chymotryp-sin were unable to hydrolyze the substrate. Cholesterol esterase, another known hydrolase impurity in crude steapsin capable of accepting bulky substrates, was able to hydrolyze the ester but with low selectivity. Finally, a novel hydrolase which was isolated from crude PPL proved to be the enzyme responsible for the highly selective transformation. 

KINETIC RESOLUTION OF 1,1-BINAPHTHOL BY CHOLESTEROL ESTERASE CATALYZED HYDROLYSIS OF THE PENTANOATE DIESTER[27]... 

In this two-step resolution, the enzyme catalyzes the hydrolysis of the diester to monoester, then the monoester to the diol. Each step contributes to the overall enantioselectivity. The reaction mixture is biphasic buffered water containing the cmde enzyme and an ether solution of the diester. The cmde enzyme is bovine pancreas acetone powder that contains many enz5mies, but the substrate rearts only with the cholesterol esterase so the other enzymes do not interfere, other than to make the work-up messy. The enz5mie cholesterol esterase requires a bile salt, taurocholate, for fiiU activity. This bile salt helps emulsify the two phases. Cholesterol esterase seems to behave more like a lipase than an esterase in this example since it works with the substrate in the organic phase. The dipentanoate ester is chosen to simplify the separation of starting diester and product diol (Figure 5.7). 

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