Study On The Effects Of Solvent And Acylating Agent On Lipase-catalyzed Resolution Of Chiral Alkyl Alcohols

Enantioselectivity is one of important characteristics displayed by lipases, of which the prominent status was enhanced in asymmetric organic synthesis by the foundation and development of non-aqueous enzymology. The synthesis of chiral alkyl alcohols of enantiomeric purity through lipase-catalyzed resolution of racemates, has been a significant means for the preparation of intermediates for chiral drugs and bioactive natural products. A practical resolution requires good enantioselectivity as well as easy separation of target product from reaction mixture. To meet this requirement, the work is carried out in this thesis.1. Improving the enantioselectivity of thermophilic lipase QLM for the resolutions of (R, S)-2-octanol and (R, S)-2-pentanolWith the resolution (R, S)-2-octanol via transesterification catalyzed by lipase QLM as model reaction, the work was carried out to optimize the enantioselectivity of lipase QLM for the resolution of (R, S)-2-octanol from the following respects: acylating agent, water activity, temperature and reaction medium (organic solvent). Vinyl acetate was effective acylating agent among the enol esters screened. According to the fact that the E value of the resolution decreased as temperature increased, 25℃was selected as optimal temperature for the resolution. The water activity in reaction system had no effect on E value though reaction conversion decreased slightly as it increased. Although log P and dielectric constant had significant effects on the enzyme activity, it was obvious that no trend was achieved in the influence of log P and dielectric constantεon E values. Instead, a significant correlation of E values to the size of solvent molecule was obtained. The enantioselectivity increased as the size of solvent molecule decreased, in particular to the value below 160 ?3. The similar correlation was observed in both cases when the resolution of (R, S)-2-pentanol with the same acyl donor and that of (R, S)-2-octanol with isopropenyl acetate as acyl donor were carried out in the same organic solvents. Therefore, the molecular size was suggested as a key factor governing the solvent effect on enantioselectivity in the resolution of sec-alcohols catalyzed by thermophilic lipase QLM. The E value for the resolution of (R, S)-2-octanol was improved to 21 of practicability by using molecularly small solvent dichloromethane with low boiling point from 11 in solvent-free system, which is of benefit to the separation of product from solvent. Thermodynamic analysis on enantioselectivity at different temperature in the solvents selected indicated that the solvent effect on enantioselectivity was affected by both enthalpic and entropic components. The enthalpic component was affected mainly by the polarity of solvent molecule by the way that the enthalpic component increased with the decrease in polarity of solvent molecule, and the effect of the size of molecule was less. The entropic component depended on the size of solvent molecule in which the entropic component increased with the increase in the size of solvent molecule. Entropic component was much more than enthalpic component and they compensated each other, which resulted in the similar correlations of enantioselectivity and entropic component to the size of solvent molecule and the exception in the correlation of E value to the size of solvent molecule. Excessive large size of molecule caused the exceptions in the correlation of enthalpic component to the polarity of solvent, and the correlation of entropic component to the size of solvent molecule. The solvent effect on enantioselectivity of resolution of sec-alcohols was reflected mainly in the solvent molecular size. This investigation provided a reference model for understanding solvent effect on enantioselectivity of lipase-catalyzed resolution through thermodynamic ananlysis , and an instruction to optimize the enantioselectivity of lipase by medium engineering for a target reaction.2. Lipase-catalyzed resolution of molecularly small chiral alky alcohols via transesterificationThrough transesterification with economically available enol esters (vinyl acetate, isopropenyl acetate and vinyl propionate), the enantioselectivity of commercial lipases PCL, PFL and CALB was investigated for the resolution of the following substrates: (R,S)-2-butanol,1; (R,S)-2-pentanol,2; (R,S)-2-hexanol,3; (R,S)-2-heptanol,4; (R,S)-2-octanol,5; (R,S)-3-hexanol,6; (R,S)-3-methyl-2-butanol,7; (R,S)-6-methyl-5-heptene-2-ol,8; (R,S)-3-methyl-2-cyclohexene-1-ol,9; (R,S)-2-methyl-1- pentanol, 10. Optimal technicse was screened out for resolution of every substrate. The results indicated that lipases PCL and PFL showed similar enantioselectivity for all substrates selected and that the E values for most substrates could not meet practical requirement (E>20). Instead, lipase CALB showed excellent E values (E>200) for substrate 2-5 and 8, and satisfactory E values (5099% and yield of 30%, while fast enantiomer (R)-2-pentanol was achieved merely with purity of ee=91% and yield of 20%. Results show that tranesterification technics involvs onerous operation, vast quantity of organic solvent and low optical purity of product, and this technics is not suitable for the preparation of preparative scale for pure enantiomers of these alcohols.3. Optimizing the separation of product from reaction mixture for lipase-catalyzed resolution of chiral alkyl alcoholsTo overcome the disadvantages in dissatisfatoey enantioselectivity for some substrates, onerous operation for separation, low purity and yield of product in transesterification technics with enol esters, the resolution with succinic anhydride as acylating agent for every substrate alcohol was investigated in organic solvent. The results showed that good enantioselectivities were obtained for the resolution of most substrates by proper choice of lipase. It is worth noting that lipase PCL showed much higher E value for substrate 7 than that shown by itself and CALB in transesterification with enol esters (173>76, 173>95). Also, CALB showed higher enantioselectivity for substrate 6 than that in transesterification with enol esters (115>76). Moreover, the enantioselectivities of lipases for substrates 1 and 10 were higher than the E values in transesterification technics.The technics with succinic anhydride as acylating agent was firstly used for the preparative resolution of (R, S)-2-pentanol with high enantioselectivity. Slowly reacting enantiomer (S)-2- pentanol was obtained with purity of ee>99% and yield of 45%, fast enantiomer (R)-2-pentanol was achieved with purity of ee=95% and yield of 40%. Compared with transesterification technics, acylation with succinic anhydride improved obviously the yield of both enantiomers and the optical purity of fast reacting enantiomer.Secondly, this technics was also used for the preparative resolution of (R, S)-2-methyl-1-pentanol with low E value to certify the practicality of this method. Slowly reacting enantiomer (R)-2- methyl-1-pentanol was obtained with purity of ee>98% and yield of 25%. All results indicates that target product could be obtained with high yield, regardless of its quantity in reaction mixture, for which the reason is that the technics via acylation with succinic anhydride simplifies the separation of product from reaction mixture. This method for the resolution of chiral alkyl alcohols shows obvious practicality compared with transesterification technics.