Hydrolase Catalyzed Michael Addition Reaction And Direct Asymmetric Mannich Reaction

Biocatalysts as environmentally friendly, mild conditions, highly stereoselective and economically feasible catalysts have expanded rapidly for the asymmetric synthesis of chiral intermediates for chemical, pharmaceutical and agrochemical. However, there is only limited availability of enzymes, and enzymes only feature limited substrate specificity. Therefore, it is significant to widen the applicability of existing enzymes by exploring novel unnatural activities of enzymes. Nowadays, enzyme promiscuity, namely the possibility that one active site of an enzyme can catalyze several different chemical transformations. It greatly expands the application of existing enzymes and provides green tools for organic synthesis.In this thesis, we reported Micheal addition of C-C bond formation catalyzed by acid proteinase. We described here the readily available acidic protease from aspergillus usamii (AUAP) catalyzed Michael addition of ketones to nitroolefins. The reaction conditions including organic solvents, solvent volume, water content, temperature, substrate stoichiometry were optimized. The scope of the reaction was tested by varying the nitroalkenes and ketones. For most of aromatic and heteroaromatic nitroalkenes, satisfied yields were obtained and the good yield of up to87%. Besides, we performed some control experiments including the completely denatured enzymes and inhibition of the catalytic activity. But the mechanism is uncertain, and further research is needed.We aslo report the first biocatalytic direct asymmetric three-component Mannich reaction. Crude protein and α-amylase were respectively used to catalyze three-component Mannich reaction between aromatic aldehydes, aromatic amines, and ketones. A commercially available crude protein from porcine pancreas mainly composed of a-amylase, lipase and trypsin. The main components of the crude protein were also used to catalyze the Mannich reaction separately, and the results indicated that a-amylase played important role in the direct asymmetric Mannich reaction. Several important factors including solvent, water content, temperature, enzyme loading, the molar ratio of substrates and pH were examined to optimize the biocatalytic process. A wide range of substrates could be accepted by the crude protein to give Mannich products in satisfactory yields with moderate to good enantioselectivity and diastereoselectivity. The crude protein was used to catalyze the Mannich reaction in1,4-dioxane, and the good yields of up to87%, enantioselectivities of up to82%ee and diastereoselectivities of up to84:16(syn/anti) were achieved. The results indicate that the temperature and optimum water content are different for one enzyme in different solvents. α-Amylase was used to catalyze the direct asymmetric Mannich reaction in1,4-dioxane, a wide range of substrates could be transformed into the corresponding Mannich products up to87%yields, enantiomeric excess (ee) up to76%and diastereoselectivities up to78:22(syn/anti).