Imobilização de enzimas em nanotubos de titanato e sua aplicação como catalisadores nas reações multicomponentes de Mannich

Abstract

The development of new sinthesis routes for the production of optically active compounds has gained attention in the recent years and so multicomponent reactions are very attractive because they consist in the interaction of three or more molecules in only one reaction step and result in the formation of carbon-carbon bonds. Optically active organic compounds can be obtained using biocatalysis such as enzymes since they can be employed in both aqueous and organic solutions, act in milder conditions of temperature and pressure, are chemo-, regio-, and enantioselective and reduce the formation of by-products. However they have some drawback such as decrease or loss of catalytic activity and stability over time or in organic solvents. To overcome these difficulties the enzyme usually are immobilized on different kinds of supports which facilitate their recuperation and reutilization. Considering that, this work aims to immobilize different kinds of enzymes on titanate nanotubes, use them in Mannich reactions to produce chiral β-amino carbonylated compounds and evaluate different reaction conditions. The titanate nanotubes were sinthetized through the hydrothermal method using TiO2 P25 Degussa and NaOH, and washed with acid and aqueous solutions. The ideal condition for its sinthesis was found to be 0,5g of TiO2 and 100mL of 10M NaOH at 120°C for 24h and washed with 0,1M HCl. The formation of titanate nanotubes were proven using SEM, TEM, XRD, FTIR, and Raman Spectroscopy, through which the peaks of Ti-OH, Ti-O-Ti and OH present in their surface. Furthermore, it was seen that the nanotube are multi walled and have 8-9 nm of diameter and molecular formula of Na2-xHxTi3O7·nH2O. Fourteen different enzymes of the hydrolases class were tested, showing yields of 18 to 13 to 67%. α-chymotrypsin stood out because it has yield of 67% and diastereoisomer ratio of 25:75, determinated by 1H NMR. Trypsin, a protease that acts in tandem with α- chymotrypsin in the digestive system but different mechanism has shown diastereoisomeric inversion comparing to the other enzyme and had yield of 42% and sin:anti proportion of 59:41. Thus both enzymes were chosen to be immobilized. Their immobilization on the nanotubes was in a 1:1 ratio using phosphate buffer for pH values of 6, 7 and 8. The immobilization efficiency was analysed by the Bradford method and the enzyme-nanotube system was characterized through SEM, TEM and FTIR. The immobilized enzymes were used as catalysts in the Mannich reaction, however there was a reduction in the yield and diastereoisomeric ratio for α- chymotrypsin while for trypsin there was also a reduction in the yield but the immobilized trypsin has shown diastereoisomeric inversion.