Estudos de imobilização da enzima (S)-norcoclaurina sintase e potencial de aplicação à síntese de tetraidroisoquinolinas

Abstract

Enzymes are macromolecules capable of catalyzing chemical reactions in mild conditions and with high chemo-, regium- and stereoselectivity, providing advantages over many conventional non-enzymatic methods. The enzyme (S)-norcoclaurine synthase ((S) -NCS), catalyzes, in nature, the reaction whose chemical mechanism was proposed by Pictet-Spengler. The reaction catalyzed by (S) -NCS occurs between dopamine and carbonyl compounds, such as aldehydes and ketones, forming tetrahydroisoquinolines, an important class of compounds with diverse pharmaceutical applications. The objective of the present work was to carry out an immobilization study of (S) -NCS, applied to the synthesis of a tetrahydroisoquinoline resulting from the reaction between dopamine and hexanal. For this, the enzyme was produced via heterologous protein expression, purified and immobilized in a series of matrices, resulting in biocatalysts evaluated for reuse and performance at different temperatures. Chromatography techniques were used for purification, UV / Visible spectroscopy and circular dichroism spectroscopy in the distant UV, for structural analysis of (S) -NCS and NMR spectroscopy to map the reaction by quantifying the concentration of reagents and product. Of all matrices, Diaion HP-2MG presented the highest value of immobilization yield (99%) followed by Celite 545 (86%), Ni-NTA (65%), Caolin (64%) and lastly Dowex 1X4 (53%). On the other hand, as for the reaction between dopamine and hexanal, the sample of Diaion HP-2MG presented a lower performance than the others, while Ni-NTA, Celite 545 and Caolin, were superior in this aspect, in addition to presenting good performance in refers to reusability. Finally, the possibility of carrying out further experiments involving higher reaction temperatures and higher concentrations of organic co-solvent in the reaction was identified in order to assess the stability gain provided by the immobilization in the matrices used in this work.