DEVELOPMENT OF SUSTAINABLE AND EFFICIENT TAILOR-MADE PROCESSES FOR PHARMA AND FOOD APPLICATIONS

The ecological burdens of chemical processes could not be underestimate anymore and a change of direction is required from the industry as well as from academia. In this scenario, my Doctoral Thesis is focused on the development of tailor-made processes for the production of fine chemicals relevant for food and pharma applications which could be considered efficient and sustainable thanks to the employment of techniques with a low environmental impact (i.e., continuous synthesis, biocatalysis, new solvent systems). The main focus of this project was to display how versatile and tuneable the combination of biocatalysis and flow chemistry could be for the synthesis of different chemical compounds, improving established classic but polluting chemical approaches or clearing the way for larger scale manufacturing. Where possible media engineering was applied to further improve the sustainability of the newly designed protocols. In all the processes studied, I also focused my attention on a crucial point of process optimisation: the development of in-line purification steps. The aim was to reduce manual downstream processes by including continuous purification steps, i.e., extraction, catch and release strategies, and/or suitable immobilised scavengers. The Doctoral work developed during these years could be divided into three major area based on the main reactions that have been explored, which were catalysed by different classes of enzymes. In the Thesis I presented and discussed: i) reactions of functional group transfer (from one molecule to another), ii) redox reactions and iii) condensation reactions for the synthesis of different functional groups (i.e., esters, carbonate, carbamate). These reactions were selected due to their wide use in classical organic synthesis that usually required harsh conditions, expensive or toxic reactants, to obtain at the end the desired product with low selectivity and high environmental impact. In some cases, a combination of more than one biocatalyst was used in the same protocol to perform further modifications of the high value chemicals obtained. Biocatalysts were employed in different physical forms (immobilised or free), as isolated enzyme or whole cells. The best approach was evaluated case by case. For example, in the transfer reaction, acyl transferase from Mycobacterium smegmatis was immobilised on glyoxyl agarose by our research group while lipase-mediated reactions were performed using the commercial preparation Novozyme® 435. Redox reactions were mainly catalysed by commercial enzymes as free biocatalyst (i.e., tyrosinase from Agaricus bisporus and laccase from Trametes versicolor). Moreover, the advantages of a self-sufficient system as whole cells were also investigated providing the opportunity to implement the immobilisation procedure of Rhodotorula rubra cells for continuous applications. Where possible the stability or the possibility to reuse the biocatalyst was evaluated in order to reduce the costs of the procedure. The choice of the right solvent was a key point in all the protocols developed and the replacement of classical organic solvents was the path followed whenever the experimental condition allowed it. This also allowed me to evaluate the operational conditions of well-known enzymes in unconventional reactions media (e.g., lipase in tert-amyl alcohol or cyclopenthylmethyl ether) expanding their possible synthetic applications.