Biocatalysis and flow chemistry are ideal partners for accessing novel chemical spaces and define sustainable and efficient synthetic tools with high level of intensification.1 Biocatalytic redox reactions are very attractive topic since they often occur with high stereo- and regio-selectivity under mild and environmentally friendly conditions. The aim of our work was to develop greener and scalable routes for the synthesis of high value chemicals, using both whole cells and isolated enzymes. Moreover, new solvent systems such as Natural Deep Eutectic Solvent (NADES) have been investigated as green co-solvents in enzymatic transformations. The processes have been designed integrating in-line workups and purification procedures to reduce manual handling, downstream processes and costs. For this purpose, meso reactors were employed in order to demonstrate that with a biocatalytic approach was also possible to obtain high amount of pure final compound improving the total productivity of the system. i) Enantioselective reduction of β-ketonitriles Immobilised whole cells of Rhodotorula rubra MIM 147, a widespred yeast genus, have been exploited in a fully automated system for the enantioselective synthesis of β-hydroxynitriles, starting from β-ketonitriles (Scheme 1).2 We selected a natural deep eutectic solvent made by choline chloride/glucose with a dual purpose as a co-solvent and as an internal source of glucose, fundamental for the cofactor regeneration. With the aim of developing a fully automated procedure for the synthesis of a key building block for the synthesis of the antidepressant drug duloxetine, i.e., (S)-3-hydroxy-3-(thiophen-2-yl)propanenitrile, an in-line purification procedure has been developed. An in-line extraction of the desired product was performed adding a flow stream of ethyl acetate followed by a liquid/liquid separation. The organic stream with any unreacted substrate and the desired product was further purified by flowing it through a column packed with polymer supported benzylamine (PS-BZA), allowing the recovery of the pure final product after simple solvent evaporation. The optimised protocol allowed the isolation of the desired product in 80 minutes of residence time with >90% conversion and >99% e.e. and resulted to be versatile for the enantioselective reduction of different β-ketonitriles. ii) Oxidation of tyrosol to hydroxytyrosol Tyrosol (Ty) and hydroxytyrosol (HTy) are valuable dietary phenolic compounds present in olive oil and wine and possess a range of biological effects.3 The availability of gram amounts of pure Ty, HTy, their metabolites and derivatives are highly appealing for a deep biological evaluation. Furtrhermore, it must be considered that the price of HTy compared to the cost of Ty is more than 1000 higher. In this context, we performed the oxidation of easily accessible Ty using a free tyrosinase from Agaricus bisporus in presence of oxygen and ascorbic acid. The aqueous flow stream was then extracted in-line with ethyl acetate and the aqueous layer, containing the biocatalyst and the excess of ascorbic acid, was recirculated to improve the overall efficiency in a self-sufficient closed-loop system. An in-line purification for the collection of pure HTy was also designed (Scheme 2). Then, a bioreactor packed with an immobilised acyltransferase from Mycobacterium smegmatis (MsAcT) was used to produce acetate derivatives (Scheme 3).4 With this modular set up, HTy was obtained with an isolated yield of 75%, whereas the acetate metabolites showed yields up to 80% in 10 minutes. References 1. Tamborini, L.; Fernandes, P.; Paradisi, F.; Molinari, F. Trends Biotechnol. 2018, 36, 73-88. 2. Facchetti, G.; Gandolfi, R.; Fusè, M.; Zerla, D.; Cesarotti, E.; Pellizzoni, M.; Rimoldi, New J. Chem. 2015, 39, 3792 – 3800 3. Karkovíc Markovíc, A.; Toríc, J.; Barbaríc, M.; Jakobušíc Brala,C. Molecules, 2019, 24, 2001. 4. Contente, M.L.; Farris, S.; Tamborini, L.; Molinari, F.; Paradisi,F. Green Chem. 2019, 21, 3263-3266. he solutions of Ty and tyrosinase with ascorbic acid were prepared in sodium phosphate
Continuous biocatalysed redox reactions for the ecofriendly and improved synthesis of pharmaceutically relevant compounds / F. Annunziata, M.L. Contente, R. Gandolfi, P. Conti, C. Pinna, A. Pinto, S. Dallavalle, L. Tamborini. ((Intervento presentato al convegno Biotrans 2023 tenutosi a La Rochelle nel 2023.
Continuous biocatalysed redox reactions for the ecofriendly and improved synthesis of pharmaceutically relevant compounds
F. Annunziata;M.L. Contente;R. Gandolfi;P. Conti;C. Pinna;A. Pinto;S. Dallavalle;L. Tamborini
2023
Abstract
Biocatalysis and flow chemistry are ideal partners for accessing novel chemical spaces and define sustainable and efficient synthetic tools with high level of intensification.1 Biocatalytic redox reactions are very attractive topic since they often occur with high stereo- and regio-selectivity under mild and environmentally friendly conditions. The aim of our work was to develop greener and scalable routes for the synthesis of high value chemicals, using both whole cells and isolated enzymes. Moreover, new solvent systems such as Natural Deep Eutectic Solvent (NADES) have been investigated as green co-solvents in enzymatic transformations. The processes have been designed integrating in-line workups and purification procedures to reduce manual handling, downstream processes and costs. For this purpose, meso reactors were employed in order to demonstrate that with a biocatalytic approach was also possible to obtain high amount of pure final compound improving the total productivity of the system. i) Enantioselective reduction of β-ketonitriles Immobilised whole cells of Rhodotorula rubra MIM 147, a widespred yeast genus, have been exploited in a fully automated system for the enantioselective synthesis of β-hydroxynitriles, starting from β-ketonitriles (Scheme 1).2 We selected a natural deep eutectic solvent made by choline chloride/glucose with a dual purpose as a co-solvent and as an internal source of glucose, fundamental for the cofactor regeneration. With the aim of developing a fully automated procedure for the synthesis of a key building block for the synthesis of the antidepressant drug duloxetine, i.e., (S)-3-hydroxy-3-(thiophen-2-yl)propanenitrile, an in-line purification procedure has been developed. An in-line extraction of the desired product was performed adding a flow stream of ethyl acetate followed by a liquid/liquid separation. The organic stream with any unreacted substrate and the desired product was further purified by flowing it through a column packed with polymer supported benzylamine (PS-BZA), allowing the recovery of the pure final product after simple solvent evaporation. The optimised protocol allowed the isolation of the desired product in 80 minutes of residence time with >90% conversion and >99% e.e. and resulted to be versatile for the enantioselective reduction of different β-ketonitriles. ii) Oxidation of tyrosol to hydroxytyrosol Tyrosol (Ty) and hydroxytyrosol (HTy) are valuable dietary phenolic compounds present in olive oil and wine and possess a range of biological effects.3 The availability of gram amounts of pure Ty, HTy, their metabolites and derivatives are highly appealing for a deep biological evaluation. Furtrhermore, it must be considered that the price of HTy compared to the cost of Ty is more than 1000 higher. In this context, we performed the oxidation of easily accessible Ty using a free tyrosinase from Agaricus bisporus in presence of oxygen and ascorbic acid. The aqueous flow stream was then extracted in-line with ethyl acetate and the aqueous layer, containing the biocatalyst and the excess of ascorbic acid, was recirculated to improve the overall efficiency in a self-sufficient closed-loop system. An in-line purification for the collection of pure HTy was also designed (Scheme 2). Then, a bioreactor packed with an immobilised acyltransferase from Mycobacterium smegmatis (MsAcT) was used to produce acetate derivatives (Scheme 3).4 With this modular set up, HTy was obtained with an isolated yield of 75%, whereas the acetate metabolites showed yields up to 80% in 10 minutes. References 1. Tamborini, L.; Fernandes, P.; Paradisi, F.; Molinari, F. Trends Biotechnol. 2018, 36, 73-88. 2. Facchetti, G.; Gandolfi, R.; Fusè, M.; Zerla, D.; Cesarotti, E.; Pellizzoni, M.; Rimoldi, New J. Chem. 2015, 39, 3792 – 3800 3. Karkovíc Markovíc, A.; Toríc, J.; Barbaríc, M.; Jakobušíc Brala,C. Molecules, 2019, 24, 2001. 4. Contente, M.L.; Farris, S.; Tamborini, L.; Molinari, F.; Paradisi,F. Green Chem. 2019, 21, 3263-3266. he solutions of Ty and tyrosinase with ascorbic acid were prepared in sodium phosphatePubblicazioni consigliate
I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.