Periodic Reporting for period 1 - SusECAS - DemoPlant (Metaraminol Demonstration Plant as Commercialisation Blueprint for Sustainable Production of Pharmaceuticals using Enzyme Cascades)
Periodo di rendicontazione: 2022-10-01 al 2024-03-31
Amino alcohols and 2-hydroxy ketones are valuable, optically active, compounds with various applications in the chemical and pharmaceutical industry with market prices >1000 €/kg, but challenging synthesis strategies. As a result of classical synthesis options with disastrous footprints, these molecules are often produced in countries with poor working conditions and significantly lower safety and environmental standards than those accepted in the EU.
The idea of 'SusECAS - DemoPlant' was to enable an economic and ecological competitive process for the production sustainable production of 2-hydroxy ketones and thus amino alcohols in Europe. In the ERC Starting Grant project 'LightCas', Prof. Rother laid the base with an atomic and step-efficient enzymatic 2-step reaction for amino alcohols from inexpensive starting materials (<10-100 €/kg). In 'SusECAS' these innovative synthesis idea should be brought to a new level of industrial application. A multi-gram demonstration process to the drug metaraminol should enable a clear indication of the ecological and economic potential.
In order to go one step further in value creation, not only the synthesis of the intermediate 3-OH-PAC was considered, but also the synthesis of metaraminol itself. In the literature, purely chemical routes starting from (R)-3-OH-PAC are known and established in pharmaceutical manufacturing companies. We looked at the enzymatic synthesis of (R)-3-OH-PAC to metaraminol and wanted to demonstrate this first on a laboratory scale and then optimize and scale it up analogously to the synthesis of (R)-3-OH-PAC. Theoretically, this type of reaction can be carried out by so-called amine transaminases. We tested a large number of enzymes and initially found some positive screening hits. The results of the screening rounds gradually showed that there are enzymes that can produce metaraminol, but only product concentrations of less than 20mM could be achieved. Even after a multi-parameter optimization of the best screening hits, the product concentration could not be further increased. We initially thought of a classical enzyme inhibition caused by the substrate and/or the product. After various investigations, it turned out that the co-factor required for these enzymes (PLP) is irreversibly destroyed by the substrate (R)-3-OH-PAC and the product metaraminol after a short time and the reaction therefore comes to a standstill. The problem could not be solved by adding new enzymes or addition of new co-factor. We also tried to precipitate the metaraminol product from the reaction while it was still forming (in-situ product isolation) and/or to find other enzymes. So far, no option has led to success. The low product concentrations of approx. 10mM are not competitive (in coordination with our industrial partners) compared to the existing chemical synthesis routes.
In contrast to the (R)-3-OH-PAC synthesis, the enzymatic production of Metaraminol from (R)-3-OH-PAC was demonstrated within 'SusECAS' only in principle. Although we have found enzymes that perform the reaction in principle, we have not managed to establish the process at a level that would be industrially competitive. Unfortunately, it has been shown that the co-factor is irreversibly damaged by the substrate and the product. This is a new phenomenon for this type of, which had not been described before. In order to be able to produce Metaraminol in an industrially competitive manner, the problem needs to be better understood and solutions developed for it. Contrary to expectations, more research is needed here. In order to draw attention to this new phenomenon, we will publish our data to sensitize other researchers to this problem and perhaps find a solution in future.