The amino alcohol Metaraminol is a valuable drug required to treat hypotension, which is a risk factor for serious diseases. An intermediate of the Metaraminol production is (R)-3-hydroxy-phenylacetylcarbinol ((R)-3-OH-PAC). In 'SusECAS', the synthesis of (R)-3-OH-PAC was developed based on the inexpensive starting materials 3-OH-benzaldehyde (3-OH-BA) and pyruvate. In principle, this type of reaction can be carried out by enzymes called carboligases. In the beginning of the project, we screened and identified a carboligase that could do this job. Once the feasibility had been fundamentally demonstrated, an important focus was on optimizing the reaction from an economic and ecological point of view. In order to survive in a possible industrial competition, the highest possible product concentrations and yields had to be achieved in the process. Thus, we performed a performed a muti-parameter reaction optimization and compared the several aspects. On the one hand, experiments with different catalyst production and formulation strategies were performed. Moreover, reactions with substrates from different manufacturers and reactions with different substrate feeding strategies have been carried out. After optimization of all these parameters, we could increase the substrate (and thus the product) concentration to >0.3-0.4M with a very high conversion value of up to >95%. Moreover, by establishing a downstream processing protocol, we could isolate the product with a very high chemical purity of >99 % and an extremely high enantiomeric purity of >99.9%. In order to show the scalability of this reaction, a synthesis with a reaction volume of 2.5 L has been successfully conducted. In discussion with our industrial partners, our developed process represents a very good and competitive technology to produce this valuable intermediate (R)-3-OH-PAC of metaraminol synthesis.
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.
