Methoxamine Synthesis in a Biocatalytic 1-Pot 2-Step Cascade Approach

Abstract

Due to its function as a vasopressor, the vicinal amino alcohol methoxamine is a potential candidate for the treatment of hypotension and incontinence or has applications in ophthalmology. In this study, a biocatalytic method was developed to produce each of the four stereoisomers of methoxamine in a sequential 1-pot 2-step cascade reaction, starting from readily available pyruvate and 2,5-dimethoxybenzaldehyde without intermediate isolation. All four isomers are accessible with high conversions and very good stereoselectivities through the modular combination of carboligases and transaminases with high stereoselectivities. The development of these cascades was made possible, among other factors, by the integration of a recently engineered triple variant of the pyruvate decarboxylase from Acetobacter pasteurianus (ApPDC-E469G-I468A-W543F), which provided access to the intermediate (S)-1-hydroxy-1-(2,5-dimethoxyphenyl)propan-2-one with a high enantiomeric excess of 98%. For the amination of these sterically demanding 2-hydroxy ketones, Bacillus megaterium transaminase in particular has proven to be a highly active and selective catalyst. All four methoxamine stereoisomers were achieved with isomeric contents between 94% and 99% and total conversions of both steps between 59% and 80%. Separation of each isomer without derivatizing was possible using supercritical fluid chomratographie with two columns connected in a row. A preparative scale (75 mL) of the 1-pot 2-step cascade to (1S,2R)-methoxamine including nonoptimized product isolation showed 85 mg HCl salt (46% isolated yield) with 94% purity (NMR) and an isomeric content of 98%.

The Effect of Visible Light on the Catalytic Activity of PLP‐Dependent Enzymes

“…The vectors for protein expression pET29a_CvATA, pASK-IBA-35_ VfATA, pET21a_BmATA and pET21a_AsATAmut11 were created by Erdmann, et al [51] For LDC expression, the vector pkk32_EcLDC was used, which was created by Kira Küsters (group of Marco Oldiges; IBG-1, Forschungszentrum Jülich GmbH, Jülich, Germany and ABBt, RWTH Aachen University, Aachen, Germany) and contained the LDCc gene from E. coli employed by Kloss, et al [52] and the vector pET22b_SrLDC, which was constructed by Baud, et al [53] Vector details are given in the Supporting Information. E. coli BL21(DE3) cells (Merck, Darmstadt, Germany) were transformed with the respective expression plasmid by adding 1 μL plasmid solution (100 ng/μL) to 100 μL bacterial solution (OD 600 =~12, in 80 mM CaCl 2 , 20 % (w/v) glycerol).…”

Biocatalysis as Useful Tool in Asymmetric Synthesis: An Assessment of Recently Granted Patents (2014–2019)

“…The technology has been recently extended to other substrates (e.g. 2, 5-dimethoxybenzaldehyde) to afford other synthetically relevant building blocks [334].…”

“…This has enabled the (re)use of pyruvic acid generated as by-product during the transaminase reaction when using L-alanine as amino donor, leading to a fully integrated multi-step enzymatic process with high selectivity and productivity to the desired compound [332,333]. The technology has been recently extended to other substrates (e.g., 2, 5-dimethoxybenzaldehyde) to afford other synthetically relevant building blocks [334].…”

Chemo‐ and Stereoselective Synthesis of Fluorinated Amino Alcohols through One‐pot Reactions using Alcohol Dehydrogenases and Amine Transaminases

“…[6] Amino alcohol asymmetric multienzymatic syntheses have not gone unnoticed, and selected combinations of enzymes have been successfully developed in recent years. Thus, bienzymatic synthesis combining transketolases, [7] pyruvate aldolases, [8] pyruvate decarboxylases [9] or alcohol dehydrogenases (ADHs) [10] with amine transaminases (ATAs) have provided efficient and selective access to amino alcohol diastereoisomers, and even the possibility of using three enzymes (epoxide hydrolases, ADHs and ATAs) in a linear cascade set-up has been successfully reported. [11] The synthesis of various families of fluorinecontaining compounds has attracted great attention in recent decades [12] due to the importance of organofluorinated compounds in medicinal chemistry [13] and asymmetric catalysis.…”