2.2 Enzymatic approach
In 2018, Turner et al. reported a chemo-enzymatic synthesis of substituted pyrazines using an amino transaminase (S-selective, ATA-113, Codexis [24]) in the presence of a suitable amine donor, which mediated the key amination of the 1,2-diketone precursor to α-aminoketones that underwent oxidative dimerization to the final product (Figure 2)[25]. In the case of pyrazines, the chirality of the amine group is irrelevant for the synthesis of the aromatic heterocycle core. All reactions were carried out at room temperature with isopropyl amine as the amine donor. Substrates were exhausted after 72 hours and pyrazine was extracted in pure form from the aqueous phase, but the yield of pyrazine was still moderate at 50-65% for symmetric11, deriving from cyclohexane-1,2-dione and 8 deriving from diacetyl, and 32% for the non-symmetric 12 from pentane-2,3-dione (Figure 1). The explanation for by-product formation requires further studies e.g. identification of possible double aminated by-products in the aqueous phase or extraction problems. The question whether dimerization to the pyrazine core occurred in the aqueous buffer or in organic solvent after extraction remains elusive and needs further investigations.
On the one hand, diketones can be produced by different chemical steps from various building blocks (Figure 2). On the other hand, there are also biological options, since the biological route is known to be via acetoin 10 (3-hydroxy-butan-2-one, Figure 2). Recent developments have been made with a ’new’ ThDP-dependent lyase, which is able to synthesize acetoin building blocks from smaller subunits e.g. pyruvate and activated acetaldehyde enzymatically. Pohl et al.showed with engineered PDC from Zymomonas mobilis (Zm PDC) and Acetobacter pasteurianus (Ap PDC) they can obtain excellent yields of 61-98% from the combinations of arylated aldehydes with 1) 3-oxobutanoic acid and 2) alkyl aldehydes. At the same time, it was published how to obtain excellent pyrazine yields from either acetaldehyde with SucA [ThDP-dependent E1 subunit of the α-ketoglutarate dehydrogenase complex from Escherichia coli(E.coli )) ], or from pyruvate (3-oxopropionic acid) with an cellobiose dehydrogenase (CDH )[26].
In industrial processes, cost is one of the determining factors. The price of ATAs (e.g. ATA-113) is much higher than chemical amination with simple ammonia. The enantiomeric integrity of the amination is of great importance e.g. for pharmaceuticals; however, for the production of planar heterocycles the enantioselectivity of the amination is redundant, whereas high regioselectivity of the condensation is essential.
In this context, non-symmetric pyrazines could be synthesized regioselective with ATA- 113 [25] in an one-pot approach, whereas a standard chemical synthesis would only allow access to symmetric or a mixture of non-symmetric pyrazines.