2.8 General Experimental Procedure for the NerA Catalyzed Tandem
Reaction
A 25 mL round-bottomed flask was charged with NerA (3 mg
mL-1), GDH (1.5 mg mL-1),
NAD+ (1 mM), deionized water (10 mL), to which the
aldehydes (50 mM) and 1, 3-dicarbonyl compound (100 mM) were introduced.
The resulting mixture was stirred for the specified amount of time at 25
°C. After completion, the product was extracted into EtOAc (4 × 10 mL).
The combined organic layers were dried over anhydrous
Na2SO4 and evaporated under a vacuum.
The residue was purified by flash column chromatography with petroleum
ether/ethyl acetate (4:1) to afford the desired products ethyl
2-(4-chlorobenzyl)-3-oxobutanoate and ethyl 2-benzyl-3-oxobutanoate.