Kerstin Spornhauer (geb. Würges), Katerina Petruevska-Seebach, Martin Peter Elsner, Stephan Lütz:
Enzyme-assisted physicochemical enantioseparation processes - part III: Overcoming yield limitations by dynamic kinetic resolution of asparagine via preferential crystallization and enzymatic racemization
Biotechnology and Bioengineering, 2009, 104, 1235-1239
The application of enantioseparation methods alone can only yield up to 50% of the desired chiral product. Thus enantioseparation becomes more attractive when accompanied by the racemization of the counter-enantiomer. Here we present first results of dynamic kinetic resolution of Lasparagine (L-Asn) via preferential crystallization and enzymatic racemization from a racemic, supersaturated solution on a 20mL scale. An enzyme lyophilisate (WT amino acid racemase from P. putida KT2440 (E.C. 184.108.40.206), overexpressed in E. coli BL21(DE3)) was used for in situ racemization (enzyme concentrations varying from 0 to 1mg/mL). When preferential crystallization was applied without any enzyme, a total of 31mg of L-Asn monohydrate could be crystallized, before crystal formation of D-Asn started. Crystallization experiments accompanied by enzymatic racemization led to a significant increase of crystallized L-Asn (198mg L-Asn monohydrate; >92%ee) giving the first experimental proof for this new process concept of dynamic kinetic resolution via preferential crystallization and enzymatic racemization. Measurements of the racemase activity before and after the crystallization process showed no significant differences, which would allow for enzyme recovery and recycling.
amino acid racemase; dynamic kinetic resolution; preferential crystallization; integrated enantioseparation process; asparagine; in situ racemization; production of L-amino acids