Determination of the absolute configuration of alpha-hydroxyglycine derivatives by enzymatic conversion and chiral high-performance liquid chromatography.

We describe an approach for facile determination of the absolute configuration of enantiomerically chromatographed racemates by combining enzymatic conversion and chiral chromatography. The method involves initial rapid development of chiral HPLC methods using polar organic eluents with polysaccharide chiral phases. We present here evidence for using the stereospecific peptidylamidoglycolate lyase (PGL, E.C. 4.3.2.5) to determine the absolute configuration of alpha-hydroxyglycine derivatives. The racemic solute was incubated with PGL, lyophilized and then enantiomerically chromatographed using the CHIRALPAK AD column. Based on the specificity of the enzyme reaction, the unreacted enantiomer was assigned the absolute configuration R.