We can offer three different methods for determination of enantiomeric purity of the amino acids in different matrices:

FID-Method:

For free amino acids the method is appropriate because no hydrolysis is performed and no byproducts are to expect.

Also for most of the amino acid derivates the method is suitable. If however the cleavage leads to high concentration of byproduct or the sample itself is contaminated, co-elution with any contamination is possible and leads to incorrect results. Seldom racemization is observed during sample preparation. These possible problems are minimized by the GC-MS (A.0.3.) analysis.

For determination of enantiomeric purity of peptides the method can give an estimation if racemization is possible. However reliable determination <1%, for Cysteine <10% is not possible with this method.

GC-MS-Method:

Racemization during this sample preparation is accompanied by deuterium exchange in the a-position (deuterium label). The proportion of D-amino acid originally present in the peptide is thus represent by the relative amounts of the unlabeled form which is monitored by mass spectrometry. This method is also indicated if the enantiomeric purity of amino acid derivatives is needed with low standard deviation (eg. for derivatives with a specification <0.3%)

HPLC method:

The third method uses chiral columns for HPLC. A couple of derivatives can be separated on chiral HPLC columns without derivatization, if they have a chromophoric group like Fmoc. The lower selectivity of the chromatographic column is often compensated by the high selectivity of the detector.