US EPA

7860079 

Journal Article 

HPLC SEPARATION OF DL-AMINO ACIDS DERIVATIZED WITH N2-(5-FLUORO-2,4-DINITROPHENYL)-L-AMINO ACID-AMIDES 

Bruckner, H; Kellerhoehl, C 

1990 

Yes 

Chromatographia
ISSN: 0009-5893
EISSN: 1612-1112 

FRIEDR VIEWEG SOHN VERLAG GMBH 

WIESBADEN 1 

30 

11-12 

621-629 

English 

10.1007/BF02269735 

WOS:A1990EQ22100003 

Neutral, acidic, and basic protein DL-amino acids (DL-AA) have been separated by HPLC as diastereomeric derivatives obtained after derivatization with N2-(5-fluoro-2,4-dinitrophenyl)-L-alanine amide (FDNP-Ala-NH2, "Marfey's reagent"). HPLC was performed on 3 μm Spherisorb ODS II as the stationary phase with gradient elution using mixtures of triethylammonium phosphate buffer (pH 3) and acetonitrile. The differences in retention times (ΔtR) of diastereomers were compared with those obtained by derivatization of DL-AA with the novel FDNP-reagents FDNP-Val-NH2, FDNP-Phe-NH2, and FDNP-Pro-NH2. FDNP-reagents were synthesized by reaction of 1,5-difluoro-2,4-dinitrobenzene with 0.5 equivalent of the respective L-AA amide in mixtures of aqueous NaHCO3 and acetone at 40-50°C. All FDNP-reagents made possible the resolution of DL-AA. However, FDNP-Val-NH2, gave the largest ΔtR-values in most cases. Large ΔtR-values mainly arise by the formation of an intramolecular hydrogen bond between the carboxy and carboxamide group in the L-L diastereomers and the non-formation of this hydrogen bond in the D-L diastereomer (the first letter refers to the configuration of the AA to be analysed, the second to that of the reagent AA amide) as well as by the low conformational freedom of amino acid residues in diastereomers. © 1990 Friedr. Vieweg & Sohn Verlagsgesellschaft mbH. 

COLUMN LIQUID CHROMATOGRAPHY; AMINO ACID DIASTEREOMER SEPARATION; MARFEY REAGENT; SANGER REAGENT; SEPARATION MECHANISM OF DIASTEREOMERS