Amino acid analysis by high-performance liquid chromatography after derivatization with diethyl ethoxymethylenemalonate.
Amino acids were determined by precolumn derivatization with diethyl ethoxymethylenemalonate and reversed-phase high-performance liquid chromatography (HPLC) with spectrophotometric detection at 280 nm. The reaction time was 50 min and the derivatives were stable at room temperature. Chromatographic resolution of a mixture of the derivatives of seventeen amino acids, including proline and cystine, was achieved within 35 min using a binary gradient system. The detection limit was 3 pmol. Amino acid analyses of acid hydrolysates of two proteins gave results equivalent to those obtained by conventional ion-exchange-based amino acid analysis. The simplicity of the procedure allows its use on any multi-purpose HPLC system.
Alaiz
M
M
Instituto de la Grasa y sus Derivados (CSIC), Seville, Spain.
Navarro
J L
JL
Girón
J
J
Vioque
E
E
D000596:Amino Acids / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D006868:Hydrolysis; D008314:Malonates / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet
D000596:Amino Acids; D008314:Malonates; C044472:diethyl (ethoxymethylene)malonate