Determination of beta-N-oxalyl-L-alpha,beta-diaminopropionic acid in Lathyrus sativus by precolumn derivatization with 1-fluoro-2,4-dinitrobenzene

An isocratic high performance liquid chromatographic (HPLC) method was developed to analyze neurotoxin,beta-N-oxalyl-L-alpha,beta -diaminopropionic acid (beta-ODAP) and its innoxious isomer, alpha-N-oxalyI-L-alpha,beta-diaminopropionic acid (alpha-ODAP) in Lathyrus sativus by precolumn derivatization with 2, 4-dinitrofluorobenzene (DNFB). The chromatographic conditions were optimized including solid phase extraction (SPE) pretreatment and pH of the buffer in mobile phase. For further explanation of the retention behavior of two isomers in theory, Hyperchem software was used to obtain the optimal structures and Gaussian 98 W to calculate the polarity of two isomers. Under the optimized conditions such as column of C-18 (5 mu m, 15 mm x 4.6 mm, mobile phase of 25 mmol/L KH2PO4-acetonitrile (90: 10, V/V), flow rate of 0.8 mL/min and room temperature condition, the contents and ratio of the two isomers from different plant developing procedures of Lathyrus sativus in special environment were obtained.