HPLC ANALYSIS OF A BACKBONE COMPOUND, BETA-BENZYL-N-CARBOBENZOXY-L-ASPARTYL-D-ALANINE, OF INTENSE PEPTIDE SWEETENERS BELONGING TO L-ASPARTYL-D-ALANINE AMIDES

A simple and reliable HPLC method for determining a backbone compound, beta-benzyl-N-carbobenzoxy-L-aspartyl-D-alanine, from intense peptide sweeteners belonging to L-aspartyl-D-alanine amides is reported. The backbone compounds synthesized showed high enough purity to be used as standards. The most effective eluent composition and flow rate for the HPLC assay of the backbone compound were CH3CN: 0.2 m triethylamine-H3PO4 (pH 3) in 1:1 ratio and 0.8 ml min-1, respectively. The HPLC method established here was found to have excellent reproducibility and favourable recovery for the backbone compound. When analysed by this method, the optimum reaction time between the silylated D-alanine and mixed anhydride in the synthesis of the backbone compound was found to be 80 min.