Quantitative analysis of N-phenylpropenoyl-L-amino acids in roasted coffee and cocoa powder by means of a stable isotope dilution assay

Since recent reports on the role of N-phenylpropenoyl-L-amino acids as powerful antioxidants and key contributors to the astringent taste of cocoa nibs, there is an increasing interest in the concentrations of these phytochemicals in plant-derived foods. A versatile analytical method for the accurate quantitative analysis of N-phenylpropenoyl-L-amino acids in plant-derived foods by means of HPLC-MS/MS and synthetic stable isotope labeled N-phenylpropenoyl-L-amino acids as internal standards was developed. By means of the developed stable isotope dilution assay (SIDA), showing recovery rates of 95-102%, 14 N-phenylpropenoyl-L-amino acids were quantified for the first time in cocoa and coffee samples. On the basis of the results of LC-MS/MS experiments as well as cochromatography with the synthetic reference compounds N-[3′,4′-dihydroxy-(E)-cinnamoyl]-L-tryptophan, N-[4′-hydroxy- (E)-cinnamoyl]-L-tryptophan, and N-[4′-hydroxy-3′-methoxy-(E)- cinnamoyl]-L-tyrosine, respectively, were detected for the first time in cocoa powder, and (-)-N-[4′-hydroxy-(E)-cinnamoyl]-L-tyrosine, (-)-N-[3′,4′-dihydroxy-(E)-cinnamoyl]-L-tyrosine, N-[4′-hydroxy-3′-methoxy-(E)-cinnamoyl]-L-tyrosine, (+)-N-[3′,4′-dihydroxy-(E)-cinnamoyl]-L-aspartic acid, (+)-N-[4′-hydroxy-(E)-cinnamoyl]-L-aspartic acid, N-[3′,4′- dihydroxy-(E)-cinnamoyl]-L-tryptophan, N-[4′-hydroxy-(E)-cinnamoyl]-L- tryptophan, and N-[4′-hydroxy-3′-methoxy-(E)-cinnamoyl]-L- tryptophan, respectively, were detected for the first time in coffee beverages.