Chiral analysis of aromatic amino acids in food supplements using subcritical fluid chromatography and Chirobiotic T2 column

Abstract

The enantiomeric recognition capability of the teicoplanin-based Chirobiotic T2 column for the analysis of amino acids using subcritical fluid chromatography is presented in this work. The chiral separation of the aromatic protein amino acids phenylalanine, tyrosine and tryptophan was studied employing an elevated percentage of organic modifier (ranging from 35 to 60% v/v), and a high percentage of water as additive (2–10%, v/v). Baseline enantioseparation of the individual amino acids was achieved in 7min (in 15min for their simultaneous separation by serially coupling of chiral/achiral columns) using 40% of modifier composed of a mixture methanol/water (90:10 v/v) without the necessity of other acidic or basic additives. The proposed method was validated obtaining suitable levels of recovery, precision and linearity. The detection and quantification limits ranged from 0.5 to 2.0μg/mL and from 1.7 to 6.7μg/mL, respectively, which allowed the determination of low amounts of d-amino acids (up to 0.2% for tryptophan) in the presence of a large excess of corresponding l-enantiomers. Finally, the method was applied to the determination of l-phenylalanine, l-tyrosine and l-tryptophan in five commercial food supplements, and the quality of these food supplements was confirmed through the absence of enantiomeric impurity in all of them.