BACKGROUND: Ultraperformance® Liquid Chromatography (UPLC) is increasingly used for quantitative amino acid screening. The Waters MassTrak™ UPLC Amino Acid Analysis (AAA) Solution kit offers rapid analysis with minimal sample preparation. We describe a simple modification of this method enabling enhanced chromatographic separation of previously problematic analytes with improvements in quantification. METHODS: The commercial UPLC method was compared with our modified version of the same method. The modification incorporates eluent buffer of increased organic content run at reduced column temperature. UPLC methods were compared by analyzing amino acids from 57 plasma samples. A comparison (n=131) between the modified UPLC method and ion-exchange chromatography was also carried out. RESULTS: The commercial method produced a large negative bias for Tyrosine (-22.72%±14.10) and ornithine (-15.02%±10.07). Assay imprecision of Tyrosine using the commercial method (mean Tyrosine: 72.58 and 31.17μmol/l) produced values of 10.86% and 21.12% respectively, compared with the modified method (3.39% and 4.47%). The comparison of modified UPLC and ion-exchange methods was favorable, validating the improvements observed in amino acid quantification. CONCLUSION: The modified UPLC method has eliminated significant bias associated with the commercially available method. The modification is simple, robust and readily adaptable to the current MassTrak™ AAA Solution kit for clinical applications.
BACKGROUND: Ultraperformance® Liquid Chromatography (UPLC) is increasingly used for quantitative amino acid screening. The Waters MassTrak™ UPLC Amino Acid Analysis (AAA) Solution kit offers rapid analysis with minimal sample preparation. We describe a simple modification of this method enabling enhanced chromatographic separation of previously problematic analytes with improvements in quantification. METHODS: The commercial UPLC method was compared with our modified version of the same method. The modification incorporates eluent buffer of increased organic content run at reduced column temperature. UPLC methods were compared by analyzing amino acids from 57 plasma samples. A comparison (n=131) between the modified UPLC method and ion-exchange chromatography was also carried out. RESULTS: The commercial method produced a large negative bias for Tyrosine (-22.72%±14.10) and ornithine (-15.02%±10.07). Assay imprecision of Tyrosine using the commercial method (mean Tyrosine: 72.58 and 31.17μmol/l) produced values of 10.86% and 21.12% respectively, compared with the modified method (3.39% and 4.47%). The comparison of modified UPLC and ion-exchange methods was favorable, validating the improvements observed in amino acid quantification. CONCLUSION: The modified UPLC method has eliminated significant bias associated with the commercially available method. The modification is simple, robust and readily adaptable to the current MassTrak™ AAA Solution kit for clinical applications.
Authors: Kirsten K Ahring; Frederik Dagnæs-Hansen; Annemarie Brüel; Mette Christensen; Erik Jensen; Thomas G Jensen; Mogens Johannsen; Karen S Johansen; Allan M Lund; Jesper G Madsen; Karen Brøndum-Nielsen; Michael Pedersen; Lambert K Sørensen; Mads Kjolby; Lisbeth B Møller Journal: PLoS One Date: 2022-01-11 Impact factor: 3.240