title: Development of a Postcolumn Infused-Internal Standard Liquid Chromatography Mass Spectrometry Method for Quantitative Metabolomics Studies.
abstract: Quantitative metabolomics has become much more important in clinical research in recent years. Individual differences in matrix effects (MEs) and the injection order effect are two major factors that reduce the quantification accuracy in liquid chromatography-electrospray ionization-mass spectrometry-based (LC-ESI-MS) metabolomics studies. This study proposed a postcolumn infused-internal standard (PCI-IS) combined with a matrix normalization factor (MNF) strategy to improve the analytical accuracy of quantitative metabolomics. The PCI-IS combined with the MNF method was applied for a targeted metabolomics study of amino acids (AAs). D8-Phenylalanine was used as the PCI-IS, and it was postcolumn-infused into the ESI interface for calibration purposes. The MNF was used to bridge the AA response in a standard solution with the plasma samples. The MEs caused signal changes that were corrected by dividing the AA signal intensities by the PCI-IS intensities after adjustment with the MNF. After the method validation, we evaluated the method applicability for breast cancer research using 100 plasma samples. The quantification results revealed that the 11 tested AAs exhibit an accuracy between 88.2 and 110.7%. The principal component analysis score plot revealed that the injection order effect can be successfully removed, and most of the within-group variation of the tested AAs decreased after the PCI-IS correction. Finally, targeted metabolomics studies on the AAs showed that tryptophan was expressed more in malignant patients than in the benign group. We anticipate that a similar approach can be applied to other endogenous metabolites to facilitate quantitative metabolomics studies.
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University , Taipei 10051, Taiwan.|The Metabolomics Core Laboratory, Center of Genomic Medicine, National Taiwan University , Taipei 10055, Taiwan.
authors.forename: Hsiao-Wei
authors.initials: HW
authors.lastname: Liao
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University , Taipei 10051, Taiwan.|The Metabolomics Core Laboratory, Center of Genomic Medicine, National Taiwan University , Taipei 10055, Taiwan.
authors.forename: Guan-Yuan
authors.initials: GY
authors.lastname: Chen
authors.affiliation: Department of Internal Medicine, National Taiwan University Hospital , Taipei 10051, Taiwan.
authors.forename: Ming-Shiang
authors.initials: MS
authors.lastname: Wu
authors.affiliation: Department of Internal Medicine, National Taiwan University Hospital , Taipei 10051, Taiwan.
authors.forename: Wei-Chih
authors.initials: WC
authors.lastname: Liao
authors.affiliation: Department of Internal Medicine, National Taiwan University Hospital , Taipei 10051, Taiwan.|Department of Oncology, National Taiwan University Hospital , Taipei 10051, Taiwan.|Oncology Center, National Taiwan University Hospital Hsin-Chu Branch , Hsinchu City 300, Taiwan.
authors.forename: Ching-Hung
authors.initials: CH
authors.lastname: Lin
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University , Taipei 10051, Taiwan.|The Metabolomics Core Laboratory, Center of Genomic Medicine, National Taiwan University , Taipei 10055, Taiwan.|Department of Pharmacy, National Taiwan University Hospital , Taipei 10051, Taiwan.
authors.forename: Ching-Hua
authors.identifier: 0000-0001-5722-0360
authors.initials: CH
authors.lastname: Kuo
mesh_terms: D001943:Breast Neoplasms / Q000097:blood* / Q000473:pathology; D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods / Q000592:standards*; D005260:Female; D017022:Flow Injection Analysis / Q000295:instrumentation; D006801:Humans; D055432:Metabolomics / Q000379:methods / Q000592:standards*; D015588:Observer Variation; D010649:Phenylalanine / Q000097:blood; D025341:Principal Component Analysis; D012015:Reference Standards; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods / Q000592:standards*; D014364:Tryptophan / Q000097:blood
chemical_list: D010649:Phenylalanine; D014364:Tryptophan
keywords: LC−ESI−MS; amino acids; breast cancer; matrix normalization factor (MNF); metabolomics; postcolumn infused-internal standard (PCI-IS)