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Development and validation of an ultra-performance liquid chromatography quadrupole time of flight mass spectrometry method for rapid quantification of free amino acids in human urine
Journal article   Open access   Peer reviewed

Development and validation of an ultra-performance liquid chromatography quadrupole time of flight mass spectrometry method for rapid quantification of free amino acids in human urine

Richard Joyce, Viktorija Kuziene, Xin Zou, Xueting Wang, Frank Pullen and Ruey Leng Loo
Amino acids, Vol.48(1), pp.219-234
2016
DOI: https://doi.org/10.1007/s00726-015-2076-0
PMCID: PMC4710665
PMID: 26319643
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Published (Version of Record)CC BY V4.0,  Open Access

Abstract

Absolute quantification Free amino acids HILIC-UPLC-qTOF-MS Human urine
An ultra-performance liquid chromatography quadrupole time of flight mass spectrometry (UPLC-qTOF-MS) method using hydrophilic interaction liquid chromatography was developed and validated for simultaneous quantification of 18 free amino acids in urine with a total acquisition time including the column re-equilibration of less than 18 min per sample. This method involves simple sample preparation steps which consisted of 15 times dilution with acetonitrile to give a final composition of 25 % aqueous and 75 % acetonitrile without the need of any derivatization. The dynamic range for our calibration curve is approximately two orders of magnitude (120-fold from the lowest calibration curve point) with good linearity (r 2 ≥ 0.995 for all amino acids). Good separation of all amino acids as well as good intra- and inter-day accuracy (<15 %) and precision (<15 %) were observed using three quality control samples at a concentration of low, medium and high range of the calibration curve. The limits of detection (LOD) and lower limit of quantification of our method were ranging from approximately 1–300 nM and 0.01–0.5 µM, respectively. The stability of amino acids in the prepared urine samples was found to be stable for 72 h at 4 °C, after one freeze thaw cycle and for up to 4 weeks at −80 °C. We have applied this method to quantify the content of 18 free amino acids in 646 urine samples from a dietary intervention study. We were able to quantify all 18 free amino acids in these urine samples, if they were present at a level above the LOD. We found our method to be reproducible (accuracy and precision were typically <10 % for QCL, QCM and QCH) and the relatively high sample throughput nature of this method potentially makes it a suitable alternative for the analysis of urine samples in clinical setting.

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Collaboration types
Domestic collaboration
Citation topics
2 Chemistry
2.211 Mass Spectrometry
2.211.990 Metabolomics
Web Of Science research areas
Biochemistry & Molecular Biology
ESI research areas
Biology & Biochemistry
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