Abstract
Amino acids in human tears play certain physiological roles and their determination is challenging due to complicated chemical properties. This study described a fast and sensitive method for the simultaneous determination of 15 amino acids (AAs) in tears by hydrophilic interaction liquid chromatography and quadrupole orbitrap mass spectrometry (HILIC-Q-Orbitrap-MS). Amino acids in tears were extracted by methanol, and then cleaned up with a solid phase extraction (SPE) cartridge. Chromatographic separation was performed on a 1.7 μm BEH Amide column within 8 min. Tear samples spiked with free AAs were tested in terms of linearity, sensitivity, repeatability, and recovery. Two stable isotope-labeled amino acids were used as internal standards to improve the method performance. Recoveries for all analytes ranged from 89 to 107%. Intra-day and inter-day precision, expressed as relative standard deviations, were all below 10%, and the method detection limits ranged from 0.02 μmol L-1 to 0.11 μmol L-1. The developed method with high throughput and high analyte specificity shows good promise for consistent analysis of free amino acids in tears.