Microscale analysis of amino acids using gas chromatography-mass spectrometry after methyl chloroformate derivatization.
To conduct studies of stable isotope incorporation and dilution in growing plants, a rapid microscale method for determination of amino acid profiles from minute amounts of plant samples was developed. The method involves solid-phase ion exchange followed by derivatization and analysis by gas chromatography-mass spectrometry (GC-MS). The procedure allowed the eluent to be derivatized directly with methyl chloroformate without sample lyophilization or other evaporation procedures. Sample extraction and derivatization required only ca. 30min and quantification of the 19 amino acids eluted from the cation exchange solid-phase extraction step from a single cotyledon (0.4mg fresh weight) or three etiolated 7-day-old Arabidopsis seedlings (0.1mg fresh weight) was easily accomplished in the selected ion monitoring mode. This method was especially useful for monitoring mass isotopic distribution of amino acids as illustrated by Arabidopsis seedlings that had been labeled with deuterium oxide and (15)N salts. Sample preparation was facile, rapid, economical, and the method is easily modified for integration into robotic systems for analysis with large numbers of samples.
Chen
Wen-Ping
WP
Department of Horticultural Science, University of Minnesota, Saint Paul, MN 55108, United States. chenx071@umn.edu
Yang
Xiao-Yuan
XY
Hegeman
Adrian D
AD
Gray
William M
WM
Cohen
Jerry D
JD
D000596:Amino Acids / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D017360:Arabidopsis / Q000737:chemistry / Q000378:metabolism; D002138:Calibration; D016009:Chi-Square Distribution; D018548:Cotyledon / Q000737:chemistry / Q000378:metabolism; D017666:Deuterium Oxide; D005561:Formates / Q000737:chemistry* / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D007553:Isotope Labeling / Q000379:methods; D009587:Nitrogen Isotopes; D055550:Protein Stability; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D036226:Seedlings / Q000737:chemistry / Q000378:metabolism; D012680:Sensitivity and Specificity; D052616:Solid Phase Extraction
D000596:Amino Acids; D005561:Formates; D009587:Nitrogen Isotopes; D017666:Deuterium Oxide; C014667:methyl chloroformate