Negative chemical ionization gas chromatography/mass spectrometry to quantify urinary 3-methylhistidine: application to burn injury.
A rapid method for measuring 3-methylhistidine (3MH) in rat and human urine with higher sensitivity and precision than any previously reported method is described using internal standard [1-(13)C]3MH (M+1) and negative chemical ionization (NCI) gas chromatography/mass spectrometry (GC/MS). Internal standard [1-(13)C]3MH (M+1) was added to rat and human urine samples, hydrolyzed, and absorbed onto cation exchange columns. The column eluent was dried and derivatized for GC/MS analysis. Quantification of 3MH levels was accomplished by monitoring the m/z 204 fragment. The m/z 204 fragment was chosen due to the fragment's abundance and stability as determined by analysis of [methyl-(2)H(3), (18)O(2)]3MH (M+7) and [methyl-(13)C]3MH (M+1) fragmentation patterns under NCI conditions. This method shows excellent linearity (0.9989) over the range studied (0-0.5 mol), high recovery (95.9%), and low coefficient of variation (4.7%). The described method is sensitive enough to detect 6.8 pmol amount of urinary 3MH with a precision of 9.1%. The in vivo utility of this method to quantify urinary 3MH was tested in a burn injury rat model and on urine specimens from pediatric burn patients. Data obtained from the urine of burn-injured rats and pediatric burn patients match previously reported trends and validate the in vivo utility of this method.
Beffa
David C
DC
Shriners Burns Hospital, Boston, MA 02114, USA. dbeffa@uic.edu
Carter
Edward A
EA
Lu
Xiao-Ming
XM
Yu
Yong-Ming
YM
Prelack
Kathrina
K
Sheridan
Robert L
RL
Young
Vernon R
VR
Fischman
Alan J
AJ
Tompkins
Ronald G
RG
D000293:Adolescent; D000818:Animals; D002056:Burns / Q000652:urine*; D002648:Child; D002675:Child, Preschool; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D007223:Infant; D008762:Methylhistidines / Q000592:standards / Q000652:urine*; D015394:Molecular Structure; D051381:Rats; D012015:Reference Standards; D015203:Reproducibility of Results
D008762:Methylhistidines