An optimized procedure for metabonomic analysis of rat liver tissue using gas chromatography/time-of-flight mass spectrometry.
In this paper, we present a tissue metabonomic method with an optimized extraction procedure followed by instrumental analysis with gas chromatography/time-of-flight mass spectrometry (GC/TOFMS) and spectral data analysis with multivariate statistics. Metabolite extractions were carried out using three solvents: chloroform, methanol, and water, with design of experiment (DOE) theory and multivariate statistical analysis. A two-step metabolite extraction procedure was optimized using a mixed solvent of chloroform-methanol-water (1:2:1, v/v/v) and then followed by methanol alone. This approach was subsequently validated using standard compounds and liver tissues. Calibration curves were obtained in the range of 0.50-125.0mug/mL for standards and 0.02-0.25g/mL acceptable for liver tissue samples. For most of the metabolites investigated, relative standard deviations (RSD) were below 10% within a day (reproducibility) and below 15% within a week (stability). Rat liver tissues of carbon tetrachloride-induced acute liver injury models (n=10) and healthy control rats (n=10) were analyzed which demonstrated the applicability of the developed procedure for the tissue metabonomic study.
Pan
Li
L
Shanghai Center for Systems Biomedicine, College of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, PR China.
Qiu
Yunping
Y
Chen
Tianlu
T
Lin
Jinchao
J
Chi
Yi
Y
Su
Mingming
M
Zhao
Aihua
A
Jia
Wei
W
D000818:Animals; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D008099:Liver / Q000737:chemistry* / Q000378:metabolism*; D008297:Male; D055432:Metabolomics / Q000379:methods; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley