Isotopomer measurement techniques in metabolic flux analysis II: mass spectrometry.
Mass spectrometry (MS) offers a sensitive, reliable, and highly accurate method for measurement of isotopic labeling, which is required for generating comprehensive flux maps using metabolic flux analysis (MFA). We present protocols for assessing isotope labeling in a wide range of biochemical species, including proteinogenic amino acids, free organic and amino acids, sugar phosphates, lipids, starch-glucose, and RNA-ribose. We describe the steps of sample preparation, MS analysis, and data handling required to obtain high-quality isotope labeling measurements that are applicable to MFA. By selecting target analytes that maximize identifiability of the key fluxes of interest, MS measurements of isotope labeling can provide a powerful platform for assessing metabolic fluxes in complex biochemical networks.
Vanderbilt University, Nashville, TN, USA.
Jamey D
JD
Young
Douglas K
DK
Allen
John A
JA
Morgan
D000596:Amino Acids / Q000737:chemistry; D005227:Fatty Acids / Q000737:chemistry; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D005947:Glucose / Q000737:chemistry; D006868:Hydrolysis; D007553:Isotope Labeling; D013058:Mass Spectrometry / Q000379:methods*; D064688:Metabolic Flux Analysis / Q000379:methods*; D011506:Proteins / Q000737:chemistry; D013213:Starch / Q000737:chemistry; D053719:Tandem Mass Spectrometry / Q000379:methods
D000596:Amino Acids; D005227:Fatty Acids; D011506:Proteins; D013213:Starch; D005947:Glucose