title: Comparison of different mass spectrometry techniques in the measurement of L-[ring-(13)C6]phenylalanine incorporation into mixed muscle proteins.
abstract: Precise measurement of low enrichment of stable isotope labeled amino-acid tracers in tissue samples is a prerequisite in measuring tissue protein synthesis rates. The challenge of this analysis is augmented when small sample size is a critical factor. Muscle samples from human participants following an 8 h intravenous infusion of L-[ring-(13)C(6)]phenylalanine and a bolus dose of L-[ring-(13)C(6)]phenylalanine in a mouse were utilized. Liquid chromatography tandem mass spectrometry (LC/MS/MS), gas chromatography (GC) MS/MS and GC/MS were compared to the GC-combustion-isotope ratio MS (GC/C/IRMS), to measure mixed muscle protein enrichment of [ring-(13)C(6)]phenylalanine enrichment. The sample isotope enrichment ranged from 0.0091 to 0.1312 molar percent excess. As compared with GC/C/IRMS, LC/MS/MS, GC/MS/MS and GC/MS showed coefficients of determination of R(2)= 0.9962 and R(2) = 0.9942, and 0.9217 respectively. However, the precision of measurements (coefficients of variation) for intra-assay are 13.0%, 1.7%, 6.3% and 13.5% and for inter-assay are 9.2%, 3.2%, 10.2% and 25% for GC/C/IRMS, LC/MS/MS, GC/MS/MS and GC/MS, respectively. The muscle sample sizes required to obtain these results were 8 µg, 0.8 µg, 3 µg and 3 µg for GC/C/IRMS, LC/MS/MS, GC/MS/MS and GC/MS, respectively. We conclude that LC/MS/MS is optimally suited for precise measurements of L-[ring-(13)C(6)]phenylalanine tracer enrichment in low abundance and in small quantity samples.
authors: Division of Endocrinology, Diabetes, Metabolism and Nutrition, Mayo Clinic, 200 First St. SW, 5-194 Joseph, Rochester, MN 55905, USA.
authors: Piotr
authors: P
authors: Zabielski
authors: G Charles
authors: GC
authors: Ford
authors: X Mai
authors: XM
authors: Persson
authors: Abdul
authors: A
authors: Jaleel
authors: Jerry D
authors: JD
authors: Dewey
authors: K Sreekumaran
authors: KS
authors: Nair
mesh_terms: D000818:Animals; D002247:Carbon Isotopes / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D002853:Chromatography, Liquid / Q000379:methods; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D007553:Isotope Labeling; D016014:Linear Models; D008297:Male; D009124:Muscle Proteins / Q000737:chemistry* / Q000378:metabolism; D018482:Muscle, Skeletal / Q000737:chemistry; D010649:Phenylalanine / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D011849:Radioactive Tracers; D051381:Rats; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D002247:Carbon Isotopes; D009124:Muscle Proteins; D011849:Radioactive Tracers; D010649:Phenylalanine
references: Diabetes. 2010 Oct;59(10):2366-74
references: 20622162
references: J Chromatogr B Analyt Technol Biomed Life Sci. 2009 Jan 1;877(1-2):43-9
references: 19036645
references: J Mass Spectrom. 1996 Apr;31(4):367-76
references: 8799283
references: Am J Physiol Endocrinol Metab. 2009 Oct;297(4):E889-97
references: 19654285
references: Am J Physiol Endocrinol Metab. 2006 Oct;291(4):E729-36
references: 16705065
references: Biomed Mass Spectrom. 1985 Aug;12(8):432-6
references: 2931135
references: Am J Physiol Endocrinol Metab. 2001 Feb;280(2):E203-8
references: 11158921
references: Metabolism. 1997 Aug;46(8):943-8
references: 9258279
references: J Clin Invest. 1988 Jul;82(1):198-205
references: 3392207
references: J Clin Endocrinol Metab. 1996 Oct;81(10):3469-75
references: 8855787
references: Clin Sci Mol Med. 1975 Dec;49(6):581-90
references: 1204289
references: Am J Physiol. 1997 Sep;273(3 Pt 1):E564-70
references: 9316447
references: J Lipid Res. 2010 Sep;51(9):2761-5
references: 20526002
references: Am J Physiol Endocrinol Metab. 2006 Jul;291(1):E190-7
references: 16449301
references: Proc Nutr Soc. 1981 Sep;40(3):321-24
references: 7029552
references: Rapid Commun Mass Spectrom. 1992 Jul;6(7):421-4
references: 1638043
references: Anal Biochem. 1994 Aug 15;221(1):72-7
references: 7985807
references: Am J Physiol Endocrinol Metab. 2004 Jan;286(1):E92-101
references: 14506079
references: Am J Physiol Endocrinol Metab. 2008 Nov;295(5):E1255-68
references: 18765679
references: Am J Physiol. 1988 Feb;254(2 Pt 1):E208-13
references: 3279803
references: Am J Physiol. 1994 Aug;267(2 Pt 1):E203-9
references: 8074199
references: Clin Physiol. 1983 Oct;3(5):387-98
references: 6357592
references: J Clin Invest. 1998 May 1;101(9):2000-7
references: 9576765