Measurement of muscle protein fractional synthetic rate by capillary gas chromatography/combustion isotope ratio mass spectrometry.
The measurement of skeletal muscle protein fractional synthetic rate using an infusion of (1-13C)leucine and measuring the isotopic abundance of the tracer in skeletal muscle protein by preparative gas chromatography (GC)/ninhydrin isotope ratio mass spectrometry (IRMS) is laborious and subject to errors owing to contamination by 12C. The purpose of this study was to compare muscle (13C)leucine enrichment measured with the conventional preparative GC/ninhydrin IRMS approach to a new, continuous-flow technique using capillary GC/combustion IRMS. Quadriceps muscles were removed from four Sprague-Dawley rats after each was infused at a different rate with (1-13C)leucine for 6-8 h. Muscle leucine enrichment (at. % excess) measured by both methods differed by less than 4%, except at low (13C)leucine enrichments (less than 0.03 at. % excess). In addition, capillary GC/combustion IRMS was used to assess muscle (13C)leucine enrichment and fractional muscle protein synthesis rate in ten normal young men and women infused with (1,2-13C2)leucine for 12-14 h. This approach reduced the variability of the isotope abundance measure and gave estimates of muscle protein synthesis rate (0.050 +/- 0.011% h-1 (mean +/- SEM); range = 0.023-0.147% h-1) that agree with published values determined using the standard analytical approach. The measurement of (13C)leucine enrichment from skeletal muscle protein by capillary GC/combustion IRMS provides a simple, acceptable and practical alternative to preparative GC/ninhydrin IRMS.
Yarasheski
K E
KE
Metabolism Division, Washington University School of Medicine, St Louis, Missouri 63110.
Smith
K
K
Rennie
M J
MJ
Bier
D M
DM
D000328:Adult; D000818:Animals; D002849:Chromatography, Gas; D005260:Female; D006801:Humans; D007930:Leucine / Q000737:chemistry; D008297:Male; D013058:Mass Spectrometry; D009124:Muscle Proteins / Q000032:analysis / Q000096:biosynthesis*; D009132:Muscles / Q000737:chemistry / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
D009124:Muscle Proteins; D007930:Leucine