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Bioanalysis. 2014 Mar;6(6):745-59. doi: 10.4155/bio.13.323.

A clinical biomarker assay for the quantification of d3-creatinine and creatinine using LC-MS/MS.

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GlaxoSmithKline, Research Triangle Park, NC 27709, USA.



Current methods to measure skeletal muscle mass are not practical in a clinical setting. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods that measure the amount of urinary d3-creatinine enrichment after a single tracer dose of d3-creatine was developed.


The biomarkers d3-creatinine and creatinine were detected in human urine using LC-MS/MS. In this assay a surrogate analyte, d3-creatinine, was used to quantify endogenous creatinine. However, since endogenous concentrations of creatinine were orders of magnitude higher than d3-creatinine, the peak area of a less intense isotope of creatinine was acquired. A response factor is used to correct for using a less intense isotope multiple reaction monitoring transition.


Novel LC-MS/MS assays were developed that quantify the biomarkers d3-creatinine, creatinine and d3-creatine in urine. This method allows the estimation of total body creatine pool size and subsequent calculation of muscle mass. This assay was originally validated as fit-for-purpose and was followed by full validation.

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