Simultaneous quantification of labeled 2H5-glycerol, 13C6-glucose, and endogenous D-glucose in mouse plasma using liquid chromatography tandem mass spectrometry
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Monitoring the level of glucose and glycerol or their labeled derivatives in biological fluid for kinetic studies has always been challenging, especially in mice, because of the limited volume in addition to the complexity of plasma. For such application, we developed a simple, fast, and sensitive method for the simultaneous measurement of absolute concentrations of labeled 2H5-glycerol and 13C6-glucose as well as endogenous D-glucose using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). In our study, 15.0 μL of mouse plasma was processed by a one-step protein precipitation, followed by LC-MS/MS analysis. The quantification of the analytes was carried out by monitoring the product ion scan of their corresponding deprotonated molecular ions and constructing the extracted ion fragmentogram by choosing a specific product ion for each analyte (equivalent to precursor ion to product ion transitions). The limit of detection (LOD) was evaluated to be 1.0 μM for both 2H5-glycerol and 13C6-glucose, and the limit of quantitation (LOQ) was observed to be 5.0 μM for both 2H5-glycerol and 13C6-glucose in diluted mice plasma that corresponds to 50 μM in plasma or 4.60 and 9.01 mg/dL of glycerol and glucose in plasma, respectively. The extraction recoveries are 81.9 % (CV = 8.1 %) for 2H5-glycerol and 26.2 % (CV = 13.6 %) for 13C6-glucose.
KeywordsLC-MS/MS Glucose Glycerol Stable isotope tracer Plasma Mouse
This work was supported by the National Institutes of Health NS061777. The authors thank Ms Georgia Dolios for proofreading the manuscript.
Conflict of interest
The authors declare that they have no conflict of interest.
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