Precise determination of glucose-d(2)/glucose ratio in human serum and plasma by APCI LC-MS/MS.
BACKGROUND
Determinations of glucose turnover based on infusion of deuterium-labelled glucose are frequently undertaken in studies involving the pathophysiology of diabetes or obesity or the metabolic response to physical exercise.


METHODS
A liquid chromatography tandem mass spectrometry (LC-MS/MS) method was developed for determination of the glucose-d(2)/glucose ratio in human serum and plasma following the intra-venous infusion of 6,6-d(2)-glucose. Atmospheric pressure chemical ionization measuring negative ions in selected reaction monitoring mode (product ions at m/z 179-->89 and m/z 181-->89) was used. The serum or plasma samples (50 microL) were prepared by protein precipitation with acetonitrile and the resulting supernatant was directly used for the LC-MS/MS analysis. The chromatography was performed on a Luna 3 micron NH2 100A column (100 x 2 mm) using a mobile phase containing 85% acetonitrile with 20 mmol/L of ammonium acetate at a flow rate of 400 microL/min and an oven temperature of 40 degrees C.


RESULTS
A linear response was obtained for the glucose-d(2)/glucose ratio over the relative concentration range of 0-10%, r(2)>0.999. The peak area ratio was determined with an imprecision of 1.20-8.19% (coefficient of variation). The ion suppression from matrix compared to water was in the order of 55%. Chromatographic retention time was between 4 and 5 min and the total analysis time was 10 min. The validated method was successfully applied for the analysis of human serum samples from a clinical study involving infusion of 6,6-d(2)-glucose and evaluation of endogenous glucose production.


CONCLUSIONS
It is concluded that the described method provides an easy and precise technique for the determination of serum and plasma glucose-d(2)/glucose ratios in clinical studies.
Ullah
Shahid
S
Department of Medicine, Division of Clinical Pharmacology, Karolinska Institute and University Hospital, Stockholm, Sweden.
Wahren
John
J
Beck
Olof
O
D001274:Atmospheric Pressure; D001786:Blood Glucose / Q000032:analysis* / Q000737:chemistry; D002853:Chromatography, Liquid / Q000379:methods*; D003903:Deuterium / Q000032:analysis*; D005615:Freezing; D006801:Humans; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D011786:Quality Control; D015203:Reproducibility of Results; D012996:Solutions; D055815:Young Adult
D001786:Blood Glucose; D012996:Solutions; D003903:Deuterium