Plasma citrulline measurement using UPLC tandem mass-spectrometry to determine small intestinal enterocyte pathology.
Citrulline is a nonessential free amino acid, detectable in various biological fluids such as plasma, urine and cerebrospinal fluid. The plasma citrulline concentration is increasingly considered to be a reliable biomarker of enterocyte function. Current analysis usually involves lengthy HPLC separations as a part of classical amino acid profiling, or mass spectrometry usually in combination with derivatization. We employed UPLC-HILIC-tandem mass-spectrometry (MS/MS) of acetonitrile-derived supernatants from plasma samples of control subjects and of patients who had received myeloablative chemotherapy. Detection was achieved by the selected reaction monitoring of transitions: m/z 176-->70 and 180-->74 (for the deuterated standard), respectively. The method was precise and accurate with inter-day CV<3.9% (n=30), recoveries ranging from 98.0 to 100.3% and high linearity from 0.3 to at least 2,000 micromol/L. The results for 202 plasma samples agreed well with those obtained by the classical HPLC-fluorescence method. By a simple protein precipitation/extraction step and the UPLC separation the result can be available within 30 min of receipt with a capacity of at least 12 assays per hour. Citrulline in blood and plasma or serum was stable for at least 2 days at room temperature which would permit postal transport to the laboratory. The UPLC-MS/MS method for measuring plasma citrulline concentrations is fast and robust and is therefore an ideal tool for monitoring the intestinal enterocyte capacity of patients with various pathological conditions.
Demacker
Pierre N M
PN
Department of Clinical Chemistry, Canisius Wilhelmina Hospital, Weg door Jonkerbos 100, 6532 SZ Nijmegen, The Netherlands.
Beijers
Antonius M
AM
van Daal
Henny
H
Donnelly
J Peter
JP
Blijlevens
Nicole M A
NM
van den Ouweland
Johannes M W
JM
D016022:Case-Control Studies; D002853:Chromatography, Liquid / Q000379:methods*; D002956:Citrulline / Q000097:blood*; D006801:Humans; D007421:Intestine, Small / Q000473:pathology*; D012016:Reference Values; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*
D002956:Citrulline