Liquid chromatography-tandem mass spectrometry method for mycophenolic acid and its glucuronide determination in saliva samples from children with nephrotic syndrome

BackgroundSaliva sampling is one of the methods of therapeutic drug monitoring for mycophenolic acid (MPA) and its metabolite, mycophenolic acid glucuronide (MPAG). The study describes the liquid chromatography tandem mass spectrometry (LC-MS/MS) method developed for saliva MPA and MPAG determination in children with nephrotic syndrome.MethodsThe mobile phase consisted of methanol and water at gradient flow, both with 0.1% formic acid. Firstly, 100 mu L of saliva was evaporated at 45 degrees C for 2 h to dryness, secondly, it was reconstituted in the mobile phase, and finally 10 mu L was injected into the LC-MS/MS system. Saliva from ten children with nephrotic syndrome treated with mycophenolate mofetil was collected with Salivette (R).ResultsFor MPA and MPAG, within the 2-500 ng/mL range, the method was selective, specific, accurate and precise within-run and between-run. No carry-over and matrix effects were observed. Stability tests showed that MPA and MPAG were stable in saliva samples if stored for 2 h at room temperature, 18 h at 4 degrees C, and at least 5 months at - 80 degrees C as well as after three freeze-thaw cycles, in a dry extract for 16 h at 4 degrees C, and for 8 h at 15 degrees C in the autosampler. The analytes were not adsorbed onto Salivette (R) cotton swabs. For concentrations above 500 ng/mL, the samples may be diluted twofold. In children, saliva MPA and MPAG were within the ranges of 4.6-531.8 ng/mL and 10.7-183.7 ng/mL, respectively.ConclusionsThe evaluated LC-MS/MS method has met the validation requirements for saliva MPA and MPAG determination in children with nephrotic syndrome. Further studies are needed to explore plasma-saliva correlations and assess their potential contribution to MPA monitoring.