Mass spectrometry assay method for detection and quantitation of kidney function metabolites
A method for determining in a sample, by mass spectrometry, the amount of one or more analytes selected from the group consisting of N-acetylthreonine, TMAP, phenylacetylglutamine, tryptophan, creatinine, meso-erythritol, arabitol, myo-inositol, N-acetyl serine, N-acetylalanine, 3-methylhistidine, trans-4-hydroxyproline, kynurenine, urea, C-glycosyltryptophan, 3-indoxyl sulfate, pseudouridine, and combinations thereof is described. The method comprises subjecting the sample to an ionization source under conditions suitable to produce one or more ions detectable by mass spectrometry from each of the one or more of the analytes; measuring, by mass spectrometry, the amount of the one or more ions from each of the one or more analytes; and using the measured amount of the one or more ions to determine the amount of each of the one or more analytes in the sample. Also described is a kit comprising one or more isotopically labeled analogues as internal standards for each of the one or more analytes.
1. A method for detecting and determining in a sample the amount of five or more kidney function analytes, the method comprising:
a) purifying the sample by liquid chromatography;
b) subjecting the purified sample to an ionization source under conditions suitable to produce one or more ions detectable by mass spectrometry from each of the five or more of the analytes selected from the group consisting of N-acetylthreonine, TMAP, phenylacetylglutamine, tryptophan, creatinine, N-acetylalanine, 3-methylhistidine, trans-4-hydroxyproline, kynurenine, urea, and pseudouridine, wherein the analytes are not derivatized prior to ionization;
c) measuring, by tandem mass spectrometry operated in positive mode in a single injection, the amount of the one or more ions from each of the five or more analytes, wherein the one or more ions used to determine the amount of each of the five or more analytes is one or more ions selected from the ions in Tables 3 and 5; wherein an eluent from the liquid chromatography step is directly and automatically introduced into an electrospray source of the mass spectrometer; and
d) using the measured amount of the one or more ions to determine the amount of each of the five or more analytes in the sample.
2. The method of claim 1 , wherein the amount of six or more analytes is determined.
3. The method of claim 1 , wherein said liquid chromatography is selected from the group consisting of high performance liquid chromatography, ultra high performance liquid chromatography, and turbulent flow liquid chromatography.
4. The method of claim 1 , wherein an internal standard is used to determine the amount of the five or more analytes in the sample.
5. The method of claim 4 , wherein the internal standard comprises an isotopically labeled analogue of at least one of the five or more analytes to be measured.
6. The method of claim 1 , wherein the run time is 7 minutes or less.
7. The method of claim 6 , wherein the run time is 4 minutes or less.
8. The method of claim 3 , wherein said ultra high performance liquid chromatography is conducted using hydrophilic interaction chromatography (HILIC).
9. The method of claim 1 , wherein five of the five or more analytes comprise N-acetylthreonine, phenylacetylglutamine, tryptophan, creatinine, and pseudouridine.
10. The method of claim 9 , wherein the coefficient of variation over 40 samples is equal to or less than 6.5% for each of the five analytes when the sample is plasma.
11. The method of claim 1 , wherein nine of the five or more analytes comprise N-acetylthreonine, phenylacetylglutamine, tryptophan, creatinine, N-acetylalanine, 3-methylhistidine, trans-4-hydroxyproline, kynurenine, and urea.
12. The method of claim 11 , wherein the coefficient of variation over 12 samples is equal to or less than 14.5% for each of the nine analytes when the sample is plasma.