Comprehensive detection of disorders of purine and pyrimidine metabolism by HPLC with electrospray ionization tandem mass spectrometry.
BACKGROUND
Clinical presentation and disease severity in disorders of purine and pyrimidine metabolism vary considerably. We present a method that allows comprehensive, sensitive, and specific diagnosis of the entire spectrum of abnormalities in purine and pyrimidine metabolism in 1 analytical run.


METHODS
We used reversed-phase HPLC electrospray ionization tandem mass spectrometry to investigate 24 metabolites of purine and pyrimidine metabolism in urine samples from healthy persons and from patients with confirmed diagnoses of inherited metabolic disorders. Urine samples were filtered and diluted to a creatinine concentration of 0.5 mmol/L. Stable-isotope-labeled internal standards were used for quantification. The metabolites were analyzed by multiple-reaction monitoring in positive and negative ionization modes.


RESULTS
Total time of analysis was 20 min. Recovery (n = 8) of a compound after addition of a known concentration was 85%-133%. The mean intraday variation (n = 10) was 12%. The interday variation (n = 7) was < or =17%. Age-related reference intervals were established for each compound. Analysis of patient urine samples revealed major differences in tandem mass spectrometry profiles compared with those of control samples. Twelve deficiencies were reliably detected: hypoxanthine guanine phosphoribosyl transferase, xanthine dehydrogenase, purine nucleoside phosphorylase, adenylosuccinate lyase, uridine monophosphate synthase, adenosine deaminase, adenine phosphoribosyl transferase, molybdenum cofactor, thymidine phosphorylase, dihydropyrimidine dehydrogenase, dihydropyrimidinase, and beta-ureidopropionase.


CONCLUSIONS
This method enables reliable detection of 13 defects in purine and pyrimidine metabolism in a single analytical run.
Hartmann
Susen
S
Division of Metabolic Diseases, Department of General Pediatrics, University Children's Hospital Heidelberg, Heidelberg, Germany. Susen.Hartmann@med.uni-heidelberg.de
Okun
Jürgen G
JG
Schmidt
Christiane
C
Langhans
Claus-Dieter
CD
Garbade
Sven F
SF
Burgard
Peter
P
Haas
Dorothea
D
Sass
Jörn Oliver
JO
Nyhan
William L
WL
Hoffmann
Georg F
GF
D000293:Adolescent; D000367:Age Factors; D002648:Child; D002675:Child, Preschool; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D007223:Infant; D011686:Purine-Pyrimidine Metabolism, Inborn Errors / Q000175:diagnosis* / Q000652:urine; D011687:Purines / Q000652:urine*; D011743:Pyrimidines / Q000652:urine*; D012016:Reference Values; D021241:Spectrometry, Mass, Electrospray Ionization
D011687:Purines; D011743:Pyrimidines