Defects in pyrimidine degradation identified by HPLC-electrospray tandem mass spectrometry of urine specimens or urine-soaked filter paper strips

H. Van Lenthe, A. B.P. Van Kuilenburg, Tetsuya Ito, A. H. Bootsma, A. Van Cruchten, Y. Wada, A. H. Van Gennip

Research output: Contribution to journalArticle

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Abstract

Background: Urinary concentrations of thymine, uracil, and their degradation products are useful indicators of deficiencies of enzymes of the pyrimidine degradation pathway. We describe a rapid, specific method to measure these concentrations to detect inborn errors of pyrimidine metabolism. Methods: We used urine or urine-soaked filter-paper strips as samples and measured thymine, uracil and their degradation products dihydrothymine, dihydrouracil, N-carbamyl-β-aminoisobutyric acid and N-carbamyl-β-alanine. Reversed-phase HPLC was combined with electrospray ionization tandem mass spectrometry, and detection was performed by multiple-reaction monitoring. Stable-isotope-labeled reference compounds were used as internal standards. Results: All pyrimidine degradation products could be measured in one analytical run of 15 min. Detection limits were 0.4-4 μmol/L. The intraassay imprecision (CV) of urine samples with added compounds was 1.3-12% for liquid urines and 1.0-10% for filter-paper extracts of the urines. The interassay imprecision (CV) was 3-11% (100-200 μmol/L). Recoveries were 89-99% at 100-200 μmol/L and 95-106% at 1 mmol/L in liquid urines and 93-103% at 100-200 μmol/L and 100-106% at 1 mmol/L in filter-paper samples. Correct identifications of deficiencies of the pyrimidine-degrading enzymes were readily made with urine samples from patients with known defects. Conclusions: HPLC with electrospray ionization tandem mass spectrometry allows rapid testing for disorders of the pyrimidine degradation pathway and filter-paper samples allow easy collection, transport and storage of urine samples. (C) 2000 American Association for Clinical Chemistry.

Original languageEnglish
Pages (from-to)1916-1922
Number of pages7
JournalClinical Chemistry
Volume46
Issue number12
Publication statusPublished - 27-12-2000
Externally publishedYes

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Tandem Mass Spectrometry
Mass spectrometry
High Pressure Liquid Chromatography
Urine
Degradation
Defects
Electrospray ionization
Thymine
Uracil
Electrospray Ionization Mass Spectrometry
Aminoisobutyric Acids
Liquids
Enzymes
Metabolism
Isotopes
Alanine
Inborn Errors Metabolism
pyrimidine
Recovery
Monitoring

All Science Journal Classification (ASJC) codes

  • Clinical Biochemistry
  • Biochemistry, medical

Cite this

Van Lenthe, H., Van Kuilenburg, A. B. P., Ito, T., Bootsma, A. H., Van Cruchten, A., Wada, Y., & Van Gennip, A. H. (2000). Defects in pyrimidine degradation identified by HPLC-electrospray tandem mass spectrometry of urine specimens or urine-soaked filter paper strips. Clinical Chemistry, 46(12), 1916-1922.
Van Lenthe, H. ; Van Kuilenburg, A. B.P. ; Ito, Tetsuya ; Bootsma, A. H. ; Van Cruchten, A. ; Wada, Y. ; Van Gennip, A. H. / Defects in pyrimidine degradation identified by HPLC-electrospray tandem mass spectrometry of urine specimens or urine-soaked filter paper strips. In: Clinical Chemistry. 2000 ; Vol. 46, No. 12. pp. 1916-1922.
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abstract = "Background: Urinary concentrations of thymine, uracil, and their degradation products are useful indicators of deficiencies of enzymes of the pyrimidine degradation pathway. We describe a rapid, specific method to measure these concentrations to detect inborn errors of pyrimidine metabolism. Methods: We used urine or urine-soaked filter-paper strips as samples and measured thymine, uracil and their degradation products dihydrothymine, dihydrouracil, N-carbamyl-β-aminoisobutyric acid and N-carbamyl-β-alanine. Reversed-phase HPLC was combined with electrospray ionization tandem mass spectrometry, and detection was performed by multiple-reaction monitoring. Stable-isotope-labeled reference compounds were used as internal standards. Results: All pyrimidine degradation products could be measured in one analytical run of 15 min. Detection limits were 0.4-4 μmol/L. The intraassay imprecision (CV) of urine samples with added compounds was 1.3-12{\%} for liquid urines and 1.0-10{\%} for filter-paper extracts of the urines. The interassay imprecision (CV) was 3-11{\%} (100-200 μmol/L). Recoveries were 89-99{\%} at 100-200 μmol/L and 95-106{\%} at 1 mmol/L in liquid urines and 93-103{\%} at 100-200 μmol/L and 100-106{\%} at 1 mmol/L in filter-paper samples. Correct identifications of deficiencies of the pyrimidine-degrading enzymes were readily made with urine samples from patients with known defects. Conclusions: HPLC with electrospray ionization tandem mass spectrometry allows rapid testing for disorders of the pyrimidine degradation pathway and filter-paper samples allow easy collection, transport and storage of urine samples. (C) 2000 American Association for Clinical Chemistry.",
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Van Lenthe, H, Van Kuilenburg, ABP, Ito, T, Bootsma, AH, Van Cruchten, A, Wada, Y & Van Gennip, AH 2000, 'Defects in pyrimidine degradation identified by HPLC-electrospray tandem mass spectrometry of urine specimens or urine-soaked filter paper strips', Clinical Chemistry, vol. 46, no. 12, pp. 1916-1922.

Defects in pyrimidine degradation identified by HPLC-electrospray tandem mass spectrometry of urine specimens or urine-soaked filter paper strips. / Van Lenthe, H.; Van Kuilenburg, A. B.P.; Ito, Tetsuya; Bootsma, A. H.; Van Cruchten, A.; Wada, Y.; Van Gennip, A. H.

In: Clinical Chemistry, Vol. 46, No. 12, 27.12.2000, p. 1916-1922.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Defects in pyrimidine degradation identified by HPLC-electrospray tandem mass spectrometry of urine specimens or urine-soaked filter paper strips

AU - Van Lenthe, H.

AU - Van Kuilenburg, A. B.P.

AU - Ito, Tetsuya

AU - Bootsma, A. H.

AU - Van Cruchten, A.

AU - Wada, Y.

AU - Van Gennip, A. H.

PY - 2000/12/27

Y1 - 2000/12/27

N2 - Background: Urinary concentrations of thymine, uracil, and their degradation products are useful indicators of deficiencies of enzymes of the pyrimidine degradation pathway. We describe a rapid, specific method to measure these concentrations to detect inborn errors of pyrimidine metabolism. Methods: We used urine or urine-soaked filter-paper strips as samples and measured thymine, uracil and their degradation products dihydrothymine, dihydrouracil, N-carbamyl-β-aminoisobutyric acid and N-carbamyl-β-alanine. Reversed-phase HPLC was combined with electrospray ionization tandem mass spectrometry, and detection was performed by multiple-reaction monitoring. Stable-isotope-labeled reference compounds were used as internal standards. Results: All pyrimidine degradation products could be measured in one analytical run of 15 min. Detection limits were 0.4-4 μmol/L. The intraassay imprecision (CV) of urine samples with added compounds was 1.3-12% for liquid urines and 1.0-10% for filter-paper extracts of the urines. The interassay imprecision (CV) was 3-11% (100-200 μmol/L). Recoveries were 89-99% at 100-200 μmol/L and 95-106% at 1 mmol/L in liquid urines and 93-103% at 100-200 μmol/L and 100-106% at 1 mmol/L in filter-paper samples. Correct identifications of deficiencies of the pyrimidine-degrading enzymes were readily made with urine samples from patients with known defects. Conclusions: HPLC with electrospray ionization tandem mass spectrometry allows rapid testing for disorders of the pyrimidine degradation pathway and filter-paper samples allow easy collection, transport and storage of urine samples. (C) 2000 American Association for Clinical Chemistry.

AB - Background: Urinary concentrations of thymine, uracil, and their degradation products are useful indicators of deficiencies of enzymes of the pyrimidine degradation pathway. We describe a rapid, specific method to measure these concentrations to detect inborn errors of pyrimidine metabolism. Methods: We used urine or urine-soaked filter-paper strips as samples and measured thymine, uracil and their degradation products dihydrothymine, dihydrouracil, N-carbamyl-β-aminoisobutyric acid and N-carbamyl-β-alanine. Reversed-phase HPLC was combined with electrospray ionization tandem mass spectrometry, and detection was performed by multiple-reaction monitoring. Stable-isotope-labeled reference compounds were used as internal standards. Results: All pyrimidine degradation products could be measured in one analytical run of 15 min. Detection limits were 0.4-4 μmol/L. The intraassay imprecision (CV) of urine samples with added compounds was 1.3-12% for liquid urines and 1.0-10% for filter-paper extracts of the urines. The interassay imprecision (CV) was 3-11% (100-200 μmol/L). Recoveries were 89-99% at 100-200 μmol/L and 95-106% at 1 mmol/L in liquid urines and 93-103% at 100-200 μmol/L and 100-106% at 1 mmol/L in filter-paper samples. Correct identifications of deficiencies of the pyrimidine-degrading enzymes were readily made with urine samples from patients with known defects. Conclusions: HPLC with electrospray ionization tandem mass spectrometry allows rapid testing for disorders of the pyrimidine degradation pathway and filter-paper samples allow easy collection, transport and storage of urine samples. (C) 2000 American Association for Clinical Chemistry.

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M3 - Article

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VL - 46

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