Assessment of thymidine phosphorylase function: measurement of plasma thymidine (and deoxyuridine) and thymidine phosphorylase activity

Methods Mol Biol. 2012;837:121-33. doi: 10.1007/978-1-61779-504-6_8.


We describe detailed methods to measure thymidine (dThd) and deoxyuridine (dUrd) concentrations and thymidine phosphorylase (TP) activity in biological samples. These protocols allow the detection of TP dysfunction in patients with mitochondrial neurogastrointestinal encephalomyopathy (MNGIE). Since the identification of mutations in TYMP, the gene encoding TP, as the cause of MNGIE (Nishino et al. Science 283:689-692, 1999), the assessment of TP dysfunction has become the best screening method to rule out or confirm MNGIE in patients. TYMP sequencing, to find the causative mutations, is only needed when TP dysfunction is detected. dThd and dUrd are measured by resolving these compounds with high-performance liquid chromatography (HPLC) followed by the spectrophotometric monitoring of the eluate absorbance at 267 nm (HPLC-UV). TP activity can be measured by an endpoint determination of the thymine formed after 1 h incubation of the buffy coat homogenate in the presence of a large excess of its substrate dThd, either spectrophotometrically or by HPLC-UV.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Analytic Sample Preparation Methods
  • Deoxyuridine / blood*
  • Deoxyuridine / urine
  • Enzyme Assays / methods*
  • Humans
  • Intestinal Pseudo-Obstruction / blood
  • Intestinal Pseudo-Obstruction / enzymology
  • Intestinal Pseudo-Obstruction / urine
  • Mitochondrial Encephalomyopathies / blood
  • Mitochondrial Encephalomyopathies / enzymology
  • Mitochondrial Encephalomyopathies / urine
  • Muscular Dystrophy, Oculopharyngeal
  • Ophthalmoplegia / congenital
  • Thymidine / blood*
  • Thymidine / urine
  • Thymidine Phosphorylase / blood*
  • Thymidine Phosphorylase / metabolism*


  • Thymidine Phosphorylase
  • Thymidine
  • Deoxyuridine

Supplementary concepts

  • Visceral myopathy familial external ophthalmoplegia