HPLC MEASUREMENT OF PURINE NUCLEOTIDE DEGRADATION PRODUCTS IN AMNIOTIC FLUID AND ITS USE IN PRENATAL DIAGNOSIS

Abstract

We describe a convenient method for the separation and quantification of xanthine, hypoxanthine, and uric acid in 20 μL samples of amniotic fluid using HPLC and UV detection. The analysis is performed on a Sepharon SGX C₁₈ column and the elution system consists of a 60-mmol/L potassium phosphate buffer, pH 4.6. The limit of detection is 7 pmol for hypoxanthine and xanthine and 10 pmol for uric acid. The intra- and interassay coefficients of variation are less than 5%. The applicability of the method is illustrated with the determination of micromolar concentrations of oxopurines in amniotic fluid samples obtained from 204 trimester pregnancies with an apparent increased risk of chromosome disorders.

Keywords:

• HPLC
• Amniotic fluid
• Xanthine
• Hypoxanthine
• Uric acid
• Oxopurines

ACKNOWLEDGMENTS

This study was supported by a grant (N^o 1/4132/97) of the Ministry of Education of the Slovak Republic.