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Rapid determination of the hypoxanthine increase in ischemic exercise tests
P A Bolhuis1, R Zwart, P R Bär
1Department of Neurology, Academic Medical Centre, Amsterdam, The Netherlands.
Clinical Chemistry
|August 1, 1988
Summary
A new enzymatic method offers a faster alternative for detecting hypoxanthine, aiding in the diagnosis of myoadenylate deaminase deficiency. This method simplifies the process compared to traditional HPLC analysis.
Area of Science:
- Biochemistry
- Enzymology
- Clinical Diagnostics
Background:
- Ischemic exercise tests are used to diagnose glycogenoses and myoadenylate deaminase deficiency.
- Serum lactate and ammonia levels are commonly measured, but hypoxanthine determination is an alternative for myoadenylate deaminase deficiency.
- High-performance liquid chromatography (HPLC) for hypoxanthine is effective but time-consuming.
Purpose of the Study:
- To develop a rapid, indirect enzymatic equilibrium method for quantifying hypoxanthine.
- To establish an alternative diagnostic tool for myoadenylate deaminase deficiency.
Main Methods:
- An indirect enzymatic equilibrium method was developed, coupling a chromogenic system with xanthine oxidase.
- The method utilizes 3,5-dichloro-2-hydroxy-benzenesulfonic acid and 4-aminophenazone to detect hydrogen peroxide formation.
- Assay parameters optimized include pH 7.8 and absorbance maximum at 510 nm.
Main Results:
- The calibration curve was linear from 0 to 100 mumol/L with a detection limit of 0.9 mumol/L.
- Analytical variability (CV) ranged from 1.5% to 3.6% within-run and 4.5% to 8.5% between-run.
- A coefficient of correlation of 0.68 was observed between plasma hypoxanthine and ammonia increases in controls.
Conclusions:
- The developed indirect enzymatic method provides a viable and efficient alternative for hypoxanthine determination.
- This assay can be performed using standard spectrophotometry or centrifugal analyzers, offering practical advantages in clinical settings.