A simple quantitative assay for urinary adenosine using column-switching high-performance liquid chromatography

Area of Science:

• Biochemistry
• Analytical Chemistry
• Physiology

Background:

• Adenosine is a crucial local regulator of diverse cellular functions.
• Measuring adenosine levels in biological fluids aids in understanding its physiological and pathophysiological roles.
• Accurate quantification methods are essential for reliable adenosine level assessment.

Purpose of the Study:

• To develop and validate a precise, high-performance liquid chromatography (HPLC) method for quantifying urinary adenosine.
• To establish age-related reference values for urinary adenosine excretion in healthy individuals.
• To investigate the association between urinary adenosine levels and specific pathophysiological conditions.

Main Methods:

• Development of an automated HPLC method for urinary adenosine analysis.
• Method validation including linearity, recovery, and precision (intra- and inter-assay coefficients of variation).
• Analysis of urinary adenosine in healthy infants, children, adults, and patients with metabolic disease or respiratory failure.

Main Results:

• A rapid (35 min) and accurate HPLC method for urinary adenosine was established, requiring minimal sample preparation.
• Significant differences in urinary adenosine excretion were observed across age groups, with higher levels in infants and children compared to adults.
• Elevated urinary adenosine levels were detected in patients with metabolic disease and severe respiratory failure.

Conclusions:

• The developed HPLC method provides a reliable tool for urinary adenosine quantification.
• Urinary adenosine excretion varies significantly with age, with higher levels in younger individuals.
• Increased urinary adenosine is associated with adverse pathophysiological conditions, highlighting its potential as a biomarker.