Direct clean-up and analysis of ribonucleosides in physiological fluids.

Area of Science:

• Biochemistry
• Analytical Chemistry
• Molecular Biology

Background:

• Ribonucleosides are crucial molecules involved in various cellular processes.
• Quantifying modified and unmodified ribonucleosides in physiological fluids is challenging.
• Understanding ribonucleoside metabolism is vital for diagnosing and treating diseases.

Purpose of the Study:

• To develop and validate a novel method for the selective separation and quantification of ribonucleosides.
• To analyze the excretion patterns of various ribonucleosides in human urine and serum.
• To explore the potential applications of this method in clinical diagnostics and research.

Main Methods:

• On-line multidimensional high-performance affinity chromatography (HPAC) coupled with reversed-phase liquid chromatography (RPLC).
• Group-selective separation of unmodified, modified, and hypermodified ribonucleosides.
• Quantification of specific ribonucleosides including N1-methyladenosine, N1-methylinosine, N2-methylguanosine, N2-dimethylguanosine, N6-carbamoylthreonyladenosine, and 2-pyridone-5-carboxamido-N-ribofuranoside.

Main Results:

• The HPAC-RPLC method successfully separated and quantified various ribonucleosides in urine and serum.
• Excretion levels and patterns of key ribonucleosides were determined in control subjects and patients with different diseases.
• The study established reference ranges and identified potential disease-specific alterations in ribonucleoside profiles.

Conclusions:

• The developed HPAC-RPLC method is a powerful tool for analyzing ribonucleosides in physiological fluids.
• This technique offers potential as a non-invasive screening test for diseases related to ribonucleoside metabolism.
• Applications include monitoring nucleoside chemotherapy and studying renal ribonucleoside reutilization.