Improved method for simultaneous determination of ascorbic acid and dehydroascorbic acid, isoascorbic acid and...

Area of Science:

• Analytical Chemistry
• Biochemistry
• Food Science

Background:

• Vitamin C, comprising ascorbic acid and dehydroascorbic acid, is a crucial nutrient with antioxidant properties.
• Accurate quantification of both forms is essential for nutritional assessment and health studies.
• Existing methods may lack the sensitivity or specificity to differentiate and quantify both active forms simultaneously.

Purpose of the Study:

• To develop and validate a rapid, dual-detection High-Performance Liquid Chromatography (HPLC) method for the simultaneous determination of ascorbic acid and dehydroascorbic acid.
• To assess the method's accuracy and reliability in various food and plasma matrices.

Main Methods:

• HPLC separation coupled with a dual detection system: direct detection of ascorbic acid and indirect fluorimetric detection of dehydroascorbic acid post-derivatization.
• Post-column derivatization using O-phenyldiamine for enhanced dehydroascorbic acid detection.
• Analysis of diverse food samples (fruits, vegetables) and plasma.

Main Results:

• Simultaneous separation of ascorbic acid, dehydroascorbic acid, and their D-isomers within 10 minutes.
• Excellent repeatability for vitamin C quantification in fruits and vegetables (0.3–1.9% RSD).
• Good reproducibility across different matrices (1.9–3.6% RSD for vitamin C; 2.4–5.8% RSD for individual forms in reference materials).

Conclusions:

• The developed HPLC method provides a reliable and efficient approach for total vitamin C analysis.
• The method's accuracy and precision make it suitable for routine analysis in food and clinical settings.
• This technique facilitates comprehensive nutritional evaluation by quantifying both active forms of vitamin C.