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Related Experiment Videos

Spectrophotometric determination of ascorbic acid and dehydroascorbic acid

T Moeslinger1, M Brunner, I Volf

  • 1Institute for Medical Physiology, Vienna, Austria.

Clinical Chemistry
|August 1, 1995
PubMed
Summary
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This study introduces a rapid spectrophotometric method for quantifying ascorbic acid in human plasma. The new assay is accurate, precise, and suitable for routine clinical analysis.

Area of Science:

  • Biochemistry
  • Analytical Chemistry
  • Clinical Diagnostics

Background:

  • Accurate measurement of ascorbic acid (vitamin C) is crucial for assessing nutritional status and diagnosing related health conditions.
  • Existing methods for ascorbic acid quantification can be time-consuming or require specialized equipment, limiting their widespread clinical application.

Purpose of the Study:

  • To develop and validate a rapid, spectrophotometric method for the quantitative determination of ascorbic acid in human plasma.
  • To establish the sensitivity, linearity, accuracy, and precision of the proposed assay.

Main Methods:

  • Human plasma samples were extracted using methanol/trichloroacetic acid.
  • Ascorbic acid was enzymatically oxidized to dehydroascorbic acid using ascorbate oxidase.

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  • Dehydroascorbic acid concentration was measured spectrophotometrically by monitoring absorbance changes.
  • Ascorbic acid levels were calculated as the difference between total ascorbic acid and dehydroascorbic acid.

    • Main Results:

    • The method demonstrated a low detection limit (< 0.5 mumol/L) and excellent linearity (r > 0.995) over a wide concentration range (0-1000 mumol/L).
    • Analytical recovery of ascorbic acid added to plasma ranged from 93-105%, with between-day variance < 7%.
    • Comparison with a chromatographic method showed strong agreement (y = 1.02x - 0.653).

    Conclusions:

    • The developed spectrophotometric method provides a fast (< 10 min/sample), accurate, and reliable means for measuring ascorbic acid in human plasma.
    • This assay is suitable for simultaneous analysis of multiple samples, offering a practical alternative for clinical laboratories.
    • The method's high precision and accuracy support its use in nutritional assessment and clinical diagnostics.