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A specific method for determination of total ascorbic acids in urine by the alpha,alpha'- dipyridyl method
Clinica Chimica Acta; International Journal of Clinical Chemistry
|September 1, 1981
Summary
This study details a method for measuring ascorbic acid in urine using the alpha,alpha-dipyridyl technique. The optimized procedure effectively removes interfering substances and accurately quantifies vitamin C levels in urine samples.
Area of Science:
- Biochemistry
- Analytical Chemistry
- Clinical Chemistry
Background:
- Accurate determination of ascorbic acid (vitamin C) in biological samples is crucial for assessing nutritional status and health.
- Existing methods for urine ascorbic acid analysis may be affected by interfering substances.
- The alpha,alpha'-dipyridyl method offers a potential approach for sensitive ascorbic acid quantification.
Purpose of the Study:
- To describe and validate the application of the alpha,alpha'-dipyridyl method for the determination of ascorbic acid in human urine.
- To optimize the method for removing interfering substances commonly found in urine.
- To ensure accurate and reliable quantification of ascorbic acid in urine samples.
Main Methods:
- Urine samples were acidified with trichloroacetic acid and treated with activated carbon to remove interfering compounds.
- Dehydroascorbic acid was reduced to ascorbic acid using dithiothreitol.
- Excess dithiothreitol was removed with N-ethylmaleimide.
- Ascorbic acid was quantified by measuring the reduction of ferric ions to ferrous ions, which complex with alpha,alpha'-dipyridyl in the presence of phosphoric acid.
- Interference from endogenous ferrous ions was mitigated using sodium hydrogen phosphate and phosphoric acid.
Main Results:
- The described method effectively removes common interfering substances from urine samples.
- The reduction of dehydroascorbic acid to ascorbic acid was achieved using dithiothreitol.
- The alpha,alpha'-dipyridyl complex formation allowed for sensitive detection of ascorbic acid.
- Interference from endogenous ferrous ions was successfully addressed, ensuring accurate measurements.
Conclusions:
- The alpha,alpha'-dipyridyl method, with modifications, is suitable for the accurate determination of ascorbic acid in urine.
- The described sample preparation and interference removal steps enhance the reliability of the assay.
- This method provides a valuable tool for clinical and research settings requiring urine vitamin C level assessment.