Cited by 92Vitamin C and Antioxidants ResearchAntioxidant Activity and Oxidative StressClinical Laboratory Practices and Quality Control

Appropriate Handling, Processing and Analysis of Blood Samples Is Essential to Avoid Oxidation of Vitamin C to Dehydroascorbic Acid

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Pullar, Juliet, Bayer, Simone, Carr, Anitra·2018·Antioxidants

Abstract

Vitamin C (ascorbate) is the major water-soluble antioxidant in plasma and its oxidation to dehydroascorbic acid (DHA) has been proposed as a marker of oxidative stress in vivo. However, controversy exists in the literature around the amount of DHA detected in blood samples collected from various patient cohorts. In this study, we report on DHA concentrations in a selection of different clinical cohorts (diabetes, pneumonia, cancer, and critically ill). All clinical samples were collected into EDTA anticoagulant tubes and processed at 4 °C prior to storage at −80 °C for subsequent analysis by HPLC with electrochemical detection. We also investigated the effects of different handling and processing conditions on short-term and long-term ascorbate and DHA stability in vitro and in whole blood and plasma samples. These conditions included metal chelation, anticoagulants (EDTA and heparin), and processing temperatures (ice, 4 °C and room temperature). Analysis of our clinical cohorts indicated very low to negligible DHA concentrations. Samples exhibiting haemolysis contained significantly higher concentrations of DHA. Metal chelation inhibited oxidation of vitamin C in vitro, confirming the involvement of contaminating metal ions. Although EDTA is an effective metal chelator, complexes with transition metal ions are still redox active, thus its use as an anticoagulant can facilitate metal ion-dependent oxidation of vitamin C in whole blood and plasma. Handling and processing blood samples on ice (or at 4 °C) delayed oxidation of vitamin C by a number of hours. A review of the literature regarding DHA concentrations in clinical cohorts highlighted the fact that studies using colourimetric or fluorometric assays reported significantly higher concentrations of DHA compared to those using HPLC with electrochemical detection. In conclusion, careful handling and processing of samples, combined with appropriate analysis, is crucial for accurate determination of ascorbate and DHA in clinical samples.

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Crossref · 10.3390/antiox7020029

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Authors
Pullar, J., Bayer, S., & Carr, A.
Affiliations
University of Otago (NZ)
Corresponding author
Carr, Anitra
Venue
Antioxidants
Publisher
MDPI AG
Published
2018-02-11
Volume
7
Issue
2
Pages
29
ISSN
2076-3921 (ISSN-L)
OpenAlex
W2794422490
Cited by
92
References
49
Citations by year
2026: 3 · 2025: 7 · 2024: 12 · 2023: 15 · 2022: 5 · 2021: 20 · 2020: 13 · 2019: 8 · 2018: 9
FWCI
10.34
Type
journal article
Language
en
Open access
Locations
  • Antioxidants (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • Health Research Council of New Zealand (16/037)
  • Canterbury Medical Research Foundation (PRO15/06)
Topics
  • Health Sciences › Nursing › Nutrition and Dietetics › Vitamin C and Antioxidants Research
  • Health Sciences › Medicine › Biochemistry › Antioxidant Activity and Oxidative Stress
  • Health Sciences › Medicine › Physiology › Clinical Laboratory Practices and Quality Control
Keywords
vitamin C, dehydroascorbic acid, ascorbic acid oxidation, blood sample handling, EDTA anticoagulant, metal chelation, metal ion contamination, oxidative stress biomarkers, hemolysis, sample storage temperature, plasma, whole blood, diabetes mellitus, pneumonia, cancer, critically ill patients, HPLC-electrochemical detection

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