Vitamin C – or ascorbic acid – exists in the circulation partially bound to albumin. Roles of ascorbic acid include collagen synthesis, carnitine synthesis, bile salt production, haemostasis, hormone synthesis, optimal leukocyte function, and enhancing iron absorption. Humans receive vitamin C from diet (fruit and vegetables) and supplements. Ascorbic acid is one of the most effective water soluble antioxidants in biological fluids preventing free radical mediated oxidative damage to biological macromolecules including DNA, lipids and proteins. A second important role includes regenerating other antioxidants including the lipid soluble antioxidant α-tocopherol (vitamin E). There is epidemiological evidence that a healthy intake of fruit and vegetables provides enough vitamin C to protect against stroke, cardiovascular disease, and most forms of cancer.
Deficiency and Toxicity
Frank vitamin C deficiency, which leads to the classical disease of scurvy, is rare in developed countries, although suboptimal vitamin C status is common. However, with abnormal diets it may occur in the very young and very old. Other at-risk population groups include alcoholics, smokers, cancer patients, and renal dialysis patients.
Early symptoms of deficiency are of fatigue followed by easy bruising, bleeding, hair loss, joint pain and swelling.
Vitamin C is essentially non-toxic with the possible exception of high-dose supplementation. Rare adverse effects include uricosuria, oxaluria and kidney stone formation, excess iron absorption and pro-oxidant effects in the presence of transition metals such as free copper or iron.
Assessment of status
The nutritional status of vitamin C can be assessed by its measurement in plasma. It is important to note that concentrations are influenced by recent dietary intake and particularly by the systemic inflammatory response which lowers levels substantially independent of its nutritional status. After elective surgery for knee arthroplasty, plasma ascorbic acid concentration was significantly and transiently reduced by 74% during the evolution of the systemic inflammatory response within 48 h of injury (see Table 1).81 This reduction is dependent on the magnitude of the systemic inflammatory response (Table 2).28 Plasma ascorbic acid concentrations, when adjusted for albumin, remained significantly lower. Therefore plasma ascorbic acid concentration, whether or not adjusted for binding and redistribution, is unlikely to be a reliable measurement of vitamin C status where there is evidence of a systemic inflammatory response (CRP ≥5 mg/L).
Recommended Daily Allowance
Adults: 90 mg/day.
Effect of Systemic Inflammatory Response on Plasma Vitamin C Concentrations
Table 1: Baseline, peak/trough and day 90 concentrations of CRP, vitamin C (ascorbic acid) and albumin following elective surgery for hip arthroplasty (n = 11).81 Median (range).
|CRP (mg/L)||<6 (<6–<6)||169 (92-273)||<6 (<6–<6)||<0.001|
|Plasma ascorbic acid (µmol/L)||61 (23-127))||16 (9-47)||44 (9-97)||<0.001|
|45 (39-48)||32 (25-37)||43 (34-45)||<0.001|
|Ascorbic acid/albumin||1.36 (0.52-2.82)||0.52 (0.28-1.66)||1.10 (0.21-2.16)||<0.001|
Table 2: Distribution of median plasma vitamin C concentrations according to increments of CRP concentrations (n = 516).28
Median plasma vitamin C concentration
Sample Requirements and Reference Ranges for Vitamin C
|Sample Type||Plasma (fasting sample preferred*)|
|Container||Lithium heparin (non-gel) or EDTA. Lithium heparin gel tubes unsuitable.|
|Precautions||Vitamin C is very unstable82 and so samples require to be treated within 4 hours of collection (See below for details).|
|Minimum volume||250 µL** stabilised plasma|
15 to 90 µmol/L
< 10 µmol/L (at risk of deficiency)
|Mean turnaround time||3.5 days|
|Method||HPLC with electrochemical detection (In-house)82|
|Traceability||This assay is not currently traceable.|
|Intermediate Precision (CV)||14.6% at 12.8 μmol/L, 10.6% at 93.5 μmol/L|
|Measurement Uncertainty, U||12.8 ± 3.7 μmol/L, 93.5 ± 19.9 μmol/L|
|Analytical Goals (CV)||Acceptable 15%, Desirable 10%***|
|EQA Scheme||No EQA scheme available.|
* Ideally a fasting sample should be collected, especially if the patient is receiving oral or parenteral vitamin C supplementation. If this is not possible, sample should be taken at least 8 hours post treatment for patients receiving oral supplementation or TPN.
** Absolute minimum volume; this volume is insufficient to carry out repeat analysis if analysis fails.
*** Goal origin: biological variation.71
Preparation of Samples for Vitamin C Analysis
Vitamin C is very unstable82 and so samples require to be treated within 4 hours of collection.
If the sample cannot be delivered to Glasgow Royal Infirmary Clinical Biochemistry department (in working hours) within four hours of collection, add equal volumes of plasma and 6% metaphosphoric acid (VWR catalogue no. 291904A or similar product). Mix for 30 seconds.
Send to Glasgow Royal Infirmary within 24 hours (no ice required).
If delivery to Glasgow is outwith 24 hours, freeze the treated plasma at -70°C (for 4 days maximum) or -20°C (for 1 day maximum) until sending and then send by first class post (no ice required).
Please note 6% metaphosphoric acid should be stored at 4ºC and remains stable for 3 weeks following preparation.