Vitamin C is marketed as a dietary supplement,
partly because of its ‘antioxidant’
properties. However, we report here that
vitamin C administered as a dietary supplement
to healthy humans exhibits a prooxidant,
as well as an antioxidant, effect in
vivo.
We conducted a study1 involving 30
healthy volunteers (16 females and 14 males
aged between 17 and 49) whose diets were
supplemented with 500 milligrams per day
of vitamin C (ascorbic acid) for 6 weeks. We
assessed the levels of oxidative damage to
peripheral blood lymphocytes in terms of
modified DNA bases. The level of 8-
oxoguanine was found to decrease on supplementation
relative to both placebo
(calcium carbonate; 500 mg per day for 6
weeks) and baseline measurements, whereas
the level of 8-oxoadenine increased.
For each volunteer, blood was collected
at 3-weekly intervals for up to 12 weeks (6
weeks on placebo, 6 weeks on vitamin C) and then another sample was taken 7
weeks after completion of the vitamin C
course (washout period). In each case,
levels of plasma ascorbate were determined,
lymphocytes were isolated and
their DNA extracted before analysing
oxidative damage.
Supplementation of diets with 500 mg
per day of vitamin C resulted in a significant
increase in ascorbate levels in the plasma
(about 60%) compared with both
pre-supplementation and placebo (data not
shown). Following the washout period, vitamin
C levels returned to the concentrations
observed at baseline and in the
placebo. In contrast, there was no change in
ascorbate concentrations during oral placebo
treatment in comparison with the baseline
value.
We used gas chromatography–mass
spectrometry (GC–MS)2–4 to assess the levels
of 8-oxoguanine and 8-oxoadenine,
which are markers for DNA damage mediated
by oxygen radicals. Supplementation
of diets with 500 mg per day of vitamin C
resulted in a significant decrease (P<0.01 by
ANOVA compared with both baseline and
placebo) in 8-oxoguanine levels (Fig. 1). No
significant differences from baseline damage
to DNA bases was observed during the
placebo treatment. Furthermore, following
the 7-week ‘washout’ period, levels of 8-
oxoguanine returned to those observed at
baseline and during placebo treatment.
In contrast, supplementation with vitamin
C resulted in a significant increase
(P<0.01 by ANOVA compared with baseline
and with placebo values) in 8-oxoadenine
levels in DNA isolated from
lymphocytes. Again, there were no significant
differences during the placebo treatment.
During the washout, the mean level
of 8-oxoadenine returned to that observed
at baseline or during placebo, such that
there was a statistically significant decrease
in 8-oxoadenine compared with the vitamin
C supplementation period (P<0.01).
There were no changes in either lymphocytes
or neutrophil counts during either
placebo or vitamin C treatments (data not
shown), confirming that the alterations
noted in 8-oxopurine levels in lymphocyte
DNA were not due to gross changes in cell
type.
We established baseline values for the 8-
oxoguanine and 8-oxoadenine lesions in
human lymphocyte DNA. Mean values
obtained for 8-oxoguanine (30 lesions per
105 guanine bases) and 8-oxoadenine (8
lesions per 105 adenine bases) were remarkably
similar to those found in other in vivo
systems5–7. These results re-emphasize that
GC–MS is a powerful technique for measuring
oxidative DNA damage as it can
simultaneously quantify more than one
modified DNA base.
Endogenous oxygen radicals are capable
of damaging cellular biomolecules such
as DNA8. Most of these species produced
in vivo are quenched by antioxidant
defences. However, a fine balance exists
that may be disrupted in favour of oxidants
(oxidative stress), giving rise to an
accumulation of biomolecular damage,
which in turn may play a role in major diseases
such as cancer, rheumatoid arthritis
and atherosclerosis9. It has been postulated
that dietary intervention with vitamin C
may reduce oxidative stress and thereby
prevent such diseases.
Although the antioxidant nature in vivo
of vitamin C has been questioned10, it is
nonetheless marketed as supplements in
doses of 500 mg or more per day as an
‘antioxidant’. Our discovery of an increase
in a potentially mutagenic lesion, 8-oxoadenine11,12,
following a typical vitamin C
supplementation should therefore be of
some concern, although at doses of less
than 500 mg per day the antioxidant effect
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