25th Anniversary Linus Pauling Institute at Oregon State University banner


Does Vitamin C Cause Genetic Damage?

Balz Frei, Ph.D.
Director, Linus Pauling Institute
Professor of Biochemistry and Biophysics

photo of Balz Frei

In a study published in April in the respected scientific journal Nature, Ian Podmore and colleagues in England claimed that supplementation with 500 milligrams (mg) per day of vitamin C causes genetic damage in humans and, therefore, may cause cancer. The study involved 30 healthy volunteers who were given a placebo for six weeks, followed by 500 mg of vitamin C per day for another six weeks, and finally neither placebo nor vitamin C for an additional seven weeks ("washout period"). Blood was taken from the volunteers at the beginning of the study, at three week intervals during the placebo and vitamin C supplementation periods, and again at the end of the study. White blood cells called lymphocytes were extracted from the blood samples and analyzed for two different types of oxidative DNA damage: 8-oxoadenine and 8-oxoguanine. During the placebo phase, no changes occurred in either 8-oxoadenine or 8-oxoguanine levels. However, with vitamin C supplementation, 8-oxoadenine levels increased, indicative of increased oxidative DNA damage, while 8-oxoguanine levels decreased, indicative of decreased oxidative DNA damage. At the end of the washout period, both 8-oxoadenine and 8-oxoguanine returned to levels observed before vitamin C supplementation (see graph on next page). The authors concluded that vitamin C at a daily dose of 500 mg "exhibits pro-oxidant properties" in humans that cause genetic damage, "although at doses of less than 500 mg per day the antioxidant effect [of vitamin C] may predominate."

The data from the study, however, raise more questions then they answer. Which is more important: vitamin C's beneficial, antioxidant effect of lowering 8-oxoguanine levels, or the adverse, pro-oxidant effect of increasing 8-oxoadenine levels? Does supplemental vitamin C really influence these levels? What are the biological consequences of decreased 8-oxoguanine and increased 8-oxoadenine levels? A highly respected group of scientists at the Massachusetts Institute of Technology has shown that 8-oxoguanine is at least one order of magnitude more mutagenic than 8-oxoadenine, which means that 8-oxoguanine is at least ten times more likely than 8-oxoadenine to cause genetic damage that could lead to cancer. Therefore, the Nature study indicates that vitamin C supplementation is actually beneficial, because the benefits of decreased 8-oxoguanine levels outweigh any adverse effects of increased 8-oxoadenine levels.

However, the study is seriously flawed in several crucial aspects. First, the study design is very poor and, in fact, unacceptable by current standards used by the U.S. Food and Drug Administration to evaluate the efficacy and safety of a drug. The study was neither randomized nor double-blinded; the volunteers and the scientists conducting the study knew what supplements were being used at what time. This can affect the volunteers' compliance and possibly bias interpretation of the data by the scientists. Furthermore, despite the authors' claims, the study is not truly placebo-controlled. During the vitamin C supplementation period, there was no control group receiving a placebo. Let's assume, for example, that the assay used for measuring 8-oxoguanine and 8-oxoadenine shows some variability from one batch of samples to the next. How can we determine whether an increase or decrease in 8-oxoguanine and 8-oxoadenine levels is due to the variability of the assay or truly reflects changes occurring in the volunteers? A placebo group must be included simultaneously with the vitamin C supplemented group. When the samples are analyzed, there should be a difference between the two groups if vitamin C produces an effect. A placebo group is also required for comparison with the supplemented group to account for any significant changes in the diet or the environment (e.g. exposure to cigarette smoke or sunlight) of the volunteers over the 19 week study period that may result in changes in 8-oxoguanine and 8-oxoadenine levels unrelated to vitamin C supplementation. Such changes can only be detected in a group given a placebo over the entire study period, and this was not done.

More problems with the study

The authors used a method called "gas chromatography-mass spectrometry" to measure 8-oxoguanine and 8-oxoadenine levels. This method is known to cause ex vivo artifacts, meaning that 8-oxoguanine and 8-oxoadenine levels increase many-fold during the sample preparation and analytical procedures after the blood is taken from the subjects. Consequently, the authors report levels for 8-oxoguanine that are much higher than those currently considered the actual values for 8-oxoguanine in human lymphocytes obtained by more accurate methodologies. Podmore and colleagues report "30 [8-oxoguanine] lesions per 105 guanine bases [of DNA]", whereas the majority of the experts in the field report values in the range of 0.4 to 4 8-oxoguanine lesions per 105 guanine bases of DNA. In fact, at a recent scientific conference chaired by LPI faculty member Maret Traber and myself in Colorado, Dr. Andrew Collins from the DNA Instability Group of the Rowett Research Institute in Aberdeen, U.K., reported levels as low as 0.04 8-oxoguanine lesions per 105 guanine bases, using a technology that eliminates ex vivo artifacts. That's a whopping 750-fold difference from what was reported in the Nature study! What does this mean? In all likelihood, 99.9% of the 8-oxoguanine levels measured by Podmore and colleagues were not actually formed in the volunteers, but instead in the samples after they were collected from the volunteers. Much less is known about the measurement of 8-oxoadenine by different methods, but it is likely that these levels, too, were artificially and massively elevated by ex vivo oxidation. Although there is no explanation for the movement of 8-oxoguanine and 8-oxoadenine levels in opposite directions during the vitamin C supplementation period (see graph), this question becomes moot in light of the existence of artifactual oxidation. My Ph.D. advisor constantly reminded me "not to waste clean thinking on dirty data", and these data are a prime example. The authors failed to take the necessary precautions to prevent ex vivo artifacts in their study, and as a result their data are uninterpretable.

Modified and reprinted by permission from Nature 392:599, 1998©
1998 Macmillan Magazines Ltd.

Even more problems!

Another important objection has been raised by Dr. Mark A. Levine, head of the Molecular and Clinical Nutrition Section of the National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health. Dr. Levine, one of the world's experts on vitamin C metabolism, noticed that the plasma vitamin C levels of the 30 healthy volunteers in the Podmore study were 50 ÁM at the beginning of the study before any supplemental vitamin C was given. This plasma level has been shown to be associated with maximal vitamin C levels attainable in lymphocytes (published in the Proceedings of the National Academy of Sciences in 1996). Therefore, supplementation with 500 mg per day of vitamin C could not have further increased lymphocyte vitamin C levels. How, then, could vitamin C have influenced oxidative DNA damage in these cells, if vitamin C levels remained unchanged? Unfortunately, the authors of the Nature study did not measure lymphocyte vitamin C levels in their volunteers.

The study by Podmore and colleagues and their conclusion that vitamin C is harmful are compromised by unproved assumptions, an unacceptable study design, and highly inaccurate measurements. The data presented do not advance our understanding of vitamin C's anti- or pro-oxidant function, and the authors' warning about vitamin C supplements is unwarranted. Hundreds of scientific studies strongly suggest that vitamin C helps reduce the risk of cancer and has important health benefits beyond preventing scurvy. Additional studies are likely to find more benefits. It would be tragic if people stopped taking vitamin C supplements, or decreased their intake of fruits and vegetables, based on this single, flawed study.

For information on vitamin C and health, see the Linus Pauling Institute's Micronutrient Information Center.

Last updated November, 1998

Honoring a Scientific Giant with Research Toward Longer, Better Lives

Return to Fall/Winter 1998 Table of Contents Return to LPI Home Page Please send any comments, suggestions, or questions about The Linus Pauling Institute to lpi@oregonstate.edu