Ascorbate (ascorbic acid, vitamin C), in pharmacologic concentrations easily achieved in humans by i.v. administration, selectively kills some cancer cells but not normal cells. We proposed that pharmacologic ascorbate is a prodrug for preferential steady-state formation of ascorbate radical (Asc •− ) and H 2 O 2 in the extracellular space compared with blood. Here we test this hypothesis in vivo . Rats were administered parenteral (i.v. or i.p.) or oral ascorbate in typical human pharmacologic doses (≈0.25–0.5 mg per gram of body weight). After i.v. injection, ascorbate baseline concentrations of 50–100 μM in blood and extracellular fluid increased to peaks of &gt;8 mM. After i.p. injection, peaks approached 3 mM in both fluids. By gavage, the same doses produced ascorbate concentrations of &lt;150 μM in both fluids. In blood, Asc •− concentrations measured by EPR were undetectable with oral administration and always &lt;50 nM with parenteral administration, even when corresponding ascorbate concentrations were &gt;8 mM. After parenteral dosing, Asc •− concentrations in extracellular fluid were 4- to 12-fold higher than those in blood, were as high as 250 nM, and were a function of ascorbate concentrations. By using the synthesized probe peroxyxanthone, H 2 O 2 in extracellular fluid was detected only after parenteral administration of ascorbate and when Asc •− concentrations in extracellular fluid exceeded 100 nM. The data show that pharmacologic ascorbate is a prodrug for preferential steady-state formation of Asc •− and H 2 O 2 in the extracellular space but not blood. These data provide a foundation for pursuing pharmacologic ascorbate as a prooxidant therapeutic agent in cancer and infections.

Ascorbate in pharmacologic concentrations selectively generates ascorbate radical and hydrogen peroxide in extracellular fluid in vivo