Purpose. The effects of ascorbic acid on Stratum corneum lipid models following ultraviolet irradiation were studied adding iron ions as transition metal catalysts.
  Methods. Lipid peroxidation was quantified by the thiobarbituric acid assay. The qualitative changes were studied on a molecular level by mass spectrometry. To elucidate the nature of free radical involvement we carried out electron paramagnetic resonance studies. The influence of ascorbic acid on the concentration of hydroxyl radicals was examined using the spin trapping technique. Moreover, we checked the vitamin's ability to react with stable radicals.
  Results. Ascorbic acid was found to have prooxidative effects in all lipid systems in a concentration dependent manner. The degradation products of ascorbic acid after its prooxidative action were detected. The concentration of the hydroxyl radicals in the Fenton assay was decreased by ascorbic acid. The quantification assay of 2,2-diphenyl-1-picrylhydracyl hydrate showed reduced concentration levels of the stable radical caused by ascorbic acid.
  Conclusions. Considering human skin and its constant exposure to UV light and oxygen, an increased pool of iron ions in irradiated skin and the depletion of co-antioxidants, the administration of ascorbic acid in cosmetical formulations or in sunscreens could unfold adverse effects among the Stratum corneum lipids.

KEY WORDS: ascorbic acid; thiobarbituric acid assay; mass spectrometry; EPR; oxidative stress.

Introduction

  Ascorbic acid is a hydrophilic hexuronic acid lacton. Because humans have lost the ability to synthesize ascorbic acid, it must be provided as an essential micronutrient from food.
  The most well-known biochemical function of ascorbic acid is the protection of the prolyl and lysyl hydroxylases. This seems to be essential for the synthesis of stable collagen molecules. Furthermore, ascorbic acid has been shown to be a neuromodulator, an antiviral substance and immunostimulant, and a radical scavenger.
  Most of the chemical and biochemical properties of ascorbic acid are related to its oxoenediol structure and can be explained by its participation in biochemical redox processes. Ascorbic acid is a diabasic acid and in aqueous solution a strong reducing compound.
  Despite the high number of publications dealing with the vitamin, and the relatively simple chemical structure of the molecule, other properties of the molecule are not yet fully understood. Conflicting data has lead to controversy in the literature, and at present even the daily human intake is still under debate. Further sources of controversy are the interactions of vitamin C with reactive oxygen species (ROS), as both antioxidant and prooxidant properties have been reported. Recently, genotoxic effects were suggested to be a result of the ability of ascorbic acid to decompose lipid hydroperoxides to DNA damaging secondary products raising the question why add ascorbic acid to drug formulation for topical application.
  In the skin, ascorbic acid is part of the antioxidative network of the Stratum corneum and therefore, along with lipophilic antioxidants, proteins, and other biomolecules, a target to the oxidative stressors of the environment. A decrease in the concentration of ascorbic acid was measured in murine Stratum corneum following ozone exposure.
  In this paper, therefore, the mechanism of the redox behavior of ascorbic acid is studied using Stratum corneum lipid model systems to test its actions on exposure to UV light. As the biggest organ of the human body, the skin is constantly exposed to both ultraviolet radiation and oxygen. The degradation of ascorbic acid caused by UV-light was studied at a molecular level using electrospray ionization mass spectrometry (ESI-MS). The advantages of the soft electrospray ionization (ESI) avoiding early fragmentation and the ion trap possibilities of full scan mass spectrometry (MS), MA/MS, and multiple state MS experiments allowed the identification of the ascorbic acid decomposition products as well as the lipid peroxidation products.
  Electron paramagnetic resonance (EPR) spectroscopy is being used more frequently in pharmaceutical research because of the unique information which can be obtained by EPR spectrums of both in vivo and in vitro experiments. We used the spin trapping method by 5,5-dimethyl-1-pyrroline-1-oxide (DMPO) to detect the highly reactive hydroxyl radicals. The influence of vitamin C on the concentration of these free radicals generated by the Fenton system, asa well as the vitamin's properties regarding the stable 2,2-diphenyl-1-picrylhydrazyl hydrate (DPPH) radical, were studied.