Vitamin C metabolomic mapping in experimental diabetes with 6-deoxy-6-fluoro-ascorbic acid and high resolution 19F-nuclear magnetic resonance spectroscopy.

Metabolomic mapping is an emerging discipline geared at providing information on a large number of metabolites as a complement to genomics and proteomics. Here we have probed ascorbic acid homeostasis and degradation in diabetes using 6-deoxy-6-fluoro ascorbic acid (F-ASA) and 750 MHz (19)F-nuclear magnetic resonance (NMR) spectroscopy with proton decoupling In vitro, Cu(2+)-mediated degradation of F-ASA revealed the formation of 4 major stable degradation products at 24 hours. However, when normal or diabetics rats were injected with F-ASA intraperitoneally (IP) for 4 days, up to 20 fluorine-labeled compounds were observed in the urine. Their composition resembled, in part, metal catalyzed degradation of F-ASA and was not explained by spontaneous degradation in the urine. Diabetes led to a dramatic increase in urinary F-ASA loss and a relative decrease in most other urinary F-compounds. Diabetes tilted F-ASA homeostasis toward oxidation in liver (P <.01), kidney (P <.01), spleen (P <.01), and plasma (P <.01), but tended to decrease oxidation in brain, adrenal glands, and heart. Surprisingly, however, besides the major oxidation product fluoro-dehydroascorbic acid (F-DHA), no F-ASA advanced catabolites were detected in tissues at 5 micromol/L sensitivity. These findings not only confirm the key role of the kidney in diabetes-mediated loss of ascorbic acid, but demonstrate that only selected tissues are prone to increased oxidation in diabetes. While the structure of most degradation products needs to be established, the method illustrates the power of high resolution (19)F-NMR spectroscopy for the mapping of complex metabolomic pathways in disease states.

An electrospray ionization ion trap mass spectrometric (ESI-MS-MSn) study of dehydroascorbic acid hydrolysis at neutral pH.

The hydrolysis of dehydroascorbic acid (DAAH) at neutral pH and 27 degrees C was investigated by direct infusion electrospray ionisation ion trap mass spectrometry (ESI-MS). This approach permitted derivatisation and elution procedures to be avoided, reducing to the minimum extent sample manipulation and allowing a rapid and direct observation of the species involved in the reaction. Six main peaks, related to hydrated dehydroascorbate (HyDAA-) and diketogulonate (HyDKG-) anions, were observed in the mass spectra of DAAH solutions at different times of incubation and were characterised by MSn experiments. The relevant signal intensities changed with time and a model, based on the irreversible pseudo-first order HyDAA(-)-->HyDKG- conversion, fitted successfully the data obtained for dehydroascorbate. The kinetic constant of the process was (3.2 +/- 0.5) x 10(-2) min-1. The influence of metal ion traces on the hydrolysis rate was also checked, performing experiments in the presence of EDTA, and was found to be negligible.