ABSTRACT
BACKGROUND: Low dose X-irradiation (IR) from computer tomography (CT) can generate free radicals, which can damage biologically relevant molecules and ultimately lead to cancer. These effects are especially concerning for children owing to their higher radiosensitivity and longer life expectancy than adults. The lipid phase micronutrients (LPM) coenzyme Q10, carotenoids, E vitamers, and vitamin A are potent radical scavengers that can act as intracellular antioxidants. METHODS: We investigated changes in circulating levels of these LPM in 17 children (0.25-6 y) undergoing medically indicated CT scans involving relatively low IR doses. Blood was drawn before and 1h after CT scans and analyzed using HPLC with electrochemical and UV/VIS detection. RESULTS: We found significant decreases (p<0.05) in post-CT plasma levels in several LPM which suggests that these LPM can serve as biodosimeters and may protect against damage from IR during clinical procedures such as CT. The strongest predictors for pre- to post-CT changes for many LPM were their baseline levels. CONCLUSION: Future larger studies are warranted to confirm our findings and to test whether high circulating antioxidant levels protect against IR damage in vivo with an ultimate goal of establishing prophylactic modalities for CT-induced IR damage.
Subject(s)
Carotenoids/blood , Tocopherols/blood , Tomography, X-Ray Computed/adverse effects , Ubiquinone/analogs & derivatives , Vitamin A/blood , Vitamins/blood , Child , Child, Preschool , Female , Humans , Infant , Male , Ubiquinone/bloodABSTRACT
Circulating lipid-phase micronutrients (LPM) such as 25-hydroxylated D vitamers, retinol, tocopherols, carotenoids including their isomers, and coenzyme Q10 play important roles in health maintenance and disease prevention and can serve as useful biomarkers. We developed fast, affordable, and accurate HPLC assays that simultaneously measured all above LPM in a single run using UV/VIS detection at 265nm, 295nm, and 480nm with (1) a C18 column alone; (2) a C30 column alone; or (3) each of these columns connected in series. The C18 column alone could separate all major LPM of interest in less than 17min but insufficiently resolved the lycopene isomers, the 25-hydroxylated D vitamers, lutein from zeaxanthin and ß- from γ-tocopherol. The C30 column alone separated all LPM of interest including many isomeric analytes but failed to resolve the Q10 compounds, which co-eluted with carotenoids. Connecting the C18 and C30 columns in series with a detector after the C30 column and a pressure resistant detector between the columns resulted in ideal resolution and accurate quantitation of all LPM of interest but required software capable of processing the acquired data from both detectors. Connecting the C18 and C30 columns in series with exclusively one detector after the C30 column resulted in carotenoid-Q10 interferences, however, this was remedied by heart-cutting 2D-LC with a 6-port valve between the columns, which resolved all analytes in 42min. Faster run times led to some analytes not being resolved. Many variations of these methods are possible to meet the needs of individual requirements while minimizing sample material and turn-around-times.
Subject(s)
Carotenoids/blood , Cholecalciferol/blood , Chromatography, High Pressure Liquid/methods , Tocopherols/blood , Ubiquinone/analogs & derivatives , Vitamin A/blood , Adult , Carotenoids/chemistry , Carotenoids/isolation & purification , Cholecalciferol/chemistry , Cholecalciferol/isolation & purification , Chromatography, High Pressure Liquid/instrumentation , Female , Humans , Limit of Detection , Male , Micronutrients/blood , Micronutrients/chemistry , Micronutrients/isolation & purification , Reproducibility of Results , Spectrophotometry, Ultraviolet , Tocopherols/chemistry , Tocopherols/isolation & purification , Ubiquinone/blood , Ubiquinone/chemistry , Ubiquinone/isolation & purification , Vitamin A/chemistry , Vitamin A/isolation & purificationABSTRACT
Coenzyme Q10 (Q10) is present in the circulation mainly in its reduced form (ubiquinol-10; UL10), but oxidizes quickly ex vivo to ubiquinone-10 (UN10). Therefore, native UL10:UN10 ratios, used as markers of redox status and disease risk, are difficult to measure. We established an RP-(U)HPLC method with coulometric detection to measure natively circulating UL10 and UN10 concentrations by adding a ubiquinol/ubiquinone mixture as an internal standard immediately after plasma preparation. This allowed adjustment for unavoidable artificial UL10 oxidation as well as for total losses (or gains) of analytes during sample storage, processing, and analysis because the internal standards exactly paralleled the chemical behavior of Q10. This technique applied to blood (n = 13) revealed Q10 levels of 680-3300 nM with a mean UL10:UN10 ratio of 95:5, which was inversely associated with total Q10 (r=-0.69; p=0.004). The oxidation of UL10 to UN10 was equimolar, increased by O(2), and decreased by lower temperatures or various degassing methods. Although UL10 was stable in blood or when pure in organic solvents at 22 degrees C, its oxidation was catalyzed dose dependently by alpha-tocopherol and butylated hydroxytoluene, particularly when present in combination. Key structural features for the catalytic pro-oxidant properties of phenolic antioxidants included two substituents vicinal to the phenolic hydroxyl group.
