Vitamin E (α-tocopherol) is required to prevent fetal resorption in rodents. To study α–tocopherol’s role in fetal development, a non-placental model is required. Therefore, the zebrafish, an established developmental model organism, was studied by feeding the fish a defined diet with or without added α–tocopherol. Zebrafish (age: 4–6 w) were fed the deficient (E-), sufficient (E+), or lab diet up to 1 y. All groups showed similar growth rates. The exponential rate of α–tocopherol depletion up to ~80 day in E- zebrafish was 0.029 ± 0.006 nmol/g, equivalent to a depletion half-life of 25 ± 5 days. From age ~80 d, the E- fish (5 ± 3 nmol/g) contained ~50 times less α–tocopherol than the E+ or lab diet fish (369 ± 131 or 362 ± 107, respectively, P<0.05). E-depleted adults demonstrated decreased startle response suggesting neurologic deficits. Expression of selected oxidative stress and apoptosis genes from livers isolated from the zebrafish fed the three diets were evaluated by quantitative polymerase chain reaction and were not found to vary with vitamin E status. When E-depleted adults were spawned, they produced viable embryos with depleted α–tocopherol concentrations. The E- embryos exhibited a higher mortality (P<0.05) at 24 h post fertilization (hpf) and a higher combination of malformations and mortality (P<0.05) at 120 hpf than embryos from parents fed E+ or lab diets. This study documents for the first time that vitamin E is essential for normal zebrafish embryonic development.

Background:We investigated the effects of vitamin C status on the metabolome of adult zebrafish. Results: Levels of inosine monophosphate (IMP) and AMP deaminase (AMPD) activity were enhanced in vitamin C-deficient zebrafish. Conclusion: Vitamin C deficiency activates the purine nucleotide cycle in zebrafish. Significance: The link between vitamin C deficiency and elevated AMPD activity is relevant to metabolic diseases.

Thus, urinary α-CEHC is a valid biomarker of α-tocopherol status that can be used to set a value for the Estimated Adequate Requirement of vitamin E.

To test the hypothesis that embryogenesis depends upon α-tocopherol (E) to protect embryo polyunsaturated fatty acids (PUFAs) from lipid peroxidation, new methodologies were applied to measure α-tocopherol and fatty acids in extracts from saponified zebrafish embryos. A solid phase extraction method was developed to separate the analyte classes, using a mixed mode cartridge (reverse phase, π–π bonding, strong anion exchange), then α-tocopherol and cholesterol were measured using standard techniques, while the fatty acids were quantitated using a novel, reverse phase liquid chromatography–mass spectrometry (LC–MS) approach. We also determined if α-tocopherol status alters embryonic lipid peroxidation products by analyzing 24 different oxidized products of arachidonic or docosahexaenoic (DHA) acids in embryos using LC with hybrid quadrupole-time of flight MS. Adult zebrafish were fed E− or E+ diets for 4 months, and then were spawned to obtain E− and E+ embryos. Between 24 and 72 hours post-fertilization (hpf), arachidonic acid decreased 3-times faster in E− (21 pg/h) compared with E+ embryos (7 pg/h, P<0.0001), while both α-tocopherol and DHA concentrations decreased only in E− embryos. At 36 hpf, E− embryos contained double the 5-hydroxy-eicosatetraenoic acids and 7-hydroxy-DHA concentrations, while other hydroxy-lipids remained unchanged. Vitamin E deficiency during embryogenesis depleted DHA and arachidonic acid, and increased hydroxy-fatty acids derived from these PUFA, suggesting that α-tocopherol is necessary to protect these critical fatty acids.

α-Tocopherol is a required, lipid-soluble antioxidant that protects PUFA. We hypothesized that α-tocopherol deficiency in zebrafish compromises PUFA status. Zebrafish were fed for 1 y either an α-tocopherol-sufficient (E+; 500 mg α-tocopherol/kg) or -deficient (E-; 1.1 mg α-tocopherol/kg) diet containing α-linolenic (ALA) and linoleic (LA) acids but without arachidonic acid (ARA), EPA, or DHA. Vitamin E deficiency in zebrafish decreased by ~20% (n-6) (P < 0.05) and (n-3) (P < 0.05) PUFA and increased the (n-6):(n-3) PUFA ratio (P < 0.05). In E- compared to E+ females, long chain-PUFA status was impaired, as assessed by a ~60% lower DHA:ALA ratio (P < 0.05) and a ~50% lower ARA:LA ratio (P < 0.05). fads2 (P < 0.05) and elovl2 (P < 0.05) mRNA expression was doubled in E- compared to E+ fish. Thus, inadequate vitamin E status led to a depletion of PUFA that may be a result of either or both increased lipid peroxidation and an impaired ability to synthesize sufficient PUFA, especially (n-3) PUFA.