Cutaneous Vitamin D Synthesis in Carnivorous Species

The aim of this study was to investigate the differences of the ability to synthesize sufficient amounts of vitamin D in the skin of different carnivorous species. To this endskin tissue of 22 different carnivorous species were collected from dead animals from zoo’s and our pathology department. Wis tar rat skin served as a positive control.Cholesterol, 7-DHC, and vitamin D content was determined after UVB exposure at 37°C, and compared to non-irradiated skin. Overall, there was a significant effect of species and skin thickness, but not of UVB irradiation, on 7-DHC and vitamin D concentrations of the skin.The relatively low cutaneous levels of the vitamin D precursor 7-DHC observed in this study suggest that most terrestrial carnivores are unable to synthesize sufficient amounts of vitamin D. The results have to be taken into account when preparing food for these species when held under captive conditions. Cite this article: Corbee RJ, Vaandrager AB, Kik MJL, MolenaarMR, Hazewinkel HAW (2015) Cutaneous Vitamin D Synthesis in Carnivorous Species. J Vet Med Res 2(4): 1031. Central Corbee (2015) Email: J Vet Med Res 2(4): 1031 (2015) 2/4 mL millipure water and 3 mL chloroform:methanol 1/2, v/v, were added to the tubes and were mixed for 40 min by regular vortexing. Then the tubes were centrifuged for 2 min at 2000 rpm. The supernatant was transferred to new glass tubes and 2 mL millipure water and 2 mL chloroform were added. After vortexing for 30 sec, the tubes were centrifuged for 5 min at 2000 rpm. The lower layer was transferred to a new tube and evaporated under N2 gas. These samples were stored at -20°C prior to MS-analysis. Before MS-analysis, the samples were resuspended in 500 μL chloroform:methanol 1/1, v/v containing 0.002% butylated hydroxytoluene (BHT) as an anti-oxidant, and 20 μL was injected on a Lichrospher RP18-e column. A gradient was generated from acetonitrile:water 95/5, v/v, to acetone/ chloroform 85/15, v/v, at a constant flow rate of 1 mL/min. Total run time per sample was 13 min. MS of lipids was performed using Atmospheric Pressure Chemical Ionization (APCI) on a Biosystems API-4000 Q-trap (MDS Sciex, Concord, ON, Canada). The system was controlled by Analyst version 1.4.2 software (MDS Sciex, Concord, ON, Canada) and operated in positive ion mode and in the multiple reaction monitoring (MRM) mode using the following settings: source temperature 420°C, nebulizer gas (GS1) 5, nebulizer current 3 μA, curtain gas 10, collision gas. High and declustering potential and collision energy were empirically optimized for each compound. The MRM transitions (m/z) used were: 367.3 → 159.1 for 7-DHC, desmosterol and vitamin D (species were identified with regard to retention times 7.2, 7.5, and 6.45 min, respectively), and 369.3 → 287.3 for cholesterol. Data analysis was performed using Analyst 1.4.2 software (MDS Sciex, Concord, ON, Canada). Quantitation was done relative to standards run separately (all steroid standards were from Sigma-Aldrich (St. Louis, MO, USA)). As cholesterol is expected to be extracted with similar efficiency as vitamin D and 7-DHC, and cholesterol is a good indicator of the amount of cellular material present in the skins, the data are expressed as a ratio to cholesterol. All skin samples were analysed in duplicate. In case of high variations (i.e. >20%) the skin samples were analysed for another two times. The average levels of two samples are demonstrated.